Fermentations

J Biol Chem, 1999 Dec 10, 274(50), 35950 - 4
Amplification of signaling activity of the arc two-component system of Escherichia coli by anaerobic metabolites . An in vitro study with different protein modules; Georgellis D et al.; In Escherichia coli, changes in redox condition of growth are sensed and signaled by the Arc two-component system . This system consists of ArcB as the membrane-associated sensor kinase and ArcA as the cytoplasmic response regulator . ArcB is a tripartite kinase, possessing a primary transmitter, a receiver, and a secondary transmitter domain that catalyzes the phosphorylation of ArcA via a His --> Asp --> His --> Asp phosphorelay, as well as the dephosphorylation of ArcA-P by a reverse phosphorelay . When ArcA and ArcB were incubated with ATP, the peak levels of phosphorylated proteins increased in the presence of the fermentation metabolites D-lactate, acetate, or pyruvate . In this study, we report that these effectors accelerate the autophosphorylation activity of ArcB and enhance the transphosphorylation of ArcA, but have no effect on the dephosphorylation of ArcA-P . Moreover, the presence of the receiver domain of ArcB is essential for the effectors to influence the autophosphorylation rate of the primary transmitter domain of ArcB.

Biotechnol Prog, 1999 Nov-Dec, 15(6), 1095 - 105
An integrated process for biomolecule isolation and purification; Dai XP et al.; Biomolecule isolation and purification from a fermentation broth usually involve centrifugation, filtration, adsorption, and chromatography steps . Each step contributes to the product cost and product loss . In this research, a cyclic process integrating commercially available ultrafiltration membranes and chromatographic resin beads was developed to achieve the same goal in one device . The device consisted of ion exchange beads on the shell side of a hollow fiber ultrafiltration module . Loading of proteins on the stationary phase on the shell side was carried out for a period of 5-20 min from the permeate on the shell side produced from tube-side feed in ultrafiltration . The eluent was then introduced either from the shell-side inlet or tube-side inlet; the chromatographic fractions were collected from the shell-side outlet . The column was regenerated/washed next to start a new cycle . Systems studied in this cyclic process include the following binary mixtures: myoglobin and beta-lactoglobulin; hemoglobin and bovine serum albumin; and myoglobin and alpha-lactalbumin . Excellent resolutions of the proteins were obtained . A yeast-based cellular suspension containing a mixture of myoglobin and alpha-lactalbumin was also applied to this device . The target proteins were recovered and purified successfully . The cyclic process-based device integrates clarification, concentration, and chromatographic purification of biomolecules and is suitable for both extracellular and intracellular products.

J Appl Microbiol, 1999 Oct, 87(4), 511 - 9
Production of Aspergillus xylanase by lignocellulosic waste fermentation and its application; Gawande PV et al.; Strains of Aspergillus terreus and A . niger, known to produce xylanase with undetectable amounts of cellulase, were studied for xylanase (EC 3.2.1.8) production on various lignocellulosic substrates using solid state fermentation . Of the lignocellulosic substrates used, wheat bran was the best for xylanase production . The effects of various parameters, such as moistening agent, level of initial moisture content, temperature of incubation, inoculum size and incubation time, on xylanase production were studied . The best medium for A . terreus was wheat bran moistened with 1:5 Mandels and Strenberg mineral solution containing 0.1% tryptone, at 35 degrees C, and at inoculum concentration 2x107-2x108 spores 5 g-1 substrate; for A . niger, the best medium was wheat bran moistened with 1:5 Mandels and Strenberg mineral solution containing 0.1% yeast extract, at 35 degrees C, and at an inoculum concentration of 2x107-2x108 spores 5 g-1 substrate . Under these conditions, A . terreus produced 68.9 IU ml-1 of xylanase, and A . niger, 74.5 IU ml-1, after 4 d of incubation . A crude culture filtrate of the two Aspergillus strains was used for the hydrolysis of various lignocellulosic materials . Xylanase preparations from the two strains selectively removed the hemicellulose fraction from all lignocellulosic materials tested.

Eur J Biochem, 1999 Dec, 266(3), 1158 - 65
Biochemical and molecular characterization of the {NiFe} hydrogenase from the hyperthermophilic archaeon, Thermococcus litoralis; Rakhely G et al.; Thermococcus litoralis is a hyperthermophilic archaeon that grows at temperatures up to 98 degrees C by fermentative metabolism and reduces elemental sulfur (S0) to H2S . A {NiFe} hydrogenase, responsible for H2S or H2 production, has been purified and characterized . The enzyme is composed of four subunits with molecular mass 46, 42, 34 and 32 kDa . Elemental analyses gave approximate values of 22 Fe, 22 S and 1 Ni per hydrogenase . EPR spectra at 70 and 5 K indicated the presence of four or five {4Fe-4S} and one {2Fe-2S} type clusters . The optimal temperature for both H2 evolution and oxidation, using artificial electron carriers, was around 80 degrees C . The operon encoding the T . litoralis enzyme is composed of four genes forming one transcriptional unit, and transcription is not regulated by S0 . An unusual transcription-initiation site is located 139 bp upstream from the translational start point . Sequence analyses indicated the presence of new putative nucleotide-binding domains . Upstream from the hydrogenase operon, ORFs probably encoding a molybdopterin oxidoreductase enzyme have been identified . Based on sequence, biochemical and biophysical analyses, a model of the enzyme and the pathway of electron flow during catalysis is proposed.

J Antibiot (Tokyo), 1999 Aug, 52(8), 689 - 94
Roselipins, inhibitors of diacylglycerol acyltransferase, produced by Gliocladium roseum KF-1040; Tomoda H et al.; Gliocladium roseum KF-1040, a marine isolate, was found to produce a series of new inhibitors of diacylglycerol acyltransferase (DGAT) . Four active compounds, designated roselipins 1A, 1B, 2A and 2B, were isolated from the fermentation broth of the producing strain by solvent extraction, ODS column chromatography and preparative HPLC . The highest production of roselipins was observed when cultured in the medium containing natural sea water . Roselipins inhibit DGAT activity with IC50 values of 15 approximately 22 microM in an enzyme assay system using rat liver microsomes.

Atherosclerosis, 2000 Jan, 148(1), 67 - 73
Inhibitory effect of Chinese green tea on endothelial cell-induced LDL oxidation; Yang TT et al.; Green tea has been shown to inhibit Cu(2+)-induced LDL oxidation and suppress lipoxygenase activity . Since LDL oxidation is a characteristic feature of atherogenesis and lipoxygenase is involved in the disease process, the effect of Lung Chen Tea, a non-fermented Chinese green tea, on LDL oxidation induced by human umbilical cord vascular endothelial cell was investigated in the present study . Lung Chen Tea was extracted with methanol and the dried powder was redissolved in water before extraction with chloroform and then ethyl acetate . Lung Chen Tea, chloroform and ethyl acetate fractions dose-dependently reduced LDL oxidation and decreased its relative electrophoretic mobility (P<0.001) when compared to the oxidized LDL . The lipid peroxidation products, thiobarbituric acid reactive substances, and cellular cholesterol were also significantly lowered by 5 and 10 microg/ml Lung Chen Tea (P<0.001) in a dose-dependent manner . The remaining aqueous layer, which was devoid of catechins after chloroform and ethyl acetate extractions, did not prevent LDL oxidation . The results of this study demonstrated that Lung Chen Tea and catechin-rich fractions significantly prevented endothelial cell induced LDL oxidation . The consumption of Lung Chen Tea may therefore lower the risk of coronary heart diseases.

Bioorg Med Chem, 1999 Oct, 7(10), 2163 - 8
Novel screening methods--the key to cloning commercially successful biocatalysts; Taylor SJ et al.; Providing sufficient biocatalyst to support the demands of multi tonne product supply can be problematical . Here we describe how screening for and cloning a gamma-lactamase overcame biocatalyst supply issues, and greatly improved the actual biocatalytic process . The isolation of an expressing gamma-lactamase clone from a gene library necessitated a combination of classical molecular biology techniques together with innovative screening methods to identify a functional clone . Once isolated the enzyme was characterised with regard to its process performance and proved to be active at 500 g L(-1) substrate . Further development of the recombinant fermentation and downstream processing has resulted in the ability to produce sufficient biocatalyst from one 5001 fermentation to resolve 5 metric tonnes of (+/-)-lactam, whilst simplifying the process chemistry greatly.

Biotechnol Bioeng, 1999, 66(4), 203 - 10
Effect of organic acids on the growth and fermentation of ethanologenic Escherichia coli LY01; Zaldivar J et al.; Hemicellulose residues can be hydrolyzed into a sugar syrup using dilute mineral acids . Although this syrup represents a potential feedstock for biofuel production, toxic compounds generated during hydrolysis limit microbial metabolism . Escherichia coli LY01, an ethanologenic biocatalyst engineered to ferment the mixed sugars in hemicellulose syrups, has been tested for resistance to selected organic acids that are present in hemicellulose hydrolysates . Compounds tested include aromatic acids derived from lignin (ferulic, gallic, 4-hydroxybenzoic, syringic, and vanillic acids), acetic acid from the hydrolysis of acetylxylan, and others derived from sugar destruction (furoic, formic, levulinic, and caproic acids) . Toxicity was related to hydrophobicity . Combinations of acids were roughly additive as inhibitors of cell growth . When tested at concentrations that inhibited growth by 80%, none appeared to strongly inhibit glycolysis and energy generation, or to disrupt membrane integrity . Toxicity was not markedly affected by inoculum size or incubation temperature . The toxicity of all acids except gallic acid was reduced by an increase in initial pH (from pH 6.0 to pH 7.0 to pH 8.0) . Together, these results are consistent with the hypothesis that both aliphatic and mononuclear organic acids inhibit growth and ethanol production in LY01 by collapsing ion gradients and increasing internal anion concentrations .

Biotechnol Bioeng, 1999, 66(3), 195 - 201
Rapid quantitation and monitoring of plasmid DNA using an ultrasensitive DNA-binding dye; Noites IS et al.; A sensitive fluorescence-based method for monitoring plasmid DNA during production was investigated . This simple method of assaying for plasmid DNA allows rapid monitoring of plasmid yields from a recombinant Escherichia coli fed-batch fermentation . The assay has several advantages over traditional methods of plasmid DNA measurement . The fluorescent dye is highly specific and can measure total plasmid DNA concentration in about 5 min . The assay is sensitive over a wide range of plasmid concentrations of between 15 and 280 ng/mL, even in the presence of impurities that occur within alkaline lysate preparations . The technique can also be applied to monitoring fermentation and downstream purification steps .

Chem Pharm Bull (Tokyo), 1999 Jul, 47(7), 1032 - 4
Futalosine and its derivatives, new nucleoside analogs; Hosokawa N et al.; Futalosine, a new nucleoside analog, was isolated from a fermentation broth of Streptomyces sp . MK359-NF1 . Some chemical derivatives of futalosine were prepared . 6-O-Methylfutalosine methylester inhibited growth of HeLa-S3 cells in vitro (IC50 = 19.5 micrograms/ml) in contrast to the weak activity of futalosine . 6-O-Methylfutalosine methylester at concentrations higher than 10 micrograms/ml inhibited incorporation of 3H-TdR and 3H-UR but not 3H-Leu in the acid-soluble fractions of HeLa-S3 cells.

Dent Clin North Am, 1999 Oct, 43(4), 615 - 33
Diet and dental caries; Jensen ME; Dental caries is a diet-related disease that continues to be a problem for certain dental patients . Frequent consumption of fermentable carbohydrates that have low oral clearance rates increases the risk for enamel caries and perhaps is even more dangerous for root surfaces . Highly acidogenic snack foods should be consumed at mealtimes to reduce the risk, and between-meal snacks should be either nonacidogenic (such as xylitol products) or hypoacidogenic (such as sorbitol and HSH products) . Cheeses present a naturally occurring situation that may provide anticariogenic effects from the diet . Certain additives as well as sugar substitutes show great promise for the provision of between-meal snack foods that reduce the risk of dental caries . The dental team should thoroughly understand the relationship of diet to caries and conscientiously apply that knowledge to educate the patients in general as well as counsel specific high-risk individuals . Further emphasis should be placed on the acquisition of sound scientific data for counseling caries patients concerning diet and dental caries.

Plant Foods Hum Nutr, 1999, 53(4), 367 - 80
The effect of processing on total organic acids content and mineral availability of simulated cassava-vegetable diets; Adewusi SR et al.; Changes in pH, titratable acidity and mineral content (Ca, Fe, Mg, Zn) were estimated in processed cassava products while the mineral content of raw and blanched amaranthus vegetable was determined . pH of fresh cassava (6.5) decreased as total organic acid (0.07%) increased with fermentation period . Fufu and lafun had the lowest pH and the highest total organic acids contents . Fermentation of cassava increased the total calcium and iron contents, reduced magnesium level while zinc remained fairly constant in grated cassava but was reduced when soaked in water (for the preparation of fufu and lafun) . Fermentation also increased the availability of these selected minerals in both cassva products and simulated cassava-vegetable diets . Blanching reduced the mineral content of amaranthus vegetable but increased mineral availability . Fermentation of cassava and blanching of vegetables play an important role in making minerals more available and these processing methods should be encouraged to potentially ameliorate the disease states associated with mineral deficiency.

Plant Foods Hum Nutr, 1999, 53(4), 285 - 96
Effect of blanching time and salt concentration on pectolytic enzymes, texture and acceptability of fermented green beans; Mnkeni AP et al.; Effect of blanching time, salt concentration and fermentation on texture of green beans (var . Tuf) was determined . Beans were blanched at 90 degrees C for 1, 10 or 20 minutes and fermented with brine containing 3.08 and 4.6% salt (corresponding to 1.2 and 1.8% salt in the finished product) . The beans were assessed for texture mechanically using Universal Testing Machine (Model 1011, Instron) . Sensory evaluation was carried out to assess the taste, texture and overall acceptability of the products . Pectolytic enzyme (pectinestarase and exopolygalacturonase) activities were determined in fresh and fermented beans . Increase in blanching time improved the texture significantly . Significant (p<0.05) decreases in shearing energy of the beans were observed in the first two days of fermentation and, thereafter, there were no significant (p>0.05) changes except for samples blanched for one minute . Salt concentration showed a small but significant effect on texture only in samples blanched for one minute and this was probably due to activation by salt of residual pectolytic enzymes in the beans . Most acceptable beans were those blanched for 20 minutes and fermented with 1.2% salt brine.

Khirurgiia (Mosk), 1999, (10), 13 - 5
{Debatable questions of surgical sepsis treatment and diagnosis}; Reshetnikov EA et al.; On the basis of critical analysis of the results of combined treatment of 73 patients, the authors suggested that sepsis could be diagnosed only in the presence of systemic inflammatory response, multiorganic insufficiency and obligatory isolation of hemoculture . The microorganisms' specimen from the infected area and the blood are not identical both in primary and repeated inoculation . Impossibility to verify the pathogenic microorganism in the blood prevents from carrying out adequate and purposeful treatment . The absence of correlation between the severity of clinical course of sepsis, microflora species and laboratory immunography data, confirms the opinion that the organism responds to the inflammatory process by complex general physiologic reaction of defence, which includes, besides immunologic system, fermentative, endocrine, vegetative and other systems of homeostasis.

J Microbiol Methods, 2000 Jan, 39(2), 121 - 6
A polymerase chain reaction based method for detecting Mycoplasma/Acholeplasma contaminants in cell culture; Tang J et al.; A detection system that utilizes a primer mixture in a nested polymerase chain reaction for detecting Mycoplasma contaminants in cell cultures is described . Primers were designed to amplify the spacer regions between the 16S and 23S ribosomal RNA genes of Mycoplasma and Acholeplasma . This detection system was able to detect 20-180 colony forming units per milliliter of sample . Eight commonly encountered Mycoplasma and Acholeplasma contaminants, which include Mycoplasma (M.) arginini, M . fermentans, M . hominis, M . hyorhinis, M . orale, M . pirum, M . salivarium, and Acholeplasma laidlawii, were consistently amplified . Mycoplasma contaminants generated a single DNA band of 236-365 base pairs (bp), whereas A . laidlawii produced a characteristic two-band pattern of 426 and 219 bp amplicons . Species identification could be achieved by size determination and restriction enzyme digestion . Minor cross-reactions were noted with a few closely related gram positive bacteria and DNA from rat cell lines . A Mycoplasma Detection Kit for detecting Mycoplasma contaminants in cell cultures has been developed based on this approach.

J Dairy Sci, 1999 Nov, 82(11), 2486 - 96
Influence of carbohydrate source and buffer on rumen fermentation characteristics, milk yield, and milk composition in early-lactation Holstein cows; Kennelly JJ et al.; The effects of concentrate to forage ratio and sodium bicarbonate (buffer) supplementation on intake, ruminal fermentation characteristics, digestibility coefficients, milk yield, and milk composition were examined in 4 cannulated Holstein cows (100 +/- 20 d in milk) . A 4 x 4 Latin square design with 2 x 2 factorial arrangement of treatments was implemented for 3-wk experimental periods . The 4 treatments were a 50:50 concentrate to forage ratio with 1.2% of dry matter (DM) and without added buffer and a 75:25 concentrate to forage ratio with (1.2% of DM) and without (0% of DM) buffer . The forage component of the ration was a 50:50 mixture of alfalfa and barley and triticale silage, and diets were fed ad libitum as a total mixed ration . Although feed intake was not influenced by treatments, substantial treatment differences were observed for milk yield and milk composition . Cows fed high-concentrate diet had lower ruminal pH, ruminal acetate, and butyrate concentrations, whereas propionate concentrations were significantly elevated . The addition of buffer, at both levels of concentrate inclusion, resulted in elevated total volatile fatty acids and acetate concentrations . We concluded that altering the forage concentrate ratio in the diet of lactation cows influenced milk yield and milk composition, but the addition of buffer to the diet prevented the elevation in trans-C18:1 fatty acids in milk fat, and related milk fat depression, associated with feeding high-concentrate diets.

J Dairy Sci, 1999 Nov, 82(11), 2472 - 85
Effect of feeding macerated alfalfa silage on nutrient digestibility and milk yield in lactating dairy cows; Broderick GA et al.; Five feeding studies were conducted with 141 lactating Holstein cows comparing macerated and control alfalfa silage harvested at two cuttings in each of 2 yr . Overall, silage made from macerated alfalfa contained more ash (suggesting improved soil contamination); greater fiber and lower nonprotein nitrogen (NPN) content suggested greater fermentation in the silo . In a digestion study, two diets were fed containing {dry matter (DM) basis} 72% of either control or macerated second-cutting alfalfa . Apparent digestibility of neutral detergent fiber and acid detergent fiber (ADF) was increased by maceration, and similar changes in digestibility were observed with Yb or indigestible ADF as marker; indigestible ADF was used as a marker in later studies . Lactation trials were conducted with first- and second-cutting alfalfa from each year . In each study, diets were formulated from alfalfa silage plus concentrate based on processed high moisture ear corn; mean compositions were (DM basis): negative control (61% control alfalfa silage), macerated (61% macerated alfalfa silage), and positive control (50% control alfalfa silage) . All diets contained 2% crude protein from either roasted soybeans or low-solubles fish meal; soybean meal was added to make the positive control isonitrogenous (but not equal in ruminal undergraded protein) . Milk yield was greater on macerated than negative control in two of four trials but not different in the other two trials . Yields of milk and milk components were not different between macerated and positive control in one of four trials . Versus the negative control, milk fat synthesis was depressed on macerated alfalfa in one trial . Overall performance on macerated versus negative control indicated greater apparent digestibility of organic matter (OM), greater yield of milk, protein, and solids not fat, but lower milk fat content . Yields of milk and milk components were greater overall on positive control versus macerated . Estimation of net energy for lactation (NEL) from maintenance, milk yield, and body weight gain indicated that control and macerated alfalfa silage contained, respectively, 1.36 and 1.42 Mcal of NEL of OM, an increase of about 5% due to maceration of alfalfa in these trials.

Eksp Klin Farmakol, 1999 Sep-Oct, 62(5), 59 - 62
{The antioxidant effects of amtizol and trimetazidine}; Smirnov AV et al.; Experiments were conducted on rats to study the antioxidant effects of amtisol and trimethasidin (25 mg/kg) in incomplete cerebral ischemia complicated by hypoxia . Amtisol as well as trimethasidin inhibited activation of lipid peroxidation (LPO) and prevented decrease in activity of the antioxidant system . In in vitro experiments amtisol and trimethasidin blocked epinephrine auto-oxidation . Amtisol inhibited ascorbate-dependent LPO in the liposomes and metabolizing model system, inhibited fermentative NADP-dependent POL in brain tissue homogenates.

J Bacteriol, 1999 Dec, 181(23), 7228 - 34
Sulfate transport in Penicillium chrysogenum: cloning and characterization of the sutA and sutB genes; van de Kamp M et al.; In industrial fermentations, Penicillium chrysogenum uses sulfate as the source of sulfur for the biosynthesis of penicillin . By a PCR-based approach, two genes, sutA and sutB, whose encoded products belong to the SulP superfamily of sulfate permeases were isolated . Transformation of a sulfate uptake-negative sB3 mutant of Aspergillus nidulans with the sutB gene completely restored sulfate uptake activity . The sutA gene did not complement the A . nidulans sB3 mutation, even when expressed under control of the sutB promoter . Expression of both sutA and sutB in P . chrysogenum is induced by growth under sulfur starvation conditions . However, sutA is expressed to a much lower level than is sutB . Disruption of sutB resulted in a loss of sulfate uptake ability . Overall, the results show that SutB is the major sulfate permease involved in sulfate uptake by P . chrysogenum.

Appl Microbiol Biotechnol, 1999 Oct, 52(4), 608 - 11
Fixation of spent Saccharomyces cerevisiae biomass for lead sorption; Ashkenazy R et al.; Spent Saccharomyces cerevisiae cells from a beer fermentation process were evaluated for lead cation sorption . The crude biomass was washed with water and acetone prior to any other treatment . Although the washed biomass showed substantial lead ion sorption it was susceptible to microbial spoilage . Different aldehydes were tested as chemical fixation agents; however, most of them caused drastic lowering of the metal uptake capacity . However, benzaldehyde was not only an excellent fixation agent, but the biomass treated with it also retained its original lead sorption capacity . A mechanism for the fixation process is suggested.

Appl Microbiol Biotechnol, 1999 Oct, 52(4), 534 - 40
Independent production of two molecular forms of a recombinant Rhizopus oryzae lipase by KEX2-engineered strains of Saccharomyces cerevisiae; Takahashi S et al.; A mixture of rProROL having the full-length prosequence (97 amino acids) for a recombinant lipase of Rhizopus oryzae (rROL) and r28ROL having 28 amino acids of the same prosequence has been produced as active forms by Saccharomyces cerevisiae {Takahashi et al . (1998) J Ferment Bioeng 86: 164-168} . However, the separation of rProROL and r28ROL has not been successful due to their identical behavior on column chromatographs, presumably because of the similarity of their surface properties . The independent production of two different molecular forms of rROL was carried out using KEX2-engineered strains of S . cerevisiae, since r28ROL was predicted to be a product from rProROL by a Kex2-like protease . rProROL was successfully obtained by expression of the ROL gene in the S . cerevisiae kex2 strain in which the KEX2 gene encoding Kex2p was disrupted, while r28ROL was obtained by co-expression of the gene (KEX2 delta 613) encoding the soluble form of the C-terminal truncated Kex2 protease (sKex2p) . The specific lipase activities of rProROL and r28ROL were 92.9 U/mg and 140 U/mg, respectively . rProROL was stable at pH 2.2-8.0, and showed the optimal reaction temperature to be 30-35 degrees C with a T50 of 55 degrees C (T50 is the temperature resulting in 50% loss of activity) . The values for r28ROL were pH 3.0-10.0, 25-30 degrees C, and 40 degrees C, respectively . rProROL was an N-linked glycosylated form, but r28ROL was not . The enhanced thermostability of rProROL did not seem to be due to the N-linked glycosylation, as judged by the results of the Endo H treatment . rProROL had the highest esterase activity toward p-nitrophenyl laurate (C12), whereas r28ROL had the highest esterase activity toward p-nitrophenyl caprylate (C8) and stearate (C18) . These results suggest that the distinct properties of these two forms of lipase are caused by the different length of the ROL prosequence.

Appl Microbiol Biotechnol, 1999 Oct, 52(4), 502 - 7
Monitoring of low concentrations of glucose in fermentation broth; Nandakumar MP et al.; A highly sensitive glucose sensor, operating in flow-injection analysis (FIA) mode, was developed for the detection of glucose in fermentation broth . The assay system is based upon the post-column reaction of the peroxide formed in the glucose-oxidase-catalysed reaction and subsequent spectrophotometric detection of the coloured product formed . The sensor system was characterised and calibrated using standard solutions, and later used for quantification of glucose in fermentation media . Two types of enzyme column were used: one operated in packed-bed mode and the other in expanded-bed mode . Both columns were integrated into a FIA system and were found to give good analytical results . Glucose concentrations as low as 0.1 mg/l and 5 mg/l could be detected in packed- and expanded-bed modes respectively . Glucose concentrations were measured during typical fed-batch fermentation conditions in this system, and the results are presented.

J Agric Food Chem, 1999 Feb, 47(2), 633 - 6
Total antioxidant capacity of teas by the ferric reducing/antioxidant power assay; Benzie IF et al.; This study aimed to compare in vitro antioxidant power of different types of tea (Camellia sinensis) . The ferric reducing/antioxidant power (FRAP) assay was used to measure the total antioxidant power of freshly prepared infusions of 25 types of teas . Results showed that different teas had widely different in vitro antioxidant power and that the antioxidant capacity was strongly correlated (r = 0 . 956) with the total phenolics content of the tea . Expressed as micromol of antioxidant power/g of dried tea leaves, values ranged as 132-654 micromol/g for black ("fermented") teas, 233-532 micromol/g for Oolong ("semifermented") teas, and 272-1144 micromol/g for green ("nonfermented") teas . One cup of tea of usual strength (1-2%), therefore, can provide the same potential for improving antioxidant status as around 150 mg of pure ascorbic acid (vitamin C).

Infect Immun, 1999 Dec, 67(12), 6303 - 8
Induction of cytokines and chemokines in human monocytes by Mycoplasma fermentans-derived lipoprotein MALP-2; Kaufmann A et al.; Bacterial infections are characterized by strong inflammatory reactions . The responsible mediators are often bacterially derived cell wall molecules, such as lipopolysaccharide or lipoteichoic acids, which typically stimulate monocytes and macrophages to release a wide variety of inflammatory cytokines and chemokines . Mycoplasmas, which lack a cell wall, may also stimulate monocytes very efficiently . This study was performed to identify mycoplasma-induced mediators . We investigated the induction of cytokines and chemokines in human monocytes exposed to the Mycoplasma fermentans-derived membrane component MALP-2 (macrophage-activating lipopeptide 2) by dose response and kinetic analysis . We found a rapid and strong MALP-2-inducible chemokine and cytokine gene expression which was followed by the release of chemokines and cytokines with peak levels after 12 to 20 h . MALP-2 induced the neutrophil-attracting CXC chemokines interleukin-8 (IL-8) and GRO-alpha as well as the mononuclear leukocyte-attracting CC chemokines MCP-1, MIP-1alpha, and MIP-1beta . Production of the proinflammatory cytokines tumor necrosis factor alpha and IL-6 started at the same time as chemokine release but required 10- to 100-fold-higher MALP-2 doses . The data show that the mycoplasma-derived lipopeptide MALP-2 represents a potent inducer of chemokines and cytokines which may, by the attraction and activation of neutrophils and mononuclear leukocytes, significantly contribute to the inflammatory response during mycoplasma infection.

Infect Immun, 1999 Dec, 67(12), 6281 - 5
Effect of MALP-2, a lipopeptide from Mycoplasma fermentans, on bone resorption in vitro; Piec G et al.; Mycoplasmas may be associated with rheumatoid arthritis in various animal hosts . In humans, mycoplasma arthritis has been recorded in association with hypogammaglobulinemia . Mycoplasma fermentans is one mycoplasma species considered to be involved in causing arthritis . To clarify which mycoplasmal compounds contribute to the inflammatory, bone-destructive processes in arthritis, we used a well-defined lipopeptide, 2-kDa macrophage-activating lipopeptide (MALP-2) from M . fermentans, as an example of a class of macrophage-activating compounds ubiquitous in mycoplasmas, to study its effects on bone resorption . MALP-2 stimulated osteoclast-mediated bone resorption in murine calvaria cultures, with a maximal effect at around 2 nM . Anti-inflammatory drugs inhibited MALP-2-mediated bone resorption by about 30% . This finding suggests that MALP-2 stimulates bone resorption partially by stimulating the formation of prostaglandins . Since interleukin-6 (IL-6) stimulates bone resorption, we investigated IL-6 production in cultured calvaria . MALP-2 stimulated the liberation of IL-6, while no tumor necrosis factor was detectable . Additionally, MALP-2 stimulated low levels of NO in calvaria cultures, an effect which was strongly increased in the presence of gamma interferon, causing an inhibition of bone resorption . MALP-2 stimulated the bone-resorbing activity of osteoclasts isolated from long bones of newborn rats and cultured on dentine slices without affecting their number . In bone marrow cultures, MALP-2 inhibited the formation of osteoclasts . It appears that MALP-2 has two opposing effects: it increases the bone resorption in bone tissue by stimulation of mature osteoclasts but inhibits the formation of new ones.

Mol Cell Biol, 1999 Dec, 19(12), 7995 - 8002
Mycoplasmal infections prevent apoptosis and induce malignant transformation of interleukin-3-dependent 32D hematopoietic cells; Feng SH et al.; 32D cells, a murine myeloid cell line, rapidly undergo apoptosis upon withdrawal of interleukin-3 (IL-3) supplement in culture . We found that 32D cells, if infected by several species of human mycoplasmas that rapidly activated NF-kappaB, would live and continue to grow in IL-3-depleted culture . Mycoplasma-infected cells showed no evidence of autocrine production of IL-3 . Pyrrolidine dithiocarbamate (PDTC) blocked activation of NF-kappaB and led to prominent cell death . Heat-killed mycoplasmas or mycoplasmal membrane preparations alone could support continued growth of 32D cells in culture without IL-3 supplement for a substantial period of time . However, upon removal of heat-inactivated mycoplasmas, 32D cells quickly became apoptotic . In comparison, live Mycoplasma fermentans or M . penetrans infection for 4 to 5 weeks induced malignant transformation of 32D cells . Transformed 32D cells grew autonomously and no longer required support of growth-stimulating factors including IL-3 and mycoplasmas . The transformed 32D cells quickly formed tumors when injected into nude mice . Karyotyping showed that development of chromosomal changes and trisomy 19 was often associated with malignant transformation and tumorigenicity of 32D cells . Mycoplasmal infections apparently affected the fidelity of genomic transmission in cell division as well as checkpoints coordinating the progression of cell cycle events.

Biotechnol Bioeng, 1999, 66(2), 104 - 13
Metabolic shifts by nutrient manipulation in continuous cultures of BHK cells; Cruz HJ et al.; The present work aims at characterizing the regulatory mechanisms of metabolism and product formation of BHK cells producing a recombinant antibody/cytokine fusion protein . This work was carried out through the achievement of several steady-states in chemostat cultures, corresponding to different glucose and glutamine levels in the feed culture medium . Results obtained indicate that both glucose and glutamine consumptions show a Michaelis-Menten dependence on residual glucose and glutamine concentrations, respectively . Similar dependence was also observed for lactate and ammonia productions . K(Glc)(Glc) and K(Gln)(Gln) were estimated to be 0.4 and 0.15 mM, respectively, while q(max)(Glc) and q(max)(Gln) were estimated to be 1.8 and 0.55 nmol 10(-6)cells min(-1), respectively . At very low glucose concentrations, the glucose-to-lactate yield decreased markedly showing a metabolic shift towards lower lactate production; also, the glucose-to-cells yield was increased . At very low-glutamine concentrations, the glutamine-to-ammonia and glutamine-to-cells yields increased, showing a more efficient glutamine metabolism . Overall, amino acid consumption was increased under low glucose or glutamine concentrations . Metabolic-flux analysis confirmed the metabolic shifts by showing increases in the fluxes of the more energetically efficient pathways, at low-nutrient concentrations . No effect of glucose or glutamine concentrations on the cell-specific productivity was observed, even under metabolically shifted metabolism; therefore, it is possible to confine the cells to a more efficient metabolic state maintaining the productivity of the recombinant product of interest, and consequently, increasing final product titers by increasing cell concentration and culture length . This work is intended to be a model approach to characterize cell metabolism in an integrated way; it is highly valuable for the establishment of operating strategies in mammalian cell fermentations in which cell metabolism is to be confined to a desired state .

Inflammation, 1999 Dec, 23(6), 495 - 505
Mycoplasma fermentans-induced inflammatory response of astrocytes: selective modulation by aminoguanidine, thalidomide, pentoxifylline and IL-10; Gallily R et al.; Exposure of primary rat glial cells, mostly astrocytes, to heat-inactivated Mycoplasma fermentans triggers the production of tumor necrosis factor alpha (TNFalpha) nitric oxide (NO) and prostaglandin E2 (PGE2) . To attenuate the production of these proinflammatory mediators, four agents: aminoguanidine, pentoxifylline, thalidomide and IL-10 were added to astrocyte cultures . Aminoguanidine (1 and 3 mM), an inhibitor of inducible nitric oxide synthase (iNOS), suppressed the production of the three mediators . TNFalpha was the most sensitive to thalidomide, showing dose-response inhibition at concentrations of 20 microg/ml, 50 microg/ml and 250 microg/ml . PGE2 was affected only by concentrations of 50 microg/ml and 250 microg/ml, whereas NO responded solely to the highest amount of this inhibitor . The cytokine IL-10, at 10 U and 50 U, inhibited only TNFalpha production . Our results imply that selective suppression of proinflammatory mediators by various agents may prove feasible for amelioration of central nervous system inflammatory diseases.

Scand J Gastroenterol, 1999 Oct, 34(10), 967 - 73
Ethanol oxidation and acetaldehyde production in vitro by human intestinal strains of Escherichia coli under aerobic, microaerobic, and anaerobic conditions; Salaspuro V et al.; BACKGROUND: Many human colonic facultative anaerobic and aerobic bacteria are capable of alcohol dehydrogenase (ADH)-mediated ethanol oxidation . In this bacteriocolonic pathway for ethanol oxidation intracolonic ethanol is first oxidized by bacterial ADHs to acetaldehyde, which is further oxidized by either colonic mucosal or bacterial aldehyde dehydrogenases to acetate . The produced acetaldehyde is a highly toxic and carcinogenic agent . This study was aimed to investigate the ethanol oxidation capability and acetaldehyde formation of Escherichia coli IH 50546 and IH 50817 . These intestinal E . coli strains expressed either high (IH 50546) or low (IH 50817) ADH activity . METHODS: Strains were cultured for 48 h on agar plates supplemented with ethanol under aerobic, microaerobic (6% O2), and anaerobic conditions . RESULTS: Under aerobic conditions both E . coli strains oxidized ethanol . The ethanol consumption rates (ECR) were 1.046+/-0.025 mM/h and 0.367+/-0.148 mM/h with IH 50546 and IH 50817, respectively . In the case of IH 50546 this was associated with significant acetaldehyde production (418+/-13 microM), suggesting ADH-mediated ethanol oxidation . Under microaerobic conditions only IH 50546 was able to oxidize ethanol (ECR, 0.498+/-0.074 mM/h) and to produce acetaldehyde (up to 440+/-76 microM) to significant extents . Under anaerobic conditions both strains fermented glucose to ethanol . CONCLUSIONS: This study experimentally shows the potential of certain bacteria representing normal human colonic flora to produce acetaldehyde under various atmospheric conditions that may prevail in different parts of the GI tract . This bacterial adaptation may be an essential feature of the bacteriocolonic pathway to produce toxic and carcinogenic acetaldehyde from either endogenous or exogenous ethanol.

Gut, 1999 Dec, 45(6), 840 - 7
Wheat bran affects the site of fermentation of resistant starch and luminal indexes related to colon cancer risk: a study in pigs; Govers MJ et al.; BACKGROUND: Recent studies suggest that resistant starch (effective in producing butyrate and lowering possibly toxic ammonia) is rapidly fermented in the proximal colon; the distal colon especially would, however, benefit from these properties of resistant starch . AIMS: To determine whether wheat bran (a rich source of insoluble non-starch polysaccharides), known to hasten gastrointestinal transit, could carry resistant starch through to the distal colon and thus shift its site of fermentation . METHODS: Twenty four pigs were fed four human type diets: a control diet, or control diet supplemented with resistant starch, wheat bran, or both . Intestinal contents and faeces were collected after two weeks . RESULTS: Without wheat bran, resistant starch was rapidly fermented in the caecum and proximal colon . Supplementation with wheat bran inhibited the caecal fermentation of resistant starch, resulting in an almost twofold increase (from 12.9 (2.5) to 20.5 (2.1) g/day, p<0.05) in resistant starch being fermented between the proximal colon and faeces . This resulted in higher butyrate (133%, p<0.05) and lower ammonia (81%, p<0.05) concentrations in the distal colonic regions . CONCLUSIONS: Wheat bran can shift the fermentation of resistant starch further distally, thereby improving the luminal conditions in the distal colonic regions where tumours most commonly occur . Therefore, the combined consumption of resistant starch and insoluble non-starch polysaccharides may contribute to the dietary modulation of colon cancer risk.

Trends Biotechnol, 1999 Dec, 17(12), 499 - 505
The development and applications of thermal biosensors for bioprocess monitoring; Ramanathan K et al.; Enzyme thermistors are biosensors that use thermal resistors to measure the heat change caused by an enzymatic reaction . They combine the selectivity of enzymes with the sensitivity of biosensors and allow continuous analysis in a flow-injection mode . They can be used to monitor fermentation systems, biocatalysis, enzyme-catalysed synthesis and clinical and food technology . This article gives an overview of the general principles of enzyme thermistors, the sampling process and the ongoing developments in the field of bioprocess monitoring.

Eur J Clin Nutr, 1999 Nov, 53(11), 844 - 8
Comparison of starch digestibility of a blended food prepared with and without extrusion cooking; Meance S et al.; OBJECTIVE: To compare starch digestibility from a maize-soy blended food with and without extrusion cooking . DESIGN: Resistant starch, soluble and insoluble dietary fibres were measured in vitro before and after extrusion . Starch digestibility was assessed in 8 volunteers who took, in a randomised order, a test meal with either 100 g extruded (EF) or non-extruded (NEF) blended flour cooked 15 min at 80 degrees C in 500 ml of water . SETTING: Research ward for healthy volunteers . SUBJECTS: Healthy volunteers . MAIN OUTCOME MEASURES: Starch digestibility was measured by 13C enrichment of breath samples for 8 h . Breath H2 concentration was measured during 12 h to assess bacterial fermentation in the colon . Volunteers reported hunger on a visual scale every hour for 8 h . RESULTS: In vitro resistant starch, soluble and insoluble dietary fibers were higher in NEF than in EF (5.4 vs 1.1, 0.7 vs 0.5, and 13.3 vs 10.4% dw respectively) . In vivo, the area under curve (AUC) for 13CO2 excretion during 8 h was not significantly different for NEF and EF (10.3+/-1.3 vs 9.1+/-0.5 mmol/min, respectively) . AUC for H2 excretion during 12 h was significantly higher for EF than for NEF (26.9+/-5.6 vs 14.1+/-4.7 mL, P<0.05) . AUC for satiety was marginally higher with EF (12.6+/-1.6) than for NEF (10.0+/-2.1) (P=0.06) . CONCLUSIONS: Extrusion cooking does not seem to substantially improve blended foods digestibility . Extrusion of high fibre flours may promote carbohydrate fermentation in the colon and increase satiety.

Angew Chem Int Ed Engl, 1999 Nov 2, 38(21), 3197 - 3201
Discovery Through Total Synthesis-Epimerization at C7 in the CP Compounds: Is (7S)-CP-263,114 a Fermentation Product?
Meng D, Tan Q, Danishefsky SJ.
Sometimes a surprise can occur in a total synthesis . The preparative investigation of the title compound CP-263,114 and of CP-225,917 (1)-both natural products that were first isolated in 1997 by Pfizer-has shown that CP compounds of the 7R series can be converted into the 7S series (for example, 2); (7S)-CP-263,114 is itself apparently a fermentation product.

Biotechnol Bioeng, 1999, 66(1), 42 - 50
Steady-state and transient-state analysis of growth and metabolite production in a Saccharomyces cerevisiae strain with reduced pyruvate-decarboxylase activity; Flikweert MT et al.; Pyruvate decarboxylase is a key enzyme in the production of low-molecular-weight byproducts (ethanol, acetate) in biomass-directed applications of Saccharomyces cerevisiae . To investigate whether decreased expression levels of pyruvate decarboxylase can reduce byproduct formation, the PDC2 gene, which encodes a positive regulator of pyruvate-decarboxylase synthesis, was inactivated in the prototrophic strain S . cerevisiae CEN . PK113-7D . This caused a 3-4-fold reduction of pyruvate-decarboxylase activity in glucose-limited, aerobic chemostat cultures grown at a dilution rate of 0.10 h(-1) . Upon exposure of such cultures to a 50 mM glucose pulse, ethanol and acetate were the major byproducts formed by the wild type . In the pdc2Delta strain, formation of ethanol and acetate was reduced by 60-70% . In contrast to the wild type, the pdc2Delta strain produced substantial amounts of pyruvate after a glucose pulse . Nevertheless, its overall byproduct formation was ca . 50% lower . The specific rate of glucose consumption after a glucose pulse to pdc2Delta cultures was about 40% lower than in wild-type cultures . This suggests that, at reduced pyruvate-decarboxylase activities, glycolytic flux is controlled by NADH reoxidation . In aerobic, glucose-limited chemostat cultures, the wild type exhibited a mixed respiro-fermentative metabolism at dilution rates above 0.30 h(-1) . Below this dilution rate, sugar metabolism was respiratory . At dilution rates up to 0.20 h(-1), growth of the pdc2Delta strain was respiratory and biomass yields were similar to those of wild-type cultures . Above this dilution rate, washout occurred . The low micro(max) of the pdc2Delta strain in glucose-limited chemostat cultures indicates that occurrence of respiro-fermentative metabolism in wild-type cultures is not solely caused by competition of respiration and fermentation for pyruvate . Furthermore, it implies that inactivation of PDC2 is not a viable option for reducing byproduct formation in industrial fermentations .

Biotechnol Bioeng, 1999, 66(1), 1 - 16
Mathematical modeling and optimization of cellulase protein production using Trichoderma reesei RL-P37; Tholudur A et al.; The enzyme cellulase, a multienzyme complex made up of several proteins, catalyzes the conversion of cellulose to glucose in an enzymatic hydrolysis-based biomass-to-ethanol process . Production of cellulase enzyme proteins in large quantities using the fungus Trichoderma reesei requires understanding the dynamics of growth and enzyme production . The method of neural network parameter function modeling, which combines the approximation capabilities of neural networks with fundamental process knowledge, is utilized to develop a mathematical model of this dynamic system . In addition, kinetic models are also developed . Laboratory data from bench-scale fermentations involving growth and protein production by T . reesei on lactose and xylose are used to estimate the parameters in these models . The relative performances of the various models and the results of optimizing these models on two different performance measures are presented . An approximately 33% lower root-mean-squared error (RMSE) in protein predictions and about 40% lower total RMSE is obtained with the neural network-based model as opposed to kinetic models . Using the neural network-based model, the RMSE in predicting optimal conditions for two performance indices, is about 67% and 40% lower, respectively, when compared with the kinetic models . Thus, both model predictions and optimization results from the neural network-based model are found to be closer to the experimental data than the kinetic models developed in this work . It is shown that the neural network parameter function modeling method can be useful as a "macromodeling" technique to rapidly develop dynamic models of a process .

Int J Syst Bacteriol, 1999 Oct, 49 Pt 4, 1631 - 43
Psychrophilic sulfate-reducing bacteria isolated from permanently cold arctic marine sediments: description of Desulfofrigus oceanense gen . nov., sp . nov., Desulfofrigus fragile sp . nov., Desulfofaba gelida gen . nov., sp . nov., Desulfotalea psychrophila gen . nov., sp . nov . and Desulfotalea arctica sp . nov; Knoblauch C et al.; Five psychrophilic, Gram-negative, sulfate-reducing bacteria were isolated from marine sediments off the coast of Svalbard . All isolates grew at the in situ temperature of -1.7 degrees C . In batch cultures, strain PSv29T had the highest growth rate at 7 degrees C, strains ASv26T and LSv54T had the highest growth rate at 10 degrees C, and strains LSv21T and LSv514T had the highest growth rate at 18 degrees C . The new isolates used the most common fermentation products in marine sediments, such as acetate, propionate, butyrate, lactate and hydrogen, but only strain ASv26T was able to oxidize fatty acids completely to CO2 . The new strains had growth optima at neutral pH and marine salt concentration, except for LSv54T which grew fastest with 1% NaCl . Sulfite and thiosulfate were used as electron acceptors by strains ASv26T, PSv29T and LSv54T, and all strains except PSv29T grew with Fe3+ (ferric citrate) as electron acceptor . Chemotaxonomy based on cellular fatty acid patterns and menaquinones showed good agreement with the phylogeny based on 16S rRNA sequences . All strains belonged to the delta subclass of Proteobacteria but had at least 9% evolutionary distance from known sulfate reducers . Due to the phylogenetic and phenotypic differences between the new isolates and their closest relatives, establishment of the new genera Desulfotalea gen . nov., Desulfofaba gen . nov . and Desulfofrigus gen . nov . is proposed, with strain ASv26T as the type strain of the type species Desulfofrigus oceanense sp . nov., LSv21T as the type strain of Desulfofrigus fragile sp . nov., PSv29T as the type strain of the type species Desulfofaba gelida sp . nov., LSv54T as the type strain of the type species Desulfotalea psychrophila sp . nov . and LSv514T as the type strain of Desulfotalea arctica sp . nov.

Nahrung, 1999 Oct, 43(5), 320 - 4
Physico-chemical properties of Azizi' green pickled olives as affected by alkali process; el-Makhzangy A et al.; This work was carried out to study the effect of lye treatment at different concentrations (1.0, 2.0 and 2.5%) on some physical-chemical changes of green table olive 'Azizi' cultivar during fermentation process . Results indicated that lye treatment at 1% and 2% using sodium hydroxide were the more suitable concentration for processing green table olive 'Azizi' cultivar under Egyptian condition, where the quality in texture, colour, flavour and appearance was recorded . Statistical analysis revealed that there were significant differences in pH, NaCl content and acidity of green olive during the fermentation period as affected by lye treatment . A positive correlation was found between the sensorial properties of pickled olives.

Mol Med, 1999 Sep, 5(9), 631 - 40
Butyrate switches the pattern of chemokine secretion by intestinal epithelial cells through histone acetylation; Fusunyan RD et al.; BACKGROUND: Butyrate, a fermentation product of intestinal bacteria, modifies chromatin structure through histone acetylation, thereby altering gene transcription . IL-8 and MCP-1 are chemokines, expressed by intestinal epithelial cells, which attract neutrophils and monocytes, respectively . We hypothesized that butyrate may alter IL-8 and MCP-1 expression by intestinal epithelial cells through histone acetylation . MATERIALS AND METHODS: IL-8 and MCP-1 expression was measured by ELISA and RNA transfer blots . Acetylated histones were separated on acetic acid-urea-triton gels . Butyrate was compared to Trichostatin-A, a specific inhibitor of histone deacetylase and to other short chain fatty acids . RESULTS: Caco-2 cells constitutively secreted MCP-1 but not IL-8 . Butyrate reversibly decreased MCP-1 secretion . In contrast, butyrate increased IL-8 production . The effects of butyrate and Trichostatin-A were greater when cells were stimulated with IL-1beta . Butyrate and Trichostatin-A both increased histone acetylation . Trichostatin-A and other short chain fatty acids altered chemokine secretion according to their effect on histone acetylation . CONCLUSIONS: Butyrate reversibly switches chemokine secretion by epithelial cells through histone acetylation . We speculate that butyrate carries information from resident bacteria to epithelial cells . Epithelial cells transduce this signal through histone acetylation, modulating the secretion of chemokines.

Planta, 1999 Oct, 209(4), 547 - 50
Poly(beta-hydroxybutyrate) production in oilseed leukoplasts of brassica napus
Houmiel KL, Slater S, Broyles D, Casagrande L, Colburn S, Gonzalez K, Mitsky TA, Reiser SE, Shah D, Taylor NB, Tran M, Valentin HE, Gruys KJ.
Polyhydroxyalkanoates (PHAs) comprise a class of biodegradable polymers which offer an environmentally sustainable alternative to petroleum-based plastics . Production of PHAs in plants is attractive since current fermentation technology is prohibitively expensive . The PHA homopolymer poly(beta-hydroxybutyrate) (PHB) has previously been produced in leaves of Arabidopsis thaliana (Nawrath et al., 1994, Proc Natl Acad Sci USA 91: 12760-12764) . However, Brassica napus oilseed may provide a better system for PHB production because acetyl-CoA, the substrate required in the first step of PHB biosynthesis, is prevalent during fatty acid biosynthesis . Three enzymatic activities are needed to synthesize PHB: a beta-ketothiolase, an acetoacetyl-CoA reductase and a PHB synthase . Genes from the bacterium Ralstonia eutropha encoding these enzymes were independently engineered behind the seed-specific Lesquerella fendleri oleate 12-hydroxylase promoter in a modular fashion . The gene cassettes were sequentially transferred into a single, multi-gene vector which was used to transform B . napus . Poly(beta-hydroxybutyrate) accumulated in leukoplasts to levels as high as 7.7% fresh seed weight of mature seeds . Electron-microscopy analyses indicated that leukoplasts from these plants were distorted, yet intact, and appeared to expand in response to polymer accumulation.

Arch Tierernahr, 1999, 52(1), 75 - 93
{Investigations on the fermentation quality of silage from low-nitrate green forage}; Kaiser E et al.; In 3-year-experiments green forage from Dactylis glomerata and Festuca species, produced by differentiated cutting frequencies, was ensiled under laboratory conditions . The forage was almost without nitrate . Because of the higher content of water-soluble carbohydrates the fermentability coefficients of Festuca species were above of Dactylis glomerata . The fermentability coefficients of both plant species were inferior in the second and third cut, even if the first cut was delayed until July . Despite of similar fermentability coefficients of the two plant species silages contained different amounts of lactic and butyric acid . The portion of lactic acid amounted to 52 to 57% and 60 to 73% of total acids, the proportion of butyric acid 30 to 35% and 18 to 25% for Dactylis glomerata and Festuca species, respectively . According to the high fermentability coefficients silages free of butyric acid would be expected, however, by both species butyric acid always was formed, in some cases at high concentrations . In the contrary the amounts of ammonia, acetic acid and alcohol as well as the pH-value were almost extremely low . It can be concluded that the quality of silages produced of green forages with low nitrate cannot be assessed adequately by the current evaluation key.

Arch Tierernahr, 1999, 52(1), 1 - 14
Biochemical characteristics of non starch polysaccharide hydrolyzing enzyme preparations designed as feed additives for poultry and piglet nutrition; Vahjen W et al.; Non starch polysaccharide hydrolyzing enzyme preparations analyzed in this study were composed of up to nine (1-3, 1-4)-beta-glucanase activities and up to six xylanase activities with different molecular weights in the range from 100 kD down to 18 kD as determined with SDS/PAGE zymograms . Partially purified enzyme fractions differed in terms of pH-optima, isoelectric point and thermal stability in aquaeous solutions . Different beta-glucanase activities were found in different production strains, although some enzymes were conserved over genus boundaries . Enzyme preparations from the same or related strains exhibited different patterns of enzyme activity, indicating modification of strain and/or fermentation conditions . Some enzyme preparations contained significant amounts of polygalacturonase and/or galactomannase activity . The pH profiles of whole enzyme preparations resulted from pH optima of isoenzyme fractions . Temperature optima for all preparations were between 50 and 60 degrees C . Thermal stability of high molecular weight components tended to be lower than for low molecular weight fractions . Fractions with cellulase activity were most stable, followed by (1-3, 1-4)-beta-glucanase activities, while fractions with xylanase activities exhibited low thermal stabilities . Incubation of enzyme preparations and their respective active fractions in digesta supernatants revealed only small differences in residual xylanase activity . Digesta from gizzard samples led to the highest inactivation . It is concluded that commercial enzyme preparations display different modes of action and that the development of improved enzyme preparations depends not only on thermal stability, but also on pH profile, substrate specificity and proteolytic stability within the digestive tract.

Arch Tierernahr, 1999, 52(2), 195 - 201
Age and incubation time effects on in vitro caecal fermentation pattern in rabbits before and after weaning; Marounek M et al.; The caecal fermentation pattern was studied in four litters of rabbits . Rabbits were sequentially slaughtered at the age of 4 (before weaning), 6, 8 and 11 weeks . Their caecal contents were analyzed and incubated in vitro at 39 degrees C for 6 and 12 h . Net productions of short-chain fatty acids (SCFA), hydrogen and methane were determined . The average caecal weight increased three-times within two weeks after weaning, from 31.4 to 93.7 g . At the end of the experiment, the caecal weight was on average 134.4 g . A large variability of the SCFA concentration observed before weaning decreased after weaning . Measurements of caecal metabolite profiles and results of in vitro experiments indicated a certain decrease of propionate in favour of butyrate, associated with the weaning . The establishment of methanogens in rabbits was slow . Methanogenesis started in one out of four rabbits at the age of 6 weeks . Five weeks later, one of four rabbits still did not produce methane . The hydrogen recovery decreased between the 4th and the 6th week of age, due to the increase of the butyrate/propionate ratio . After the 6th week, the hydrogen recovery increased with age, apparently because of the increase in methane production . Hydrogen recovery tended to increase during incubation, suggesting a decrease of reductive acetogenesis . This increase was observed both in methanogenic and non-methanogenic rabbit caecal cultures . In former cultures, the ratio CH4/SCFA rose with time of incubation.

Arch Tierernahr, 1999, 52(2), 167 - 84
{Effect of hay particle size at different concentrations and feeding levels on digestive processes and feed intake in ruminants . 1 . Chewing activity and fermentation in the rumen}; Tafaj M et al.; In order to study the main effects of particle size three ruminally fistulated cows were fed a hay rich in fibre in long (28.7 mm), chopped (9.2 mm) and ground (2.9 mm) form in a 3 x 3 Latin square design . In another three factorial experiment with 8 wethers (4 animals were ruminally fistulated) the main effects and interactions of the above mentioned hay particle size at two concentrate levels (10.4 to 13.3 and 29.5 to 40.1% in the ration) and at two feeding levels (restricted and ad libitum) were investigated . There was no effect of chopped hay (9.2 mm) on chewing activity, whereas ground hay (2.9 mm) reduced rumination time (52% in sheep and 36% in dairy cows) and chewing expense (time/unit of DM or NDF intake) . These effects were more pronounced at high concentrate intake . Sheep increased feed intake more through reduction of rumination expense than through the extension of rumination time . pH-value, concentration of bicarbonate and C2:C3-ratio indicate favourable fermentation conditions in long (28.7 mm) and chopped hay (9.2 mm), whereas a negative influence of ground hay (2.9 mm) or ruminal fermentation could be observed.

Biochem J, 1999 Nov 15, 344 Pt 1, 77 - 84
Competition between Escherichia coli strains expressing either a periplasmic or a membrane-bound nitrate reductase: does Nap confer a selective advantage during nitrate-limited growth?
Potter LC, Millington P, Griffiths L, Thomas GH, Cole JA.
The physiological role of the periplasmic nitrate reductase, Nap, one of the three nitrate reductases synthesized by Escherichia coli K-12, has been investigated . A series of double mutants that express only one nitrate reductase were grown anaerobically in batch cultures with glycerol as the non-fermentable carbon source and nitrate as the terminal electron acceptor . Only the strain expressing nitrate reductase A grew rapidly under these conditions . Introduction of a narL mutation severely decreased the growth rate of the nitrate reductase A strain, but enhanced the growth of the Nap(+) strain . The ability to use nitrate as a terminal electron acceptor for anaerobic growth is therefore regulated primarily by the NarL protein at the level of transcription . Furthermore, the strain expressing nitrate reductase A had a substantial selective advantage in competition with the strain expressing only Nap during nitrate-sufficient continuous culture . However, the strain expressing Nap was preferentially selected during nitrate-limited continuous growth . The saturation constants for nitrate for the two strains (which numerically are equal to the nitrate concentrations at half of the maximum specific growth rate and therefore reflect the relative affinities for nitrate) were estimated using the integrated Monod equation to be 15 and 50 microM for Nap and nitrate reductase A respectively . This difference is sufficient to explain the selective advantage of the Nap(+) strain during nitrate-limited growth . It is concluded that one physiological role of the periplasmic nitrate reductase of enteric bacteria is to enable bacteria to scavenge nitrate in nitrate-limited environments.

Biochem J, 1999 Nov 15, 344 Pt 1, 69 - 76
Essential roles for the products of the napABCD genes, but not napFGH, in periplasmic nitrate reduction by Escherichia coli K-12; Potter LC et al.; The seven nap genes at minute 47 on the Escherichia coli K-12 chromosome encode a functional nitrate reductase located in the periplasm . The molybdoprotein, NapA, is known to be essential for nitrate reduction . We now demonstrate that the two c-type cytochromes, the periplasmic NapB and the membrane-associated NapC, as well as a fourth polypeptide, NapD, are also essential for nitrate reduction in the periplasm by physiological substrates such as glycerol, formate and glucose . None of the three iron-sulphur proteins, NapF, NapG or NapH, are essential, irrespective of whether the bacteria are grown anaerobically in the presence of nitrate or fumarate as a terminal electron acceptor, or by glucose fermentation . Mutation of napD resulted in the total loss of Methyl Viologen-dependent nitrate reductase activity of the molybdoprotein, NapA, consistent with an earlier suggestion by others that NapD might be required for post-translational modification of NapA.

Nutr Hosp, 1999 May, 14 Suppl 2, 22S - 31S
{Dietary fiber: concept, classification and current indications}; Garcia Peris P et al.; Fiber is a concept that refers to or encompasses several carbohydrates and lignine that resist hydrolysis by human digestive enzymes and that are fermented by the microflora of the colon . From a practical point of view, fibers can be divided into soluble and insoluble . There is general acceptance of the concepts soluble fiber, fermentable, viscous and insoluble fiber, and non-viscous and barely fermentable fiber . The physiological effects and therefore the clinical applications of both fibers are different . In general, the insoluble fiber is barely fermentable and has a marked laxative and intestinal regulatory effect . Soluble fiber is fermented to a high degree, showing a powerful trophic effect at the colon level . Soluble fiber is also attributed a positive role in the carbohydrate and lipid metabolism due to the effects that this has at the intestinal and the systemic level on the glucose and the cholesterol metabolism . The goal of this article is to review the current concept of fiber based on the existing bibliography (it is thought that perhaps the current classification should be changed and that fiber should be talked about depending on its degree of polymerization), its physiologic effects and the possible indications that this may have from a clinical point of view, be this at the level of oral or enteral nutrition.

J Bacteriol, 1999 Nov, 181(21), 6806 - 13
Role of crotonyl coenzyme A reductase in determining the ratio of polyketides monensin A and monensin B produced by Streptomyces cinnamonensis; Liu H et al.; The ccr gene, encoding crotonyl coenzyme A (CoA) reductase (CCR), was cloned from Streptomyces cinnamonensis C730.1 and shown to encode a protein with 90% amino acid sequence identity to the CCRs of Streptomyces collinus and Streptomyces coelicolor . A ccr-disrupted mutant, S . cinnamonensis L1, was constructed by inserting the hyg resistance gene into a unique BglII site within the ccr coding region . By use of the ermE* promoter, the S . collinus ccr gene was expressed from plasmids in S . cinnamonensis C730 . 1/pHL18 and L1/pHL18 . CCR activity in mutant L1 was shown to decrease by more than 90% in both yeast extract-malt extract (YEME) medium and a complex fermentation medium, compared to that in wild-type C730.1 . Compared to C730.1, mutants C730.1/pHL18 and L1/pHL18 exhibited a huge increase in CCR activity (14- and 13-fold, respectively) in YEME medium and a moderate increase (3.7- and 2 . 7-fold, respectively) in the complex fermentation medium . In the complex fermentation medium, S . cinnamonensis L1 produced monensins A and B in a ratio of 12:88, dramatically lower than the 50:50 ratio observed for both C730.1 and C730.1/pHL18 . Plasmid (pHL18)-based expression of the S . collinus ccr gene in mutant L1 increased the monensin A/monensin B ratio to 42:58 . Labeling experiments with {1, 2-(13)C(2)}acetate demonstrated the same levels of intact incorporation of this material into the butyrate-derived portion of monensin A in both C730.1 and mutant C730.1/pLH18 but a markedly decreased level of such incorporation in mutant L1 . The addition of crotonic acid at 15 mM led to significant increases in the monensin A/monensin B ratio in C730.1 and C730.1/pHL18 but had no effect in S . cinnamonensis L1 . These results demonstrate that CCR plays a significant role in providing butyryl-CoA for monensin A biosynthesis and is present in wild-type S . cinnamonensis C730.1 at a level sufficient that the availability of the appropriate substrate (crotonyl-CoA) is limiting.

Nucleic Acids Res, 1999 Nov 15, 27(22), 4391 - 8
Expression of GUT1, which encodes glycerol kinase in Saccharomyces cerevisiae, is controlled by the positive regulators Adr1p, Ino2p and Ino4p and the negative regulator Opi1p in a carbon source-dependent fashion; Grauslund M et al.; In Saccharomyces cerevisiae glycerol utilization is mediated by two enzymes, glycerol kinase (Gut1p) and mitochondrial glycerol-3-phosphate dehydrogenase (Gut2p) . The carbon source regulation of GUT1 was studied using promoter-reporter gene fusions . The promoter activity was lowest during growth on glucose and highest on the non-fermentable carbon sources, glycerol, ethanol, lactate, acetate and oleic acid . Mutational analysis of the GUT1 promoter region showed that two upstream activation sequences, UAS(INO) and UAS(ADR1), are responsible for approximately 90% of the expression during growth on glycerol . UAS(ADR1) is a presumed binding site for the zinc finger transcription factor Adr1p and UAS(INO) is a presumed binding site for the basic helix-loop-helix transcription factors Ino2p and Ino4p . In vitro experiments showed Adr1 and Ino2/Ino4 protein-dependent binding to UAS(ADR1) and UAS(INO) . The negative regulator Opi1p mediates repression of the GUT1 promoter, whereas the effects of the glucose repressors Mig1p and Mig2p are minor . Together, the experiments show that GUT1 is carbon source regulated by different activation and repression systems.

J Gastroenterol Hepatol, 1999 Sep, 14(9), 880 - 8
Butyrate from bacterial fermentation of germinated barley foodstuff preserves intestinal barrier function in experimental colitis in the rat model; Kanauchi O et al.; BACKGROUND AND AIMS: The consumption of germinated barley foodstuff (GBF) prevents inflammation and diarrhoea in a colitis model . In this study we investigated the mechanism of the preventative effect of GBF on experimental colitis in rats, in view of production of bacterial butyrate and preservation of intestinal barrier function . METHODS: Sprague-Dawley rats administered with diets supplemented with 3.5% dextran sodium sulphate were used as an experimental colitis model . Butyrate was given to rats orally or intracaecally . Intestinal barrier function was estimated by light microscopic observation of the mucosa, intestinal permeability and bacterial translocation . RESULTS: Mucosal damage was reduced by intracaecal administration of butyrate, but not by oral administration . Bacterial butyrate production and reduction of mucosal damage depended on the dose of GBF in diets . The action of endogenous bacterial butyrate, including the reduction of intestinal permeability and bacterial translocation, was inhibited by administration of an inhibitor of beta-oxidation of short-chain fatty acids . CONCLUSIONS: The feeding of GBF promotes bacterial butyrate production and improves intestinal barrier function in rats, resulting in mitigation of experimental colitis.

Can J Vet Res, 1999 Oct, 63(4), 248 - 52
Evaluation of lansoprazole (an H+/K+-ATPase inhibitor) and azithromycin (an antibiotic) for control of gastric ulceration in swine during periods of feed deprivation; Melnichouk S et al.; Helicobacter-like organisms as well as fermentative bacteria have been implicated in gastric ulcer production in swine . Irregular feeding schedules are also considered a major risk factor . A research trial was conducted to determine whether medication with an acid secretion inhibitor (lansoprazole), either alone or in combination with an antibiotic (azithromycin), would protect pigs from gastric ulceration if the animals were subjected to a 48 h period of fasting . In a 2 x 3 factorial design, 48 pigs were fasted, while an equal number were fed ad libitum . Within these 2 study groups, pigs were randomly assigned to 1 of 3 treatments: control, 30 mg lansoprazole s.i.d . for 7 d, or lansoprazole (30 mg s.i.d . for 7 d) and azithromycin (500 mg s.i.d . for 3 d) . Overall, fasted pigs were 1.9 times more likely to develop erosive or ulcerative lesions of the pars esophagea (chi2 = 9.89, P < 0.002) . Treatment with an acid secretion inhibitor alone or in combination with an antibiotic did not protect pigs from developing gastric lesions . Helicobacter-like organisms were not detected in any of the stomachs . Possibly, the lansoprazole dose of 30 mg given once per day was insufficient to prevent pH levels from becoming low enough to cause damage to epithelial tissue . Alternatively other substances such as bile acids may have caused the ulcerative lesions, even though stomach acid production was suppressed.

Adv Biochem Eng Biotechnol, 1999, 65, 243 - 80
Production of multifunctional organic acids from renewable resources; Tsao GT et al.; Recently, the microbial production of multifunctional organic acid has received interest due to their increased use in the food industry and their potential as raw materials for the manufacture of biodegradable polymers . Certain species of microorganisms produce significant quantities of organic acids in high yields under specific cultivation conditions from biomass-derived carbohydrates . The accumulation of some acids, such as fumaric, malic and succinic acid, are believed to involve CO2-fixation which gives high yields of products . The application of special fermentation techniques and the methods for downstream processing of products are described . Techniques such as simultaneous fermentation and product recovery and downstream processing are likely to occupy an important role in the reduction of production costs . Finally, some aspects of process design and current industrial production processes are discussed.

Adv Biochem Eng Biotechnol, 1999, 65, 163 - 92
Successful design and development of genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose from cellulosic biomass to fuel ethanol; Ho NW et al.; Ethanol is an effective, environmentally friendly, nonfossil, transportation biofuel that produces far less pollution than gasoline . Furthermore, ethanol can be produced from plentiful, domestically available, renewable, cellulosic biomass . However, cellulosic biomass contains two major sugars, glucose and xylose, and a major obstacle in this process is that Saccharomyces yeasts, traditionally used and still the only microorganisms currently used for large scale industrial production of ethanol from glucose, are unable to ferment xylose to ethanol . This makes the use of these safest, most effective Saccharomyces yeasts for conversion of biomass to ethanol economically unfeasible . Since 1980, scientists worldwide have actively been trying to develop genetically engineered Saccharomyces yeasts to ferment xylose . In 1993, we achieved a historic breakthrough to succeed in the development of the first genetically engineered Saccharomyces yeasts that can effectively ferment both glucose and xylose to ethanol . This was accomplished by carefully redesigning the yeast metabolic pathway for fermenting xylose to ethanol, including cloning three xylose-metabolizing genes, modifying the genetic systems controlling gene expression, changing the dynamics of the carbon flow, etc . As a result, our recombinant yeasts not only can effectively ferment both glucose and xylose to ethanol when these sugars are present separately in the medium, but also can effectively coferment both glucose and xylose present in the same medium simultaneously to ethanol . This has made it possible because we have genetically engineered the Saccharomyces yeasts as such that they are able to overcome some of the natural barrier present in all microorganisms, such as the synthesis of the xylose metabolizing enzymes not to be affected by the presence of glucose and by the absence of xylose in the medium . This first generation of genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts relies on the presence of a high-copy-number 2 mu-based plasmid that contains the three cloned genetically modified xylose-metabolizing genes to provide the xylose-metabolizing capability . In 1995, we achieved another breakthrough by creating the super-stable genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts which contain multiple copies of the same three xylose-metabolizing genes stably integrated on the yeast chromosome . This is another critical development which has made it possible for the genetically engineered yeasts to be effective for cofermenting glucose and xylose by continuous fermentation . It is widely believed that the successful development of the stable glucose-xylose-cofermenting Saccharomyces yeasts has made the biomass-to-ethanol technology a step much closer to commercialization . In this paper, we present an overview of our rationales and strategies as well as our methods and approaches that led to the ingenious design and successful development of our genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose to biofuel ethanol.

Adv Biochem Eng Biotechnol, 1999, 65, 117 - 61
Genetic engineering for improved xylose fermentation by yeasts; Jeffries TW et al.; Xylose utilization is essential for the efficient conversion of lignocellulosic materials to fuels and chemicals . A few yeasts are known to ferment xylose directly to ethanol . However, the rates and yields need to be improved for commercialization . Xylose utilization is repressed by glucose which is usually present in lignocellulosic hydrolysates, so glucose regulation should be altered in order to maximize xylose conversion . Xylose utilization also requires low amounts of oxygen for optimal production . Respiration can reduce ethanol yields, so the role of oxygen must be better understood and respiration must be reduced in order to improve ethanol production . This paper reviews the central pathways for glucose and xylose metabolism, the principal respiratory pathways, the factors determining partitioning of pyruvate between respiration and fermentation, the known genetic mechanisms for glucose and oxygen regulation, and progress to date in improving xylose fermentations by yeasts.

Microbiol Immunol, 1999, 43(7), 711 - 6
Phylogenetic analysis of saccharolytic oral treponemes isolated from human subgingival plaque; Sakamoto M et al.; A total of 74 strains of oral treponemes, which were isolated from subgingival plaque samples from patients with periodontitis, were taxonomically studied on the basis of biochemical characteristics, DNA-DNA hybridization, and 16S rRNA gene sequences . These organisms fermented carbohydrates and required rumen fluid or short-chain volatile fatty acids for growth . The isolates were divided into seven subgroups based on their biochemical characteristics . The levels of DNA relatedness among the representative strains of each subgroup and Treponema socranskii (including three subspecies) were greater than 78%, while the levels of DNA relatedness among these strains and other Treponema species, including T . denticola and "T . vincentii", were less than 15% . DNA-DNA hybridization indicated that all subgroups belonged to T . socranskii . This result correlated well with the cluster on the phylogenetic trees based on 16S rRNA sequences.

Alcohol Alcohol, 1999 Sep-Oct, 34(5), 773 - 85
Patterns of alcohol consumption in the Seychelles Islands (Indian Ocean); Perdrix J et al.; Self-reported drinking habits were examined in a random sample of 1067 persons aged 25-64 years in the Seychelles, a country in epidemiological transition where consumption of home-brewed, mostly unregistered beverages has been traditionally high . Alcohol consumption was calculated from respondents reporting at least one drink per week ('regular drinkers') . Among men, 51.1% were regular drinkers and had average intake of 112.1 ml alcohol a day . Among women, 5.9% were regular drinkers and had 49.7 ml alcohol a day . Frequency of drinking, but not amount per drinker, was slightly less in the 25-34-year than older-age categories . Home-brews (mostly palm toddy and fermented sugar cane juice) were consumed by 52% of regular drinkers and accounted for 54% of the total alcohol intake reported by all regular drinkers . Based on the reported consumption by regular drinkers only, the average annual alcohol consumption amounted respectively to 20.7 litres and 1.2 litres per man and woman aged 25-64 years, or, using extrapolation, 13.2 litres and 0.8 litres per man and woman respectively of the total population . These values may underestimate the true figures by half, since reported beer consumption accounted for 53% of beer sales . Socio-economic status was associated strongly and inversely with home-brew consumption, but slightly and positively with consumption of commercially marketed beverages . Alcohol intake was associated with smoking, high-density lipoprotein cholesterol, carbohydrate-deficient transferrin and blood pressure, but not with age and body mass index . In conclusion, these data show high alcohol consumption in the Seychelles with an important gender difference, a large proportion of alcohol derived from home-brews, and opposite tendencies for the relationships between socio-economic status and home-made or commercially marketed beverages.

J Food Prot, 1999 Oct, 62(10), 1115 - 22
Occurrence and survival of verocytotoxin-producing Escherichia coli O157 in meats obtained from retail outlets in The Netherlands; Heuvelink AE et al.; In 1996 and 1997, 2,941 fresh and processed meat products obtained from supermarkets and butcher shops in The Netherlands were examined for the presence of verocytotoxin-producing Escherichia coli of serogroup O157 (O157 VTEC) . Additionally, the fate of O157 VTEC in raw meat products stored at low temperatures and the effect of different additives were evaluated . O157 VTEC strains were isolated from 6 (1.1%) of 571 samples of raw minced beef, 2 (0.5%) of 402 samples of raw minced mixed beef and pork, 1 (1.3%) of 76 samples of raw minced pork, 1 (0.3%) of 393 samples of other raw pork products, and 1 (0.3%) of 328 samples of cooked or fermented ready-to-eat meats . Other raw beef products (n = 223) and meat samples originating from poultry (n = 819), sheep or lamb (n = 46), or wild animals (n = 83) were all found to be negative for O157 VTEC . For the survival experiments we used tartaar (minced beef with a fat content of less than 10%) and filet americain (tartaar mixed with a mayonnaise-based sauce {80 to 20%}) . The O157 VTEC strain tested was able to survive in tartaar and filet americain stored at -20, 0, 5, or 7 degrees C for 3 days . At both 7 and at 15 degrees C, O157 VTEC counts in tartaar and filet americain remained virtually unchanged throughout a storage period of 5 days . Addition of acetic acid (to pH 4.0), sodium lactate (1 and 2% {wt/wt}), or components of the lactoperoxidase-thiocyanate-hydrogen peroxide system to filet americain did not result in a reduction of viable O157 VTEC cells during storage at 7 or 15 degrees C . It was concluded that raw meat contaminated with O157 VTEC will remain a hazard even if the meat is held at low or freezing temperatures.

Crit Rev Microbiol, 1999, 25(3), 229 - 43
Mycopesticide production by fermentation: potential and challenges; Deshpande MV; The agriculture industry is in need of novel biopesticides and development of large-scale production of mycopesticide, either fungal cells themselves or cell-free fungal components . The identification of a fungal strain with pesticide activity, and its improvement, is the primary step in developing infective propagules such as conidia, blastospores, chlamydospores, oospores, and zygospores as well as in preparing hydrolytic enzyme mixtures . This review discusses various parameters for submerged and solid state fermentation to produce fungal structures, particularly of mycoparasitic and entomopathogenic species that are prospective candidates for use as mycopesticides . The understanding of the molecular aspects of fungus-fungus and fungus-insect interactions, the role of hydrolytic enzymes especially chitinases in the killing process, and the possible use of chitin synthesis inhibitors are the prime areas of research aimed at making fungi more effective either singly or in combination as mycopesticides.

J Clin Microbiol, 1999 Nov, 37(11), 3491 - 6
Escherichia coli O157:H7 and O157:H(-) strains that do not produce Shiga toxin: phenotypic and genetic characterization of isolates associated with diarrhea and hemolytic-uremic syndrome; Schmidt H et al.; We have isolated one sorbitol-nonfermenting (SNF) Escherichia coli O157:H7 isolate and five sorbitol-fermenting (SF) E . coli O157:H(-) isolates that do not contain Shiga toxin (Stx) genes (stx) . Isolates originated from patients with diarrhea (n = 4) and hemolytic-uremic syndrome (HUS) (n = 2) . All isolates harbored a chromosomal eae gene encoding gamma-intimin as well as the plasmid genes E-hly and etp . The E . coli O157:H7 isolate was katP and espP positive . Respective sera obtained from the patient with HUS contained antibodies to the O157 lipopolysaccharide antigen . The stx-negative E . coli O157:H7 isolate is genetically related to stx-positive SNF E . coli O157:H7 . All stx-negative SF E . coli O157:H(-) isolates belong to the same genetic cluster and are closely related to stx-positive SF E . coli O157:H(-) isolates . Our data indicate that stx-negative E . coli O157:H7/H(-) variants may occur at a low frequency and cannot be recognized by diagnostic methods that target Stx.

Indian J Exp Biol, 1999 Jul, 37(7), 731 - 3
Production of calcium gluconate by fermentation; Tripathi CK et al.; Calcium gluconate production by Aspergillus niger was investigated in shake flask, rolling shaker, air-lift reactor and stirred reactor . Growth pattern of the organism and fermentation conditions determined the yield of the product . High calcium gluconate production was achieved in air-lift reactor with pellet form of cell growth at moderate specific growth rate and biomass concentration . In another variation of air-lift reactor, when calcium carbonate was confined to a cellulose membrane, calcium gluconate production was maximum (149 g/L) . At higher specific growth rate, obtained in shake flask, despite the formation of cell pellets, product formation was low . Physical separation of particulate calcium carbonate and growing cells favoured product formation . In stirred reactor pulpy mycelial growth was obtained and calcium gluconate production was poor.

J Biol Chem, 1999 Oct 22, 274(43), 30794 - 8
Involvement of small GTPases in Mycoplasma fermentans membrane lipoproteins-mediated activation of macrophages; Rawadi G et al.; Mycoplasma fermentans lipoproteins (LAMPf) are capable of activating macrophages and inducing the secretion of proinflammatory cytokines . We have recently reported that mitogen-activated protein kinase (MAPK) pathways and NF-kappaB and activated protein 1 (AP-1) play a crucial role in the activation induced by this bacterial compound . To further elucidate the mechanisms by which LAMPf mediate the activation of macrophages, we assessed the effects of inhibiting small G proteins Rac, Cdc42, and Rho . The Rho-specific inhibitor C3 enzyme completely abolished the secretion of tumor necrosis factor alpha by macrophages stimulated with LAMPf and also inhibited the activation of extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase (JNK), and p38 kinase . In addition, we have shown that LAMPf stimulate Cdc42 and that inhibition of Cdc42 or Rac by dominant negative mutants abrogates LAMPf-mediated activation of JNK and transactivation of NF-kappaB and AP-1 in the murine macrophage cell line RAW 264.7 . These results indicate that small G proteins Rho, Cdc42, and Rac are involved in the cascade of events leading to the macrophage activation by mycoplasma lipoproteins.

J Anim Sci, 1999 Oct, 77(10), 2817 - 23
Feeding value of wheat-based thin stillage: in vitro protein degradability and effects on ruminal fermentation; Iwanchysko P et al.; Two experiments were conducted to evaluate the nutritive value of wheat-based thin stillage as a fluid source for ruminants . In vitro CP degradability of thin stillage was estimated relative to canola meal and heated canola meal in a completely randomized design . Four ruminally cannulated steers were used in a double cross-over design to determine the effects of consuming thin stillage or water as drinking sources on ruminal fermentation traits . The in vitro CP degradability of thin stillage (55.4%) was lower (P<.05) than that of canola meal (59.4%) and higher than that of heated canola meal (31.6%) . Ruminal pH for steers consuming thin stillage was higher (P<.05) at 1000 and 1100 and lower (P<.05) at 1900 and 2000 than that for steers consuming water . Total VFA followed a pattern that was the reverse of that reported for pH . Ruminal NH3 N levels were higher (P<.05) for steers fed thin stillage than for water-fed steers through most of the collection period . Ruminal fluid and particulate matter passage rates were not affected by treatment and averaged .165 and .06 /h, respectively . The amount of thin stillage and water that did not equilibrate with the ruminal fluid and, thus, was considered to bypass the rumen was estimated to be 51.9 and 59.2% of total fluid consumed, respectively . Feeding wheat-based thin stillage had no adverse effects on ruminal metabolism.

J Anim Sci, 1999 Oct, 77(10), 2793 - 802
Effects of different supplemental sugars and starch fed in combination with degradable intake protein on low-quality forage use by beef steers; Heldt JS et al.; Twenty ruminally fistulated steers (Exp . 1, 448 kg and Exp . 2, 450 kg) were used in two consecutive randomized complete block experiments with five treatments in each experiment . The purpose was to evaluate the impact of feeding different supplemental sugars or starch in combination with supplemental degradable intake protein (DIP) on the utilization of low-quality tallgrass-prairie hay . In Exp . 1, steers were given ad libitum access to forage and, except for the negative control (NC), received a limited supply (insufficient to maximize forage use) of supplemental DIP (.031% BW/d, DM basis) . In addition to the NC, this experiment included four supplementation treatments in which one of four carbohydrate (CHO) sources (starch, glucose, fructose, or sucrose) was fed at .30% BW of DM/d . In Exp . 2, the treatment structure was identical except that the supplemental DIP level (.122% BW, DM basis) was near the level needed to maximize forage use . Forage OM intake (FOMI) was not affected (P> or =.26) by supplementation in Exp . 1 but was increased (P = .05) in Exp . 2 . However, no difference (P> or =.46) in FOMI occurred among CHO sources in either experiment . Total OM and digestible OM intakes were increased (P<.01) by supplementation in both experiments . In Exp . 1, no difference (P>.26) in OM digestion (OMD) occurred among treatments . In Exp . 2, supplementation increased (P<.01) OMD . Additionally, sugars yielded a higher (P = .04) OMD than starch, and the monosaccharides yielded a higher (P = .02) OMD than sucrose . In Exp . 1, NDF digestion (NDFD) was decreased (P = .02) by supplementation, but no differences (P> or =.21) occurred among CHO sources . In Exp . 2, NDFD was increased (P = .03) by supplementation . Additionally, sugars led to higher (P = .05) NDFD than starch, and the monosaccharides led to higher (P = .03) NDFD than sucrose . In both experiments, discernible patterns were observable with regard to the effects of supplementation and type of supplemental CHO on ruminal fermentation characteristics . In conclusion, even though some consistency in fermentation profiles for different carbohydrate sources was evident in both experiments, forage intake and digestion responses were not consistent across experiments . This raises the possibility that carbohydrate source may interact with the amount of supplemental DIP fed and, as such, deserves additional investigation.

J Anim Sci, 1999 Oct, 77(10), 2781 - 92
Influence of postruminal supplementation of methionine and lysine, isoleucine, or all three amino acids on intake and chewing behavior, ruminal fermentation, and milk and milk component production; Robinson PH et al.; Four multiparous Holstein cows were fed a basal diet balanced with the Cornell Net Protein and Carbohydrate System (CNCPS) . Diets were formulated to be co-limiting in intestinally absorbable supplies of methionine, lysine, and isoleucine . Cows were supplemented with no amino acids (control); lysine and methionine in a ruminally protected form; isoleucine by abomasal infusion; or lysine, methionine, and isoleucine in a 4x4 Latin square arrangement of treatments with 28-d periods . Performance of cows on all treatments was lower than expected due to low intake of DM that could have been caused by the high fiber level of the basal diet . This high fiber level was likely responsible for the high daily chewing times for cows fed all diets, which was consistent with the high ruminal pH values . Intake of DM and its components were not influenced by any treatment . Milk protein percentage tended to be higher when cows were fed diets supplemented with ruminally protected lysine and methionine; however, production of milk, milk fat, and milk lactose were not affected by any treatment . Cows tended to have a higher milk lactose proportion and tended to produce more milk and milk lactose when they were abomasally infused with isoleucine alone . However, when cows were supplemented with all three amino acids, milk production and composition did not differ from that of cows fed the unsupplemented diet . Use of the CNCPS to evaluate the performance of the cows fed the unsupplemented diet suggested that these cows may have been colimited by intestinally absorbable supplies of lysine, isoleucine, and methionine in addition to metabolizable protein . Evaluation of the unsupplemented diet with an alternate model, Shield, suggested that cows fed the unsupplemented diet may have been colimited by intestinally absorbable supplies of lysine, isoleucine, and arginine . Results suggest that enhanced delivery of intestinally absorbable isoleucine may stimulate milk lactose synthesis.

J Anim Sci, 1999 Oct, 77(10), 2774 - 80
Low- and high-quality forage utilization by heifers and mature beef cows; Varel VH et al.; Eight cows (7 to 9 yr old, 522 kg) and six heifers (10 mo old, 169 kg) were fed either alfalfa hay (18.7% CP) or mature brome hay (5.1% CP) to determine the effect of cattle age on apparent forage utilization . Cattle were fitted with ruminal and duodenal cannulas and were individually fed once daily (ad libitum intake, 1000) . The split-plot design consisted of age (whole-plot) and two sampling periods feeding alfalfa or brome hay (subplot) . Each period consisted of 28 d: d 1 to 13 for adaptation, d 13 to 20 for feed intake determination, and d 20 to 28 for sampling . Nylon bags containing NDF substrate from alfalfa or brome hay were incubated ruminally for 0, 3, 6, 12, 24, 48, 96, and 192 h to determine the rate and extent of fiber degradation . Ruminal liquid dilution rate and fermentation characteristics were conducted on d 27 . Ruminal fill was determined by total evacuation at 0800 on d 28 . Cows consumed more feed (BW.75; P<.01) and had greater ruminal OM fill (P = .04) but had similar fluid fill (P = .88) compared with heifers . Ruminal liquid dilution rate was greater in cows than in heifers (P<.01) . The rate of in situ NDF degradation was 3 and .5% per hour greater in cows than in heifers when alfalfa and brome hay were fed, respectively (age x hay, P<.01) . Ruminal NDF digestibility as a percentage of intake was greater in cows than in heifers (P<.01) . Numbers of ruminal cellulolytic bacteria were not affected by treatment (P>.21) . These data indicate that mature cows have a smaller ruminal fluid fill that turns over more rapidly, and this may be responsible for a faster rate of ruminal fiber degradation in cows than in young heifers.

J Anim Sci, 1999 Oct, 77(10), 2721 - 9
Changes in gastric contents in pigs fed a finely ground and pelleted or coarsely ground meal diet; Regina DC et al.; The objective was to characterize the change in stomach contents in relation to time after feeding between pigs consuming a restricted amount of a finely ground and pelleted (FGP) or coarsely ground meal (CGM) diet . Particular interest was placed on the concentration of organic acids and ammonia, the products of microbial fermentation . Thirty barrows were ranked by weight and assigned to a postfeeding time of 2, 4, 6, 8, or 12 h and either the FGP or CGM diet . Initiation and termination of the experiment were staggered over a 2-wk period . The treatment period was 42 d . Percentage of dry matter was higher (P<.01) in the stomach contents of pigs on the CGM diet . Concentrations of pepsin and protein were higher (P<.05) and ammonia tended to be higher (P = .10) in the proximal stomach of pigs fed the FGP diet . In contrast, concentrations of acetate and L-lactate were higher (P<.05) in the proximal stomach of pigs fed the CGM diet . All pigs on the CGM diet had stomachs that graded as normal on visual inspection . There was variable damage to the stomachs of pigs on the FGP diet . Measurement of chromium concentration in the stomach after an oral dose of Cr-EDTA clearly demonstrated the mixing that occurs between the proximal and distal stomach by 2 h after feeding in pigs consuming the FGP diet, whereas a gradient was maintained in pigs consuming the CGM diet . Thus, components normally secreted in the distal stomach return to the proximal stomach . These data show that components secreted in the distal region, such as acid and pepsin, may play a role in initiating damage to the stratified squamous mucosa . High concentrations of organic acids in the stomach of pigs on the CGM diet were not associated with damage to the stratified squamous mucosa in the esophageal region.

Curr Med Chem, 1999 Dec, 6(12), 1197 - 212
Chemistry and structure activity relationships of bafilomycin A1, a potent and selective inhibitor of the vacuolar H+-ATPase; Gagliardi S et al.; Bafilomycin A1, a macrolide antibiotic isolated from the fermentation of Streptomyces spp., is a potent and selective inhibitor of vacuolar-type proton translocating ATP-ases (V-ATPases) and was used to study the physiological role of this class of enzymes . An extensive chemical effort on the unusual structure of this macrolide led to the synthesis of significantly different bafilomycin derivatives . None of the new analogues was more potent than the parent compound but provided a significant amount of information about the structural requirements for the inhibitory activity of bafilomycin A1 in particular on chicken osteoclast (cOc) ATPase . The vinylic methoxy group adjacent to a carbonyl function, the dienic system and the hydroxy group at position 7 were recognized to be essential features for bafilomycin V-ATPase-inhibitory activity . This information was utilized to design simplified novel derivatives as inhibitors of bone resorption.

Microbiology, 1999 Sep, 145 ( Pt 9), 2569 - 76
Oxalic acid production by Aspergillus niger: an oxalate-non-producing mutant produces citric acid at pH 5 and in the presence of manganese; Ruijter GJ et al.; The external pH appeared to be the main factor governing oxalic acid production by Aspergillus niger . A glucose-oxidase-negative mutant produced substantial amounts of oxalic acid as long as the pH of the culture was 3 or higher . When pH was decreased below 2, no oxalic acid was formed . The activity of oxaloacetate acetylhydrolase (OAH), the enzyme believed to be responsible for oxalate formation in A . niger, correlated with oxalate production . OAH was purified from A . niger and characterized . OAH cleaves oxaloacetate to oxalate and acetate, but A . niger never accumulated any acetate in the culture broth . Since an A . niger acuA mutant, which lacks acetyl-CoA synthase, did produce some acetate, wild-type A . niger is apparently able to catabolize acetate sufficiently fast to prevent its production . An A . niger mutant, prtF28, previously isolated in a screen for strains deficient in extracellular protease expression, was shown here to be oxalate non-producing . The prtF28 mutant lacked OAH, implying that OAH is the only enzyme involved in oxalate production in A . niger . In a traditional citric acid fermentation low pH and absence of Mn2+ are prerequisites . Remarkably, a strain lacking both glucose oxidase (goxC) and OAH (prtF) produced citric acid from sugar substrates in a regular synthetic medium at pH 5 and under these conditions production was completely insensitive to Mn2+.

Int J Biol Macromol, 1999 Nov, 26(2-3), 201 - 11
Characterization by mass spectrometry of poly(3-hydroxyalkanoates) produced by Rhodospirillum rubrum from 3-hydroxyacids; Ballistreri A et al.; The sequence distributions of two microbial copolyesters obtained by fermentation of Rhodospirillum rubrum, grown with 3-hydroxyhexanoic or 3-hydroxyheptanoic acids, were determined by analyzing the oligomers prepared by partial pyrolysis or partial methanolysis of these copolyesters using fast atom bombardment mass spectrometry (FAB-MS) . Oligomers up to pentamers were identified in the case of partial pyrolysis and up to tetradecamers in the case of partial methanolysis . The comparison between the experimental and calculated peak intensities of FAB mass spectra allows the calculation of compositions and sequence distributions, which in these copolyesters follow Bernoullian statistics, indicating that they are random terpolyesters.

Plant Foods Hum Nutr, 1999, 53(3), 199 - 208
Studies on the production of bambara groundnut (Vigna subterranea) tempe; Amadi EN et al.; Bambara groundnut, an indigenous African legume, was subjected to fermentation by three strains of Rhizopus . One strain B . arrhizus could not ferment the substrate . Mycelial penetration and binding was good when strains NRRL 2710 (R . oligosporus) and NRRL 1477 (R . stolonifer) were used . Fermentation by both strains resulted in increases of pH, moisture, protein and fat while total carbohydrate decreased by 50% . Sensory evaluation showed that bambara groundnut tempes rated similar (p>0.5) in taste and texture and higher (p<0.05) in color and flavor than soybean tempe . Bambara groundnut would be an acceptable food product in the diet as a good protein supplement.

Biotechnol Bioeng, 1999 Dec 5, 65(5), 550 - 7
Visualizing integrated bioprocess designs through "windows of operation"; Zhou YH et al.; This paper demonstrates a simple graphical approach for the design and analysis of a bioprocess flowsheet in which process interactions are significant . Results are presented showing how the feasible space for operation can be simulated and used both to address key design and operating decisions and to identify suitable trade-offs between operating variables, such as fermentation growth rate and disruption conditions, in order to achieve prespecified levels of process performance . Using verified models to describe the production and isolation of an intracellular protein alcohol dehydrogenase (ADH) in yeast as a test bed, a series of so-called "windows of operation" are developed at growth rates in the range of 0.06-0.28 h(-1) and for a range of overall process specifications . The effects of altering the process design performance specification as defined by the level of cell debris removal and the overall process productivity on the size and position of the feasible space were investigated to demonstrate the sensitivity of the flowsheet to changes in process objectives . Using the approach it has been possible to visualise the processing trade-offs required to increase performance in terms of the level of cell debris removal by 50% and the overall process productivity by 400% from a defined base level . The approach provides a convenient tool when designing integrated bioprocesses by enabling process options to be compared visually and can help in achieving better process designs and accelerating process development for the biological process industry .

Biotechnol Bioeng, 1999 Dec 5, 65(5), 537 - 41
Dielectric analysis for estimation of oil content in the mycelia of Mortierella alpina; Higashiyama K et al.; The dielectric behavior of the filamentous fungi Mortierella alpina SAM2104 and 1S-4, which produce polyunsaturated fatty acid enriched oil in the mycelia, was investigated . During the cultivation carried out in a 10-kL fermentor for 12-15 days, the relative permittivity and conductivity of the broth were measured in the frequency range of 100 kHz to 30 MHz . The dielectric parameters, i.e., the amplitude of dielectric relaxation (Deltaepsilon) and the characteristic frequency (f(c)), were obtained by fitting the Cole-Cole equation to the observed dielectric relaxation, and the conductivity of the medium (kappa(a)) was also measured . The value of Deltaepsilon gradually increased from the second day through the end of cultivation, suggesting that volume fraction of the cell increased with oil accumulation . The conductivity of the cytoplasm (kappa(i)) was calculated from the experimental values of f(c) and kappa(a), using a theoretical equation based on an ellipsoidal cell model . As a result, good correlation between the calculated kappa(i) and the oil content was obtained . These findings indicate that dielectric analysis enables us to estimate the oil content in the mycelia of oleaginous fungi and also provides a useful tool for monitoring cell growth and for controlling the cultivation process .

Biotechnol Bioeng, 1999 Dec 5, 65(5), 512 - 22
Macroscopic growth of filamentous fungi on solid substrate explained by a microscopic approach; Ferret E et al.; A quantitative model predicting biomass growth on solid media has been developed . The model takes into account steric interactions between hyphae and tips at the microscopic level (competition for substrate and tip-hypha collisions) . These interactions effect a slowing down of the hyphal, population-averaged extension rate and are responsible, at the microscopic level, for the distribution of tip orientations observed at the colony border . At the macroscopic level, a limiting value of the colony radial extension rate is attained . A mathematical model that combines hyphal branching, tip diffusion, and biomass growth was proposed to explain such behavior . Experiments using Gibberella fujikuroi were performed to validate the model; good agreement between experiments and simulations was achieved . Most parameters can be measured by simple image analysis on the peripheral growth zone, and they have clear physical meaning; that is, they correspond to properties of single, leading hyphae . The model can be used to describe two-dimensional (2D) solid media fermentation experiments under varying culture conditions; the model can also be extended to consider growth in three-dimensional (3D), complex geometry substrates .

Biotechnol Bioeng, 1999 Dec 5, 65(5), 500 - 11
Influence of fermentation conditions and microfiltration processes on membrane fouling during recovery of glucuronane polysaccharides from fermentation broths; Harscoat C et al.; We have investigated the recovery of exopolysaccharides produced by Sinorhizobium meliloti M5N1 CS bacteria from fermentation broths using different membrane filtration processes: cross-flow filtration with a 7 mm i.d . tubular ceramic membrane of 0.5-microm pores under fixed transmembrane pressure or fixed permeate flux and dynamic filtration with a 0.2 microm nylon membrane using a 16-cm rotating disc filter . With the tubular membrane, the polysaccharide mass flux was mainly limited by polymer transmission that decayed to 10% after 90 min . The mass flux of polymer produced under standard fermentation conditions (70 h at 30 degrees C) stabilized after 70 min to 15 g/h/m(2) . This mass flux rises to 36 g/h/m(2) when the mean stirring speed during fermentation is increased and to 123 g/h/m(2) when fermentation is extended to 120 h . In both cases, the mean molecular weight of polysaccharides drops from 4.0 10(5) g/mol under standard conditions to 2.7 10(5) g/mol . A similar reduction in molecular weight was observed when the fermentation temperature was raised to 36 degrees C without benefit to the mass flux . These changes in fermentation conditions have little effect on stabilized permeate flux, but raise significantly the sieving coefficient, due probably to molecular weight reduction and the filamentous aspect of the polymer as observed from SEM photographs . The polymer-mass flux was also increased by reducing transmembrane pressure (TMP) and raising the shear rate by inserting a rod in the membrane lumen . Operation under fixed permeate flux instead of constant TMP inhibited fouling during the first 4 h, resulting in higher sieving coefficients and polymer mass fluxes . The most interesting results were obtained with dynamic filtration because it allows operation at high-shear rates and low TMP . Sieving coefficients remained between 90 and 100% . With a smooth disc, the polysaccharide mass flux remained close to 180 g/h/m(2) at 1500 rpm and cell concentrations from 1 to 3 g/L . When radial rods were glued to the disc to increase wall shear stress and turbulence, the mass flux rose to 275 g/h/m(2) at the same speed and cell concentration .

Yeast, 1999 Oct, 15(14), 1555 - 64
Lipid composition of subcellular membranes of an FY1679-derived haploid yeast wild-type strain grown on different carbon sources; Tuller G et al.; The aim of the project EUROFAN (European Functional Analysis Network) is to elucidate the function of unknown genes of the yeast Saccharomyces cerevisiae at a large scale . Functional analysis is based on general and specific tests with yeast deletion strains . A prerequisite for these studies is a profound knowledge of the biochemistry and cell biology of the corresponding wild-type strain FY1679 . As a contribution from our laboratory we present here a systematic lipid analysis of the major organelles isolated from FY1679 grown in the presence of different carbon sources . Phospholipid, sterol and fatty acid composition are characteristic for each organelle . Moreover, growth of the yeast on glucose, ethanol or lactate causes in some cases marked changes of the organelle lipid pattern . As the most prominent example, cultivation of the yeast on non-fermentable carbon sources results in an increase of mitochondrial cardiolipin . As another example, the ratio of unsaturated to saturated fatty acids is enhanced in cells grown on ethanol or lactate as compared to glucose . Thus, the lipid composition of yeast subcellular membranes reflects in a significant way the nutrient conditions caused by variation of the carbon source .

Biotechnol Prog, 1999 Oct 1, 15(5), 949 - 952
Simultaneous Optimization of Feeding Rate and Operation Parameters for Fed-Batch Fermentation Processes; Wang FS et al.; An efficient method is introduced for simultaneously determining optimal polices of feeding rate and operation parameters for a fermentation process . Such an optimization problem is converted into the finite dimensional optimization problem using the control parametrization technique . The hybrid differential evolution is introduced to solve the converted problem . The optimal production rate obtained by the simultaneous optimization approach could be significantly improved with comparison to a simplified optimization problem, which is considered the optimal feed control only, as observed from the simulation results.

Biotechnol Prog, 1999 Oct 1, 15(5), 923 - 927
Estimation of O(2) and CO(2) Solubility in Microbial Culture Media; Gros JB et al.; A simple model for predicting gas solubilities of O(2) and CO(2) at low pressure and near ambient temperature in solutions of salts, sugars, and organic solvents (alcohols, ketones, ethers, aldehydes, etc.) is proposed . It is derived from the Van Laar assumptions and takes account of size differences between molecules in solution by their volumetric fraction . It is a group contribution model where anions and cations are considered as groups and other molecules are treated as in the UNIFAC (UNIQUAC Functional group Activity Coefficients) procedure . Pseudo-Henry's constants for groups were determined using solubility data in aqueous solutions containing only one salt, one sugar, or one organic compound at 25 degrees C . The predictive performance of the model was evaluated by comparison with experimental data using multicomponent aqueous salt-sugar mixtures . The model was used to estimate solubilities of oxygen and carbon dioxide in fermentation media.

Biotechnol Prog, 1999 Oct 1, 15(5), 911 - 918
The Effect of Rapeseed Oil Uptake on the Production of Erythromycin and Triketide Lactone by Saccharopolyspora erythraea; Mirjalili N et al.; Saccharopolyspora erythraea was grown in an oil-based process medium at two different laboratory scales . The initial concentration of rapeseed oil in the medium was shown not to affect the growth, while addition of oil significantly increased erythromycin A production . Increasing the agitation speed at the 2 L scale increased the growth of the culture and the production of erythromycin A but had little effect on the level of oil remaining in the fermentation . Maximum oil utilization of 50% (w/w) was obtained in both 2 and 7 L cultures . Gas chromatography analysis of the process medium showed that there was no accumulation of fatty acids and glycerides during the fermentations . The specific oil utilization by the recombinant strain of S . erythraea was significantly lower compared to the wild-type strain grown at the same scale and initial oil concentration; the recombinant strain also produced lower concentrations of the novel polyketide, triketide lactone.

Biotechnol Prog, 1999 Sep-Oct, 15(5), 876 - 83
Microbial synthesis of 3-dehydroshikimic acid: a comparative analysis of D-xylose, L-arabinose, and D-glucose carbon sources; Li K et al.; 3-Dehydroshikimic acid is a hydroaromatic precursor to chemicals ranging from L-phenylalanine to adipic acid . The concentration and yield of 3-dehydroshikimic acid microbially synthesized from various carbon sources has been examined under fed-batch fermentor conditions . Examined carbon sources included D-xylose, L-arabinose, and D-glucose . A mixture consisting of a 3:3:2 molar ratio of glucose/xylose/arabinose was also evaluated as a carbon source to model the composition of pentose streams potentially resulting from the hydrolysis of corn fiber . Escherichia coli KL3/pKL4.79B, which overexpresses feedback-insensitive DAHP synthase, synthesizes higher concentrations and yields of 3-dehydroshikimic acid when either xylose, arabinose, or the glucose/xylose/arabinose mixture is used as a carbon source relative to when glucose alone is used as a carbon source . E . coli KL3/pKL4.124A, which overexpresses transketolase and feedback-insensitive DAHP synthase, synthesizes higher concentrations and yields of 3-dehydroshikimic acid when the glucose/xylose/arabinose mixture is used as the carbon source relative to when either xylose or glucose is used as a carbon source . Observed high-titer, high-yielding synthesis of 3-dehydroshikimic acid from the glucose/xylose/arabinose mixture carries significant ramifications relevant to the employment of corn fiber in the microbial synthesis of value-added chemicals.

Biotechnol Prog, 1999 Oct 1, 15(5), 845 - 854
Catalytic Upgrading of Fermentation-Derived Organic Acids; Varadarajan S et al.; The production of organic acids in low-cost, high-efficiency fermentation processes makes available a new route to chemical production from biomass . Because of their multiple functional groups and high reactivity, organic acids can undergo a variety of reactions that are effectively catalyzed by inorganic heterogeneous or homogeneous catalysts . Lactic acid and succinic acid, in particular, are approaching large-scale production via fermentation and show excellent promise as feedstocks for catalytic conversion routes such as hydrogenation, dehydration, or condensation . A number of catalytic conversion pathways of organic acids are potentially competitive with petroleum-based routes in the current economic environment, particularly when integrated into existing biomass/crop processing schemes . This article reviews some of the key reaction pathways available using fermentation-derived organic acids as feedstocks and presents recent results from the authors' lab on succinate hydrogenation to 1,4-butanediol and tetrahydrofuran . By a judicious choice of support properties and reaction conditions, it is possible to achieve yields of either of these two products in excess of 80%.

Biotechnol Prog, 1999 Oct 1, 15(5), 777 - 793
Biocommodity Engineering; Lynd LR et al.; The application of biotechnology to the production of commodity products (fuels, chemicals, and materials) offering benefits in terms of sustainable resource supply and environmental quality is an emergent area of intellectual endeavor and industrial practice with great promise . Such "biocommodity engineering" is distinct from biotechnology motivated by health care at multiple levels, including economic driving forces, the importance of feedstocks and cost-motivated process engineering, and the scale of application . Plant biomass represents both the dominant foreseeable source of feedstocks for biotechnological processes as well as the only foreseeable sustainable source of organic fuels, chemicals, and materials . A variety of forms of biomass, notably many cellulosic feedstocks, are potentially available at a large scale and are cost-competitive with low-cost petroleum whether considered on a mass or energy basis, and in terms of price defined on a purchase or net basis for both current and projected mature technology, and on a transfer basis for mature technology . Thus the central, and we believe surmountable, impediment to more widespread application of biocommodity engineering is the general absence of low-cost processing technology . Technological and research challenges associated with converting plant biomass into commodity products are considered relative to overcoming the recalcitrance of cellulosic biomass (converting cellulosic biomass into reactive intermediates) and product diversification (converting reactive intermediates into useful products) . Advances are needed in pretreatment technology to make cellulosic materials accessible to enzymatic hydrolysis, with increased attention to the fundamental chemistry operative in pretreatment processes likely to accelerate progress . Important biotechnological challenges related to the utilization of cellulosic biomass include developing cellulase enzymes and microorganisms to produce them, fermentation of xylose and other nonglucose sugars, and "consolidated bioprocessing" in which cellulase production, cellulose hydrolysis, and fermentation of soluble carbohydrates to desired products occur in a single process step . With respect to product diversification, a distinction is made between replacement of a fossil resource-derived chemical with a biomass-derived chemical of identical composition and substitution of a biomass-derived chemical with equivalent functional characteristics but distinct composition . The substitution strategy involves larger transition issues but is seen as more promising in the long term . Metabolic engineering pursuant to the production of biocommodity products requires host organisms with properties such as the ability to use low-cost substrates, high product yield, competitive fitness, and robustness in industrial environments . In many cases, it is likely to be more successful to engineer a desired pathway into an organism having useful industrial properties rather than trying to engineer such often multi-gene properties into host organisms that do not have them naturally . Identification of host organisms with useful industrial properties and development of genetic systems for these organisms is a research challenge distinctive to biocommodity engineering . Chemical catalysis and separations technologies have important roles to play in downstream processing of biocommodity products and involve a distinctive set of challenges relative to petrochemical processing . At its current nascent state of development, the definition and advancement of the biocommodity field can benefit from integration at multiple levels . These include technical issues associated with integrating unit operations with each other, integrating production of individual products into a multi-product biorefinery, and integrating biorefineries into the broader resource, economic, and environmental systems in which they function . We anticipate that coproduction of multiple products, for example, production of fuels, chemicals, power, and/or feed, is likely to be essential for economic viability . Lifecycle analysis is necessary to verify the sustainability and environmental quality benefits of a particular biocommodity product or process . We see biocommodity engineering as a legitimate focus for graduate study, which is responsive to an established personnel demand in an industry that is expected to grow in the future . Graduate study in biocommodity engineering is supported by a distinctive blend of intellectual elements, including biotechnology, process engineering, and resource and environmental systems.

J Antibiot (Tokyo), 1999 Jul, 52(7), 607 - 12
TMC-66, a new endothelin converting enzyme inhibitor produced by Streptomyces sp . A5008; Asai Y et al.; A new endothelin converting enzyme (ECE) inhibitor, TMC-66 was isolated from the fermentation broth of Streptomyces sp . A5008 . The structure of TMC-66 was elucidated by spectroscopic analyses to be a new member of benzo{a}naphthacenequinone class of antibiotics . TMC-66 had a highly selective inhibitory activity for ECE with an IC50 value of 2.9 microM . Taxonomy of the producing strain is also described.

AIDS, 1999 Oct 1, 13(14), 1905 - 12
Association of indicators of bacterial vaginosis with a female genital tract factor that induces expression of HIV-1; Olinger GG et al.; OBJECTIVE: The aim of this study was to determine the relationship of bacterial vaginosis and bacterial vaginosis-associated microorganisms with an HIV-inducing factor (HIF) found in cervicovaginal lavage . DESIGN: A total of 26 cervicovaginal lavage specimens collected from 17 women were used in this study to determine if HIF was significantly associated with features consistent with bacterial vaginosis . METHODS: Patients were evaluated for various clinical features including age, HIV status and stage, CD4 cell counts, clinical diagnosis of gynecological infections, vaginal pH, Gram stains of vaginal fluid, phase of menstruation, and presence of cervical dysplasia . Cervicovaginal lavage specimens were analyzed for the presence of HIF by U1 bioassay . The presence of Gardnerella vaginalis, and general Mycoplasmataceae, and specifically Mycoplasma hominis, Ureaplasma urealyticum, M . fermentans, M . genitalium in cervicovaginal lavage were determined by semiquantitative PCR . RESULTS: Eleven cervicovaginal lavage samples from seven women were HIF-positive and 15 cervicovaginal lavage samples from 11 women were HIF-negative (patient No . 8 had two HIF-negative cervicovaginal lavage and one HIF-positive cervicovaginal lavage) . The following parameters were significantly associated with HIF: abnormal vaginal fluid pH (>4.5) (P = 0.006), Gram stains indicative of bacterial vaginosis (P = 0.007), normal menstrual cycle (P = 0.0007) and PCR detection and relative quantity of M . hominis (P = 0.0003, P = 0.002) . CONCLUSIONS: This study indicates that HIF is closely associated with features of bacterial vaginosis.

Eksp Klin Farmakol, 1999 Jul-Aug, 62(4), 59 - 61
{The effect of altan on the functional activity of the liver mitochondria and microsomes from rats with toxic hepatitis}; Gordienko AD et al.; In in vitro experiments althan had no effect on the respiration of intact mitochondria in state 4 according to Chance and produced a high antioxidant effect in fermentative and ascorbate-dependent lipid peroxidation in intact microsomes isolated from the rat liver . In ethanol-induced toxic hepatitis althan restored the functional activity of mitochondria to the level of that in intact animals and increased microsome hydroxylase activity in CCl4-hepatitis.

J Dairy Sci, 1999 Sep, 82(9), 2011 - 6
Effects of a Saccharomyces cerevisiae culture on in vitro mixed ruminal microorganism fermentation; Sullivan HM et al.; Previous research has shown that Saccharomyces cerevisiae culture increases lactate utilization and cellulose digestion by pure cultures of ruminal bacteria . Based on these pure culture results, in vitro mixed ruminal microorganism fermentations were conducted to determine the effects of 0.35 and 0.73 g/L of Sacc . cerevisiae culture on the fermentation of ground corn, maltose, alfalfa hay, bermudagrass hay, and lactate . In addition, experiments were performed to evaluate the effects of Sacc . cerevisiae culture and monensin on the mixed ruminal microorganism fermentation . In the presence of ground corn, both concentrations of Sacc . cerevisiae culture had little effect on final pH or fermentation products, except the 0.35 g/L treatment increased valerate concentration . Saccharomyces cerevisiae culture had little effect on final pH or fermentation products in maltose or lactate fermentations . When alfalfa hay was the substrate, 0.73 g/L of Sacc . cerevisiae culture increased propionate concentration and both treatments decreased the acetate to propionate ratio . In the case of Coastal bermudagrass hay, 0.73 g/L Sacc . cerevisiae culture increased concentrations of acetate, propionate, CH4, butyrate, isovalerate, valerate, and decreased the acetate to propionate ratio, whereas both treatments increased total volatile fatty acid concentrations . Similar to alfalfa hay, in vitro dry matter disappearance of Coastal bermudagrass hay was numerically increased in the presence of Sacc . cerevisiae culture . Monensin altered the fermentation by decreasing concentrations of CH4 and lactate and increasing concentrations of propionate . There was no interaction between Sacc . cerevisiae culture and monensin . In conclusion, the incorporation of Sacc . cerevisiae culture into mixed ruminal microorganism fermentations of ground corn, maltose, or lactate had little effect on final pH and fermentation products . However, in the presence of alfalfa hay or Coastal bermudagrass hay Sacc . cerevisiae culture increased concentrations of several fermentation products and numerically increased in vitro dry matter disappearance of forage fiber.

J Dairy Sci, 1999 Sep, 82(9), 1950 - 9
Invited review: summary of steam-flaking corn or sorghum grain for lactating dairy cows; Theurer CB et al.; Nineteen lactation trials (43 grain processing comparisons) are summarized, in addition to digestibility and postabsorptive metabolism studies . The net energy for lactation (NEL) of steam-flaked corn or sorghum grain is about 20% greater than the NEL for dry-rolled corn or sorghum . Based on lactational performance, steam-flaked sorghum grain is of equal value to steam-flaked corn, and steam-flaked corn is superior to steam-rolled corn . Steam-flaking of corn or sorghum compared to steam-rolling of corn or dry-rolling of corn or sorghum consistently improves milk production and milk protein yield . This result is because of a much greater proportion of dietary starch fermented in the rumen, enhanced digestibility of the smaller fraction of dietary starch reaching the small intestine, and increased total starch digestion . Steam-flaking increases cycling of urea to the gut, microbial protein flow to the small intestine, and estimated mammary uptake of amino acids . Steam-rolling compared to dry-rolling of barley or wheat did not alter total starch digestibilities in two trials, one with each grain source . Lactation studies with these processing comparisons have not been reported . Most cited studies have been with total mixed rations (TMR) and alfalfa hay as the principal forage . Additional studies are needed with lactating cows fed steam-flaked corn or sorghum in TMR containing alfalfa or corn silage . Optimal flake density of steam-processed corn or sorghum grain appears to be about 360 g/L (approximately 28 lb/bu).

FEMS Microbiol Ecol, 1999 Oct 1, 30(2), 147 - 155
Anaerobic conversion of carbon dioxide to methane, acetate and propionate on washed rice roots; Conrad R et al.; Washed excised roots of rice (Oryza sativa) produced H(2), CH(4), acetate, propionate and butyrate when incubated under anoxic conditions . Acetate production was most pronounced with a maximum rate (mean+/-standard error; four different root preparations) of 3.4+/-0.6 micromol h(-1) g-dry weight(-1) roots, compared to 0.45+/-0.13, 0.06+/-0.03, and 0.04+/-0.01 micromol h(-1) g-dw(-1) for propionate, butyrate and CH(4)1 kPa after one day of incubation . Then it decreased and reached more or less constant concentrations of about 50-80 Pa after about 7-8 days . Hydrogen partial pressures were always high enough to allow exergonic methanogenesis (DeltaG=-67 to -98 kJ mol(-1) CH(4)) and exergonic homoacetogenesis (DeltaG=-18 to -48 kJ mol(-1) acetate) from H(2) plus CO(2) . Radioactive bicarbonate/CO(2) was incorporated into CH(4), acetate and propionate . The specific radioactivities of the products indicated that CH(4) was exclusively produced from H(2)/CO(2) confirming a previous study . The contribution of CO(2) to the production of acetate and propionate was 32-39% and 42-61%, respectively, assuming that each carbon atom was equally labeled . Propionate also became radioactively labeled, when the roots were incubated with either {1-(14)C}acetate or {2-(14)C}acetate accounting for 60-76% of total propionate production . Reductive formation of propionate was thermodynamically favorable both from H(2) plus acetate plus CO(2) (DeltaG=-15 to -38 kJ mol(-1) propionate) and from H(2) plus CO(2) (DeltaG=-34 to -85 kJ mol(-1) propionate) . A substantial fraction of propionate was apparently reductively formed from acetate and/or CO(2) . In conclusion, our results demonstrate an intensive anaerobic dark metabolism of CO(2) on washed rice roots with reduction of CO(2) contributing significantly to the production of acetate, propionate and CH(4) . The CO(2) reduction seemed to be driven by decay and fermentation of root material.

Nat Biotechnol, 1999 Oct, 17(10), 1011 - 6
Metabolic engineering of Arabidopsis and Brassica for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production; Slater S et al.; Poly(hydroxyalkanoates) are natural polymers with thermoplastic properties . One polymer of this class with commercial applicability, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) can be produced by bacterial fermentation, but the process is not economically competitive with polymer production from petrochemicals . Poly(hydroxyalkanoate) production in green plants promises much lower costs, but producing copolymer with the appropriate monomer composition is problematic . In this study, we have engineered Arabidopsis and Brassica to produce PHBV in leaves and seeds, respectively, by redirecting the metabolic flow of intermediates from fatty acid and amino acid biosynthesis . We present a pathway for the biosynthesis of PHBV in plant plastids, and also report copolymer production, metabolic intermediate analyses, and pathway dynamics.

Appl Environ Microbiol, 1999 Oct, 65(10), 4484 - 9
Registered designation of origin areas of fermented food products defined by microbial phenotypes and artificial neural networks; Lopes MF et al.; Cheese produced from raw ewes' milk and chourico, a Portuguese dry fermented sausage, are still produced in a traditional way in certain regions of Portugal by relying on colonization by microbial populations associated with the raw materials, equipment, and local environments . For the purpose of describing the product origins and types of these fermented foods, metabolic phenotypes can be used as descriptors of the product as well as to determine the presence of compounds with organoleptic value . The application of artificial neural networks to the metabolic profiles of bacterial isolates was assayed and allowed the separation of products from different regions . This method could then be used for the Registered Designation of Origin certification process of food products . Therefore, besides test panel results for these traditionally produced food products, another tool for validating products for the marketplace is available to the producers . The method can be improved for the detection of counterfeit products.

Biotechnol Bioeng, 1999 Nov 20, 65(4), 371 - 81
Purification of recombinant and human apolipoprotein A-1 using surfactant micelles in aqueous two-phase systems: recycling of thermoseparating polymer and surfactant with temperature-induced phase separation; Persson J et al.; An effective system has been developed for purification of apolipoprotein A-1 from Escherichia coli fermentation solution and human plasma using aqueous two-phase extraction and thermal-phase separation . The system included non-ionic surfactants (Triton or Tween) and as top phase-forming polymer a random copolymer of ethylene oxide (50%) and propylene oxide (50%), Breox PAG 50A 1000, was used . The bottom phase-forming polymer was either hydroxypropyl starch, Reppal PES 100 and PES 200, or hydroxyethyl starch, Solfarex A 85 . The top-phase-forming polymer and the surfactants are thermoseparating in water solution, i.e., when heated a water phase and a polymer/surfactant phase are formed . Recombinant apolipoprotein A-1, the Milano variant, was extracted from E . coli fermentation solution in a primary Breox-starch phase system followed by thermal separation of the Breox phase where the target protein was recovered in the water phase . Both in the Breox-starch system and in the water-Breox system Triton X-100 was partitioned to the Breox phase . The addition of non-ionic surfactants to the Breox-starch system had strong effect on the purification and yield of the amphiphilic apolipoprotein A-1 . In a system containing 17% Breox PAG 50A 1000, 12% Reppal PES 100 and addition of 1% Triton X-100 the purification factor was 7.2, and the yield 85% after thermal separation of the Breox phase . Recycling of copolymer and surfactant was possible after thermal separation of copolymer phase . Approximately 85% of the copolymer and surfactant could be recycled in each extraction cycle . DNA could be strongly partitioned to the starch phase in the primary-phase system . This resulted in a 1000-fold reduction of E . coli DNA in the apolipoprotein A-1 solution obtained after thermoseparation . In extraction from human plasma containing low concentrations of apolipoprotein A-1, it was possible to reach a purification factor of 420 with 98% yield . By reducing the volume ratio to 0.1 Apo A-1 could be concentrated in a small volume of top phase (concentration factor 10) with a yield of 85% and a purification factor of 110 .

Carcinogenesis, 1999 Oct, 20(10), 1953 - 6
T cell status influences colon tumor occurrence in min mice fed short chain fructo-oligosaccharides as a diet supplement; Pierre F et al.; We have previously shown that addition of short chain fructo-oligosaccharides (indigestible carbohydrates) to food prevented colon tumors in C57BL/6-Apc(Min/+) mice, a model for human colon cancer . As gut-associated lymphoid tissue was concomitantly developed, we suggested that the immune response generated by this food may interfere with carcinogenesis due to involvement of mucosal cells in the regulation of tissue homeostasis . In the present experiment, we tested whether T cell status may influence colon tumor formation in Min mice fed a food supplement of short chain fructo-oligosaccharides . Min mice depleted of CD4(+) and CD8(+) lymphocytes developed twice as many tumors as immunocompetent mice (0.8 as compared with 0.4, the mean number in 7-week-old Min mice when food supplementation began; P = 0.02) . It is concluded that food supplementation with a substrate (a known prebiotic) fermented in the colon may stimulate a mechanism of immunosurveillance that would otherwise remain inefficient.

Dig Dis Sci, 1999 Sep, 44(9), 1914 - 23
Importance of colonic bacterial fermentation in short bowel patients: small intestinal malabsorption of easily digestible carbohydrate; Olesen M et al.; The small intestine's large capacity for glucose absorption and for adaptation seems to contradict the reported importance of carbohydrate malabsorption in short bowel (SB) patients . The aim of the present study was to investigate the occurrence of malabsorption in these patients ingesting realistic amounts of carbohydrates . We performed a dose-response study {ingestion of increasing amounts of glucose and complex carbohydrates (boiled rice and wheat bread), and the nonabsorbable disaccharide lactulose} in SB patients with an intact colon . The hydrogen (H2) -breath test and changes in serum acetate were used to evaluate colonic fermentation and, thus, indirectly, the lack of small intestinal carbohydrate assimilation . Blood glucose and plasma insulin were measured to evaluate absorption . Plasma concentrations of the ileal brake hormones--glucagon-like peptide-1 (GLP-1) and peptide tyrosine tyrosine (PYY)--were measured to test whether release of these hormones was related to colonic fermentation . Significant amounts of 25 g and 50 g glucose, and of the bread and rice meals were fermented rather than absorbed, as judged by the increases in end-expiratory H2 . Serum acetate concentrations were significantly higher in SB patients than in healthy controls . The orocecal transit times of all test meals ranged from 15 to 120 min . GLP-1 and PYY releases in SB patients were significantly higher than in healthy volunteers . They were mutually parallel and paralleled the increase in insulin . They were not related to ongoing fermentation or to intraluminal carbohydrate content per se, but most probably to absorption of glucose in the distal bowel . In conclusion, well-adapted SB patients had pronounced small intestinal malabsorption of carbohydrate, even after ingestion of small amounts of easily absorbable carbohydrates . A fast small intestinal spreading of carbohydrates, once in the small intestine, and a spill-over to the colon seem to explain the data best.

Chin J Biotechnol, 1998, 14(3), 157 - 63
Production of human recombinant bone morphogenetic protein-2A by high density culture of Escherichia coli with stationary dissolved oxygen fed-batch condition; Li M et al.; The optimization of cultivation condition in 500 ml shake flasks was carried out to produce recombinant human bone morphogenetic protein-2A (BMP-2A) in recombinant Escherichia coli YK537/pDH-B2m, followed by a 5L fermenter batch and condition-controlled fed-batch culture to obtain BMP-2A . The comparison of these two methods indicated that cultivation by keeping dissolved oxygen at 30%-40% and limiting glucose concentration could obtain BMP-2A 2.78 g/L broth, the final cell density was OD600 53 (dry cell weight 21.2 g/L), and expressed BMP-2A was 25% of the total amount of protein in E.coli . The critical fermentation conditions included: (1) keeping appropriate dissolved oxygen concentration in the process; (2) limiting glucose concentration; (3) taking heat induction at the middle-log phase and maintaining 42 degrees C for 4 hours; (4) controlling specific growth rate around 0.3 h-1 in the duration of growth.

Adv Microb Physiol, 1999, 41, 1 - 45
Factors affecting the production of L-phenylacetylcarbinol by yeast: a case study; Oliver AL et al.; L-Phenylacetylcarbinol (L-PAC) is the precursor for L-ephedrine and D-pseudoephedrine, alkaloids possessing alpha- and beta-adrenergic activity . The most commonly used method for production of L-PAC is a biological method whereby the enzyme pyruvate decarboxylase (PDC) decarboxylates pyruvate and then condenses the product with added benzaldehyde . The process may be undertaken by either whole cells or purified PDC . If whole cells are used, the biomass may be grown and allowed to synthesize endogenous pyruvate, or the cells may be used as a catalyst only, with both pyruvate and benzaldehyde being added . Several yeast species have been investigated with regard to L-PAC-producing potential; the most commonly used organisms are strains of Saccharomyces cerevisiae and Candida utilis . It was found that initial high production rates did not necessarily result in the highest final yields . Researchers then examined ways of improving the productivity of the process . The substrate, benzaldehyde, and the product, L-PAC, as well as the by-products, were found to be toxic to the biomass . Methods examined to reduce toxicity include modification of benzaldehyde dosing regimes, immobilization of biomass or purified enzymes, modification of benzaldehyde solubility and the use of two-phase reaction systems . Various means of modifying metabolism to enhance enzyme activity, relevant metabolic pathways and yield have been examined . Methods investigated include the use of respiratory quotient to influence pyruvate production and induce fermentative activity, reduced aeration to increase PDC activity, and carbohydrate feeding to modify glycolytic enzyme activity . The effect of temperature on L-PAC yield has been examined to identify conditions which provide the optimal balance between L-PAC and benzyl alcohol production, and L-PAC inactivation . However, relatively little work has been undertaken on the effect of medium composition on L-PAC yield.

J Thromb Thrombolysis, 1999 Oct, 8(3), 197 - 205
Recombinant von Willebrand factor: potential therapeutic use; Fischer BE; Human von Willebrand factor (vWF) produced by recombinant technology offers a new perspective in treatment of von Willebrand disease (vWD) . Several limitations connected with plasma-derived vWF concentrates, such as proteolytic degradation during the manufacture process, variation in multimer composition, lack of high molecular weight multimers, and donor dependence, can be overcome by rec-vWF . Recombinant vWF (rec-vWF) is produced by continuous fermentation of transformed mammalian cells . Biotechnological processes have been developed to isolated rec-vWF fractions with low, medium, and high degrees of multimerization . Structural analysis of rec-vWF demonstrated that it undergoes post-translational modifications comparable with plasma-derived vWF, such as multimerization, pro-peptide processing, and glycosylation . Functional analysis showed that rec-vWF exhibited activities comparable with plasma-derived vWF, such as platelet binding, platelet aggregation, collagen binding, and coagulation factor VIII (FVIII) binding . Collagen binding and platelet aggregation activity increased with the increasing multimer size of rec-vWF . Infusion of rec-vWF in antibody-induced vWF-deficient mice resulted in a significant decrease in bleeding . Infusion of rec-vWF in vWF-deficient dogs and pigs with severe vWD caused an increase in the endodenous FVIII level . Stabilization of FVIII in vivo was mediated both by high and low molecular weight rec-vWF molecules . Apparently, rec-vWF resisted proteolytic degradation in the circulation and no satellite bands were formed . Functional analysis in vitro and in vivo demonstrated the therapeutic potentials of rec-vWF, correction of vWF level, and stabilization of FVIII in plasma.

Appl Microbiol Biotechnol, 1999 Aug, 52(2), 135 - 45
Chemicals from biotechnology: molecular plant genetics will challenge the chemical and the fermentation industry; Wilke D; Industrial biotechnology has evolved as a significant manufacturing tool for products like fuel-grade ethanol, organic acids and bulk amino acids, but most items are still speciality products for food and pharmaceutical applications . Current development projects within the chemical industry, including lactic acid and 1,3-propanediol based polymers and plastics, indicate that new biotechnological processes and products may soon approach the market place, clearly targeted at the leading petrochemical bulk outlets . This is flanked by a strategic shift by the major chemical companies in to "life sciences"-pharmaceuticals, agrochemicals and the seed business as well as biotech fine chemicals . The recent tremendous achievements in molecular plant genetics and transgenic crop breeding will boost agrobiotechnology, agriculture and renewable raw materials as compelling projects for chemistry and biotechnology . New plant-based production routes may challenge established chemical and biochemical domains, but at the same time open new horizons to valuable feedstocks, intermediates and end-products.

Dev Biol Stand, 1999, 98, 127 - 32; discussion 133-4
A Phase I, randomized controlled clinical trial to study the reactogenicity and immunogenicity of a new split influenza vaccine derived from a non-tumorigenic cell line; Percheson PB et al.; We have found that our MDCK-derived cell line (BV-5F1) is non-tumorigenic in tests conducted in accordance with FDA guidelines, and thus may be suitable for producing live, attenuated or inactivated vaccine . The cell line has been extensively tested for the presence of contaminating microorganisms . No infectious agents of viral or other microbial origin were present . Using the BV-5F1 cell line, we have now designed a process for the large-scale production of influenza virus for the manufacture of a vaccine . The production system involves expansion of cells anchored on a microcarrier using stirred fermenters, followed by virus infection . Viral particles are purified in a way similar to the licensed egg-derived vaccine Fluviral SF and mainly involves ultracentrifugation, ultrafiltration and formaldehyde inactivation . The final product is a split inactivated vaccine . A randomized, double-blind clinical study was made in healthy adults using the new split influenza vaccine derived from viruses grown in cell culture (bivalent formulation) . The results of this Phase I study have demonstrated that the split influenza vaccine derived from cell culture is highly immunogenic and safe in adults.

Dev Biol Stand, 1999, 98, 101 - 10; discussion 111
Development of a Vero cell-derived influenza whole virus vaccine; Kistner O et al.; Influenza vaccine production is dependent on the availability of embryonated hen eggs for virus growth . This is an extremely cumbersome system with many disadvantages with respect to selection of virus variants and the presence of adventitious viruses . We have developed an alternative cell culture system which allows rapid production of large volumes of vaccine . The WHO-approved Vero cell line was used in serum-free culture to grow many influenza strains to high titre . This system could be scaled-up to allow vaccine production with a 1200 litre fermenter . A purification scheme was developed which resulted in a high purity whole virus vaccine . This was demonstrated to be at least as immunogenic as a conventional egg-derived preparation.

Eur J Cancer Prev, 1999 Aug, 8(4), 289 - 95
Effects of soft drink and table beer consumption on insulin response in normal teenagers and carbohydrate drink in youngsters; Janssens JP et al.; There is ample evidence that breast cancer susceptibility is induced during the developmental stages of the human breast where, in a manner related to sex-steroid hormones, insulin plays an important role . In turn, nutrition might be implicated . Regular soft drinks and table beer, both carbohydrate-containing drinks, are candidates affecting insulin concentrations . Eleven teenagers, between the ages of 13 and 17 years, consumed a soft drink and a table beer in a crossover study . The blood levels of insulin and glucose were related to anthropomorphometric and endocrine factors . In contrast to table beer, consumption of regular soft drinks induced a fast and dramatic increase in both glucose and insulin concentration within a maximum 1 hour after consumption . The insulin response was linearly correlated to the body mass index (BMI) . Children with a small increase in BMI are highly sensitive to regular soft drinks with regard to glucose and insulin response . The finding suggests a vicious circle of high caloric drinks, increase in BMI and insulin response . It is one of the nutritional pathways which might affect susceptibility for breast cancer in youngsters . Table beer, a drink with fermented sugars, does not share these effects on carbohydrate metabolism.

J Anim Sci, 1999 Sep, 77(9), 2554 - 63
Effect of Yucca schidigera on ruminal fermentation and nutrient digestion in heifers; Hristov AN et al.; In a replicated 3 x 3 Latin square experiment, six heifers (443 +/- 6.1 kg) fed a 61% barley grain:39% alfalfa silage diet (DM basis) were given intraruminal doses of powdered Yucca schidigera (YS) . Doses of 0 (control), 20, or 60 g/d were given at 0800 daily . Ruminal content was sampled 0, 2, 4, and 6 h after dosing . Acidity, concentrations of reducing sugars, free amino acids, and peptides in the rumen were not affected (P > .05) by YS . Relative to control, ruminal ammonia concentration was reduced (P < .05) 2 h after YS dosing . Ruminal propionate concentration was increased (P < .05) by YS . Protozoal numbers in the rumen were lower (P < .05) with YS than without . Yucca did not affect (P > .05) rate or extent of in situ DM degradability . Fibrolytic, amylolytic, and proteolytic activities in ruminal contents were similar among treatments (P > .05) . Dry matter intake, apparent digestibilities of DM, NDF, and CP, nitrogen balance, and microbial protein synthesis in the rumen were not affected (P > .05) by treatment . The effect of YS on ruminal ammonia concentration likely resulted from a decreased concentration of protozoa and, presumably, from ammonia binding by YS . The effect on ruminal propionate was probably a result of a selective inhibitory effect of YS on rumen microbial species.

Dig Dis Sci, 1999 Aug, 44(8), 1547 - 53
Short-chain fatty acids inhibit fluid and electrolyte loss induced by cholera toxin in proximal colon of rabbit in vivo; Rabbani GH et al.; Short-chain fatty acids (SCFA), acetate, propionate, and butyrate, are produced by bacterial fermentation in the colon and stimulate Na+ and Cl- absorption . We have studied the effects of SCFA on fluid and electrolyte absorption during cholera toxin (CT) -induced colonic secretion in rabbit, using a steady-state perfusion technique with marker (PEG) dilution . Perfusion with SCFA significantly (P < 0.01) reduced colonic water secretion . Butyrate reduced water secretion the most (95%), followed by propionate (90%), and acetate (80%) . Butyrate significantly (P < 0.001) reduced secretions of Na+ (95.5%), K+ (75.2%), and Cl-(80.7%) ions but not HCO3- ions . Propionate similarly reduced secretion of HCO3-(45.3%) . Acetate significantly (P < 0.001) inhibited Na+ (76.4%) and Cl- (75.7%) secretion, but the inhibitions of K+ (23.6%) and HCO3 (28.8%) were not significantly different from the controls . We conclude that SCFA reduce CT-induced water and electrolyte secretion in the rabbit proximal colon, and its potential as an antidiarrheal agent should be further evaluated.

Can J Microbiol, 1999 Apr, 45(4), 318 - 25
Simultaneous removal of phenol, ortho- and para-cresol by mixed anaerobic consortia; Tawfiki Hajji K et al.; Two different anerobic consortia, one removing phenol and ortho (o-) cresol and other removing para(p-) cresol, were cultivated in serum bottles using whey as cosubstrate substitute for proteose peptone . Phenol and p-cresol removal with the phenol-removing consortium were the same with 0.0125% (w/v) whey as with 0.05% proteose peptone . For the other consortium, 8 days were required to decrease the p-cresol concentration from 35 to 2 mg/L with 0.025% whey, while 35 days were required to achieve a similar removal with 0.5% proteose peptone . The two consortia were mixed and cultivated with 0.025% whey . Phenolic compound removal with the mixed consortia was as good as that achieved by each of the two initial consortia against their respective substrates . This removal activity was maintained after several transfers . In a continuous upflow fixed-film reactor, the mixed consortia removed over 98% of 150 mg/L of phenol and 35 mg/L of each o- and p-cresol in the influent at 29 degrees C, with 0.025% whey as cosubstrate . The hydraulic retention time (HRT) was 0.25 day, corresponding to a phenolic compound volumic loading rate of 880 mg/(L of reactor x day) . Once the continuous flow reactor achieved constant phenolic compound removal, no intermediates were found in the effluent, while in serum bottles, m-toluic acid, an o-cresol intermediate, accumulated . Measurements of the specific activity for the uptake of different substrates demonstrated the presence of all trophic groups involved in methanogenic fermentation . These activities were, in mg of substrate/(g of volatile suspended solids x day), as follows: 849 +/- 25 for the acidogens; 554 +/- 15 for the acetogens; 934 +/- 37 for the aceticlastic methanogens; and 135 +/- 15 for the hydrogenophilic methanogens . Electron micrographs of the mixed consortia showed seven different morphological bacterial types, including Methanotrix-like bacteria.

Am J Vet Res, 1999 Sep, 60(9), 1061 - 5
Effects of tylosin on concentrations of Fusobacterium necrophorum and fermentation products in the rumen of cattle fed a high-concentrate diet; Nagaraja TG et al.; OBJECTIVE: To determine effects of tylosin on ruminal concentrations of Fusobacterium necrophorum and fermentation products in cattle during rapid adaptation to a high-concentrate diet . ANIMALS: 6 steers fitted with ruminal cannulas . PROCEDURE: Steers were assigned randomly to 2 treatment groups and switched from a 0 to an 85% concentrate diet during a 4-day period . Cattle received this diet, with or without tylosin (90 mg/steer/d), for 4 weeks . Samples of ruminal contents were collected daily beginning 2 days before the treatment protocol and in the first week of concentrate feeding . Four subsequent samples were collected at weekly intervals . Concentration of F . necrophorum in samples was determined, using the most-probable-number technique . Ruminal pH and concentrations of volatile fatty acids (VFA), lactate, and ammonia also were determined . All steers received both treatments separated by 4 weeks (cross-over design), during which time they were fed alfalfa hay only . RESULTS: In control steers, concentration of F . necrophorum increased in response to the high-concentrate diet . Tylosin-fed steers had lower concentrations of F . necrophorum than control steers at all times during concentrate feeding . However, ruminal pH and concentrations of lactate, VFA, and ammonia did not differ between treatment groups . CONCLUSIONS AND CLINICAL RELEVANCE: Tylosin caused a significant reduction in ruminal concentrations of F . necrophorum during rapid adaptation to a high-concentrate diet but had no effect on fermentation products . The reduction in ruminal concentration of F . necrophorum helps explain the reduction in prevalence of hepatic abscesses reported in tylosin-fed feedlot cattle.

Dig Dis Sci, 1999 Jul, 44(7), 1476 - 84
Lactulose feeding in piglets: a model for persistent diarrhea and colitis induced by severe sugar malabsorption; Kien CL et al.; We sought to determine, in a piglet model, whether severe sugar malabsorption causes colonic injury or inflammation . Twenty-four piglets were randomized to receive either control formula (CON) or CON supplemented with lactulose (LAC) (N = 12 each group) . After seven days, inflammation, apoptosis, and crypt cell proliferation were assessed in the proximal colon (cecum) . Lactulose feeding caused persistent diarrhea . In both groups, breath H2 concentration was low, suggesting no increased fermentation in the LAC group . Weight gain/volume formula intake was identical in the CON and LAC groups (0.09+/-0.13 and 0.09+/-0.11 g/ml) respectively . Injury to the colon did not occur, but inflammation of the colon (scale 0-5) was greater in LAC (score of 1.5+/-1.38) than in CON (0.42+/-0.79; P<0.05) . Cell proliferation at the basal 40% of the crypt was 92% increased in CON (labeling index 22.8+/-9.9 vs . 11.9+/-2.8; P<0.05) . We conclude that persistent feeding during severe sugar malabsorption permits weight gain but may cause colitis.

Z Naturforsch {C}, 1999 Jul-Aug, 54(7-8), 474 - 80
Hypoxysordarin, a new sordarin derivative from Hypoxylon croceum; Daferner M et al.; Hypoxysordarin (1), a new sordarin derivative, was isolated from the fermentation broth of the facultative marine Hypoxylon croceum together with a new gamma-lactone, hypoxylactone (2) and sordarin (3) . The structures were determined by spectroscopic methods . Sordarin (3) has previously been isolated from the terrestrial Sordaria araneosa (Sordariaceae) . Like the parent compound hypoxysordarin exhibits high antifungal activities due to a specific inhibition of protein biosynthesis.

J Clin Microbiol, 1999 Oct, 37(10), 3387 - 9
Comparison of four methodologies for rapid and cost-effective identification of Candida glabrata; Fenn JP et al.; To improve turnaround time and decrease the cost of the identification of Candida glabrata, we evaluated four methods for the detection of trehalose assimilation or fermentation . These methods were compared with the API 20C method (bioMERIEUX, Hazelwood, Mo.) to determine accuracy . We recommend the use of Remel Rapid Trehalose Assimilation Broth because of its rapid, 3-h results, reasonable sensitivity, and low number of false positives.

Yeast, 1999 Sep 15, 15(12), 1159 - 69
Saccharomyces cerevisiae colony growth and ageing: biphasic growth accompanied by changes in gene expression; Meunier JR et al.; Although colony growth and morphology are central tools in yeast genetics, little is known about the cell physiology and how it changes during the colony growth and ageing . Here we show that the growth of a well-separated Saccharomyces cerevisiae colony is biphasic; a rapid growth phase is followed by a sharp transition to a slower growth phase . In the first growth phase ( approximately 24 cell divisions) most, if not all, cells divide at a rate similar to that in liquid medium and exhibit morphological, biochemical and genetic characteristics of cells engaged in the cell cycle . During the second growth phase, cells in the centre of a colony gradually enter stationary phase, so that later in this phase the growth occurs predominantly at the periphery . Unlike the biphasic growth in rich liquid media containing a fermentable carbon source, in which the first growth is fueled by fermentation and the second by aerobic metabolism, the two phases of the colony growth can be fueled either exclusively by fermentation or exclusively by aerobic metabolism . We also describe a novel technique for in situ estimation of the transcriptional status in the colony cells, which was used to monitor transcription dynamics during the colony development . Using this technique and standard methods to determine mRNA levels, we show that the transition between the first and second growth phases is accompanied by a global change in the pattern of transcription: transcription of most genes is repressed while that of some genes is induced .

Mol Biol Evol, 1999 Sep, 16(9), 1300 - 7
Phylogenetic relationships of the enigmatic hoatzin (Opisthocomus hoazin) resolved using mitochondrial and nuclear gene sequences
Hughes JM, Baker AJ.
The hoatzin (Opisthocomus hoazin) is a bizarre, long-tailed, crested bird that inhabits the riparian lowlands of South America . Among its peculiar attributes are (1) microbial foregut fermentation to convert plant cellulose in consumed foliage into simple sugars, (2) a highly modified skeleton to accommodate its large crop, and (3) in the young of this species, wing claws at the wrist joint which are used to climb among the branches of the nest tree . Consequently, the taxonomic position of this unusual bird has perplexed systematists since its description over 200 years ago . Traditionally classified among the fowl-like birds (Galliformes), recent studies have favored its placement with the cuckoos (Cuculiformes: Cuculidae) . To help resolve this systematic uncertainty, we sequenced six mitochondrial genes (cytochrome oxidase I, II, and III, ATPase 8, ATPase 6, and cytochrome b) and one nuclear gene (c-mos), totaling 5,487 base pairs . With this large data set and an appropriate range of outgroup taxa, we demonstrate that the hoatzin should not be classified among the cuckoos or Galliformes . Instead, our analyses indicate that the hoatzin is most closely related to the turacos (Musophagiformes: Musophagidae), a small family of arboreal, frugivorous birds inhabiting sub-Saharan Africa . This phylogenetic relationship is also supported by osteological behavioral, and fossil evidence.

J Invertebr Pathol, 1999 Sep, 74(2), 143 - 8
Development of the one-on-one quality assessment assay for entomopathogenic nematodes; Converse V et al.; The one-on-one bioassay was developed using Steinernema carpocapsae (All) nematodes against the wax moth larva, Galleria mellonella . The assay was used to develop and compare virulence profiles of both in vitro- and in vivo-produced nematodes and to provide a quality assessment 'standard' for in vitro-produced nematodes . The bioassay was subsequently used to develop virulence profiles for Steinernema carpocapsae (UK), S . feltiae (UK), S . feltiae (R1.5), S . feltiae (SN), S . glaseri (NJ-43), and S . riobrave (RGV) . These profiles are unique for each species and isolate and are used as a standard of virulence in routine quality assessment of nematodes produced in liquid fermentation .

Am J Physiol, 1999 Sep, 277(3 Pt 2), R767 - 75
Potentiation of carbachol-induced amylase release by propionate in guinea pig and vole pancreatic acini; Harada E et al.; The action of propionate, one of the major end products of microbial fermentation in herbivores was investigated in isolated, perifused pancreatic acini of guinea pigs, voles, and mice . With the use of guinea pig acini, 100 microM propionate had no effect, whereas 300 and 600 microM increased amylase release by six- and ninefold, respectively . Simultaneous perifusion of carbachol (CCh) 10 microM plus propionate 100 microM in guinea pig acini produced a potentiated secretory response that was 130% higher than the summated value obtained with CCh and propionate alone . The potentiation by propionate (100 microM) of CCh (10 microM)-induced amylase release was also obtained in vole pancreatic acini, but the mouse pancreatic preparation did not exhibit a similar potentiation . In contrast to CCh, propionate (100-600 microM) alone had no significant effect on intracellular Ca2+ concentration ({Ca2+}i) and did not alter {Ca2+}i elicited by CCh . Ca ionophore A23187 (5 microM)-induced amylase release in guinea pig acini was enhanced twofold by the addition of propionate . Cellular cAMP content was increased slightly by propionate, but did not alter dose dependently . The cAMP level with combinations of CCh and propionate was almost same as that with CCh alone and propionate alone . Staurosporine did not modify amylase secretion induced by a combination of CCh and propionate . These results suggest that propionate, in addition to a direct action on amylase release, potentiates CCh-induced amylase release in guinea pig and vole acini via a secretory pathway not associated with an increase in {Ca2+}i and cellular cAMP.

JPEN J Parenter Enteral Nutr, 1999 Sep-Oct, 23(5 Suppl), S101 - 5
Colonic digestion and absorption of energy from carbohydrates and medium-chain fat in small bowel failure; Jeppesen PB et al.; Gut function and the degree of intestinal insufficiency or failure in short bowel patients can be quantified with respect to wet weight and energy absorption by the use of balance studies . This enables the physician to distinguish patients with extreme intestinal failure inconsistent with the restoration of intestinal autonomy by dietary manipulation from short bowel patients with borderline gut failure in whom dietary manipulations may result in the weaning from parenteral support . A high-carbohydrate, low long-chain fat diet and a diet where long-chain fat has been replaced by medium-chain triglycerides increase absorption of energy in patients with small bowel failure, provided that they have a preserved colon in continuity . This is due to the ability of the colonic flora to ferment carbohydrates malabsorbed in the small bowel to the short-chain fatty acids (SCFAs) . These SCFAs are easily absorbed across the colonic mucosa resulting in a salvage of carbohydrate energy that otherwise would have been lost in feces . In contrast, long-chain fatty acids are not absorbed by the colon, and long-chain fat malabsorbed in the small bowel of short bowel patients are not retained in the large bowel . Recent work has indicated that the water soluble medium-chain fatty acids are effectively absorbed in the large bowel similar to the SCFAs . This may explain an almost complete absorption of medium-chain triglycerides in short bowel patients, even in patients with virtually no absorption of long-chain fat, and why this only occurs in patients with a colon in continuity . Manipulation of the dietary fat:carbohydrate ratio is much less efficacious in short bowel patients with no colonic function, and the use of medium-chain triglycerides has no proven effect on overall energy absorption from short bowel patients without a large bowel in continuity . Hence, the colon has increasingly important digestive functions as small bowel failure proceeds, not only when it comes to absorption of water and sodium, but also of energy from carbohydrates and medium-chain fat.

Biochim Biophys Acta, 1999 Aug 4, 1412(3), 212 - 29
The hyperthermophilic bacterium, Thermotoga maritima, contains an unusually complex iron-hydrogenase: amino acid sequence analyses versus biochemical characterization; Verhagen MF et al.; The hyperthermophilic bacterium, Thermotoga maritima, grows up to 90 degrees C by fermenting carbohydrates and it disposes of excess reductant by H(2) production . The H(2)-evolving cytoplasmic hydrogenase of this organism was shown to consist of three different subunits of masses 73 (alpha), 68 (beta) and 19 (gamma) kDa and to contain iron as the only metal . The genes encoding the subunits were clustered in a single operon in the order hydC (gamma), hydB (beta), and hydA (alpha) . Sequence analyses indicated that: (a) the enzyme is an Fe-S-cluster-containing flavoprotein which uses NADH as an electron donor; and (b) the catalytic Fe-S cluster resides within the alpha-subunit, which is equivalent to the single subunit that constitutes most mesophilic Fe-hydrogenases . The alpha- and beta-subunits of the purified enzyme were separated by chromatography in the presence of 4 M urea . As predicted, the H(2)-dependent methyl viologen reduction activity of the holoenzyme (45-70 U mg(-1)) was retained in the alpha-subunit (130-160 U mg(-1)) after subunit separation . However, the holoenzyme did not contain flavin and neither it nor the alpha-subunit used NAD(P)(H) or T . maritima ferredoxin as an electron carrier . The holoenzyme, but not the alpha-subunit, reduced anthraquinone-2,6-disulfonate (apparent K(m), 690 microM) with H(2) . The EPR properties of the reduced holoenzyme, when compared with those of the separated and reduced subunits, indicate the presence of a catalytic 'H-cluster' and three {4Fe-4S} and one {2Fe-2S} cluster in the alpha-subunit, together with one {4Fe-4S} and two {2Fe-2S} clusters in the beta-subunit . Sequence analyses predict that the alpha-subunit should contain an additional {2Fe-2S} cluster, while the beta-subunit should contain one {2Fe-2S} and three {4Fe-4S} clusters . The latter cluster contents are consistent with the measured Fe contents of about 32, 20 and 14 Fe mol(-1) for the holoenzyme and the alpha- and beta-subunits, respectively . The T . maritima enzyme is the first 'complex' Fe-hydrogenase to be purified and characterized, although the reason for its complexity remains unclear.

Zentralbl Veterinarmed B, 1999 Aug, 46(6), 399 - 404
Outbreak of septicaemic colibacillosis in Japanese quail (Coturnix coturnix japonica); Arenas A et al.; We have carried out an aetiological and pathological study of an outbreak of septicaemia caused by the O165 serogroup of Escherichia coli in Japanese quail (Coturnix coturnix japonica) which resulted in high mortality (90%) . After treatment with amoxicilin in drinking water (200 mg/l), morbility and mortality rates dropped markedly . Microbiological studies showed that the organism responsible was an atypical E . coli strain, on the basis of the non-fermentation of lactose (ONPG-), which belonged to the O165 serogroup and was highly virulent for 1-day-old chickens.

Mycopathologia, 1998-99, 144(2), 73 - 5
Isolation of Hanseniaspora uvarum (Kloeckera apiculata) in humans; Garcia-Martos P et al.; Isolation of Hanseniaspora uvarum, a yeast of the ascomycetes group, whose anamorph corresponds to Kloeckera apiculata, obtained from stool and two ungual specimens from three patients, is reported . This yeast has been found in soil, water, various fruits, bivalve molluscs, crabs, prawns and fruit flies; in Spain, it has been described in the fermentation processes of some wines . In our region, it has also been found in the intestine of mackerel (Scomber scombrus) . Its finding in humans constitutes a clinical rarity.

J Biol Chem, 1999 Sep 17, 274(38), 27056 - 68
Sertolin is a novel gene marker of cell-cell interactions in the rat testis; Mruk DD et al.; A novel testicular protein designated sertolin was cloned . The full-length sertolin cDNA consists of 853 base pairs with an open reading frame of 381 base pairs coding for a 127-amino acid polypeptide that shares limited identities with antaxin/josephin and thrombospondin proteins . Sertolin (calculated molecular mass, 13,759 daltons) has two mRNA transcripts of 2.3 and 1 kilobase . A 22-amino acid peptide based on the deduced amino acid sequence of sertolin (NH(2)-KKEHFNLFKAASVSHLVQVVPQ) was synthesized and used for polyclonal antibody production . Immunoblot analysis detected a 17-kDa immunoreactive band in the Sertoli cell cytosol . Using Sertoli-germ cell cocultures, sertolin expression was found to be reduced by as much as 5-fold at the time when germ cells attach onto Sertoli cells but preceding the establishment of specialized inter-Sertoli-germ cell junctions . Neither FSH nor 17beta-hydroxy-5alpha-androstan-3-one was able to affect sertolin expression, whereas estradiol-17beta and progesterone induced a significant increase in Sertoli cell sertolin expression in vitro . In addition, interleukin-1alpha, a germ cell-derived cytokine, was also able to elicit a transient but significant increase in Sertoli cell sertolin expression . Sertolin expression was also shown to increase with testicular development and is likely to be associated with the onset of spermatogenesis . In addition, sertolin expression increased in the testis when generalized inflammation was induced in adult rats by injection of fermented yeast . These results show that sertolin will be useful in characterizing cell-cell interactions in the testis.

Lab Anim Sci, 1999 Aug, 49(4), 406 - 10
Complications of ileal cannulation in cats; Mawby DI et al.; BACKGROUND AND PURPOSE: The anatomic position of the ileum is such that use of simple, noninvasive sampling techniques is difficult . Sampling of ileal contents by use of a cannula provides information about those contents-digestibility and fermentation patterns; however, complications with use of cannulas in horses and dogs have been reported . Therefore, cats were chosen as a laboratory model in which to study ileal cannulation . METHODS: Simple T-piece cannulas were surgically placed in the ileum of six adult cats . RESULTS: Postsurgical complications included cannula displacement and leakage, with subsequent abscessation and skin inflammation . Owing to these complications, the cannulas were removed from four of the six cats . Similar complications developed after removal of the cannulas . In the only attempt to collect a sample, obtaining a 0.5-ml sample of ileal fluid from one cannula took 3 h . Of the six cats, four died or were euthanized . CONCLUSION: Use of cannulas is not a viable technique of collecting ileal fluid samples in cats.

J Antibiot (Tokyo), 1999 May, 52(5), 485 - 90
Effect of thiazinotrienomycin B, an ansamycin antibiotic, on the function of epidermal growth factor receptor in human stomach tumor cells; Hosokawa N et al.; Thiazinotrienomycin B (TT-B), an ansamycin isolated from fermentation broths of Streptomyces sp . MJ672-m3, inhibited the growth in vitro of human stomach tumor SC-6 cells over 10 times more strongly than the growth of other human tumor cells, such as HeLa (cervix), T24 (bladder) and LX-1 (lung) . The extent of growth inhibition by TT-B of SC-6, but not of LX-1 nor T24, was lowered in a competitive manner by raising serum concentrations in the culture medium . TT-B inhibited the cell cycle progression of SC-6 at an early stage of the progression from G0/G1 to S . The inhibition was again competitive with serum concentrations in the culture medium . No direct inhibition of DNA synthesis was observed at the concentration range which caused the cell cycle arrest . TT-B and anti-epidermal growth factor receptor (anti-EGFR) were antagonistic to each other in inhibiting the cell cycle progression of SC-6 from G0/G1 to S, suggesting that the two compounds share the same target, EGFR . The kinase activity of EGFR was little inhibited by TT-B in a cell-free system.

J Antibiot (Tokyo), 1999 May, 52(5), 466 - 73
Sanglifehrins A, B, C and D, novel cyclophilin-binding compounds isolated from Streptomyces sp . A92-308110 . I . Taxonomy, fermentation, isolation and biological activity; Sanglier JJ et al.; A novel class of macrolides for which the name sanglifehrins is proposed, has been discovered from actinomycete strains based on their high affinity binding for cyclophilin A (CypA), an immunophilin originally identified as a cytosolic protein binding cyclosporin A (CsA) . The sanglifehrins were produced by Streptomyces sp . A92-308110 . They were isolated and purified by extraction and several chromatographic, activity-guided steps . Sanglifehrins A and B exhibit a 10 to approximately 20 fold higher affinity for CypA than CsA, whereas the affinity of sanglifehrins C and D for CypA is comparable to that of CsA . Sanglifehrins exhibit a lower immunosuppressive activity than CsA when tested in the mixed lymphocyte reaction . Their in vitro activity indicates that they belong to a novel class of immunosuppressants.

Microbiol Immunol, 1999, 43(6), 521 - 5
Detection of Mycoplasma fermentans in saliva sampled from infants, preschool and school children, adolescents and adults by a polymerase chain reaction-based assay; Shibata K et al.; Attempts were made to detect Mycoplasma fermentans in saliva sampled from 201 subjects (108 males and 93 females) aged from 4 months to 59 years by a polymerase chain reaction-based assay . M . fermentans was detected in saliva from 110 (54.7%) of 201 subjects, and 10 (28.6%) of 35 subjects aged from 4 months to 3 years . Of ten positive subjects, three were aged from 16 to 23 months and five were from 26 to 31 months . The incidence tended to increase with age up to the teens . The incidence was significantly greater in teenagers than in subjects aged from 7 to 12 years, but there was no significant difference in the incidence between the group of teenagers and each of the groups of subjects older than the teenagers . Thus, it was suggested that M . fermentans colonized the mouth at the age of about 16 months up to the age of 19 years.

J Dairy Sci, 1999 Aug, 82(8), 1779 - 90
Synchronization of carbohydrate and protein sources on fermentation and passage rates in dairy cows; Casper DP et al.; Four ruminally cannulated Holstein cows in midlactation were randomly assigned to a 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments to evaluate two nonstructural carbohydrate sources (corn or barley) with two sources of ruminally undegradable protein (soybean meal or extruded soybean meal) on milk production, ruminal fermentation, and digesta passage rates . Milk production (25.1, 27.5, 23.8, and 23.5 kg/d for the corn and soybean meal, corn and extruded soybean meal, barley and soybean meal, and barley and extruded soybean meal, respectively) and dry matter intake per unit of body weight (3.9, 4.1, 3.7, and 3.7%) were greater for cows fed corn than for cows fed barley and were similar for cows fed soybean meal or extruded soybean meal . Concentrations of ruminal NH3-N were greater for cows fed the corn and soybean meal diet than for cows fed other diets (15.0, 10.4, 9.0, and 11.3 mg/dl) . Rumen volatile fatty acid concentrations were greater for cows fed corn than barley (133, 139, 121, and 118 mumol/ml) . Fractional passage rates of solids from the rumen were greater for cows fed the barley and soybean meal diet than cows fed the corn and soybean meal diet (3.4, 3.9, 4.2, and 3.8%/h), and ruminal liquid dilution rates were similar for cows fed all diets (11.2, 11.0, 11.1, and 11.9%/h) . The attempt to synchronize ruminal nonstructural carbohydrate and crude protein degradability produced minimal benefits for midlactation dairy cows.

J Dairy Sci, 1999 Aug, 82(8), 1765 - 78
Improving energy supply to late gestation and early postpartum dairy cows; Dann HM et al.; Sixty-five multiparous Holstein cows were used to test the effects of feeding diets of varied ruminal carbohydrate availability during the transition period on dry matter intake, blood metabolites, and lactational performance . Cows received total mixed rations containing either cracked corn or steam-flaked corn beginning 28 d prior to expected calving date . At parturition, cows were assigned to a postpartum total mixed ration that contained either cracked corn or steam-flacked corn . Diets were fed until 63 d in milk . No treatment effects on prepartum or postpartum dry matter intake, body weight, and body condition score were observed . Cows fed steam-flaked corn had lower blood urea N concentrations during the prepartum period and lower plasma nonesterified fatty acid concentrations during the prepartum and postpartum periods . Cows fed steam-flaked corn postpartum produced 2.3 kg/d more milk than cows fed cracked corn during the first 63 d in milk . Fat corrected milk showed no treatment effect . Seven cows were used to evaluate treatment effects on ruminal fermentation and digesta kinetics . Prepartum and postpartum treatments had minimal effects on ruminal fermentation . Feeding steam-flaked corn prepartum decreased apparent fiber digestibility and ruminal NH3 N . Feeding steam-flaked corn postpartum decreased the acetate to propionate ratio . Prepartum and postpartum treatments did not affect digesta kinetics . An increase in ruminal carbohydrate availability during the postpartum period enhanced milk production, but had variable results on ruminal fermentation.

J Dairy Sci, 1999 Aug, 82(8), 1595 - 611
ADSA Foundation Scholar Award . Critical issues affecting the future of dairy industry: individual contributions in the scope of a global approach; Malcata FX; Several constraints that have been affecting the dairy industry are identified in a critical fashion, and directions are given with an emphasis on food processing implemented at the postproduction level . The rationale for modifications aimed at enhancing the appeal of condensed dairy products should be consubstantiated in strengthening of organoleptic characteristics, improvement of nutraceutical impact, and reduction of polluting power . This enumeration follows an order of increasing time scale required for consumer perception and increasing size scale associated with expected impact . Pursuance of such streamlines should lead to manufacture of dairy products that resemble nature more closely in terms of milk coagulation, milk fat modification, milk fermentation, whey fermentation, and starter culture addition . Directions for research and development anticipated as useful and effective in this endeavor, and which have been previously and consistently adopted in the development of an individual research program, are characterization and development of alternative rennets from plant sources, development of starter and nonstarter cultures from adventitious microflora, utilization of probiotic strains as starter cultures, upgrading of whey via physical or fermentation routes, and modification of milk fat via lipase-mediated interesterification reactions.

Am J Clin Nutr, 1999 Sep, 70(3), 391 - 404
A randomized, community-based trial of the effects of improved, centrally processed complementary foods on growth and micronutrient status of Ghanaian infants from 6 to 12 mo of age; Lartey A et al.; BACKGROUND: Koko, a fermented maize porridge used as the primary complementary food in Ghana, has been implicated in the high prevalence of child malnutrition . Weanimix, a cereal-legume blend developed by the United Nations Children's Fund and the Ghanaian government, has been promoted as an alternative . OBJECTIVE: We evaluated the effect of feeding Weanimix and 3 other locally formulated, centrally processed complementary foods on the nutritional status of 208 breast-fed infants . DESIGN: Infants were randomly assigned to receive 1 of 4 foods from 6 to 12 mo of age: Weanimix (W), Weanimix plus vitamins and minerals (WM), Weanimix plus fish powder (WF), and koko plus fish powder (KF) . Dietary and anthropometric data were collected regularly . Blood was collected at 6 and 12 mo of age to assess iron, zinc, vitamin A, and riboflavin status . Before and after the intervention, cross-sectional data on the anthropometric status of infants not included in the intervention (NI; n = 464) were collected . RESULTS: There were no significant differences between intervention groups in weight or length gain or in hemoglobin, hematocrit, transferrin saturation, plasma zinc, or erythrocyte riboflavin values between 6 and 12 mo of age . From 9 to 12 mo of age, z scores were lower in NI infants than in the combined intervention groups {at 12 mo: -1.71 +/- 0.90 compared with -1.19 +/- 0.93 for weight and -1.27 +/- 1.02 compared with -0.63 +/- 0.84 for length (P < 0.001 for both), respectively} . The percentage of infants with low ferritin values increased significantly between 6 and 12 mo of age in groups W, WF, and KF but not in group WM . Change in plasma retinol between 6 and 12 mo of age was significantly greater in group WM than in the other 3 groups combined (0.14 +/- 0.3 compared with -0.04 +/- 0.3 micromol/L, P = 0 . 003) . CONCLUSIONS: All 4 foods improved growth relative to the NI group . Infants fed WM had better iron stores and vitamin A status than those fed nonfortified foods.

Food Chem Toxicol, 1999 Jun, 37(6), 569 - 79
Inhibition of eleven mutagens by various tea extracts, (-)epigallocatechin-3-gallate, gallic acid and caffeine; Hour TC et al.; The antimutagenic properties of various tea extracts (green tea, pauchong tea, oolong tea and black tea) and their components including (-)-epigallocatechin-3-gallate (EGCG), gallic acid and caffeine were examined by the Ames test . The antimutagenic activity of the green tea extract against N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), folpet and monocrotophos was greater than those of pouchong, oolong and black tea extracts . The antimutagenic effects of tea extracts against 2-acetylaminofluorene (AAF) decreased as follows: oolong tea > pauchong tea > black tea > green tea . Furthermore, black tea showed a greater antimutagenic activity against benzo{a}pyrene (BP) . The pauchong tea showed a stronger inhibitory effect against 9-aminoacridine (9AA) and aflatoxin B1 (AFB1) than other tea extracts . EGCG markedly suppressed the direct-acting mutagenicity of MNNG, N-nitroso-N-methylurea (MNU), captan, and folpet which were alkylating agents and fungicides . Similarly, gallic acid, the major component of black tea strongly inhibited the mutagenicity of 9AA, and moderately inhibited the mutagenicity of MNNG and folpet . The caffeine was less active . EGCG and gallic acid perhaps could act as nucleophiles to scavenge the electrophilic mutagens . Taken together, these results suggest that formation of different metabolites during various stages of tea fermentation may affect antimutagenic potencies against different types of chemical mutagens.

Microbiol Mol Biol Rev, 1999 Sep, 63(3), 675 - 707
Helicobacter pylori physiology predicted from genomic comparison of two strains; Doig P et al.; Helicobacter pylori is a gram-negative bacteria which colonizes the gastric mucosa of humans and is implicated in a wide range of gastroduodenal diseases . This paper reviews the physiology of this bacterium as predicted from the sequenced genomes of two unrelated strains and reconciles these predictions with the literature . In general, the predicted capabilities are in good agreement with reported experimental observations . H . pylori is limited in carbohydrate utilization and will use amino acids, for which it has transporter systems, as sources of carbon . Energy can be generated by fermentation, and the bacterium possesses components necessary for both aerobic and anaerobic respiration . Sulfur metabolism is limited, whereas nitrogen metabolism is extensive . There is active uptake of DNA via transformation and ample restriction-modification activities . The cell contains numerous outer membrane proteins, some of which are porins or involved in iron uptake . Some of these outer membrane proteins and the lipopolysaccharide may be regulated by a slipped-strand repair mechanism which probably results in phase variation and plays a role in colonization . In contrast to a commonly held belief that H . pylori is a very diverse species, few differences were predicted in the physiology of these two unrelated strains, indicating that host and environmental factors probably play a significant role in the outcome of H . pylori-related disease.

Microbiol Mol Biol Rev, 1999 Sep, 63(3), 642 - 74
Metabolism and genetics of Helicobacter pylori: the genome era; Marais A et al.; The publication of the complete sequence of Helicobacter pylori 26695 in 1997 and more recently that of strain J99 has provided new insight into the biology of this organism . In this review, we attempt to analyze and interpret the information provided by sequence annotations and to compare these data with those provided by experimental analyses . After a brief description of the general features of the genomes of the two sequenced strains, the principal metabolic pathways are analyzed . In particular, the enzymes encoded by H . pylori involved in fermentative and oxidative metabolism, lipopolysaccharide biosynthesis, nucleotide biosynthesis, aerobic and anaerobic respiration, and iron and nitrogen assimilation are described, and the areas of controversy between the experimental data and those provided by the sequence annotation are discussed . The role of urease, particularly in pH homeostasis, and other specialized mechanisms developed by the bacterium to maintain its internal pH are also considered . The replicational, transcriptional, and translational apparatuses are reviewed, as is the regulatory network . The numerous findings on the metabolism of the bacteria and the paucity of gene expression regulation systems are indicative of the high level of adaptation to the human gastric environment . Arguments in favor of the diversity of H . pylori and molecular data reflecting possible mechanisms involved in this diversity are presented . Finally, we compare the numerous experimental data on the colonization factors and those provided from the genome sequence annotation, in particular for genes involved in motility and adherence of the bacterium to the gastric tissue.

Br Poult Sci, 1999 Jul, 40(3), 419 - 22
Effects of a xylanase on individual bird variation, starch digestion throughout the intestine, and ileal and caecal volatile fatty acid production in chickens fed wheat; Choct M et al.; 1 . The effects of a xylanase on digesta viscosity and starch digestibility throughout the small intestine and volatile fatty acid production in the ileum and caeca were investigated in broilers fed on a low-metabolisable energy wheat diet . 2 . The xylanase reduced (P<0.01) duodenal (2.9 vs . 1.7), jejunal (4.6 vs . 2.3) and ileal (14.0 vs . 3.9) digesta viscosities (mPas) and increased AME (P<0.01) of the wheat and starch digestibility (P<0.5) in the jejunum and ileum . Between-bird variability in ME and digestibility of starch was also reduced . 3 . Enzyme supplementation reduced (P<0.05) fermentation in the ileum, but increased (P<0.05) it in the caeca . 4 . The anti-nutritive effect of soluble non-starch polysaccharides (NSP) is related to their ability to increase digesta viscosity along the gut; this in turn causes changes in gut microflora and efficiency of nutrient utilisation by the chicken . Use of appropriate enzymes is an effective way of dealing with grains with high NSP content in poultry diets.

Fiziol Zh, 1999, 45(4), 69 - 76
{The effect of melatonin on photoperiod changes in the glutathione system of the brain under acute hypoxia}; Zamors'kyi II et al.; The effect of acute hypobaric hypoxia against on the background of single-shot melatonin administration on the activity of glutathione ferments system (glutathione peroxidase, glutathione S-transferase and glucose-6-phosphate dehydrogenase) and the contents of reduced glutathione and malondialdehyde in the supernatant of juvenile male of white rats forebrain was investigated under three conditions of lighting--natural conditions of lighting in the spring-summer period of year, constant illumination and constant darkness during the one week . The constant illumination enhanced the lipid peroxidation with simultaneous decreasing in the activity of glutathione peroxidase in control animals . The constant illumination enhanced the lipid peroxidation with simultaneous decrease in the activity of glutathione peroxidase in control animals . The constant darkness reduced the intensity of free-radical oxidation improving the antioxidant neuron protection . The melatonin decreased the contents of malondialdehyde, raised the activity of glutathione enzyme system and eliminated the negative influence of constant illumination on forebrain antioxidant system in intact animals . The acute hypoxia resulted in the increase of lipid peroxidation and reduced the activity of antioxidant enzymes, which were most expressed in constant light conditions . The melatonin administration before of 30 minutes of acute hypoxia modeling reduced the intensity of oxidizing stress, which was generated by acute hypoxia.

Appl Environ Microbiol, 1999 Sep, 65(9), 4245 - 57
Temperature-dependent fermentation of D-sorbitol in Escherichia coli O157:H7; Bouvet OM et al.; The influence of growth temperature on the ability to ferment D-sorbitol was investigated in Escherichia coli O157:H7 . It was found that O157:H7 strains have a temperature-sensitive sorbitol phenotype . D-Sorbitol transport and sorbitol-6-phosphate dehydrogenase activities were expressed in sorbitol-fermenting cells grown at 30 degrees C but only at a low level at 40 degrees C . Sorbitol-positive variants able to transport D-sorbitol were easily selected at 30 degrees C from culture of Sor(-) E . coli O157:H7 strains.

Appl Environ Microbiol, 1999 Sep, 65(9), 3915 - 9
Inactivation of MXR1 abolishes formation of dimethyl sulfide from dimethyl sulfoxide in Saccharomyces cerevisiae; Hansen J; Dimethyl sulfide (DMS) is a sulfur compound of importance for the organoleptic properties of beer, especially some lager beers . Synthesis of DMS during beer production occurs partly during wort production and partly during fermentation . Methionine sulfoxide reductases are the enzymes responsible for reduction of oxidized cellular methionines . These enzymes have been suggested to be able to reduce dimethyl sulfoxide (DMSO) as well, with DMS as the product . A gene for an enzymatic activity leading to methionine sulfoxide reduction in Saccharomyces yeast was recently identified . We confirmed that the Saccharomyces cerevisiae open reading frame YER042w appears to encode a methionine sulfoxide reductase, and propose the name MXR1 for the gene . We found that Mxr1p catalyzes reduction of DMSO to DMS and that an mxr1 disruption mutant cannot reduce DMSO to DMS . Mutant strains appear to have unchanged fitness under several laboratory conditions, and in this paper I hypothesize that disruption of MXR1 in brewing yeasts would neutralize the contribution of the yeast to the DMS content in beer.

Plant Foods Hum Nutr, 1999, 53(2), 159 - 68
Effect of varied fermentation periods on the diabetogenic potential of toasted cassava granules; Ihedioha JI et al.; The effects of exclusive gari diets prepared using varied fermentation periods on the fasting blood glucose levels (FBGL) and glucose tolerance (GT) of rats were studied . Thirty growing male albino rats divided randomly into 5 groups of 6 rats each were used . The 5 groups were each fed gari diets fermented for 0, 24, 48, 72 hours; and a standard rat diet, respectively, for 8 weeks . Data on the total cyanogen content and % crude protein of the diets; FBGL, GT, body weight, water:feed consumption ratio and clinical observations of the rats were collected . Results showed that the total cyanogen content and % crude protein of the diets were depleted as fermentation periods increased . The FBGL of all the gari-fed rats were significantly elevated and their GT significantly impaired with a significant variation between the groups . The elevated FBGL and impaired GT which were found to increase with increase in fermentation period were strongly inversely correlated to the % crude protein content of the diets (r = -0.92 and -0.96 respectively) suggesting that the diabetogenic potential of gari diets strongly depended on its % crude protein content . Shortened fermentation periods leading to production of gari with high total cyanogen content did not induce higher diabetogenicity.

Plant Foods Hum Nutr, 1999, 53(2), 103 - 11
Reduction of some toxicants in Icacina mannii by fermentation with Saccharomyces cerevisiae; Antai SP et al.; The effect of fermentation on the level of some toxic components (hydrocyanic acid, phytic acid and oxalic acid) of Icacina mannii paste was investigated . The results indicate that fermentation caused a decrease in the level of these toxicants ranging from 178 mg/kg to 70 mg/kg for hydrocyanic acid, 638 mg/kg to 463 mg/kg for oxalic acid and 49 mg/kg to 21 mg/kg for phytic acid . When the paste was supplemented with active cells of Saccharomyces cerevisiae there was a marked decrease in the level of the three toxicants ranging from 84 mg/kg to 16 mg/kg for hydrocyanic acid, 374 mg/kg to 88 mg/kg for oxalic acid and 24 mg/kg to 5 mg/kg for phytic acid . Supplementing the paste with yeast cells at a concentration of 7.5 g/kg before fermenting, resulted in approximately a 90% reduction in the level of the three toxic compounds after 144 hours of fermentation.

J Antibiot (Tokyo), 1999 Jun, 52(6), 525 - 30
Scyphostatin, a neutral sphingomyelinase inhibitor from a discomycete, Trichopeziza mollissima: taxonomy of the producing organism, fermentation, isolation, and physico-chemical properties; Nara F et al.; We performed experiments to screen for neutral sphingomyelinase inhibitors using rat brain microsomes as an enzyme source . Among more than 10,000 microbial extracts tested, a mycelial extract of Trichopeziza mollissima SANK 13892 exhibited potent inhibitory activity . The active compound, scyphostatin, was purified by a series of chromatographies . Scyphostatin inhibited the enzyme with an IC50 value of 1.0 microM.

Eur J Oral Sci, 1999 Aug, 107(4), 251 - 9
Effect of in situ plaque mineral supplementation on the state of saturation of plaque fluid during sugar-induced acidogenesis; Pearce EI et al.; Dental plaque fluid is normally supersaturated with respect to enamel mineral but this may change to a state of undersaturation when plaque pH falls following sugar exposure, placing the adjacent enamel at risk of caries . We have determined the saturation status of the fluid in both resting and fermenting plaque following mineral supplementation . Eleven subjects abstained from oral hygiene and rinsed their mouth 3 times/d for 3 d with a placebo solution or with test solutions designed to enrich plaque with hydroxyapatite or fluorhydroxyapatite . On the morning of day 4, plaque samples were collected before and after exposure to 10% sucrose . Compared to the placebo, use of the test rinses resulted in significantly higher concentrations of Ca, P and F in plaque residue . In plaque fluid, higher post-sucrose Ca2+ free concentrations and saturation levels with respect to enamel mineral and fluorapatite were found after use of the hydroxyapatite rinse compared to the placebo, effects that probably resulted from the release of cell-bound Ca2+ as well as from the dissolution of apatite . Thus, some evidence was obtained that the test mouthrinses can counteract the fall in saturation level found when plaque is exposed briefly to sucrose . Potential long-term benefits of the test mouthrinses deserve further study.

Gene Ther, 1999 Aug, 6(8), 1482 - 8
pCOR: a new design of plasmid vectors for nonviral gene therapy; Soubrier F et al.; A totally redesigned host/vector system with improved properties in terms of safety has been developed . The pCOR plasmids are narrow-host range plasmid vectors for nonviral gene therapy . These plasmids contain a conditional origin of replication and must be propagated in a specifically engineered E . coli host strain, greatly reducing the potential for propagation in the environment or in treated patients . The pCOR backbone has several features that increase safety in terms of dissemination and selection: (1) the origin of replication requires a plasmid-specific initiator protein, pi protein, encoded by the pir gene limiting its host range to bacterial strains that produce this trans-acting protein; (2) the plasmid's selectable marker is not an antibiotic resistance gene but a gene encoding a bacterial suppressor tRNA . Optimized E . coli hosts supporting pCOR replication and selection were constructed . High yields of supercoiled pCOR monomers were obtained (100 mg/l) through fed-batch fermentation . pCOR vectors carrying the luciferase reporter gene gave high levels of luciferase activity when injected into murine skeletal muscle.

J Pediatr Gastroenterol Nutr, 1999 Sep, 29(3), 327 - 31
13C and H2 breath tests to study extent and site of starch digestion in children with cystic fibrosis; Amarri S et al.; BACKGROUND: Starch is an important source of energy for children with cystic fibrosis, but little is known about their capacity to digest it . METHODS: A 13C breath test was used to measure starch digestion and oxidation in 16 children with cystic fibrosis (median {range} age, 7.9 {4-15} years; 7 girls, 9 boys) and 5 normal healthy control subjects (median age, 8.3 {7-13} years; 3 girls, 2 boys) . A test meal of 13C flour and lactulose was consumed and breath samples were obtained half-hourly thereafter for 6 hours to measure 13C enrichment by isotope ratio mass spectrometry and H2 by electrochemistry . The test was repeated on 10 children with cystic fibrosis when they were taking pancreatic supplements . RESULTS: The median (range) cumulative percentage 13C dose recovery (cPDR), was 35% (18-52%) in control subjects, 18% (9-33%) in children with cystic fibrosis without enzymes, and 29% (22-51%) in those with pancreatic supplements . cPDR differed significantly between healthy control subjects and children with cystic fibrosis without enzymes (p = 0.01) and between children with cystic fibrosis with and without enzymes (p < 0.0001), but there was no difference between control subjects and children with cystic fibrosis taking enzymes (p = 0.5) . Eight children with cystic fibrosis had a cPDR within control range, and in six there was a second peak in 13CO2 enrichment coincident with an increase in H2 . CONCLUSIONS: Starch digestion and oxidation are diminished in children with cystic fibrosis, but pancreatic enzymes restored them to near normal levels . A second peak in 13CO2 enrichment, suggestive of colonic starch fermentation was absent in healthy children, but present in some children with cystic fibrosis and abolished by pancreatic enzymes.

Chem Biol, 1999 Sep, 6(9), 617 - 24
Multiple regulatory genes in the tylosin biosynthetic cluster of Streptomyces fradiae; Bate N et al.; BACKGROUND: The macrolide antibiotic tylosin is composed of a polyketide lactone substituted with three deoxyhexose sugars . In order to produce tylosin efficiently, Streptomyces fradiae presumably requires control mechanisms that balance the yields of the constituent metabolic pathways together with switches that allow for temporal regulation of antibiotic production . In addition to possible metabolic feedback and/or other signalling devices, such control probably involves interplay between specific regulatory proteins . Prior to the present work, however, no candidate regulatory gene(s) had been identified in S . fradiae . RESULTS: DNA sequencing has shown that the tylosin biosynthetic gene cluster, within which four open reading frames utilise the rare TTA codon, contains at least five candidate regulatory genes, one of which (tylP) encodes a gamma-butyrolactone signal receptor for which tylQ is a probable target . Two other genes (tylS and tylT) encode pathway-specific regulatory proteins of the Streptomyces antibiotic regulatory protein (SARP) family and a fifth, tylR, has been shown by mutational analysis to control various aspects of tylosin production . CONCLUSIONS: The tyl genes of S . fradiae include the richest collection of regulators yet encountered in a single antibiotic biosynthetic gene cluster . Control of tylosin biosynthesis is now amenable to detailed study, and manipulation of these various regulatory genes is likely to influence yields in tylosin-production fermentations.

Biol Rev Camb Philos Soc, 1999 Aug, 74(3), 259 - 76
The naturally occurring furanones: formation and function from pheromone to food; Colin Slaughter J; Three closely related 4-hydroxy-3(2H)-furanones have been found in a range of highly cooked foodstuffs where they are important flavour compounds with aroma threshold values as low as 20 micrograms kg-1 water (approximately 0.14 mumol l-1) . The compounds are formed mainly as a result of the operation of the Maillard reactions between sugars and amino acids during heating but one compound, 5-(or 2)-ethyl-2-(or 5)-methyl-4-hydroxy-3(2H)-furanone, appears in practice to be produced by yeast, probably from a Maillard intermediate, during the fermentation stages in the production of soy sauce and beer . The compounds are also important in the flavour of strawberry, raspberry, pineapple and tomato but the route of biosynthesis is unknown . Two 3-hydroxy-2(5H)-furanones, emoxyfuranone and sotolon, which are produced spontaneously from amino acids such as threonine and 4-hydroxy-L-leucine are major contributors to meaty and spicy/nutty flavours in foods . The biosynthesis of 5-(1,2-dihydroxyethyl)-3,4-dihydroxy-2(5H)-furanone (ascorbic acid, vitamin C) and 5-hydroxymethyl-3,4-dihydroxy-2(5H)-furanone (erythroascorbic acid) from sugars in plants and yeast, respectively, has been characterized to the enzymic level . After treatment with chlorine, humic waters contain a range of chloro-furanones, some of which, particularly 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), are powerful mutagens . The furanones which occur in foods are also mutagenic to bacteria and cause DNA damage in laboratory tests . However, these compounds are, in practice, very effective anti-carcinogenic agents in the diets of animals which are being treated with known cancer-inducing compounds such as benzo{alpha}pyrene or azoxymethane . Two of the food-derived furanones have antioxidant activity comparable to that of ascorbic acid . A biological function has been discovered for some of the furanones besides vitamin C . 5-Methyl-4-hydroxy-3(2H)-furanone is a male pheromone in the cockroach Eurycolis florionda (Walker) and the 2,5-dimethyl derivative deters fungal growth on strawberries and is an important component of the attractive aroma of the fruit . The red seaweed Delisea pulchra (Greville) Montagne produces a range of brominated furanones which prevent colonisation of the plant by bacteria by interfering with the acylated homoserine lactone (AHL) signalling system used by the bacteria for quorum sensing . In addition, these compounds can deter grazing by marine herbivores . It is proposed here that the evolved biological function of a number of furanones is to act as inter-organism signal molecules in several different systems . This has resulted in two coincidental effects which are important for humans . Firstly, the easily oxidized nature of the furanones in general, which is likely to be an important property in their functioning as signal molecules, results in both mutagenic and anti-carcinogenic activity . The balance of these two effects from compounds in the diet has yet to be fully established . Secondly, and more specifically, the 4-hydroxy-3(2H)-furanones associated with fruit aromas act to attract animals to the fruit, which ensures seed dispersal . In the case of humans, the coincidental synthesis of some of these compounds in foods during preparation results in these foods appearing particularly attractive through the transferred operation of the original signalling mechanisms.

Proc Nutr Soc, 1999 May, 58(2), 243 - 8
High-meat diets and cancer risk; Bingham SA; Up to 80% of breast, bowel and prostate cancers are attributed to dietary practices, and international comparisons show strong positive associations with meat consumption . Estimates of relative risk obtained from cohort investigations are in the same direction, although generally weak, and red and processed meats rather than white meat seem to be associated with elevated risk of colon cancer . In breast cancer, there are consistent associations with total meat intake and there is evidence of a dose response . Despite these associations with meat, existing studies suggest that vegetarians do not have reduced risk of breast, bowel or prostate cancer, but there are no quantitative estimates of amounts of meat consumed by meat eaters in these cohort studies . Possible mechanisms underlying epidemiological associations include the formation of heterocyclic amines in meat when it is cooked . These heterocyclic amines require acetylation by P450 enzymes, and individuals with the fast-acetylating genotype who eat high amounts of meat may be at increased risk of large-bowel cancer . NH3 and N-nitroso compounds (NOC) formed from residues by bacteria in the large bowel and probably also important . NH3 is a promotor of large-bowel tumours chemically induced by NOC, and some of the chromosomal mutations found in human colo-rectal cancer are consistent with effects of NOC and heterocyclic amines . However, the type, amount, and cooking method of meat or protein associated with increased risk are not certain . The effects of high levels of meat on NH3 and NOC output are not reduced by increasing the amount of fermentable carbohydrate in the diet, but interaction between meat, NSP and vegetable intakes on the risk of cancer has not been studied comprehensively . The interaction between dietary low-penetrance genetic polymorphic and somatic mutation factors has also been investigated to a limited extent . Current Department of Health (1998) recommendations are that meat consumption should not rise, and that consumers at the top end of the distribution should consider a reduction in intakes.

J Environ Sci Health B, 1999 Sep, 34(5), 849 - 58
Comparison of gas chromatography and immunoassay methods in quantifying fenitrothion residues in grape juice processed into alcoholic drinks; Dagher SM et al.; Wine and Arak, the national alcoholic drink in Lebanon, were prepared from grape juice fortified with fenitrothion to a concentration of 20ppm . Samples of the 11 fractions produced by the fermentation and distillation steps were analyzed for fenitrothion residues using gas chromatography (GC) and enzyme-linked immunosorbent assay (ELISA) . Results of residue analyses showed that the two techniques were highly correlated (r = 0.978) and indicated that fenitrothion was stable during the fermentation steps but not during distillation . The clarified wine 35 days later contained about 85% (15.3 ppm) of the fenitrothion concentration found in the juice as determined by GC analysis . Arak was prepared by a two-steps distillation of the clarified wine . The alcohol distillate and undistilled fraction from the first distillation contained 2.5 ppm and 5.8 ppm of fenitrothion, respectively . No fenitrothion residues were detected by both techniques in the four fractions collected from the second distillation step.

J Bacteriol, 1999 Sep, 181(17), 5309 - 16
Signal-dependent phosphorylation of the membrane-bound NarX two-component sensor-transmitter protein of Escherichia coli: nitrate elicits a superior anion ligand response compared to nitrite; Lee AI et al.; The Nar two-component regulatory system, consisting of the dual sensor-transmitters NarX and NarQ and the dual response regulators NarL and NarP, controls the expression of various anaerobic respiratory pathway genes and fermentation pathway genes . Although both NarX and NarQ are known to detect the two environmental signals nitrate and nitrite, little is known regarding the sensitivity and selectivity of ligand for detection or activation of the sensor-transmitters . In this study, we have developed a sensitive anion-specific in vitro assay for NarX autophosphorylation by using Escherichia coli membranes highly enriched in the full-length NarX protein . In this ATP- and magnesium-dependent reaction, nitrate elicited a greater signal output (i.e., NarX autophosphorylation) than did nitrite . Nitrate stimulation occurred at concentrations as low as 5 microM, and the half-maximal level of NarX autophosphorylation occurred at approximately 35 microM nitrate . In contrast, nitrite-dependent stimulation was detected only at 500 microM, while 3.5 mM nitrite was needed to achieve half-maximal NarX autophosphorylation . Maximal nitrate- and nitrite-stimulated levels of NarX phosphorylation were five and two times, respectively, over the basal level of NarX autophosphorylation . The presence of Triton X-100 eliminated the nitrate-stimulated kinase activity and lowered the basal level of activity, suggesting that the membrane environment plays a crucial role in nitrate detection and/or regulation of kinase activity . These results provide in vitro evidence for the differential detection of dual signaling ligands by the NarX sensor-transmitter protein, which modulates the cytoplasmic NarX autokinase activity and phosphotransfer to NarL, the cognate response regulator.

J Bacteriol, 1999 Sep, 181(17), 5250 - 6
Response of hya expression to external pH in Escherichia coli; King PW et al.; The hya operon of Escherichia coli is composed of the genes which synthesize uptake hydrogenase isoenzyme 1 (Hyd1) . Although hya expression and Hyd1 synthesis occur only under anaerobic conditions, Hyd1 is not essential for growth . In this study we used a hya'-'lacZ fusion to characterize parameters of anaerobic growth that maximize hya expression in an attempt to further elucidate Hyd1 function . We found that the expression pattern of hya followed a decline of external pH . In buffered media where the pH value was set, the onset of hya expression initiated earlier in growth and reached a greater peak level in acidic than in alkaline medium . When cultures expressing hya were shifted from acidic to alkaline conditions, hya expression was arrested; shifting from alkaline to acidic conditions stimulated hya expression . Maximal expression of hya under all growth conditions required the sigma factor RpoS and transcriptional regulators AppY and ArcA . In the absence of RpoS or AppY, the response of hya expression onset to external pH was evident and maximal hya levels remained greater in acidic than in alkaline medium . However, the absence of ArcA led to a diminished response of expression onset to external pH and the loss of elevated expression at an acidic external pH . The fermentation end product formate slightly altered hya expression levels but was not required for hya to respond to external pH . In contrast to hya expression, the onset of hyb operon expression, encoding uptake hydrogenase isoenzyme 2, was constitutive with respect to external pH . However, external pH did affect hyb expression levels, which, in contrast to hya, were maximal in alkaline rather than acidic medium.

Adv Biophys, 1999, 36, 159 - 206
Multiple sequence alignment: algorithms and applications; Gotoh O; Elucidation of interrelationships among sequence, structure, function, and evolution (FESS relationships) of a family of genes or gene products is a central theme of modern molecular biology . Multiple sequence alignment has been proven to be a powerful tool for many fields of studies such as phylogenetic reconstruction, illumination of functionally important regions, and prediction of higher order structures of proteins and RNAs . However, it is far too trivial to automatically construct a multiple alignment from a set of related sequences . A variety of methods for solving this computationally difficult problem are reviewed . Several important applications of multiple alignment for elucidation of the FESS relationships are also discussed . For a long period, progressive methods have been the only practical means to solve a multiple alignment problem of appreciable size . This situation is now changing with the development of new techniques including several classes of iterative methods . Today's progress in multiple sequence alignment methods has been made by the multidisciplinary endeavors of mathematicians, computer scientists, and biologists in various fields including biophysicists in particular . The ideas are also originated from various backgrounds, pure algorithmics, statistics, thermodynamics, and others . The outcomes are now enjoyed by researchers in many fields of biological sciences . In the near future, generalized multiple alignment may play a central role in studies of FESS relationships . The organized mixture of knowledge from multiple fields will ferment to develop fruitful results which would be hard to obtain within each area . I hope this review provides a useful information resource for future development of theory and practice in this rapidly expanding area of bioinformatics.

Arch Biochem Biophys, 1999 Sep 1, 369(1), 133 - 42
Human selenium-dependent thioredoxin reductase from HeLa cells: properties of forms with differing heparin affinities; Gorlatov SN et al.; The TrxRl form of thioredoxin reductase (TrxR) was the major form of the enzyme isolated from HeLa cells grown in a fermentor at 35 degrees C under controlled aeration conditions favorable to growth, nominally 30% of saturation of dissolved oxygen or 8 ml of oxygen in a liter of medium . This TrxR1 form was not retained on a heparin affinity matrix, it contained one selenium per subunit, was highly active catalytically, and showed strong cross-reactivity with anti-rat liver TrxR1 polyclonal antibodies . At higher aeration, 50% of saturation of dissolved oxygen or 12 ml of oxygen in a liter of medium, HeLa cell growth was slower and additional TrxR forms that bound to heparin were present in purified enzyme preparations . A minor component, TrxR2, the mitochondrial form of TrxR, was detected in the heparin-bound enzyme fraction . One enzyme form that contained less selenium (ca . 0.5 Se per TrxR subunit) was only about 50% as active with thioredoxin or 5,5'dithiobis(2-nitrobenzoic acid) as substrate . Cross-reactivity of this form with anti-rat liver TrxR1 polyclonal antibodies was very weak . The isoelectric point of the low Se enzyme, 5.85, was higher than that, 5.2-5.4, of normal Se content enzyme . Affinity of purified fully active TrxR1 to heparin could be induced by reduction with NADPH or tris-(2-carboxyethyl)phosphine (TCEP) . Under anaerobic conditions there was complete retention of Se indicating that an enzyme conformation change effected by reduction was involved . The TCEP-reduced enzyme form was very oxygen labile and upon exposure to air both the Se content and catalytic activity decreased by about 50% . Addition of millimolar concentrations of NADPH or NADP(+) to the TCEP-reduced enzyme gave full protection from oxygen inactivation . TrxR1 exhibited weak peroxidase activity with H(2)O(2) as substrate in the presence of an NADPH-generating system but this activity was unstable . Specific alkylation of the selenocysteine residue of TrxR1 which completely inhibits the NADPH-dependent reduction of disulfides also destroyed peroxidase activity .

J Anim Sci, 1999 Aug, 77(8), 2305 - 12
Effects of laidlomycin propionate and monensin on the in vitro mixed ruminal microorganism fermentation; Domescik EJ et al.; The objective of this study was to compare the effects of laidlomycin propionate and monensin on the in vitro fermentation of ground corn, Trypticase, or alfalfa hay by mixed ruminal microorganisms . Ruminal fluid was collected from two steers fed 9.27 kg DM of a high-concentrate (62.2% ground corn and 17.4% cottonseed hulls) diet per day and composited . In the first study, no ionophore was included in the diet; the diet in the second study contained 11.1 g of laidlomycin propionate per ton of feed . The animals were allowed an adjustment period of 14 d for each dietary treatment before samples were collected . When ruminal fluid from unadapted animals was used, both monensin and laidlomycin propionate decreased (P<.05) CH4 concentration and the acetate:propionate ratio with ground corn and alfalfa hay . Monensin reduced (P<.05) in vitro dry matter disappearance of alfalfa and increased (P<.05) final pH in the ground corn and alfalfa hay fermentations . Both laidlomycin propionate and monensin decreased (P<.05) concentrations of acetate, propionate, isobutyrate, isovalerate, CH4, and NH3 in Trypticase fermentations . When ruminal fluid from adapted animals was used, both ionophores still reduced the concentrations of most fermentation products . However, there was generally less inhibition compared with fermentations inoculated with unadapted mixed ruminal microorganisms . In the presence of 5 mM maltose, mixed ruminal bacteria produced high concentrations (10 to 11 mM) of lactate, and addition of both ionophores to these fermentations was effective in reducing (P<.05) lactate production . In conclusion, laidlomycin propionate alters the mixed ruminal microorganism fermentation in a manner similar to monensin, but, at the concentrations used in this study, monensin seemed to be a more potent inhibitor.

Biotechnol Bioeng, 1999 Oct 20, 65(2), 212 - 8
Wine yeast fermentation vigor may be improved by elimination of recessive growth-retarding alleles; Ramirez M et al.; The presence of recessive growth-retarding alleles can reduce the fitness of industrial wine yeasts . In nature, these alleles are supposed to be eliminated through "genome renewal" . We emulated this process in the laboratory to increase the fermentation vigor of wine yeasts . The procedure is simply to sporulate the yeast strains and select new homozygous single-spore descendants . Most of the yeasts achieve a faster onset of fermentation when recessive deleterious genes are eliminated . The increase of the degree of homozygosity has no relation, either direct or inverse, with the fermentation vigor of the yeasts or with the quality of the resulting wine . However, in some strains in which recessive growth-retarding alleles have been eliminated, the fermentation vigor and the quality of the wine were found to be improved simultaneously .

Ukr Biokhim Zh, 1999 Jan-Feb, 71(1), 86 - 8
{Effect of carnosine on the transformation of sodium nitrite in vitro in the presence of catalase}; Krys'ko OM et al.; It was shown that carnosine changes the compacting of ferment's globe . On the one hand, that action is causes with the connecting of haem . On the other hand, carnosine influences on the hydrate's membrane around protein . Dipeptide influences also on the interaction of catalase with sodium nitrite, which intensity depends on sequence of introducing HNO2 and carnosine into the medium.

Arh Hig Rada Toksikol, 1999 Mar, 50(1), 67 - 78
{Megamin, faith, hope and placebos---a critical review}; Momcilovic B; Megamin is a registered nutriceutical in Croatia . Allegedly, it consists mostly of a naturally occurring zeolite (aluminum silicate) ion-exchange material tagged with vitamins and minerals . Mass-media claimed that Megamin possessed strong anticancer properties although it had not been tested through an appropriate, randomized, double-blind, placebo-controlled, crossover clinical trial . The paper uses Megamin as a paradigmatic model to discuss how higher neurocognitive functions like hope and faith may be related to the placebo effect and give an illusion of symptomatic well being without supporting evidence in the objective signs of the disease . The only plausible role of Megamin is that it reduces gastrointestinal toxic burden by affecting the anaerobic fermenting processes after digestion of food and by removing harmful metabolites after medical treatment of cancer and/or liver and kidney organ failure . The psychological effect of unwarranted mass-media claims about the beneficial role of Megamin in cancer treatment is scrutinized.

FEBS Lett, 1999 Aug 6, 456(2), 232 - 8
Comparison of the effects of bax-expression in yeast under fermentative and respiratory conditions: investigation of the role of adenine nucleotides carrier and cytochrome c; Priault M et al.; A new system for bax-expression in yeast has been devised to investigate bax's effect under fermentative and respiro-fermentative conditions . This has allowed us to show unambiguously that the ability of bax to kill yeast is higher under respiratory conditions than under purely fermentative conditions . The extent of killing under respiro-fermentative conditions (non-repressive sugars) is intermediate . It has been proposed that the two proteins adenine nucleotides carrier (ANC) and cytochrome c play a crucial role in bax-induced cell death . We have investigated the effects of deletion of the genes encoding the two proteins on the toxicity induced by bax, using this new system . The absence of ANC did not modify bax-induced lethality in any way . Moreover, the absence of cytochrome c also did not prevent bax-induced death . Only the kinetics of lethality were altered . All these effects are prevented by co-expression of bcl-xL.

J Ind Microbiol Biotechnol, 1999 Jun, 22(6), 633 - 636
Production of xylitol by Candida peltata; Saha B et al.; The ability of Candida peltata NRRL Y-6888 to ferment xylose to xylitol was evaluated under different fermentation conditions such as pH, temperature, aeration, substrate concentration and in the presence of glucose, arabinose, ethanol, methanol and organic acids . Maximum xylitol yield of 0.56 g g-1 xylose was obtained when the yeast was cultivated at pH 6.0, 28 degrees C and 200 rpm on 50 g L-1 xylose . The yeast produced ethanol (0.41 g g-1 in 40 h) from glucose (50 g L-1) and arabitol (0.55 g g-1 in 87 h) from arabinose (50 g L-1) . It preferentially utilized glucose > xylose > arabinose from mixed substrates . Glucose (10 g L-1), ethanol (7.5 g L-1) and acetate (5 g L-1) inhibited xylitol production by 61, 84 and 68%, respectively . Arabinose (10 g L-1) had no inhibitory effect on xylitol production.

J Ind Microbiol Biotechnol, 1999 Jun, 22(6), 622 - 626
Production of raw-starch digesting amyloglucosidase by Aspergillussp GP-21 in solid state fermentation; Mamo G et al.; Aspergillus sp GP-21 produced a raw-starch digesting amyloglucosidase which showed optimum activity at 65 degrees C and pH 5.0-5.5 . At 50 degrees C the enzyme converted about 40% of raw corn starch to glucose within 48 h . Enzyme production was studied in solid state fermentation using wheat bran . Productivity was affected by the level of moisture, incubation temperature and the presence or absence of supplements . Maximum enzyme production was observed at a moisture level of 75% and at 30 degrees C . Enzyme production was stimulated by supplementing wheat bran with 0.25% proteose peptone, 1% trace mineral solution, 0.01% CaCl2 and 0.01% MgSO4.

J Ind Microbiol Biotechnol, 1999 Jun, 22(6), 575 - 581
Conversion of corn fiber to ethanol by recombinant E . coli strain FBR3; Dien B et al.; We have developed a novel ethanologenic Escherichia coli strain FBR3 that is an efficient biocatalyst for converting mixed sugar streams (eg, arabinose, glucose, and xylose) into ethanol . In this report, the strain was tested for conversion of corn fiber hydrolysates into ethanol . Corn fiber hydrolysates with total sugar concentrations of 7.5% (w/v) were prepared by reacting corn fiber with dilute sulfuric acid at 145 degrees C . Initial fermentations of the hydrolysate by strain FBR3 had lag times of approximately 30 h judged by ethanol production . Further experiments indicated that the acetate present in the hydrolysate could not solely account for the long lag . The lag phase was greatly reduced by growing the pre-seed and seed cultures on corn fiber hydrolysate . Ethanol yields for the optimized fermentations were 90% of theoretical . Maximum ethanol concentrations were 2.80% w/v, and the fermentations were completed in approximately 50 h . The optimal pH for the fermentation was 6.5 . Below this pH, sugar consumption was incomplete and above it, excess base addition was required throughout the fermentation . Two alternative neutralization methods (overliming and overliming with sulfite addition) have been reported for improving the fermentability of lignocellulosic hydrolysates . These methods further reduced the lag phase of the fermentation, albeit by a minor amount.

Yeast, 1999 Aug, 15(11), 1097 - 109
Deletion of SFI1, a novel suppressor of partial Ras-cAMP pathway deficiency in the yeast Saccharomyces cerevisiae, causes G(2) arrest; Ma P et al.; When glucose is added to Saccharomyces cerevisiae cells grown into stationary phase or on non-fermentable carbon sources a rapid loss of heat stress resistance occurs . Mutants that retain high stress resistance after addition of glucose are called 'fil', for deficient in fermentation induced loss of stress resistance . Transformation of the fil1 mutant, which harbours a point mutation in adenylate cyclase, with a yeast gene library on a single copy plasmid resulted in transformants that were again stress-sensitive . One of the genes isolated in this way was a gene of previously unknown function . We have called it SFI1, for suppressor of fil1 . SFI1 is an essential gene . Combination of Sfi1 and cAMP pathway mutations indicates that Sfi1 itself is not involved in the cAMP pathway . Conditional sfi1 mutants did not show enhanced heat resistance under the restrictive condition, whereas overexpression of SFI1 rendered cells heat-sensitive . Sfi1 may be a downstream target of the protein kinase A pathway, but its precise relationship with heat resistance remains unclear . Further analysis showed that Sfi1 is required for cell cycle progression, more specifically for progression through G(2)-M transition . Cells expressing SFI1 under the control of a galactose-inducible promoter arrest after addition of glucose as doublets of undivided mother and daughter cells . These doublets contain a single nucleus and lack mitotic spindles . Sfi1 shares homology with Xenopus laevis XCAP-C, a protein required for chromosome assembly . The conserved residues between these two proteins show a strong bias for charged amino acids . Hence, Sfi1 might be required for correct mitotic spindle assembly and its precise role might be in chromosome condensation . In conclusion, we have identified an essential function in the G(2)-M transition of the cell cycle for a yeast gene of previously unknown function .

Microb Pathog, 1999 Sep, 27(3), 145 - 53
Expression of the monocytic differentiation/activation factor P48 in Mycoplasma species; Hall RE et al.; P48 is a 48 kd monocytic differentiation/activation factor previously purified from the conditioned medium of the Reh human pre-B cell leukemia line . It induces differentiation of HL-60 promyelocytic leukemia cells along the monocytic pathway and production of IL1, TNF-alpha and IL6 in human monocytes and monocytic cell lines . Recently our laboratory isolated cDNA clones for P48 from Reh cells and genomic clones from Mycoplasma fermentans DNA and showed that P48 is a M . fermentans gene product . In this paper we report the analysis of P48 expression at the DNA, mRNA and protein levels in different Mycoplasma species . Polymerase Chain Reaction (PCR) analysis of extracted DNA using P48-specific oligonucleotide primers revealed P48 sequences in M . fermentans but not M . hominis, M . iowae, M . genitalium or M . capricolum . Southern analysis of Mycoplasma DNAs revealed hybridizing bands in M . fermentans and M . capricolum under low stringency, but only in M . fermentans under high stringency . Consistent with this, Northern blot studies revealed a single hybridizing transcript in M . fermentans but not in other Mycoplasma species tested . However, Western blot studies with anti-P48 antibodies revealed P48 antigenic material in M . fermentans, as well as M . hominis and M . iowae . These studies demonstrate that the gene for P48 is derived from M . fermentans or a closely related species and is absent in these other species tested . However, the P48 protein exhibits shared antigenic determinants among several Mycoplasma species which presently are of unknown function or significance . P48 is a Mycoplasma -derived immunomodulatory molecule which may be important in Mycoplasma pathophysiology and may be useful in understanding human haematopoietic differentiation and the control of cytokine biosynthesis .

Zhongguo Yi Xue Ke Xue Yuan Xue Bao, 1997 Oct, 19(5), 342 - 6
{The cloning, expression, purification and bioactivity identification of recombinant IGF-I}; Wei H et al.; The fragment containing IGF-I cDNA and cyc1 terminator from PSK-IGF-I plasmid was cloned into PSK43 SB treated by Hind III, Klenow and Cla I, generating a recombinant PSB-IGF-I (DH), which have double Hind III sites . This plasmid was treated by Hind III and Klenow, ligated to give a recombinant plasmid PSB-IGF-I(NH), which has no Hind III site . After analysed by restriction endonucleases and sequencing, the fusion site was verified to be correct . This plasmid was excised and a BamH I-Cla I fragment containing yeast alpha-factor promotor, leader sequence, IGF-I cDNA and cycl terminator was cloned into yeast episomal plasmid vector YEpHC8, generating a recombinant plasmid YEpHC8-IGF-I . Transformed this plasmid into yeast competence cells BJ1990 by using LiAc method . The expressed products were secrected into the medium broth and having the correct molecular weight of 8,100 on SDS-PAGE . The products of IGF-I were raised by using high-cell-density fermentation of Saccharomyces cerevisiae . The cleared supernatant of yeast medium was applied to Bio-Rex 70 resin, eluted with 1 mol/L ammonium acetate, pH8.0 . Main peak was pooled and applied to Bio-Gel P10 (200-400 mesh) . The second peak was collected and get relatively pure IGF-I proteins . The biological activities of IGF I product was assayed in NIH3T3 cells by using MTT method . The results show that the expressed IGF-I can obviously stimulate NIH3T3 cells to proliferate at the concentration ranging from 10 ng/ml to 50 ng/ml, suggesting that the protein has its biological activities.

Eur J Clin Nutr, 1999 Jul, 53(7), 503 - 13
Guidelines for sugar consumption in Europe: is a quantitative approach justified?
Ruxton CH, Garceau FJ, Cottrell RC.
OBJECTIVES: There is incongruity between the sugar consumption guidelines set in different European countries . A number have adopted maximum limits ranging from 10-25% energy, while others have no quantitative recommendations at all . This raises the question whether or not there should be a common European guideline for sugar consumption . DESIGN: This paper examines if such a goal for sugar is merited and reviews the published literature on associations between sugar consumption and dental caries, obesity and micronutrient dilution . RESULTS: Evidence showed that higher intakes of sugar were related to leanness, not obesity, and had no detrimental effects on micronutrient intakes in most people . In the case of dental caries, there was a relationship between frequency of sugar intake and the incidence of decay . However, in populations where fluoride use was adequate, associations between sugar intake and caries rarely reached statistical significance . CONCLUSIONS: The available evidence does not justify a common quantitative recommendation for sugar . It is suggested that dental caries merits a more integrated public health approach where advice on the frequency of foods containing fermentable-carbohydrates is placed in context alongside oral hygiene.

Biosens Bioelectron, 1999 May 31, 14(5), 481 - 93
An optical biosensor for monitoring recombinant proteins in process media; Disley DM et al.; This paper describes the construction of a sensor for the direct monitoring of a recombinant protein, the human insulin analogue (MI3) . The surface plasmon resonance (SPR) sensor incorporates an immobilised, sterilisable affinity-ligand that has been designed to bind to MI3 . In practice, gold SPR devices were fabricated with; a 2D assembly of ethanethiol-modified ligand, a 2D mixed-assembly of ethanethiol-modified ligand and mercaptoethanol, a 3D coating of ligand-modified terminal-thiolated poly(vinyl)alcohol (PVA) or a 3D hydrogel of dextran coupled to a self-assembled monolayer (SAM) of mercaptohexaneundecanl-ol . Routine measurement of the concentration MI3 in the concentration range 1-100 mg/l in pilot-scale samples of crude fermentation broth have been achieved with high sensitivity levels and a high signal-to-noise ratio . Analysis can be achieved within < 10 min with the active surface being regenerable for at least 60 cycles over a 6 month period . The coupling of a robust, sterilisable and highly-selective sensor-coating with suitable transducer technologies promises to deliver sensors that are capable of direct in situ monitoring of biopharmaceuticals in industrial bioprocesses.

Bioorg Med Chem Lett, 1999 Jul 19, 9(14), 2003 - 6
Lobophorins A and B, new antiinflammatory macrolides produced by a tropical marine bacterium; Jiang ZD et al.; Two new antiinflammatory macrolides, lobophorins A and B (1 and 2), have been isolated from fermentation broths of a marine bacterium isolated from the surface the Caribbean brown alga Lobophora variegata (Dictyotales) . The new compounds, distantly related to antibiotics of the kijanimicin class, are potent inhibitors of topical PMA-induced edema in the mouse ear assay when administered either topically or IP.

J Nutr Sci Vitaminol (Tokyo), 1999 Apr, 45(2), 173 - 81
Secretion and excretion of immunoglobulin A to cecum and feces differ with type of indigestible saccharides; Kudoh K et al.; The study was conducted to elucidate the effects of orally administered indigestible saccharides (IDS) on immunoresponses of the intestinal tracts, especially secretion and excretion of immunoglobulin A (IgA) . Male 4-week-old Sprague-Dawley rats were fed diets containing several kinds of IDS (cellulose, corn husk, glucomannan, curdlan and lactulose) at 5% for three weeks . The results indicated that the proportion of IgA-presenting lymphocytes in the cecal mucosa of the tested IDS groups increased significantly or tended to increase compared with that of the cellulose group . No significant differences among the experimental groups were observed in the CD4(+)- and CD8(+)-presenting lymphocytes and the CD4+/CD8+ ratios in the small intestine, cecum and mesenteric lymph nodes . IgA amounts in the cecal contents increased significantly in the glucomannan and curdlan groups as compared with that in the cellulose group . The inconsistent results were observed in the cecal IgA amounts of the lactulose group . Although IgA excretion into feces increased periodically in the cellulose, hardly any changes were observed in the glucomannan and curdlan groups . These results revealed that IgA secretion from cecal mucosa to contents was promoted, and its excretion to feces was decreased by the oral administration of highly fermentable IDS, respectively, while non- or low-fermentable IDS functioned adversely to IgA responses in the intestinal tract . It is suggested that the response of IgA in the intestinal immune system differs with the type of IDS ingested.

Proc Natl Acad Sci U S A, 1999 Aug 17, 96(17), 9721 - 6
Systematic changes in gene expression patterns following adaptive evolution in yeast; Ferea TL et al.; Culturing a population of Saccharomyces cerevisiae for many generations under conditions to which it is not optimally adapted selects for fitter genetic variants . This simple experimental design provides a tractable model of adaptive evolution under natural selection . Beginning with a clonal, founding population, independently evolved strains were obtained from three independent cultures after continuous aerobic growth in glucose-limited chemostats for more than 250 generations . DNA microarrays were used to compare genome-wide patterns of gene expression in the evolved strains and the parental strain . Several hundred genes were found to have significantly altered expression in the evolved strains . Many of these genes showed similar alterations in their expression in all three evolved strains . Genes with altered expression in the three evolved strains included genes involved in glycolysis, the tricarboxylic acid cycle, oxidative phosphorylation, and metabolite transport . These results are consistent with physiological observations and indicate that increased fitness is acquired by altering regulation of central metabolism such that less glucose is fermented and more glucose is completely oxidized.

Curr Opin Biotechnol, 1999 Aug, 10(4), 358 - 64
Cellulase for commodity products from cellulosic biomass
Himmel ME, Ruth MF, Wyman CE.
A vital objective for second millennium biotechnology will be the enzymatic conversion of renewable cellulosic biomass to inexpensive fermentable sugars; new and more efficient fermentation processes will convert this biological 'currency' to a variety of commodity products . Although early strides will be made using process development and engineering disciplines, mid-term and longer advances must rely heavily on insight gained through protein and metabolic engineering technologies . These challenging goals can be met most effectively by the full integration of academic, federal, and industrial efforts in teams that develop and apply new fundamental knowledge to key cost drivers.

Eur J Biochem, 1999 Aug, 264(1), 176 - 82
Acetylene hydratase of Pelobacter acetylenicus . Molecular and spectroscopic properties of the tungsten iron-sulfur enzyme; Meckenstock RU et al.; Acetylene hydratase of Pelobacter acetylenicus is a tungsten iron-sulfur protein involved in the fermentation of acetylene to ethanol and acetate . Expression of the enzyme was increased 10-fold by feeding a 50-L batch culture continuously with 104 Pa acetylene at pH 6.8-7.0 . Acetylene hydratase was purified to homogeneity by a three-step procedure in either the absence or presence of dioxygen . The enzyme was a monomer with a molecular mass of 73 kDa (SDS/PAGE) or 83 kDa (matrix-assisted laser-desorption ionization MS) and contained 0.5 +/- 0.1 W (inductively coupled plasma/MS) and 1.3 +/- 0.1 molybdopterin-guanine dinucleotide per mol . Selenium was absent . EPR spectra (enzyme as isolated, under air) showed a signal typical of a {3Fe-4S} cluster with gav = 2.01, at 10 K . In enzyme prepared under N2/H2, this signal was absent and reaction with dithionite led to a rhombic signal with gz = 2.048, gy = 1.939 and gx = 1.920 indicative of a low-potential ferredoxin-type {4Fe-4S} cluster . Upon oxidation with hexacyanoferrate(III), a new signal appeared with gx = 2.007, gy = 2.019 and gz = 2.048 (gav = 2.022), which disappeared after further oxidation . The signal was still visible at 150 K and was tentatively assigned to a W(V) center . The iron-sulfur center of acetylene hydratase (prepared under N2/H2) gave a midpoint redox potential of -410 +/- 20 mV in a spectrophotometric titration with dithionite . Enzyme activity depended on the redox potential of the solution, with 50% of maximum activity at -340 +/- 20 mV . The presence of a pterin-guanine dinucleotide cofactor differentiates acetylene hydratase from the aldehyde ferredoxin oxidoreductase-type enzymes which have a pterin mononucleotide cofactor.

Scand J Infect Dis, 1999, 31(2), 141 - 4
Haemolytic-uraemic syndrome caused by vero toxin-producing Escherichia coli serotype Rough: K-: H49; Keskimaki M et al.; The first case of haemolytic-uraemic syndrome (HUS) caused by Vero toxin-producing Escherichia coli (VTEC) which belonged to a novel serotype, Rough: K-: H49, is reported . The case was initially diagnosed as nephropathia epidemica caused by Puumala virus, but the subsequent diagnosis of HUS caused by VTEC was made after bacteriological investigation . The strain isolated fermented sorbitol produced VT2 toxin but not enterohaemolysin, nor did it carry the eaeA gene . In VTEC strains, the O antigen, the eaeA gene and enterohaemolysin production have been characterized as virulence-associated factors and believed to have an effect on pathogenesis of these strains to cause haemorrhagic colitis or HUS . The findings of this study demonstrate that there is a need for further studies to evaluate the pathogenetic mechanism of VTEC and need for easy diagnostic methods exploiting other properties than O157 antigen and non-fermentation of sorbitol to find all VTEC in human infections.

Can J Microbiol, 1999 May, 45(5), 423 - 6
The use of babassu oil as substrate to produce bioemulsifiers by Candida lipolytica; Sarubbo LA et al.; Candida lipolytica IA 1055 produced an extracellular emulsifier when using babassu oil as its sole carbon source during batch and fed batch fermentations at 27 degrees C . Emulsification activity was detected after 60 h of growth in all conditions studied . The bioemulsifier was isolated after 144 h of fermentation from the best condition studied . The biopolymer seems to be a polysaccharide-protein-lipid complex.

Can J Microbiol, 1999 May, 45(5), 418 - 22
Cloning, sequencing, and expression of cscA invertase from Escherichia coli B-62; Sahin-Toth M et al.; We have isolated a 2.5-kb DNA fragment from plasmid pST5R7 encoding a sucrose utilization system from Escherichia coli B-62 which confers a sucrose-fermenting phenotype to transformed E . coli K-12 strains . DNA-sequence determination revealed one full-length open reading frame 98% identical to cscA, the sucrose-hydrolase (invertase) gene of the csc regulon from E . coli EC3132 . Functional characterization indicates that high-level expression and limited periplasmic release of invertase is responsible for the sucrose-fermenting capacity of transformed E . coli K-12 strains carrying cscA.

Acta Odontol Scand, 1999 Apr, 57(2), 65 - 71
Dental plaque fluoride and pH in children exposed to different water fluoride levels; Sampaio FC et al.; The aims of this study were to determine plaque fluoride concentrations in children not exposed to topical fluorides but to different fluoride levels in the drinking water (0.1 and 2.0 ppm), and to observe whether plaque fluoride was related to plaque pH . Twenty-five children (6 to 7 years old) were selected from two rural villages in Brazil . A sub-set of subjects was examined for resting and fermenting plaque pH before sampling . A maximum of 14 sites was studied in each subject (vestibular and interproximal of first molars and central incisors) . Plaque fluoride was extracted and measured with an inverted fluoride electrode under oil . Amounts of plaque were determined by protein analysis . Mean values in the 0.1 ppm village were 1.3 ngF/mg of plaque wet weight (SD = 1.1) and in the 2.0 ppm village 2.5 ngF/mg (SD = 2.1) and were not statistically different (Kruskal-Wallis test, P = 0.09) . Plaque fluoride varied considerably from site to site in the same mouth . Combining sites in all subjects, plaque fluoride concentrations were positively related to resting and fermenting pH (regression analysis, P< 0.01-0.001, adjR2 = 0.12-0.31) . On an individual basis the same trend was found for fermenting, but not for resting pH . In conclusion, our findings showed a moderate influence of water fluoride upon dental plaque fluoride concentrations and give some support to the theory that low fermenting pH may contribute to the release of bound plaque fluoride.

Sci Prog, 1999, 82 ( Pt 2), 171 - 84
Engineering approaches to chewing and digestion; Alexander RM; The guts of people and animals function like industrial chemical plants . They are assemblies of tubes and tanks in which foods are hydrolysed by enzyme-catalysed reactions, or fermented by microorganisms . Raw materials enter at one end, waste matter is voided at the other, and valuable products are abstracted on the way . A mill at the entrance end reduces the raw materials to small fragments, enabling the reactions to proceed faster . This paper shows how ideas from chemical engineering are guiding research on the gut, giving much clearer understanding of how foods respond to chewing, and of how guts are designed to process different foods . We will discuss the teeth as a grinding mill, and the digestive tube as a chain of chemical reactors.

Biotechnol Prog, 1999 Jul, 15(4), 617 - 21
Predicting fermentability of wood hydrolyzates with responses from electronic noses
Mandenius CF, Lid n H, Eklov T, Taherzadeh MJ, Lid n G.
The fermentability of lignocellulose hydrolyzates have been predicted from the responses of a combination of chemical gas sensors . The hydrolyzates were prepared by dilute-acid hydrolysis of wood from pine, aspen, birch, and spruce . The volatile emission from the hydrolyzates before fermentation was measured, and the sensor array response pattern was compared with the observed fermentability of the hydrolyzates, i.e . with the final ethanol concentration after fermentation and the maximum specific ethanol production rate . Two concentration parameters in the hydrolyzates, furfural and the sum of furfural and 5-(hydroxymethyl)furfural (HMF), were also predicted from the responses . The sensors used were metal oxide semiconductor field effect transistors (MOSFET), tin oxide semiconductor devices, and conductive polymer sensors configured in two sensor arrays . The sensor array response pattern was analyzed by principal component analysis and artificial neural networks . Predictions from artificial neural networks deviated from measured values with less than 15%.

J Biotechnol, 1999 Jul 2, 72(3), 157 - 67
Rapid analysis of the expression of heterologous proteins in Escherichia coli using pyrolysis mass spectrometry and Fourier transform infrared spectroscopy with chemometrics: application to alpha 2-interferon production; McGovern AC et al.; Cell pastes and supernatant Escherichia coli samples, taken from an industrial bioprocess overproducing recombinant alpha 2 IFN were analysed using pyrolysis mass spectrometry (PyMS) and diffuse reflectance-absorbance Fourier transform infrared spectroscopy (FT-IR) . PyMS and FT-IR are physico-chemical methods which measure predominantly the bond strengths of molecules and the vibrations of bonds within functional groups, respectively . They therefore give quantitative information about the total biochemical composition of the bioprocess sample . The interpretation of these hyperspectral data, in terms of the quantity of alpha 2 IFN in the cell pastes and supernatant samples was possible only after the application of the 'supervised learning' methods of artificial neural networks (ANNs) and partial least squares (PLS) regression . Both PyMS and FT-IR are novel, rapid and economical methods for the screening and the quantitative analysis of complex biological bioprocess over producing recombinant proteins . Models established using either spectral data set had a similarly satisfactory predictive ability . This shows that whole-reaction mixture spectral methods, which measure all molecules simultaneously, do contain enough information to allow their quantification when the entire spectra are used as the inputs to methods based on supervised learning . Moreover, this is the first study where FT-IR in the mid-IR range has been used to quantify the expression of a heterologous protein directly from fermentation broths and the first study to compare the abilities of PyMS and FT-IR for the quantitative analyses of an industrial bioprocess.

Inflamm Res, 1999 Jun, 48(6), 296 - 300
Biogenic amines in foods: histamine and food processing; Bodmer S et al.; Biogenic amines, e.g . histamine, occur in many different foods . At high concentrations, they are risk factors for food intoxication, whereas moderate levels may lead to food intolerance . Sensitive persons, with insufficient diamine oxidase activity, suffer from numerous undesirable reactions after intake of histamine containing foods . Besides spoiled foodstuffs, especially fermented foods tend to contain elevated levels of biogenic amines, although their concentrations vary extensively not only between different food varieties but also within the varieties themselves . High histamine content in foods and beverages result from microbial contamination . The evidence of enteral histaminosis represents a challenge for the food industry to produce foods with histamine levels as low as possible . We therefore investigated critical steps for histamine formation during food production processes, and established production methods that include low-histamine technology.

Nat Toxins, 1999, 7(1), 1 - 23
Biological detoxification of fungal toxins and its use in plant breeding, feed and food production; Karlovsky P; Enzymatic inactivation of fungal toxins is an attractive strategy for the decontamination of agricultural commodities and for the protection of crops from phytotoxic effects of fungal metabolites . This review summarizes research on the biological detoxification of fungal toxins by microorganisms and plants and its practical applications . Some mycotoxins are detoxified during ensiling and other fermentation processes (aflatoxins, alternariol, mycophenolic acid, patulin, PR toxin) while others are transformed into toxic products or survive fermentation unchanged . Plants can detoxify fomannoxin, fusaric acid, HC-toxin, ochratoxin A and oxalate but the degradation of deoxynivalenol has yet to be proven . Microflora of the digestive tract of vertebrates and invertebrates exhibit detoxification activities towards aflatoxins, ochratoxin A, oxalate and trichothecenes . Some toxin-producing fungi are able to degrade or transform their own products under suitable conditions . Pure cultures of bacteria and fungi which detoxify mycotoxins have been isolated from complex microbial populations by screening and enrichment culture techniques . Genes responsible for some of the detoxification activities have been cloned and expressed in heterologous hosts . The detoxification of aflatoxins, cercosporin, fumonisins, fusaric acid, ochratoxin A, oxalic acid, patulin, trichothecenes and zearalenone by pure cultures is reviewed . Finally, current application of these results in food and feed production and plant breeding is summarized and expected future developments are outlined .

Biotechnol Bioeng, 1999 Oct 5, 65(1), 54 - 64
Monitoring GFP-operon fusion protein expression during high cell density cultivation of Escherichia coli using an on-line optical sensor; DeLisa MP et al.; Synthesis of an operon fusion protein was investigated in batch and fed-batch cultures at high cell densities of recombinant Escherichia coli JM105 {pBAD-GFP::CAT} . Glucose-limited growth was achieved without accumulation of inhibitory byproducts allowing high cell densities (110 g L(-1) DCW) to be attained . This was believed to be the highest reported value for dry cell mass of E . coli strain JM105 expressing two recombinant proteins . Transcription of the two reporter genes, green fluorescent protein (GFP) and chloramphenicol acetyltransferase (CAT), was under the control of the p(BAD) promoter of the araBAD (arabinose) operon . Each protein was independently translated via separate ribosome binding sites . CAT served as a model recombinant protein product to illustrate the noninvasive quantitative reporting ability of GFP during high cell density fermentations . Expression of GFP was monitored on-line using an intensity-based optical sensor . A linear correlation between the on-line GFP intensity and the enzymatic activity of CAT allowed for in vivo real-time quantitative monitoring of a fermentation product under conditions of high biomass concentration and high productivity .

Biotechnol Bioeng, 1999 Oct 5, 65(1), 24 - 33
Effect of selected aldehydes on the growth and fermentation of ethanologenic Escherichia coli; Zaldivar J et al.; Bioethanol production from lignocellulosic raw-materials requires the hydrolysis of carbohydrate polymers into a fermentable syrup . During the hydrolysis of hemicellulose with dilute acid, a variety of toxic compounds are produced such as soluble aromatic aldehydes from lignin and furfural from pentose destruction . In this study, we have investigated the toxicity of representative aldehydes (furfural, 5-hydroxymethlyfurfural, 4-hydroxybenzaldehyde, syringaldehyde, and vanillin) as inhibitors of growth and ethanol production by ethanologenic derivatives of Escherichia coli B (strains KO11 and LY01) . Aromatic aldehydes were at least twice as toxic as furfural or 5-hydroxymethylfurfural on a weight basis . The toxicities of all aldehydes (and ethanol) except furfural were additive when tested in binary combinations . In all cases, combinations with furfural were unexpectedly toxic . Although the potency of these aldehydes was directly related to hydrophobicity indicating a hydrophobic site of action, none caused sufficient membrane damage to allow the leakage of intracellular magnesium even when present at sixfold the concentrations required for growth inhibition . Of the aldehydes tested, only furfural strongly inhibited ethanol production in vitro . A comparison with published results for other microorganisms indicates that LY01 is equivalent or more resistant than other biocatalysts to the aldehydes examined in this study .

Microbiology, 1999 Jul, 145 ( Pt 7), 1547 - 62
Role of Escherichia coli RpoS, LexA and H-NS global regulators in metabolism and survival under aerobic, phosphate-starvation conditions; Gerard F et al.; It has been suggested that Escherichia coli can resist aerobic, glucose-starvation conditions by switching rapidly from an aerobic to a fermentative metabolism, thereby preventing the production by the respiratory chain of reactive oxygen species (ROS) that can damage cellular constituents . In contrast, it has been reported that E . coli cannot resist aerobic, phosphate (Pi)-starvation conditions, probably because of the maintenance of an aerobic metabolism and the continuous production of ROS . This paper presents evidence that E . coli cells starved for Pi under aerobic conditions indeed maintain an active aerobic metabolism for about 3 d, which allows the complete degradation of exogenous nutrients such as arginine (metabolized probably to putrescine via the SpeA-initiated pathway) and glucose (metabolized notably to acetate), but cell viability is not significantly affected because of the protection afforded against ROS through the expression of the RpoS and LexA regulons . The involvement of the LexA-controlled RuvAB and RecA proteins with the RecG and RecBCD proteins in metabolism and cell viability implies that DNA double-strand breaks (DSB), and thus hydroxyl radicals that normally generate this type of damage, are produced in Pi-starved cells . It is shown that induction of the LexA regulon, which helps protect Pi-starved cells, is totally prevented by introduction of a recB mutation, which indicates that DSB are actually the main DNA lesion generated in Pi-starved cells . The requirement of RpoS for survival of cells starved for Pi may thus be explained by the role played by various RpoS-controlled gene products such as KatE, KatG and Dps in the protection of DNA against ROS . In the same light, the degradation of arginine and threonine may be accounted for by the synthesis of polyamines (putrescine and spermidine) that protect nucleic acids from ROS . Besides LexA and RpoS, a third global regulator, the nucleoid-associated protein H-NS, is also shown to play a key role in Pi-starved cells . Through a modulation of the metabolism during Pi starvation, H-NS may perform two complementary tasks: it helps maintain a rapid metabolism of glucose and arginine, probably by favouring the activity of aerobic enzymes such as the NAD-dependent pyruvate dehydrogenase complex, and it may enhance the cellular defences against ROS which are then produced by increasing RpoS activity via the synthesis of acetate and presumably homoserine lactone.

FEMS Microbiol Lett, 1999 Aug 1, 177(1), 177 - 85
Saccharomyces uvarum, a distinct group within Saccharomyces sensu stricto; Rainieri S et al.; A natural subgroup (that we refer to as Saccharomyces uvarum) was identified, within the heterogeneous species Saccharomyces bayanus . The typical electrophoretic karyotype, interfertility of hybrids between strains, distinctive sugar fermentation pattern, and uniform fermentation characteristics in must, indicated that this subgroup was not only highly homogeneous, but also clearly distinguishable from other species within the Saccharomyces sensu stricto group . Investigation of the S . bayanus type strain and other strains that have been classified as S . bayanus, confirmed the apparent lack of homogeneity and, in some cases, supported the hypothesis that they are natural hybrids.

FEMS Microbiol Lett, 1999 Aug 1, 177(1), 67 - 73
Fermentative toluene degradation in anaerobic defined syntrophic cocultures; Meckenstock RU; A syntrophic coculture of a new sulfate-reducing isolate, strain TRM1, with Wolinella succinogenes degraded toluene with either fumarate or NO3- as the terminal electron acceptor . Neither strain TRM1 nor W . succinogenes could metabolise toluene under these conditions in pure culture . Syntrophic degradation was 2-3 times slower than toluene utilisation by strain TRM1 in pure culture with sulfate as electron acceptor . The culture did not produce benzoate or fatty acids like acetate or propionate in detectable amounts . An increase in biomass of the syntrophic toluene-degrading culture was shown in a growth curve with nitrate as the terminal electron acceptor . Both partner organisms were detected microscopically at the end of the growth experiment . Syntrophic degradation of toluene with W . succinogenes and fumarate as the terminal electron acceptor was also demonstrated with the iron reducer Geobacter metallireducens . The results provide the first example of a fermentative oxidation of an aromatic hydrocarbon in a defined coculture.

Appl Biochem Biotechnol, 1999 May, 80(2), 107 - 20
Dissolved oxygen concentration affects the accumulation of HIV-1 recombinant proteins in Escherichia coli; Qoronfleh MW; A central problem in aerobic growth of any culture is the maintenance of dissolved oxygen concentration (DOC) above growth-limiting levels especially in high-cell density fermentations that are usually of the fed-batch type . Fermentor studies have been conducted to determine the influence of DOC on the production of heterologous proteins in Escherichia coli . The results demonstrated that there is a significant degree of product-to-product variation in the response of heterologous protein accumulation to DOC . For translational fusions of the human immunodeficiency virus-1 (HIV-1) proteins p24Gag and Env41, the imposition of a dissolved oxygen (DO) limitation resulted in 100 and 15% increases in the respective product yields . On the other hand, the imposition of a DO limitation had no effect on the production of a similar translational fusion of the HIV-1 protein p55Gag, and a large negative effect on the production of an influenza protein (C13) . The stimulatory effects of DOC on p24Gag production were investigated further . The results of my studies suggested that the stimulatory effect observed at reduced agitation rates on p24Gag accumulation was owing to an oxygen effect and not a shear effect . Furthermore, the results of my investigations indicated that the effect a DOC had on the production of p24Gag was strongly influenced by the cell density at which the culture was induced.

Vet Hum Toxicol, 1999 Aug, 41(4), 251 - 7
An analysis of the chronic oral toxicity of polyether ionophore antibiotics in animals; Oehme FW et al.; Feeding well-mixed ionophores to adapted cattle improves ruminal fermentation and growth rates . In nonruminants, growth is improved by reducing competing gastrointestinal microorganisms . Interactions of monensin with other drugs may be beneficial or toxic . Tiamulin and furazolidone potentiate monensin's negative effects . For example, monensin produces positive inotropy and cardiomyopathy dependent on calcium and extracellular sodium . Based on available toxicity data and derived no observable effect levels (NOEL) in the same species and across species, monensin was more toxic than salinomycin, lasalocid or narasin . Lasalocid was 5- to 10-fold less toxic to horses than is monensin . Based on available toxicity data and derived NOEL, lasalocid was less toxic than all ionophores except salinomycin . Very high levels of narasin caused death in sows, leg muscle weakness in turkeys, and cardiopulmonary clinical signs in 15% of the rabbits from Brazilian rabbit farms . Only salinomycin and lasalocid were less toxic than narasin . Salinomycin was the least toxic of all the ionophores . Maduramicin was the most toxic of all the ionophores . Nearly all maduramicin fed to poultry persists in litter (manure), making this poultry litter toxic if fed to cattle as a nitrogen source . While ionophore comparative toxicity was difficult to estimate, most cross-comparisons utilized NOEL within and across species . The relative toxicities of the ionophores from lowest to highest were salinomycin < lasalocid < or = narasin < or = monensin (but lasalocid < monensin) < maduramicin.

J Appl Microbiol, 1999 Jul, 87(1), 29 - 40
Characterization of Candida krusei strains from spontaneously fermented maize dough by profiles of assimilation, chromosome profile, polymerase chain reaction and restriction endonuclease analysis; Hayford AE et al.; Several isolates of Candida krusei from indigenous spontaneously fermented maize dough have been characterized for the purpose of selecting appropriate starter cultures and methods for their subspecifies typing . The present work describes the occurrence of C . krusei in Ghanaian fermented maize dough . For detailed pheno- and genotyping, 48 representative isolates were selected and comparison was made with clinical isolates of C . krusei and reference cultures . The techniques applied included the assimilation of carbon compounds by the API ID 32 C kit, determination of chromosome profile by pulse field gel electrophoresis, polymerase chain reaction (PCR) profiles, restriction endonuclease analysis (REA) and Southern blot hybridization . For the 48 isolates tested, 82% had the same assimilation profiles, being able to assimilate N-acetyl-glucosamine, DL-lactate, glycerol and to ferment glucose . Chromosome and PCR profiles, REA and Southern blot hybridization techniques all had a high discriminatory power and revealed DNA polymorphism, which allowed for discrimination among the strains and hence subspecific typing . On the basis of PCR and REA profiles, isolates were grouped into clusters . Southern blot hybridization appeared to be the most sensitive with respect to strain specificity . Our results demonstrated that the three methods, PCR, REA and Southern blot hybridization, were suitable tools, easy to analyse, fast (with regard to PCR) and reliable methods for the typing of C . krusei isolates to species and below species level . Based on the use of these techniques, we demonstrated that several strains of C . krusei were involved in the fermentation of maize dough from the onset and remain dominant throughout the fermentation.

J Ethnopharmacol, 1999 Jul, 66(1), 11 - 7
Antioxidant and eicosanoid enzyme inhibition properties of pomegranate seed oil and fermented juice flavonoids; Schubert SY et al.; The antioxidant and eicosanoid enzyme inhibition properties of pomegranate (Punica granatum) fermented juice and seed oil flavonoids were studied . The pomegranate fermented juice (pfj) and cold pressed seed oil (pcpso) showed strong antioxidant activity close to that of butylated hydroxyanisole (BHA) and green tea (Thea sinensis), and significantly greater than that of red wine (Vitis vitifera) . Flavonoids extracted from pcpso showed 31-44% inhibition of sheep cyclooxygenase and 69-81% inhibition of soybean lipoxygenase . Flavonoids extracted from pfj showed 21-30% inhibition of soybean lipoxygenase though no significant inhibition of sheep cyclooxygenase . The pcpso was analyzed for its polyphenol content and fatty acid composition . Total polyphenols in pcpso showed a concentration by weight of approximately 0.015% . Pcpso fatty acid composition showed punicic acid (65.3%) along with palmitic acid (4.8%), stearic acid (2.3%), oleic acid (6.3%), linoleic acid (6.6%) and three unidentified peaks from which two (14.2%) are probably isomers of punicic acid (El-Shaarawy, M.I., Nahpetian, A., 1983) . Studies on pomegranate seed oil . Fette Seifen Anstrichmittel 83(3), 123-126).

Trends Plant Sci, 1999 Aug, 4(8), 320 - 325
Ethanolic fermentation: new functions for an old pathway; Tadege M et al.; Ethanolic fermentation is an ancient metabolic pathway . In plants, it is a major route of ATP production under anaerobic conditions . In addition, recent developments suggest that the pathway has important functions in the presence of oxygen . Both of the enzymes required for the production of acetaldehyde and ethanol, pyruvate decarboxylase and alcohol dehydrogenase, are highly abundant in pollen, resulting in fermentation in fully oxygenated cells . Acetaldehyde toxicity is an inevitable side effect of aerobic fermentation . Could acetaldehyde be the elusive pollen factor that contributes to male sterility in cmsT maize? The versatility of this ancient pathway is also illustrated by the induction of aerobic fermentation by environmental stress and activation of a defense response by overexpression of pyruvate decarboxylase.

Immunopharmacology, 1999 Apr, 41(3), 183 - 6
Effect of MSC on the immune response of mice; Hidvegi M et al.; The supposed immunostimulatory actions of MSC, a new fermented wheat germ extract standardized to its benzoquinone composition (trade name: AVEMAR) were studied examining blastic transformation of peripheral blood lymphocytes of mice treated with MSC . It was found that MSC significantly increased the degree of blastic transformation caused by Concanavalin A . Using the B10LP to C57Bl skin graft system, MSC (0.03 and 3.0 g kg(-1) applied orally) acted in favour of restoring the immune function . On the other hand, 2,6-dimethoxy-p-benzoquinone (DMBQ), applied in equivalent doses (0.012 and 1.2 mg kg(-l)), did not shorten the rejection time of skin grafts . The immune restoring effect, as well as the blastic transformation enhancing potential of MSC may be exploited in various cases of decreased immune response.

Antonie Van Leeuwenhoek, 1999 Apr, 75(3), 207 - 15
Genetic and phenotypic diversity of Saccharomyces sensu stricto strains isolated from Amarone wine . Diversity of Saccharomyces strains from Amarone wine; Torriani S et al.; Individual yeast strains belonging to the Saccharomyces sensu stricto complex were isolated from Amarone wine produced in four cellars of the Valpolicella area (Italy) and characterized by conventional physiological tests and by RAPD-PCR and mtDNA restriction assays . Thirteen out of 20 strains were classified as Saccharomyces cerevisiae (ex S . cerevisiae p.r . cerevisiae and p.r . bayanus) and the remaining as Saccharomyces bayanus (ex S . cerevisiae p.r . uvarum) . RAPD-PCR method proved to be a fast and reliable tool for identification of Saccharomyces sensu stricto strains and also gave intraspecific differentiation . Restriction analysis of mtDNA permitted to distinguish S . cerevisiae and S . bayanus species and to discern polymorphism among S . cerevisiae isolates . The assessment of the phenotypic diversity within the isolates by gas-chromatographic analysis of secondary fermentation products was explored . Small quantities of isobutanol were produced by most of the strains and higher amounts by some S . cerevisiae strains with phenotypes Gal- and Mel-; all S . bayanus strains produced low amounts of amilyc alcohols . From this study it appears that each winery owns particular strains, with different genetic and biochemical characteristics, selected by specific environmental pressures during the Amarone winemaking process carried out at low temperature in presence of high sugar content.

Appl Environ Microbiol, 1999 Aug, 65(8), 3738 - 41
Identification of Ruminococcus flavefaciens as the predominant cellulolytic bacterial species of the equine cecum; Julliand V et al.; Detection and quantification of cellulolytic bacteria with oligonucleotide probes showed that Ruminococcus flavefaciens was the predominant species in the pony and donkey cecum . Fibrobacter succinogenes and Ruminococcus albus were present at low levels . Four isolates, morphologically resembling R . flavefaciens, differed from ruminal strains by their carbohydrate utilization and their end products of cellobiose fermentation.

Appl Environ Microbiol, 1999 Aug, 65(8), 3298 - 303
Purification, characterization, and heterologous expression in Fusarium venenatum of a novel serine carboxypeptidase from Aspergillus oryzae; Blinkovsky AM et al.; A novel serine carboxypeptidase (EC 3.4.16.1) was found in an Aspergillus oryzae fermentation broth and was purified to homogeneity . This enzyme has a molecular weight of ca . 67,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its specific activity is 21 U/mg for carbobenzoxy (Z)-Ala-Glu at pH 4.5 and 25 degrees C . It has a ratio of bimolecular constants for Z-Ala-Lys and Z-Ala-Phe of 3.75 . Optimal enzyme activity occurs at pH 4 to 4.5 and 58 to 60 degrees C for Z-Ala-Ile . The N terminus of this carboxypeptidase is blocked . Internal fragments, obtained by cyanogen bromide digestion, were sequenced . PCR primers were then made based on the peptide sequence information, and the full-length gene sequence was obtained . An expression vector that contained the recombinant carboxypeptidase gene was used to transform a Fusarium venenatum host strain . The transformed strain of F . venenatum expressed an active recombinant carboxypeptidase . In F . venenatum, the recombinant carboxypeptidase produced two bands which had molecular weights greater than the molecular weight of the native carboxypeptidase from A . oryzae . Although the molecular weights of the native and recombinant enzymes differ, these enzymes have very similar kinetic parameters.

Am J Clin Nutr, 1999 Aug, 70(2), 240 - 6
Inositol phosphates with different numbers of phosphate groups influence iron absorption in humans; Sandberg AS et al.; BACKGROUND: Inositol hexaphosphate (IP(6)) is a well-known inhibitor of iron absorption, whereas the effects of the less-phosphorylated derivatives of IP(6) are less known . OBJECTIVES: The objective was to investigate the effects of inositol tri-, tetra-, and pentaphosphates (IP(3), IP(4), and IP(5), respectively) on iron absorption in humans . DESIGN: Iron absorption was measured in 5 experiments from single meals by extrinsic labeling with (55)Fe and (59)Fe and determination of whole-body retention and the erythrocyte uptake of isotopes . In experiments 1-3 the meals contained white-wheat rolls to which 10 mg P as IP(5), IP(4), or IP(3), respectively, was added . Inositol 1,2,6-triphosphate {Ins(1,2, 6)P(3)} and a mixture of isomers of IP(4) and IP(5) were studied . White-wheat rolls contained 10 mg P as IP(3) + IP(4) and 2 mg P as IP(5) + IP(6) in experiment 4 and 20 mg P as IP(3) + IP(4) and 3 mg P as IP(5) + IP(6) in experiment 5; inositol phosphates were obtained via fermentation of sodium phytate . Each experiment had 8-11 subjects . RESULTS: In experiment 1, iron absorption was reduced by 39%, whereas there was no significant effect on iron absorption in experiments 2 and 3 . In experiments 4 and 5, iron absorption was reduced by 54% and 64%, respectively, suggesting that IP(3) and IP(4) contributed to the inhibitory effect . CONCLUSIONS: IP(5) has an inhibitory effect on iron absorption, whereas IP(3) and IP(4) in isolated form have no such effect . IP(3) and IP(4) in processed food contribute to the negative effect on iron absorption, presumably by binding iron between different inositol phosphates . To improve iron absorption from cereals and legumes, degradation of inositol phosphates needs to be to less-phosphorylated inositol phosphates than IP(3).

Plant Foods Hum Nutr, 1998, 52(4), 337 - 51
Studies on the influence of temperature, relative humidity and microenvironment on the natural fermentation of African oil bean seeds to 'Ugba'; Isu NR et al.; Studies on the enhancement of the traditional production of 'Ugba' (a protein-rich fermented food) from African oil bean seeds were undertaken by fermenting the bean seeds at different temperatures, relative humidities (RH) and microenvironments . Fermentation was monitored by pH, texture, amino-nitrogen content and the viable cell count of the substrate . The 40 degrees C, 98% RH or the 130 microm high density polyethylene (HDPE) treatment increased the fermentation microflora from ca . 10(6) CFU/g to ca . 10(8) CFU/g with high initial changes in pH (5.8-ca . 7.9) and texture (2.0 kg/cm2 to between 1.4 kg/cm2 and 0.9 kg/cm2) in 24 hours . Products with amino-nitrogen contents of between 12.00 mg N/100 g dry matter and 14.00 mg N/100 g dry matter were obtained in 3 days . The cell count of the 30 degrees C, 80% RH or 70 microm treatment increased from 10(6) CFU/g to ca . 10(7) CFU/g and the pH increased from 5.8 to about 6.7 with a coincident decrease in the texture value from 2.0 kg/cm2 to about 1.7 kg/cm2 in 24 hours . Products with amino-nitrogen contents between 15.00 mg N/100 g dry matter and 19.2 mg N/100 g dry matter were obtained after 3 days . Changes in the fermentation indicators were not significant at p < or = 0.05 (pH and texture) and at p < or = 0.01 (amino-nitrogen) after 3 days for the 25 degrees C, 59.9% RH or 50 microm low density polyethylene (LDPE) treatment . Products of fermentation at the combined optimal conditions (80% RH, 35 degrees C, and 70 microm HDPE) compared very well with the traditionally fermented products in terms of pH, texture and amino-nitrogen content.

Comp Biochem Physiol A Mol Integr Physiol, 1999 Jun, 123(2), 129 - 35
The influence of fibre in the diet on growth rates and the digestibility of nutrients in the greater cane rat (Thryonomys swinderianus); van Zyl A et al.; The greater cane rat Thryonomys swinderianus is a coprophagous rodent in which fermentation occurs in the large caecum . The extent to which a 45% increase in the fibre component of the diet influenced growth rates of cane rats and the digestibility of nutrients and energy was investigated in two feeding trials . Higher fibre levels in the diet reduced the digestibility of dry matter, protein and fat, while animals digested fibre components (neutral-detergent fibre, acid detergent fibre, hemicellulose and cellulose) with a comparable efficiency to those maintained on a low fibre diet . In one of the trials animals fed the high fibre diet exhibited significantly lower growth rates than animals fed the low fibre diet . Digestibility coefficients of the cane rats for neutral-detergent fibre and protein seem to be intermediate to high when compared to reported values for the porcupine, guinea-pig, degu and rabbit . It is suggested that the ability of cane rats to utilise large quantities of fibre enable them to survive periods when only dry grass is available.

Rev Argent Microbiol, 1999 Apr-Jun, 31(2), 65 - 71
{Effect of trace metals on cell morphology, enzyme activation, and production of citric acid in a strain of Aspergillus wentii}; Majolli MV et al.; Data concerning the effect of very low concentrations of metals on citric acid production by microorganisms, as well as on the activity of enzymes presumptively involved in the process, are confuse . The bulk of information was obtained mainly studying selected strains of Aspergillus niger . Information concerning other citric acid producer filamentous fungi, such as A . wentii, is scanty . In the present article we report the effect of different cations on the growth pattern of A . wentii P1 as well as on the related citric acid production and the activity of several enzymes . It was found that without any addition to the culture medium the fungus developed a pelleted form of growth, pellets being about 1.5 mm in diameter . The citric acid yield was about 90% . The addition of Cu2+ impaired the sugar uptake, as well as the production of citric acid and biomass . The uptake of sugar increased in the presence of Zn2+, and there was a marked increase of the biomass production, which could account for the low citric acid production . The addition of Fe2+ impaired the citric acid production and, as sulfate, the sugar uptake . The presence of Fe3+ markedly impaired the citric acid production and increased the sugar uptake . There is no agreement about the enzymes involved in the accumulation of citric acid by microorganisms . In spite of this, aconitase (Ac), isocitrate lyase (IL), isocitrate dehydrogenase NAD(+)-dependent (ICDH- NAD+) and isocitrate dehydrogenase NADP(+)-dependent (ICDH-NADP+) are often postulated as key enzymes . In our case, these enzymes were active during the standard fermentation, although with variations, particularly concerning Ac and IL . The behavior of enzymes might be different when tested in vivo or in vitro, mainly from the quantitative point of view . Nevertheless, the activity determined in vitro might give some indication concerning the effect on fermentation of substances present in the medium . It was found that all the enzymes tested increased their activity in the presence of Fe2+ and Fe3+ . The addition of Cu2+ improved the activity of Ac, IL and ICDH-NADP+, while that of ICDH-NAD+ was impaired . The presence of Zn2+ decreased the activity of the enzymes excepting that of ICDH-NADP+ which increased . It must be pointed out that according to these results there are differences concerning the behavior of ICDH-NAD+ and ICDH-NADP+ in the presence of Cu2+ or Zn2+.

Biochemistry (Mosc), 1999 Jul, 64(7), 839 - 44
Differential changes in alpha2-macroglobulin and hemopexin in brain and liver in response to acute inflammation; Saso L et al.; Changes in serum and cerebrospinal fluid (CSF) proteins following generalized acute inflammation induced by fermented yeast in the rat was examined by concanavalin A-blotting, immunoblotting, and radioimmunoassay . Using alpha2-macroglobulin (alpha2-M) and hemopexin (HPX) as marker proteins, the concentration alpha2-M was found to increase in serum and CSF by 150- and 5-fold, respectively, whereas the concentration of HPX increased by about 4-fold in both fluids following yeast-induced inflammation . The lesser increase in alpha2-M in the CSF versus the systemic circulation is not likely to be the result of changes in the permeability of the blood--brain barrier, since no change in the total protein content of CSF was detected in inflamed rats when compared to control animals . These results, however, illustrate the regulation of the same protein, such as alpha2-M, in two separate organs within the same animal can be drastically different . These results also suggest a possible protective role of alpha2-M in the brain during acute inflammation . Moreover, these observations are consistent with the previous observation that there is a differential response in the level of alpha2-M between the testis and the systemic circulation during inflammation.

Nutr Hosp, 1999 May-Jun, 14(3), 101 - 13
{Importance of the polyamines: review of the literature}; Farriol M et al.; The polyamines, putrescine, spermidine and spermine, are low-molecular weight substances, synthesized in eucariot cells from their immediate precursor, ornithine . These nitrogen compounds are essential for growth . The organism's endogenous supply is obtained through the diet or by synthesis in the intestinal flora . The polyamines are found in fruits and vegetables, foods of animal origin (milk, eggs, fish, and meat) and fermented food products (cheese, beer, and sauerkraut) . Being nitrogenated compounds, they are considered as "minor" components of the diet . Ornithine decarboxylase, a short-half life enzyme, is the key to polyamine biosynthesis . Cellular polyamines are found in free or conjugated forms, the latter made possible, above all, by the presence of positive charges in their molecules . Their particular structure facilitates interaction with anions and binding to nuclear and membrane structures, particularly phospholipids, proteins and DNA . The organism's requirements for these substances are elevated during phases of intense growth or increased demand; thus, the nutritional supply can be crucial during the evolution of processes that involve a high degree of loss combined with deficits in endogenous biosynthesis . Increased tissue and organ polyamine concentrations correlate with diseases of neoplastic origin . It has been hypothesized that polyamine inhibition could be a therapeutic mechanism for such conditions . The supply of polyamines in artificial nutrition has not been measured and there are few data on the effects of artificial nutrition on circulating levels or the usual sites of storage in the body.

Magnes Res, 1999 Jun, 12(2), 99 - 108
Do forestomach epithelia exhibit a Mg2+/2H(+)-exchanger?
Leonhard-Marek S.
In ruminants the forestomachs (reticulum, rumen and omasum) represent the main site of Mg absorption . Readily fermentable carbohydrates enhance Mg availability for ruminants in vivo . The beneficial effect of carbohydrate addition on Mg absorption occurs preintestinally and is associated with changes in the rumen fluid (short chain fatty acids, SCFA, luminal pH, carbon dioxide, NH3, lactate, osmolarity) . SCFA and HCO3-/CO2 increase Mg efflux from the isolated reticulorumen in vivo and stimulate 28Mg flux from mucosal to serosal across rumen epithelium in vitro . The stimulatory effect of SCFA on Mg absorption differs between acids, being a function of SCFA absorption and possibly SCFA metabolism . The stimulatory effect of HCO3-/CO2 is mediated by carbonic anhydrase activity . Experimental data suggest that the availability of protons inside the cell might be involved in the stimulation of Mg absorption . The contribution of Mg/H exchange to Mg2+ uptake across the apical membrane of rumen epithelium is discussed in comparison with other possible ways of stimulation (Mg/Na exchange, Mg/anion cotransport, electrogenic and metabolic effects of SCFA) . It remains to be shown that Mg2+ uptake into rumen epithelium is directly linked to an efflux of protons . However, an apical Mg/H exchange (in contrast to other suggested Mg transporters) would explain a variety of in vivo and in vitro observations on ruminal Mg absorption.

Appl Microbiol Biotechnol, 1999 Jun, 51(6), 780 - 5
Oxygen starvation induces cell death in Candida shehatae fermentations of D-xylose, but not D-glucose; Kastner JR et al.; Candida shehatae cells, cultivated on D-glucose and D-xylose, were subjected to a shift from fully aerobic to anaerobic fermentative conditions . After anaerobic conditions were imposed, growth was limited to approximately one doubling or less as C . shehatae rapidly entered a stationary phase of growth . Following the shift to anoxia, cell viability rapidly declined and the total cell volume declined in the D-xylose fermentations . Moreover, the cell volume distribution shifted to smaller volumes . Cell viability, measured by plate counts, declined nine times faster for D-xylose fermentations than for D-glucose fermentations . Anaerobic growth did not occur on either D-glucose or D-xylose . Selected vitamins and amino acids did not stimulate anaerobic growth in C . shehatae, but did enhance anaerobic growth on D-glucose in S . cerevisiae . The decline in cell viability and lack of anaerobic growth by C . shehatae were attributed to oxygen deficiency and not to ethanol inhibition . The results shed light on why C . shehatae anaerobic fermentations are not currently practical and suggest that research directed towards a biochemical understanding of why C . shehatae can not grow anaerobically will yield significant improvements in ethanol fermentations from D-xylose.

Antibiot Khimioter, 1999, 44(6), 16 - 20
{Action of avermectins on lymphoid leukemia P-388 cells in vitro}; Mosin VA et al.; Avermectins are final products in the fermentation process with Streptomyces avermitilis . They have parasitocidic activity and are used as the main substances of insectoacaronematocides . The study of the activity of the natural avermectin complex (aversectin C) and separate avermectins A1, A2, B1 and B2 in the cell culture of lymphoid leukemia P-388 showed that within the concentrations of 0.1 to 1.0 microgram/ml aversectin C inhibited the growth of the tumor cells and induced their death . The inhibition was due to blocking the cell mitosis . The cell death was accompanied by internucleosomal degradation of the DNA nuclei i.e . the death was of the apoptosis type . The sensitivity of the cells to aversectin C was directly proportional to their initial proliferative activity . As for the separate avermectins only avermectin A1 had the cytotoxic activity within the concentrations used, avermectin A2 had the cytostatic activity and avermectins B1 showed no activity under the experimental conditions.

Nutrition, 1999 Jul-Aug, 15(7-8), 633 - 7
Management of patients with a short bowel; Nightingale JM; Short bowel syndrome most commonly results after bowel resections for Crohn's disease . The normal human small intestinal length ranges from about 3 to 8 m, thus if the initial small intestinal length is short, a relatively small resection of the intestine may result in the problems of a short bowel . Two types of patient with a short bowel are encountered in clinical practice: those with their jejunum anastomosed to a functioning colon, and those with a jejunostomy . Both types of patient have problems absorbing adequate macronutrients, and both need long-term vitamin B12 therapy . Patients with a jejunostomy also have major problems with large stomal losses of water, sodium, and magnesium . This high-volume jejunostomy output is treated by restricting oral fluids, giving a glucose-saline solution to drink, and using drugs that either reduce gastrointestinal motility (loperamide or codeine phosphate) or secretions (H2 antagonists, proton pump inhibitors, or octreotide) . Patients whose jejunal length is less than 100 cm, and whose stomal output is greater than their oral intake, benefit most from antisecretory drugs . In patients with a retained colon, bacterial fermentation of unabsorbed carbohydrate in the colon results in energy being salvaged . However, they have increased oxalate absorption and a 25% chance of developing calcium oxalate renal stones . Thus patients with a colon are advised to eat a high-energy diet rich in carbohydrate but low in oxalate . Patients with a jejunostomy need a high-energy iso-osmolar diet with added salt . Both patient types have a 45% prevalence of gallstones . With current therapy most patients with a short bowel have a normal body mass index and a good quality of life.

J Nutr, 1999 Aug, 129(8), 1579 - 84
Conjugated linoleic acids alter milk fatty acid composition and inhibit milk fat secretion in dairy cows; Chouinard PY et al.; Conjugated linoleic acids (CLA) have positive health effects in experimental models . Our objective was to determine the effect of CLA supplementation on milk of dairy cows . A commercial source of CLA was infused abomasally to by-pass rumen fermentation . The supplement contained 61.2% CLA; the major CLA isomers were cis/trans 8,10, cis/trans 9,11, cis/trans 10,12 and cis/trans 11,13 . Four Holstein cows were used in a 4 x 4 Latin square design . Treatments were 5-d infusions of 0, 50, 100 and 150 g/d of CLA supplement . Infusion increased milk fat content of CLA from 6.8 mg/g fat (zero dose) to 63.6 mg/g fat (highest dose) . All of the major CLA isomers in the supplement were transferred to milk fat in a dose-dependent manner . Apparent efficiency of transfer to milk fat was 22.5, 22.5, 10.2 and 26.3% for cis/trans 8,10, cis/trans 9,11, cis/trans 10,12 and cis/trans 11,13, respectively . CLA infusion had no effect on milk protein and little effect on milk yield (21.5, 20.4, 20.9 and 18.3 kg/d for 0, 50, 100 and 150 g/d CLA supplement, respectively) . However, CLA infusion dramatically reduced milk fat . On average, the content and yield of milk fat were reduced by 52 and 55%, respectively . The role of specific CLA isomers and mechanism(s) for the reduction in milk fat have not been established, although the pattern of milk fatty acids demonstrated effects were most pronounced on de novo fatty acid synthesis and the desaturation process . Overall, dietary supplemention of CLA increased milk fat content of CLA, altered milk fatty acid composition and markedly reduced the content and yield of milk fat.

Protein Expr Purif, 1999 Jul, 16(2), 315 - 23
Use of the green fluorescent protein variant (YFP) to monitor MetArg human proinsulin production in Escherichia coli; Daabrowski S et al.; Green fluorescent protein (GFP), a relatively new reporter gene, is making an impact on many aspects of science . The attributes of GFP could also be applied to the area of recombinant protein production . The work described here represents the first experiments using GFP as a tool to monitor recombinant protein production in real time in the fermentation process . We have constructed plasmids containing an operon fusion of the gene encoding MetArg-human proinsulin and reporter gene GFP (GFP, BFP, and YFP variants) . The MetArg-proinsulin and GFP variant reporter protein were overexpressed in Escherichia coli BL21(DE3) after isopropyl beta-d-thiogalactoside induction . The MetArg-proinsulin to YFP protein ratio did not change in the cells during the bioprocess . Since there is a quantitative relationship between the level of MetArg-proinsulin concentration and YFP fluorescence, it is possible to measure only YFP fluorescence in order to monitor the production of MetArg-proinsulin during the bioprocess . The expression level of MetArg-proinsulin could reach 20-25% . Some 140 mg recombinant MetArg-human proinsulin could be obtained easily from 1 liter of fermentation medium . The MetArg-proinsulin could simply be changed into human insulin by trypsin and carboxypeptidase B treatment in later steps . These experiments provide possibilities for using the YFP reporter gene as a convenient tool to monitor protein expression in biotechnological processes . The proposed technique could reduce the time- and labor-intensive analysis of protein production and would improve the efficiency of process development .

Food Chem Toxicol, 1999 Apr, 37(4), 297 - 305
Exposure to N-nitroso compounds in a population of high liver cancer regions in Thailand: volatile nitrosamine (VNA) levels in Thai food; Mitacek EJ et al.; The recent case-control studies in Thailand indicate that a high incidence of liver cancer in Thailand has not been associated with common risk factors such as hepatitis B infection, aflatoxin intake and alcohol consumption . While the infestation by the liver fluke Opisthorchis viverrini (OV) accounted for the high risk in north-east Thailand, there was no such exposure in the other regions of the country where the incidence of liver cancer is also high . Case-control studies suggest that exposure to exogenous and possibly endogenous nitrosamines in food or tobacco in betel nut and cigarettes may play a role in the development of hepatocellular carcinoma (HCC), while OV infestation and chemical interaction of nitrosamines may also be aetiological factors in the development of cholangiocarcinoma (CCA) . Over 1800 samples of fresh and preserved food were systematically collected and tested between 1988 and 1996 . All the food items identified by anthropological studies to be consumed frequently in four major regions of Thailand were analysed for volatile nitrosamines using gas chromatography combined with a thermal energy analyser . Relatively high levels of N-nitrosodimethylamine (NDMA), N-nitrosopiperidine (NPIP) and N-nitrosopyrrolidine (NPYR) were detected in fermented fish ("Plasalid") . NDMA was also detected at levels ranging from trace amounts to 66.5 microg/kg in several salted and dried fish ("Larb-pla" and "Pla-siu") . NDMA and NPYR were frequently detected in several vegetables, particularly fermented beans ("Tau-chiau") at levels ranging between 1 and 95.1 microg/kg and 0-146 microg/kg, respectively . The possible role of nitrosamines in Thai food in the aetiology of liver cancer (HCC, CCA) is discussed.

Vet Microbiol, 1999 Jun 30, 67(3), 203 - 11
Isolation of Verocytotoxin-producing Escherichia coli O157:H7 from cattle at slaughter in Italy; Bonardi S et al.; Cattle arriving for slaughter at a large abattoir in northern Italy between April 1997 and January 1998 were examined for intestinal carriage of Verocytotoxin-producing Escherichia coli (VTEC) O157 using an immunomagnetic separation technique . Sixty sorbitol non-fermenting VTEC O157 strains were isolated from 59 (13.1%) of the 450 cattle examined . In particular, VTEC O157 was found in 37 (16.6%) of 223 feedlot cattle and in 22 (16.1%) of 137 dairy cull cows, but not in the 90 veal calves sampled . The isolation rate was higher during warm weather (17.5%), falling to an average of 2.9% during the winter months . VT-negative, O157 latex-agglutinating E . coli strains were isolated from 23 (5.1%) of the 450 animals . PCR analysis showed that all 60 VTEC O157 strains carried the VT2 gene and that 25 strains also carried the VT1 gene . In addition, four of the VT-negative, O157 latex-agglutinating E . coli strains carried the VT2 gene . Atypical biochemical features were observed in some VTEC O157: two strains (3.3%) showed beta-glucuronidase activity, and seven (11.7%) produced urease.

Biotechnol Bioeng, 1999 Sep 20, 64(6), 722 - 8
Response of Rhizopus oligosporus to temporal temperature profiles in a model solid-state fermentation system; Ikasari L et al.; Membrane overcultures of Rhizopus oligosporus were shifted from 37 to 50 degrees C for 10 h and then returned to 37 degrees C, mimicking the temporal temperature profiles which typically occur in SSF due to heat transfer limitations . Analysis with a modified two-phase growth model suggests that the temperature upshift causes a 48% decrease in the number of actively extending hyphal tips, and that the first order death rate constant of tips increases from 0 . 059 to 0.073 h(-1) . The fungus did not immediately recover when the temperature was returned to 37 degrees C . The model assumed that the specific growth rate constant microgram(g) was not affected by the increase of temperature, although contradictory data was obtained from radial growth rate experiments .

Biotechnol Bioeng, 1999 Sep 20, 64(6), 698 - 708
An inverse correlation between stress resistance and stuck fermentations in wine yeasts . A molecular study; Ivorra C et al.; During alcoholic fermentations yeast cells are subjected to several stress conditions and, therefore, yeasts have developed molecular mechanisms in order to resist this adverse situation . The mechanisms involved in stress response have been studied in Saccharomyces cerevisiae laboratory strains . However a better understanding of these mechanisms in wine yeasts could open the possibility to improve the fermentation process . In this work an analysis of the stress response in three wine yeasts has been carried out by studying the expression of several representative genes under several stress conditions which occur during fermentation . We propose a simplified method to study how these stress conditions affect the viability of yeast cells . Using this approach an inverse correlation between stress-resistance and stuck fermentations has been found . We also have preliminary data about the use of the HSP12 gene as a molecular marker for stress-resistance in wine yeasts .

Int J Biol Macromol, 1999 Jun-Jul, 25(1-3), 31 - 6
Recent advances in polyhydroxyalkanoate production by bacterial fermentation: mini-review; Lee SY et al.; Poly(3-hydroxybutyrate) {P(3HB)} and other polyhydroxyalkanoates (PHAs) have been drawing much attention as biodegradable substitutes for conventional nondegradable plastics . For the economical production of P(3HB), various bacterial strains, either wild-type or recombinant, and new fermentation strategies were developed for the production of P(3HB) with high concentration and productivity . To reduce the cost of carbon substrate, several processes for P(3HB) production from cheap carbon sources were also developed . P(3HB) can now be produced to a content of 80% of cell dry weight with the productivity greater than 4 g/l per h . Fermentation strategy was also developed for the efficient production of medium chain length PHA by high cell density culture . With all these advances, P(3HB) and PHAs can be produced by bacterial fermentation at a cost (ca . $2/kg) similar to that of other biodegradable polymers under development.

Arch Microbiol, 1999 Aug, 172(2), 116 - 24
Chlorobium ferrooxidans sp . nov., a phototrophic green sulfur bacterium that oxidizes ferrous iron in coculture with a "Geospirillum" sp . strain; Heising S et al.; A green phototrophic bacterium was enriched with ferrous iron as sole electron donor and was isolated in defined coculture with a spirilloid chemoheterotrophic bacterium . The coculture oxidized ferrous iron to ferric iron with stoichiometric formation of cell mass from carbon dioxide . Sulfide, thiosulfate, or elemental sulfur was not used as electron donor in the light . Hydrogen or acetate in the presence of ferrous iron increased the cell yield of the phototrophic partner, and hydrogen could also be used as sole electron source . Complexed ferric iron was slowly reduced to ferrous iron in the dark, with hydrogen as electron source . Similar to Chlorobium limicola, the phototrophic bacterium contained bacteriochlorophyll c and chlorobactene as photosynthetic pigments, and also resembled representatives of this species morphologically . On the basis of 16S rRNA sequence comparisons, this organism clusters with Chlorobium, Prosthecochloris, and Pelodictyon species within the green sulfur bacteria phylum . Since the phototrophic partner in the coculture KoFox is only moderately related to the other members of the cluster, it is proposed as a new species, Chlorobium ferrooxidans . The chemoheterotrophic partner bacterium, strain KoFum, was isolated in pure culture with fumarate as sole substrate . The strain was identified as a member of the epsilon-subclass of the Proteobacteria closely related to "Geospirillum arsenophilum" on the basis of physiological properties and 16S rRNA sequence comparison . The "Geospirillum" strain was present in the coculture only in low numbers . It fermented fumarate, aspartate, malate, or pyruvate to acetate, succinate, and carbon dioxide, and could reduce nitrate to dinitrogen gas . It was not involved in ferrous iron oxidation but possibly provided a thus far unidentified growth factor to the phototrophic partner.

Math Biosci, 1999 Jul, 159(2), 145 - 63
Use of in vitro gas production models in ruminal kinetics; Pitt RE et al.; Physiological systems models for ruminant animals are used to predict the extent of ruminal carbohydrate digestion, based on rates of intake, digestion, and passage to the lower tract . Digestion of feed carbohydrates is described in these models by a first-order rate constant . Recently, an in vitro gas production technique has been developed to determine the digestion kinetics in batch fermentation, and nonlinear mathematical models have been fitted to the cumulative gas production data from these experiments . In this paper, we present an analysis that converts these gas production models to an effective first-order rate constant that can be used directly in rumen systems models . The analysis considers the digestion of an incremental mass of substrate entering the rumen . The occurrence of passage is represented probabilistically, and integration through time gives the total mass of substrate and total rate of digestion in the rumen . To demonstrate the analysis, several gas production models are fitted to a sample data set for corn silage, and the effective first-order rate constants are calculated . The rate constants for digestion depend on ruminal passage rate, an interaction that arises from the nonlinearity of the gas production models.

J Biol Chem, 1999 Jul 23, 274(30), 20772 - 8
Oxygen exchange between acetate and the catalytic glutamate residue in glutaconate CoA-transferase from Acidaminococcus fermentans . Implications for the mechanism of CoA-ester hydrolysis; Selmer T et al.; The exchange of oxygen atoms between acetate, glutaryl-CoA, and the catalytic glutamate residue in glutaconate CoA-transferase from Acidaminococcus fermentans was analyzed using {(18)O(2)}acetate together with matrix-assisted laser desorption/ionization time of flight mass spectrometry of an appropriate undecapeptide . The exchange reaction was shown to be site-specific, reversible, and required both glutaryl-CoA and {(18)O(2)}acetate . The observed exchange is in agreement with the formation of a mixed anhydride intermediate between the enzyme and acetate . In contrast, with a mutant enzyme, which was converted to a thiol ester hydrolyase by replacement of the catalytic glutamate residue by aspartate, no (18)O uptake from H(2)(18)O into the carboxylate was detectable . This result is in accord with a mechanism in which the carboxylate of aspartate acts as a general base in activating a water molecule for hydrolysis of the thiol ester intermediate . This mechanism is further supported by the finding of a significant hydrolyase activity of the wild-type enzyme using acetyl-CoA as substrate, whereas glutaryl-CoA is not hydrolyzed . The small acetate molecule in the substrate binding pocket may activate a water molecule for hydrolysis of the nearby enzyme-CoA thiol ester.

Eur J Biochem, 1999 Jul, 263(2), 305 - 11
An ethanol-inducible MDR ethanol dehydrogenase/acetaldehyde reductase in Escherichia coli: structural and enzymatic relationships to the eukaryotic protein forms; Shafqat J et al.; An ethanol-active medium-chain dehydrogenase/reductase (MDR) alcohol dehydrogenase was isolated and characterized from Escherichia coli . It is distinct from the fermentative alcohol dehydrogenase and the class III MDR alcohol dehydrogenase, both already known in E . coli . Instead, it is reminiscent of the MDR liver enzyme forms found in vertebrates and has a K(m) for ethanol of 0.7 mM, similar to that of the class I enzyme in humans, however, it has a very high k(cat), 4050 min(-1) . It is also inhibited by pyrazole (K(i) = 0.2 microM) and 4-methylpyrazole (K(i)= 44 microM), but in a ratio that is the inverse of the inhibition of the human enzyme . The enzyme is even more efficient in the reverse direction of acetaldehyde reduction (K(m) = 30 microM and k(cat) = 9800 min(-1)), suggesting a physiological function like that seen for the fermentative non-MDR alcohol dehydrogenase . Growth parameters in complex media with and without ethanol show no difference . The structure corresponds to one of 12 new alcohol dehydrogenase homologs present as ORFs in the E . coli genome . Together with the previously known E . coli MDR forms (class III alcohol dehydrogenase, threonine dehydrogenase, zeta-crystallin, galactitol-1-phosphate dehydrogenase, sensor protein rspB) there is now known to be a minimum of 17 MDR enzymes coded for by the E . coli genome . The presence of this bacterial MDR ethanol dehydrogenase, with a structure compatible with an origin separate from that of yeast, plant and animal ethanol-active MDR forms, supports the view of repeated duplicatory origins of alcohol dehydrogenases and of functional convergence to ethanol/acetaldehyde activity . Furthermore, this enzyme is ethanol inducible in at least one E . coli strain, K12 TG1, with apparently maximal induction at an enthanol concentration of approximately 17 mM . Although present in several strains under different conditions, inducibility may constitute an explanation for the fairly late characterization of this E . coli gene product.

J Biotechnol, 1999 Jun 11, 72(1-2), 13 - 20
High cytoplasmic expression in E . coli, purification, and in vitro refolding of a single chain Fv antibody fragment against the hepatitis B surface antigen; Sanchez L et al.; A single-chain Fv (scFv) antibody fragment against the hepatitis B surface antigen (HBsAg) was expressed in Escherichia coli in the form of two independent fusion proteins, with either 60 ('long') or 27 ('short') amino acid N-terminal encoding sequences related to human interleukin-2 . Both fusion proteins were expressed insolubly and at high levels in the bacterial cytoplasm (approximately 30% of total bacterial protein in MM294 cells at a laboratory scale) . When recombinant cells were cultured in 5-1 fermentors, expression and optical density increased 2- and 4-fold, respectively, compared to a previous periplasmic insoluble version of the same anti HBsAg scFv . After extraction and solubilization in urea, the cytoplasmic scFvs were purified using immobilized metal ion affinity chromatography, followed by DTT treatment, and refolding by dialysis against a basic pH buffer containing EDTA . The refolded scFvs recognized the recombinant HBsAg in ELISA . Results of an ELISA where antigen affinity chromatography repurified scFvs were used as standards, indicated that refolding efficiencies were high: 56.2% for the 'short' fusion scFv, and 50.6% for the 'long' fusion scFv . Corrected final yields of active scFv were 30.3 and 27.3 mg l-1, respectively, for the aforementioned fusion proteins, 5-6 times better than those reported for the periplasmic scFv variant.

Anal Biochem, 1999 Jul 15, 272(1), 43 - 7
A method of fast separation of lignin peroxidases using convective interaction media disks; Podgornik H et al.; The HPLC separation of lignin peroxidase isoenzymes using Convective Interaction Media disks containing quaternary amine and diethylaminoethyl ion-exchange active groups is proposed . In contrast to standard HPLC procedures the separation can be performed within a few minutes without considerably affecting the separation resolution . The method is reproducible and gives a linear response of integrated peak area to protein concentration for all measured isoenzymes . The separation resolution is retained unchanged by applying crude culture filtrate instead of a sample previously frozen and dialyzed . The optimized method might therefore be used for on-line monitoring of lignin peroxidase isoenzyme composition during fermentation . On the other hand, the proposed method is comparable in time to the original method of lignin peroxidase activity measurement (proposed by Tien and Kirk), providing additionally the isoenzyme composition .

Biotechnol Bioeng, 1999 Sep 5, 64(5), 620 - 3
Isolation of a novel microorganism, pestalotia heterocornis, producing paclitaxel
Noh MJ, Yang JG, Kim KS, Yoon YM, Kang KA, Han HY, Shim SB, Park HJ.
Pestalotia heterocornis was isolated from soil collected in yew forest and was shown to produce paclitaxel in semisynthetic liquid media . The presence of paclitaxel in the fungal extract was confirmed by FAB mass spectrometry and NMR spectroscopy . The maximum yield of paclitaxel was 31 &mgr;g per liter . Optimal paclitaxel production occurred after 5-7 days in a 20-liter scale fermentation at 23 degrees C . These results indicate that P . heterocornis is an excellent candidate for consideration in fermentation technology .

Biotechnol Bioeng, 1999 Sep 5, 64(5), 552 - 7
Scale-up of citric acid fermentation by redox potential control
Berovic M.
To obtain high citric acid productivity in Aspergillus niger fermentation on beet molasses substrate, a certain redox potential profile with two maxima (260 and 280 mV) and two minima (180 and 80 mV) must be maintained . The most effective regulation of redox potential is by regulation of aeration and agitation . It has been shown that control of redox potential by aeration and agitation is a most successful method for scale-up from 10-L laboratory scale to the 100- and 1000-L pilot-plant scale, even in geometrically dissimilar stirred-tank reactors .

Biochem Biophys Res Commun, 1999 Jul 14, 260(3), 775 - 80
Redistribution of activated caspase-3 to the nucleus during butyric acid-induced apoptosis; Mandal M et al.; The colonic epithelial cells near the top of the crypt and in the lumen have been shown to undergo apoptosis . Since butyric acid is the major short-chain fatty acid produced by fermentation of dietary fiber in the large bowel, it has been proposed that it could act as an important regulator of apoptosis in colorectal cancer . Here we report that in cells treated with butyric acid, the cleavage of DNA-PKcs was paralleled or preceded by the induction of activation of caspase-3, and these events were inhibited by Bcl-2 overexpression . We also demonstrated the redistribution of activated caspase-3 to the nuclear compartment where it locally cleaves DNA-PKcs and poly(ADP-ribose) polymerase, and cleaved fragments were released in the cytosolic compartment . The observed activation of caspase-3 and nuclear cleavage of its substrates and their subsequent release into the cytosol were inhibited by a specific caspase-3 inhibitor, the tetrapeptide DEVD-CHO . These findings suggest that relocalization of activated caspase-3 to the nucleus may constitute an important apoptotic signal during butyric acid-induction of apoptosis human colorectal cancer cells .

Ukr Biokhim Zh, 1998 Nov-Dec, 70(6), 53 - 8
{Relationship between values of antioxidant enzyme system activity in various tissues of intact rats}; Levadnaia OV et al.; It was studied the relation between activities of ferments an antioxidant system: of superoxide dismutase, catalase and GSH-peroxidase in the homogenates of livers, lungs and cerebrum of intact rats . When activities were brought to identical units of measurement, it was determined that relation of activities can see with a point to view of chemical kinetics laws for consecutively-parallel reactions . It is followed from the result that the activity of catalase livers can be explained by the participation of catalases in other reactions, which were connected with forming a hydrogen peroxide . From the relations between ferments of antioxidant system it was discovered that GSH-peroxidase is the most important antioxidant enzyme for the cerebrum . Data of the relation of activities ferments of antioxidant system are stipulated by the tissues particularities and they are reflected a contribution of every biocatalyst in that system.

Rheumatology (Oxford), 1999 Jun, 38(6), 504 - 9
Detection of mycoplasmal infections in blood of patients with rheumatoid arthritis; Haier J et al.; OBJECTIVE: Mycoplasmal infections are associated with several acute and chronic illnesses . Some mycoplasmas can enter a variety of tissues and cells, and cause system-wide or systemic signs and symptoms . METHODS: Patients (14 female, 14 male) diagnosed with rheumatoid arthritis (RA) were investigated for mycoplasmal infections in their blood leucocytes using a forensic polymerase chain reaction (PCR) procedure . Amplification was performed with genus- and species-specific primers, and a specific radiolabelled internal probe was used for Southern hybridization with the PCR product . Patients were investigated for the presence of Mycoplasma spp., and positive cases were further tested for infections with the following species: M . fermentans, M . hominis, M . pneumoniae and M . penetrans . RESULTS: The Mycoplasma spp . sequence, which is not entirely specific for mycoplasmas, was amplified from the peripheral blood of 15/28 patients (53.6%) and specific PCR products could not be detected in 13 patients (46.4%) . Significant differences (P < 0.001) were found between patients and positive healthy controls in the genus test (3/32) and in the specific tests (0/32) . Moreover, the incidence of mycoplasmal infections was similar in female and male patients . Using species-specific primers, we were able to detect infections with M . fermentans (8/28), M . pneumoniae (5/28), M . hominis (6/28) and M . penetrans (1/28) in RA patients . In 36% of the patients, we observed more than one Mycoplasma species in the blood leucocytes . All multiple infections occurred as combinations of M . fermentans with other species . CONCLUSIONS: The results suggest that a high percentage of RA patients have systemic mycoplasmal infections . Systemic mycoplasmal infections may be an important cofactor in the pathogenesis of RA, and their role needs to be explored further.

J Chromatogr A, 1999 Jun 11, 845(1-2), 171 - 9
Physico-chemical characterization of recombinant hepatitis B surface antigen by a multidimensional approach; Tleugabulova D et al.; Initially, our work was directed to respond to the question: why hepatitis B surface antigen (HBsAg) produces a very broad peak in preparative size-exclusion chromatography (SEC) . For this purpose, we used a multidimensional approach based on SEC fractionation of purified HBsAg followed by the individual analysis of SEC fractions by a battery of assays, such as SEC, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, enzyme-linked immunosorbent assay and transmission electron microscopy . As a result, HBsAg particles were shown to be heterogeneous in terms of particle assembly . In order to elucidate the origin of HBsAg heterogeneity, we included here the denaturing SEC into a multidimensional approach . The data from denaturing SEC evidenced the fragmentation of protein monomers within the HBsAg particle that, probably, occurs during fermentation broth, rather than during in vitro HBsAg processing . The fractions isolated from widely separated regions of HBsAg peak differed in the extent of protein fragmentation, suggesting that the variable extent of protein degradation within HBsAg particles may be one of the factors responsible for broadening of the HBsAg peak in SEC.

Appl Biochem Biotechnol, 1999 Spring, 77-79, 435 - 44
Bioconversion of secondary fiber fines to ethanol using counter-current enzymatic saccharification and co-fermentation; Jeffries TW et al.; This research examined several enzymatic and microbial process for the conversion of waste cellulosic fibers into ethanol . The first was a one-stage process in which pulp fines were contacted with commercial enzyme solutions . The second process used sequential, multistage saccharification . The third used sequential enzyme addition in a countercurrent mode . Experiments compared the results with various feedstocks, different commercial enzymes, supplementation with beta-glucosidase, and saccharification combined with fermentation . The highest saccharification (65%) from a 4% consistency pulp and the highest sugar concentration (5.4%) from an 8% consistency pulp were attained when 5 FPU/g plus 10 IU/g of beta-glucosidase were used . Sequential addition of enzyme to the pulp in small aliquots produced a higher overall sugar yield/U enzyme than the addition of the same total amount of enzyme in a single dose . In the saccharification and fermentation experiments, we produced 2.12% ethanol from a 5.4% sugar solution . This represents 78% of the theoretical maximum . This yield could probably be increased through optimization of the fermentation step . Even when little saccharification occurred, the enzyme facilitated separation of water, fiber, and ash, so cellulase treatment could be an effective means for dewatering pulp sludges.

Biotechnol Bioeng, 1999 Aug 5, 64(3), 310 - 21
Process intensification by direct product sequestration from batch fermentations: application of a fluidised bed, multi-bed external loop contactor
Hamilton GE, Morton PH, Young TW, Lyddiatt A.
A critical comparison has been made of the relative efficacy of the primary purification of an extracellular acid protease produced by the yeast Yarrowia lipolytica . The performance of conventional, discrete sequences of fermentation, broth clarification and fixed bed, anion exchange chromatography has been compared with fluidised bed adsorption directly interfaced with post-term fermentation broth and fluidised bed adsorption directly integrated with productive fermentations (so-called direct product sequestration; DPS) . Advantages of the latter, in terms of the improved yield and molecular quality of the protease end product are discussed in terms of the design, assembly and operation of component parts of DPS devices and their generic application to other extracellular bioproducts of microbial fermentations .

Biotechnol Bioeng, 1999 Aug 5, 64(3), 284 - 9
Fermentability of the hemicellulose-derived sugars from steam-exploded softwood (douglas fir)
Boussaid A, Robinson J, Cai Yj, Gregg DJ, Saddler JN.
Steam explosion ofDouglas fir wood chips under low-severity conditions (log Ro = 3.08 corresponding to 175 degrees C, 7.5 min, and 4.5% SO2) resulted in the recovery of around 87% of the original hemicellulose component in the water-soluble stream . More than 80% of the recovered hemicellulose was in a monomeric form . As the pretreatment severity increased from 3.08 to 3.76, hemicellulose recovery dropped to 43% of the original hemicellulose found in Douglas fir chips while the concentration of glucose originating from cellulose hydrolysis increased along with the concentration of sugar degradation products such as furfural and hydroxymethylfurfural . Despite containing a higher concentration of hexose monomers (mainly glucose originating from cellulose degradation), the water-soluble fraction prepared under high-severity conditions (log Ro = 3.73 corresponding to 215 degrees C, 2.38 min, and 2.38% SO2) was not readily fermented . Only the two hydrolyzates obtained at low and medium (195 degrees C, 4.5 min, and 4.5% SO2) severities were fermented to ethanol using a spent sulfur liquor adapted strain of Saccharomyces cerevisiae . High ethanol yields were obtained for these two hydrolyzates with 0.44 g of ethanol produced per gram of hexose utilized (86% of theoretical) . However, the best results of hemicellulose recovery and fermentability were obtained for the low-severity water-soluble fraction which was fermented significantly faster than the fraction obtained after medium-severity treatment probably because it contained higher amounts of fermentation inhibitors .

Biotechnol Bioeng, 1999 Jul 20, 64(2), 232 - 9
Imprinted polymers as tools for the recovery of secondary metabolites produced by fermentation
Ju JY, Shin CS, Whitcombe MJ, Vulfson EN.
Imprinted polymers were synthesized using a mixture of pigments, N-glutamyl-rubropuctamine, and N-glutamyl-monascorubramine (I) as the template, and 2-methacrylamido-6-picoline or 4-aminostyrene as functional monomers, to obtain recognition materials capable of forming hydrogen bonds and charge interactions, respectively, with carboxyl groups of target I in the binding sites . The polymers were prepared thermally at a template loading of 5 mol% using ethylene-glycol dimethacrylate or trimethylolpropane trimethacrylate as crosslinkers and acetonitrile or tetrahydrofuran as porogens . The selective binding of I to both types of polymer was demonstrated, although aminostyrene-based materials showed higher overall adsorption and were studied in more detail . It was shown that the kinetics of binding of I from ethyl-acetate extracts of fermented Monascus sp . was very rapid and virtually all the pigment adsorbed can be released by washing the polymer with ethanol-water mixtures . The feasibility of reusing imprinted polymer in consecutive adsorption/desorption cycles was also demonstrated .

Biotechnol Bioeng, 1999 Jul 5, 64(1), 82 - 91
Fermentation seed quality analysis with self-organising neural networks
Ignova M, Montague GA, Ward AC, Glassey J.
Industrial fermentation processes operate under well defined operating conditions to attempt to minimise production variability . Variability occurs for many reasons but a long held belief is that variation in the state of the seed is highly influential . In this paper a seed stage (a batch process) of an industrial antibiotic fermentation is considered and the performance of the main production fermentations is correlated with the quality of the seed using an unsupervised Kohonen self-organising feature map (SOM) . It is shown that using only seed information poor performance in the final stage fermentations can be predicted . Data from industrial penicillin G fermenters is used to demonstrate the procedure .

Biotechnol Bioeng, 1999 Aug 5, 64(3), 257 - 66
Bubble rise velocities and drag coefficients in non-Newtonian polysaccharide solutions; Margaritis A et al.; Microbially produced polysaccharides have properties which are extremely useful in different applications . Polysaccharide producing fermentations start with liquid broths having Newtonian rheology and end as highly viscous non-Newtonian solutions . Since aerobic microorganisms are used to produce these polysaccharides, it is of great importance to know the mass transfer rate of oxygen from a rising air bubble to the liquid phase, where the microorganisms need the oxygen to grow . One of the most important parameters determining the oxygen transfer rate is the terminal rise velocity of air bubble . The dynamics of the rise of air bubbles in the aqueous solutions of different, mostly microbially produced polysaccharides was studied in this work . Solutions with a wide variety of polysaccharide concentrations and rheological properties were studied . The bubble sizes varied between 0.01 mm3 and 10 cm3 . The terminal rise velocities as a function of air bubble volume were studied for 21 different polysaccharide solutions with different rheological properties . It was found that the terminal velocities reached a plateau at higher bubble volumes, and the value of the plateau was nearly constant, between 23 and 27 cm/s, for all solutions studied . The data were analyzed to produce the functional relationship between the drag coefficient and Reynolds number (drag curves) . It was found out that all the experimental data obtained from 21 polysaccharide solutions (431 experimental points), can be represented by a new single drag curve . At low values of Reynolds numbers, below 1.0, this curve could be described by the modofoed Hadamard-Rybczynski model, while at Re > 60 the drag coefficient was a constant, equal to 0.95 . The latter finding is similar to that observed for bubble rise in Newtonian liquids which was explained on the basis of the "solid bubble" approach .

Biotechnol Bioeng, 1999 Jul 5, 64(1), 61 - 73
Fed-batch fermentor synthesis of 3-dehydroshikimic acid using recombinant Escherichia coli; Li K et al.; 3-Dehydroshikimic acid (DHS), in addition to being a potent antioxidant, is the key hydroaromatic intermediate in the biocatalytic conversion of glucose into aromatic bioproducts and a variety of industrial chemicals . Microbial synthesis of DHS, like other intermediates in the common pathway of aromatic amino acid biosynthesis, has previously been examined only under shake flask conditions . In this account, synthesis of DHS using recombinant Escherichia coli constructs is examined in a fed-batch fermentor where glucose availability, oxygenation levels, and solution pH are controlled . DHS yields and titers are also determined by the activity of 3-deoxy-D-arabino-heptulosonic acid 7-phosphate (DAHP) synthase . This enzyme's expression levels, sensitivity to feedback inhibition, and the availability of its substrates, phosphoenolpyruvate (PEP) and D-erythrose 4-phosphate (E4P), dictate its in vivo activity . By combining fed-batch fermentor control with amplified expression of a feedback-insensitive isozyme of DAHP synthase and amplified expression of transketolase, DHS titers of 69 g/L were synthesized in 30% yield (mol/mol) from D-glucose . Significant concentrations of 3-dehydroquinic acid (6.8 g/L) and gallic acid (6.6 g/L) were synthesized in addition to DHS . The pronounced impact of transketolase overexpression, which increases E4P availability, on DHS titers and yields indicates that PEP availability is not a limiting factor under the fed-batch fermentor conditions employed .

Biotechnol Bioeng, 1999 Jun 20, 63(6), 705 - 11
Use of multi-staining flow cytometry to characterise the physiological state of Escherichia coli W3110 in high cell density fed-batch cultures; Hewitt CJ et al.; High cell density fed-batch fermentations of Escherichia coli W3110 have been carried out at specific growth rates of less than 0.3 h-1, to investigate the effect of glucose limitation on the physiological state of individual cells . After an initial exponential batch phase, the feed rate was held constant and a final dry cell weight of approximately 50 g per litre was achieved . The fermentations were monitored by mass spectrometry whilst measurements of pH, DOC, CFU/mL, TCN, OD500nm and residual glucose concentrations were made . Satisfactory and reproducible results were obtained . Flow cytometric analysis of cells in broth samples, based on either of two multi-staining protocols, revealed a progressive change in cell physiological state throughout the course of the fermentations . From these measurements it was concluded that the loss in reproductive viability towards the end of the fed-batch process is due to cell death and not due to the formation of a "viable but nonculturable state" as had previously been reported . Since the presence of a high proportion of dead or dying cells at any time during a fermentation has a detrimental effect on the synthesis of any desired product it is proposed that an on-line flow cytometric analysis and control strategy could be used as a means of increasing overall process efficiency .

J Nutr, 1999 Jul, 129(7 Suppl), 1474S - 7S
Effects of dietary inulin on serum lipids; Davidson MH et al.; Inulin is a carbohydrate belonging to a class of compounds known as fructans . Because inulin is resistant to digestion in the upper gastrointestional tract it reaches the large intestine essentially intact, where it is fermented by indigenous bacteria . Thus, it may be classified as a soluble dietary fiber . Soluble fibers have been shown to modulate serum lipids . A recent study examined the effect of consuming three servings per day of inulin-containing foods, compared with the effect of similar foods without inulin, on serum lipid profiles among hypercholesterolemic men and women . In addition, the practicality of including 18 g/d of inulin in a low fat diet was investigated . The recent study randomized, double-blind, crossover trial with two 6-wk treatment periods, separated by a 6-wk washout . Men and women (n = 21) with baseline LDL increased significantly (7.4 and 12.3%, respectively) during the control phase . There were small, nonsignificant declines in total (1.3%) and LDL-C (2.1%) during the inulin phase . Thus, differences in response between periods (inulin - control) were significant (P < 0.05) for LDL-C (-14.4%) and total cholesterol (-8.7%) . Mild gastrointestinal discomfort was more common during the inulin than the control food phase; however, the gastrointestinal side-effect profile of inulin was similar to that of other soluble fibers . Although it was not possible to draw firm conclusions, inulin may have blunted the hypercholesterolemic effects observed during consumption of control foods . Additional research will be required to confirm the possible lipid-modulating properties of dietary inulin in humans.

J Nutr, 1999 Jul, 129(7 Suppl), 1467S - 70S
Biochemical basis of oligofructose-induced hypolipidemia in animal models; Delzenne NM et al.; Oligofructose (OFS), a mixture of nondigestible/fermentable fructooligosaccharides, decreases serum triacylglycerol (TAG) when it is included in the standard, fiber-free or high fat diet of rats . This paper summarizes in vivo and in vitro data to establish a biochemical mechanism underlying the hypolipidemic effect of OFS . When OFS is added to the standard (carbohydrate-rich) diet of rats at the dose of 10 g/100 g, a TAG-lowering action occurs as a consequence of a reduction of de novo liver fatty acid synthesis . The depression in the activity of all lipogenic enzymes and fatty acid synthase mRNA suggests that OFS modifies the gene expression of lipogenic enzymes . Through its modulation of de novo lipogenesis, OFS can protect against liver lipid accumulation induced by providing 10% fructose-enriched water for 48 h . OFS also significantly decreases serum insulin and glucose, which are both known to participate in the nutritional regulation of lipogenesis . It also increases the intestinal production of incretins, namely, glucose-dependent insulinotropic peptide and glucagon-like peptide 1 . This latter phenomenon results mainly from promotion of intestinal tissue proliferation by oligofructose fermentation end-products . Collectively, a link likely exists between the modulation of hormone and incretin production by OFS, and its antilipogenic effect.

J Nutr, 1999 Jul, 129(7 Suppl), 1446S - 50S
The application of ecological principles and fermentable fibers to manage the gastrointestinal tract ecosystem; Buddington RK et al.; Because diet can influence the structure and functions of the gastrointestinal tract, there are opportunities for using diet as a "management tool" to affect the resident microbiota . Fermentable fibers increase the densities of beneficial bacteria and stimulate growth and functions of the healthy intestine . Recent findings show that after acute diarrhea, the use of an oral electrolyte solution with the fermentable fiber oligofructose accelerates recovery of beneficial bacteria, reduces the relative abundance of detrimental bacteria, stimulates mucosal growth and enhances digestive and immune functions . This review will focus on how the principles of stream ecology can be applied to better understand the distribution of bacteria along the length of the gastrointestinal tract, the effect of diarrhea on the gastrointestinal ecosystem and how fermentable fibers can be used as a "management tool" to promote gastrointestinal health in normal states and during recovery from diarrhea.

J Nutr, 1999 Jul, 129(7 Suppl), 1436S - 7S
Caloric value of inulin and oligofructose; Roberfroid MB; Dietary carbohydrates, which are absorbed as hexose, (glucose, fructose) have a caloric value of 3.9 kcal/g (16.3 kJ/g), and their cellular metabolism produces approximately 38 mol ATP/mol . However, chicory inulin and oligofructose resist digestion and they are not absorbed in the upper part of the gastrointestinal tract . After oral ingestion, they reach the colon intact where they become hydrolyzed and extensively fermented by saccharolytic bacteria, which produce short-chain carboxylic and lactic acids as electron sinks . Depending on both the degree of their colonic fermentation and the assumptions of the model used, the caloric value of such nondigested but fermented carbohydrates varies between 0 and 2.5 kcal/g . Through the catabolism of the absorbed short-chain carboxylic and lactic acids, they may produce up to 17 mol ATP/mol of fermented sugar moiety . Because the daily intake of these dietary carbohydrates is likely to remain relatively small (<10% and probably often not >5% of total daily calorie intake), it is of low relevance nutritionally to give them a precise caloric value . On the basis of biochemical balance charts for carbon atoms, metabolic pathways and energy yields to the host, the caloric value of a fructosyl residue in chicory inulin and oligofructose has been calculated to be approximately 25-35% that of a fully digested and absorbed fructose molecule . For the purpose of food labeling, it is recommended that chicory inulin and oligofructose, like all the other carbohydrates that are more or less completely fermented in the human colon, should be given a caloric value of 1.5 kcal/g (6.3 kJ/g).

J Nutr, 1999 Jul, 129(7 Suppl), 1434S - 5S
Nondigestible carbohydrates and mineral bioavailability; Greger JL; Generally, fiber and compounds associated with fiber in cereal products (e.g., phytates) have been found to reduce the apparent absorption of minerals (such as calcium, magnesium, zinc and manganese) in humans, livestock and animal models . The effects of "soluble" forms of fiber (specifically pectins, gums, resistant starches, lactulose, oligofructose and inulin) on mineral absorption are more difficult to characterize . The addition of these soluble forms of fiber has been found in various studies to add viscosity to the gut contents, promote fermentation and the production of volatile fatty acids in the cecum, have a trophic effect on the ceca of animals and increase serum enteroglucagon concentrations . Thus it is not surprising that the addition of soluble forms of fiber to diets often has been found to improve absorption of minerals . This may reflect absorption of electrolytes from the large intestine . Future work should address the mechanisms by which ingestion of nondigestible carbohydrates improves mineral absorption in humans.

J Nutr, 1999 Jul, 129(7 Suppl), 1424S - 7S
Fiber, inulin and oligofructose: similarities and differences; Schneeman BO; The biological, chemical and physical properties of dietary fibers are associated with physiologic actions in the small and large intestine that have important metabolic implications for health . These properties of fiber include dispersibility in water, bulk, viscosity, adsorption and binding of compounds and fermentability . Dietary fructans share some of the properties of dietary fiber and thus are likely to have similar metabolic effects . Within the small intestine, properties such as dispersibility in water, bulking and viscosity are associated with slowing the digestion and absorption of carbohydrate and lipid and promoting nutrient absorption along a greater length of the small intestine . Both of these actions are related to cholesterol reduction and blunting of alimentary gylcemia . Although fructans are dispersible in water and will provide some bulk because they are nondigestible in the small intestine, they do not appear to be associated with significant increases in viscosity . Thus one would predict that any immediate effects on alimentary glycemia or on cholesterol reduction are likely to be modest compared with more viscous polysaccharides . Fermentability and bulking capacity of nondigestible carbohydrates define an essential role of fiber in maintaining gastrointestinal health . Within the large intestine, carbohydrates that are not digested in the small intestine are available for fermentation by the microflora present . Carbohydrates that are dispersible in the aqueous phase are more readily digested by microbes . A large body of evidence indicates that dietary fructans are digested in the large intestine, resulting in an increase in microbial mass and production of short-chain fatty acids.

J Nutr, 1999 Jul, 129(7), 1333 - 9
Oligo-L-methionine and resistant protein promote cecal butyrate production in rats fed resistant starch and fructooligosaccharide; Morita T et al.; We examined the role of resistant protein and peptides in promoting cecal butyrate production in rats fed rapidly fermentable carbohydrates . Rats were fed diets containing raw potato starch (RPS, 200 g/kg diet) or fructooligosaccharide (FOS, 60 g/kg diet) with casein, soy or rice protein (250 g/kg diet) for 13 d . In rats fed RPS with casein, the major cecal organic acid was acetate (441 micromol), but lactate and succinate were also found in considerable amounts (324 micromol) . Succinate was the major cecal organic acid (235 micromol) in rats fed FOS with casein . When rice protein was fed with RPS, the contribution of lactate was significantly lower and that of propionate tended to be higher (P < 0.1) than in rats fed casein . In rats fed rice protein with FOS, cecal butyrate and acetate were greater and cecal succinate was lower than in rats fed casein with FOS (P < 0.05) . Despite the similar amounts of undigested protein in rice and soy proteins, soy protein did not similarly affect cecal butyrate in rats fed FOS or RPS . In another experiment, rats were fed diets containing high amylose cornstarch (HAS, 200 g/kg diet) with casein, casein + oligo-L-methionine (OM, 3 g/kg diet), soy protein, soy protein + OM (3 g/kg diet) or rice protein (250 g/kg diet) for 10 d . OM (digestibility, 31%) was substituted for the same amount of casein . Rats fed rice protein had greater cecal butyrate than rats fed casein (P < 0.05) . OM supplementation to casein or soy protein increased cecal butyrate compared with rats fed casein or soy protein alone (P < 0.05) . These data support our hypothesis that resistant protein and peptides promote cecal butyrate production and suggest that the differing potency of rice and soy proteins in promoting cecal butyrate production might be explained in part by the different amino acid composition of resistant protein.

J Nat Prod, 1999 Jun, 62(6), 889 - 90
Mathemycin B, a new antifungal macrolactone from actinomycete species HIL Y-8620959; Mukhopadhyay T et al.; A new macrocyclic lactone antibiotic mathemycin B (1) was isolated from the fermentation broth of an Actinomycete sp . culture Y-8620959 . The structure of 1 was elucidated by high-resolution MS and interpretation of 2D NMR results . Mathemycin B is active against a variety of phytopathogenic organisms.

Appl Microbiol Biotechnol, 1999 May, 51(5), 553 - 63
Recent trends in the biochemistry of surfactin; Peypoux F et al.; The name surfactin refers to a bacterial cyclic lipopeptide, primarily renowned for its exceptional surfactant power since it lowers the surface tension of water from 72 mN m-1 to 27 mN m-1 at a concentration as low as 20 microM . Although surfactin was discovered about 30 years ago, there has been a revival of interest in this compound over the past decade, triggered by an increasing demand for effective biosurfactants for difficult contemporary ecological problems . This simple molecule also looks very promising as an antitumoral, antiviral and anti-Mycoplasma agent . Structural characteristics show the presence of a heptapeptide with an LLDLLDL chiral sequence linked, via a lactone bond, to a beta-hydroxy fatty acid with 13-15 C atoms . In solution, the molecule exhibits a characteristic "horse saddle" conformation that accounts for its large spectrum of biological activity, making it very attractive for both industrial applications and academic studies . Surfactin biosynthesis is catalysed non-ribosomally by the action of a large multienzyme complex consisting of four modular building blocks, called the surfactin synthetase . The biosynthetic activity involves the multicarrier thiotemplate mechanism and the enzyme is organized in structural domains that place it in the family of peptide synthetases, a class of enzymes involved in peptidic secondary-metabolite synthesis . The srfA operon, the sfp gene encoding a 4'-phosphopantetheinyltransferase and the comA regulatory gene work together for surfactin biosynthesis, while the gene encoding the acyltransferase remains to be isolated . Concerning surfactin production, there is no indication whether the genetic regulation, involving a quorum-sensing mechanism, overrides other regulation factors promoted by the fermentation conditions . Knowledge of the modular arrangement of the peptide synthetases is of the utmost relevance to combinatorial biosynthetic approaches and has been successfully used at the gene level to modify the surfactin template . Biosynthetic and genetic rationales have been described for building variants . A fine study of the structure/function relationships associated with the three-dimensional structure has led to the recognition of the specific residues required for activity . These studies will assist researchers in the selection of molecules with improved and/or refined properties useful in oil and biomedical industries.

Appl Environ Microbiol, 1999 Jul, 65(7), 3114 - 20
Effect of growth temperature on hydrolytic and esterifying activities from pseudomonas fragi CRDA 037 grown on whey
Fonchy E, Morin A, Rodrigue N, Muller B, Chalier P.
The production of hydrolytic and esterifying activities of Pseudomonas fragi CRDA 037 grown on a whey-based medium was investigated at different temperatures over time . The optimal temperature was found to be critical and different for the production of both activities . The highest hydrolytic activity was detected with bacteria cultivated at between 24 degrees C (149.2 U/liter) and 27 degrees C (133.8 U/liter), while the highest production of ethyl valerate (esterifying activity) was observed by using biomass grown at 15 degrees C (0.75 U/liter) . When the fermentation temperature was increased, the incubation time necessary to reach the maximal concentration of both activities was reduced . Studies of the thermostability of both activities showed that the hydrolytic activity was more stable than the esterifying activity at 15 and 24 degrees C . Statistical analysis allowed the determination of the equations that predicted the production of hydrolytic and esterifying activities as a function of time and growth temperature . The optimal assay temperatures for the hydrolytic and esterifying activities were 37 degrees C and 12 to 15 degrees C, respectively.

Appl Environ Microbiol, 1999 Jul, 65(7), 3056 - 63
Microbial communities associated with anaerobic benzene degradation in a petroleum-contaminated aquifer; Rooney-Varga JN et al.; Microbial community composition associated with benzene oxidation under in situ Fe(III)-reducing conditions in a petroleum-contaminated aquifer located in Bemidji, Minn., was investigated . Community structure associated with benzene degradation was compared to sediment communities that did not anaerobically oxidize benzene which were obtained from two adjacent Fe(III)-reducing sites and from methanogenic and uncontaminated zones . Denaturing gradient gel electrophoresis of 16S rDNA sequences amplified with bacterial or Geobacteraceae-specific primers indicated significant differences in the composition of the microbial communities at the different sites . Most notable was a selective enrichment of microorganisms in the Geobacter cluster seen in the benzene-degrading sediments . This finding was in accordance with phospholipid fatty acid analysis and most-probable-number-PCR enumeration, which indicated that members of the family Geobacteraceae were more numerous in these sediments . A benzene-oxidizing Fe(III)-reducing enrichment culture was established from benzene-degrading sediments and contained an organism closely related to the uncultivated Geobacter spp . This genus contains the only known organisms that can oxidize aromatic compounds with the reduction of Fe(III) . Sequences closely related to the Fe(III) reducer Geothrix fermentans and the aerobe Variovorax paradoxus were also amplified from the benzene-degrading enrichment and were present in the benzene-degrading sediments . However, neither G . fermentans nor V . paradoxus is known to oxidize aromatic compounds with the reduction of Fe(III), and there was no apparent enrichment of these organisms in the benzene-degrading sediments . These results suggest that Geobacter spp . play an important role in the anaerobic oxidation of benzene in the Bemidji aquifer and that molecular community analysis may be a powerful tool for predicting a site's capacity for anaerobic benzene degradation.

J Dairy Sci, 1999 Jun, 82(6), 1296 - 304
Effect of coating whole cottonseed on performance of lactating dairy cows; Bernard JK et al.; Thirty-six lactating Jersey cows were used in a randomized block design to determine the effect of coating whole fuzzy cottonseed to improve handling characteristics on intake, milk yield, apparent digestibility of nutrients, and blood gossypol concentrations . Treatments included whole cottonseed at 15% of dietary dry matter either as whole cottonseed, whole cottonseed coated with 5% gelatinized corn starch, or whole cottonseed coated with 5% corn starch plus 10% maltodextrin sugar . Dry matter intake, milk yield, percentage of milk protein and lactose, and yield of milk components were not different among treatments; however, the percentage of milk fat was depressed when maltodextrin sugar was included in the coating . When in vitro fermentations of mixed ruminal microorganism were conducted, final pH was lower and concentrations of total fatty acids, propionate, and L-lactate were higher for whole cottonseed coated with starch and sugar compared with uncoated cottonseed . Nutrient intake was similar among treatments, but the apparent digestibility of acid and neutral detergent fiber was reduced when coated cottonseed were fed . Total plasma gossypol concentration was higher for the cottonseed coated with starch compared with cottonseed coated with starch and sugar, but the difference was not of biological significance . Results of this study indicate that coating whole cottonseed with starch does not alter its palatability or nutrient value for supporting milk yield, but a reduction in fiber digestibility was observed . Inclusion of 10% maltodextrin sugar in the coating altered ruminal fermentation and resulted in a depressed percentage of milk fat.

J Dairy Sci, 1999 Jun, 82(6), 1252 - 60
Effects of extrusion of grain and feeding frequency on rumen fermentation, nutrient digestibility, and milk yield and composition in dairy cows; Shabi Z et al.; The effect of corn extrusion and feeding frequency on ruminal and postruminal digestibility and milk yield was studied in cows fed a high concentrate diet . Four Israeli Holstein cows fitted with rumen and abomasal cannulas were used . The experiment was arranged as a 2 x 2 factorial design, with two diets and two feeding frequencies (two or four meals per day) . One diet contained 40% ground corn . In the second diet, half of the ground corn was replaced with extruded corn . Feeding cows the extruded versus ground corn diet decreased ruminal ammonia N and plasma urea N concentrations, increased postruminal digestibility of nonstructural carbohydrates, reduced dry matter intake, decreased yield of milk and milk components, and increased efficiency of milk energy and milk protein synthesis . The inclusion of extruded corn in the diet did not affect ruminal volatile fatty acid . Increasing the feeding frequency reduced the diurnal variation in ruminal pH, ruminal ammonia, and plasma urea, and increased dry matter intake--considerably more in the cows fed ground versus extruded corn--and improved postruminal organic matter, nonstructural carbohydrate, and crude protein digestibility . Total tract digestibility of organic matter and crude protein and milk yield and composition were also increased when cows were fed four versus two meals . Concurrent with the feeding frequency and grain processing effect, an increase in rumen-undegradable protein flow was related to increased digestion of nonstructural carbohydrate postruminally (r = 0.54) . We concluded that for cows fed high-starch diets more frequent meals are useful for improving postruminal digestibility and milk yield and composition.

Kansenshogaku Zasshi, 1999 May, 73(5), 407 - 13
{Characterization of enterohemorrhagic Escherichia coli O26 and development of its isolation media}; Hiramatsu R et al.; We studied 101 strains of Enterohemorrhagic Escherichia coli (EHEC) O26 isolated from diarrhea patients in six prefectural institutes of public health in Japan during June 1996 and December 1997 and tried to establish an isolation medium for EHEC O26 . None of the 101 EHEC O26 strains fermented rhamnose; Whereas all of the other EHEC including O157 and non-EHEC (166 strains) fermented rhamnose except 1 strain of non-EHEC . All of the randomly selected EHEC O26 (14 strains of O26:H11.2 strains of O26:H-) showed a very high resistance to potassium tellurite (Minimal Inhibitory Concentration (MIC) > or = 50 micrograms/ml), whereas all of the randomly selected non-EHEC (26 strains) but 1 showed a high sensitivity (MIC < or = 6.25 micrograms/ml) to this compound . On the basis of these results, we developed a Rhamnose MacConkey (RMAC) medium in which lactose in the MacConkey medium was replaced by rhamnose, and Cefixime-Potassium Tellurite-RMAC (CT-RMAC) medium in which Cefixime (0.05mg/l) and Potassium Tellurite (25mg/l) was added to RMAC for the isolation of EHEC O26 strains . We then evaluated the specifcity of these selective media by growing a selected number of O26 (24 strains) and 9 selected strains of bacteria . All of the EHEC O26 strains generated rhamnose non-fermented colonies (white color) on both media . In contrast to the EHEC O26, the vast majority of E . coli strains (166/167 = 99.4%) other than EHEC O26 were theoretically assumed to generate red colonies on the RMAC medium because of their rhamnose fermenting character and most of them were assumed not to grow on CT-RMAC medium because of their sensitivity to potassium tellurite . These findings and results indicate that EHEC O26 can be easily distinguished from other strains of E . coli including O157 . Although EHEC O26 strains showed somewhat poor growth on CT-RMAC medium compared with that on RMAC medium, these O26 showed almost the same degree of growth on CT-RMAC as they showed on DHL media . The results of the present study demonstrated that the use of RMAC and CT-MRAC media for the isolation of EHEC O26 is very reliable and efficient with RMAC having good sensitivity and CT-RMAC having a better specificity for the isolation of this strain of EHEC.

Hindustan Antibiot Bull, 1997 Feb-Nov, 39(1-4), 11 - 5
Auxotrophic requirement for sporangiospore--yeast transformation of Dimorphomyces diastaticus strain C 12; Omoifo CO; The filamentous microorganism tentatively known as Dimorphomyces diastaticus originally isolated from fermenting soursop extract, in contradistinction grew in the considiogeneous form in glucose-ammonium sulphate-basal salts buffered medium which induced polar budding yeast cells of another dimorphic microorganism, D . pleomorphis (1) . Further experiments using this medium showed that a number of growth factors including inositol, thymine and uracil separately incorporated inducted yeast morphology . Carbon substrates gave rise to polar budding yeast cells in the following order: galactose, malic acid, maltose, but mycelial fragments were preponderant in the reverse order . There was no growth with citric acid as substrate . With maltose-substrate and uracil incorporation in buffered basal salts medium, only polar budding yeast cells were induced, while granular particles abound with thymine supplementation . It was concluded that D . diastaticus strain C12 auxotrophically required a dissacharide, maltose, and pyrimidine base, uracil, for sporangiospore-yeast transformation in buffered ammonium sulphate-basal salts medium, pH 3.5.

Hindustan Antibiot Bull, 1997 Feb-Nov, 39(1-4), 1 - 10
Isolation and purification of penicillin G acylase obtained from Escherichia coli (NCIM-2400) and immobilisation on Eupergit C for the production of 6 amino penicillanic acid; Hegde MM et al.; Penicillin G-Acylase is produced by submerged cultivation of E . Coli (NCIM-2400) and extracted from the harvested fermented broth, purified (affinity chromatography) and immobilised on Eupergit C (Synthetic polymer in bead form) . The immobilised penicillin G acylase properties are studied and compared with soluble penicillin G-acylase . The control parameters for conversion of penicillin G-K to 6 APA are optimised {e.g . substrate (Pen G-K) concentration ratio to immobilised penicillin G-acylase, temperature, pH etc.} in a stirred tank reactor . Our findings suggest that immobilised penicillin G-acylase can be used commercially and the productivity of 1 kg . of immobilised enzyme is around 400 kg of 6 APA under given desired stipulated conditions.

Mol Microbiol, 1999 Jun, 32(6), 1198 - 211
The response to stationary-phase stress conditions in Escherichia coli: role and regulation of the glutamic acid decarboxylase system; De Biase D et al.; Inducible bacterial amino acid decarboxylases are expressed at the end of active cell division to counteract acidification of the extracellular environment during fermentative growth . It has been proposed that acid resistance in some enteric bacteria strictly relies on a glutamic acid-dependent system . The Escherichia coli chromosome contains distinct genes encoding two biochemically identical isoforms of glutamic acid decarboxylase, GadA and GadB . The gadC gene, located downstream of gadB, has been proposed to encode a putative antiporter implicated in the export of gamma-aminobutyrate, the glutamic acid decarboxylation product . In the present work, we provide in vivo evidence that gadC is co-transcribed with gadB and that the functional glutamic acid-dependent system requires the activities of both GadA/B and GadC . We also found that expression of gad genes is positively regulated by acidic shock, salt stress and stationary growth phase . Mutations in hns, the gene for the histone-like protein H-NS, cause derepressed expression of the gad genes, whereas the rpoS mutation abrogates gad transcription even in the hns background . According to our results, the master regulators H-NS and RpoS are hierarchically involved in the transcriptional control of gad expression: H-NS prevents gad expression during the exponential growth whereas the alternative sigma factor RpoS relieves H-NS repression during the stationary phase, directly or indirectly accounting for transcription of gad genes.

J Cell Sci, 1999 Jul, 112 ( Pt 14), 2347 - 54
A single external enzyme confers alternative NADH:ubiquinone oxidoreductase activity in Yarrowia lipolytica; Kerscher SJ et al.; NADH:ubiquinone oxidoreductases catalyse the first step within the diverse pathways of mitochondrial NADH oxidation . In addition to the energy-conserving form commonly called complex I, fungi and plants contain much simpler alternative NADH:ubiquinone oxido-reductases that catalyze the same reaction but do not translocate protons across the inner mitochondrial membrane . Little is known about the distribution and function of these enzymes . We have identified YLNDH2 as the only gene encoding an alternative NADH:ubiquinone oxidoreductase (NDH2) in the obligate aerobic yeast Yarrowia lipolytica . Cells carrying a deletion of YLNDH2 were fully viable; full inhibition by piericidin A indicated that complex I activity was the sole NADH:ubiquinone oxidoreductase activity left in the deletion strains . Studies with intact mitochondria revealed that NDH2 in Y . lipolytica is oriented towards the external face of the mitochondrial inner membrane . This is in contrast to the situation seen in Saccharomyces cerevisiae, Neurospora crassa and in green plants, where internal alternative NADH:ubiquinone oxidoreductases have been reported . Phylogenetic analysis of known NADH:ubiquinone oxidoreductases suggests that during evolution conversion of an ancestral external alternative NADH:ubiquinone oxidoreductase to an internal enzyme may have paved the way for the loss of complex I in fermenting yeasts like S . cerevisiae.

Can J Microbiol, 1999 Feb, 45(2), 145 - 52
Isolation of Desulfovibrio intestinalis sp . nov . from the hindgut' of the lower termite Mastotermes darwiniensis; Frohlich J et al.; A Gram-negative, anaerobic sulfate-reducing bacterium was isolated from hindgut contents of the lower termite Mastotermes darwiniensis Froggatt (strain KMS2) . Strain KMS2 is motile by a single polar flagellum . The isolate possesses desulfoviridin and catalase activity . The G+C content of its DNA is in the range of 54.5-55.5 mol% (strain KMS2) . It respires hydrogen and different low molecular weight organic compounds in the presence of sulfate, thiosulfate, and sulfite, and also oxygen . The isolated strain ferments pyruvate . Fastest growth with a doubling time of 12.5 h was obtained at 37 degrees C and not at 28 degrees C, the temperature at which the termites were grown . The isolate showed a 16S rDNA sequence homology of 95.9% to Desulfovibrio desulfuricans ATCC 27774 and a DNA-DNA homology of 44.6% to D . desulfuricans Essex 6 (type strain) . Based on its biochemical properties and 16S rDNA sequence, the isolate was assigned to a new species named Desulfovibrio intestinalis.

Biosci Biotechnol Biochem, 1999 May, 63(5), 934 - 6
Multi-enzymatic glucosylation using Eucalyptus UDP-glucosyltransferase coupled UDPglucose-fermentation by bakers' yeast; Nakajima N et al.; The enzymatic synthesis of glucoside compounds using a membrane-associated UDP-glucosyltransferase fraction from Eucalyptus perriniana cultured cells as a water-insoluble catalyst (N . Nakajima, et . al., J . Ferment . Bioeng., 84 (5), pp . 455-460, 1997) has been effectively done by coupling UDPglucose-fermentation by bakers' yeast . For example, beta-thujaplicin (hinokitiol) and p-aminobenzoic acid were converted respectively to their corresponding beta-D-monoglucosides with the conversion rate of around 24-26% by the multi-enzymatic system with UDPglucose as a glucose donor, which is produced by yeast cells from glucose and 5'-UMP . Addition of either cellobiose, a substrate of beta-glucosidase, or DL-1,2-anhydro-myo-inositol, an inhibitor for the enzyme in the reaction mixture, could increased the yield of these beta-D-monoglucosides . This new enzymatic system could also be used for the synthesis of flavonoid glucosides such as isoquercitrin (quercetin 3-O-beta-D-glucoside).

Biosci Biotechnol Biochem, 1999 May, 63(5), 805 - 12
Production and properties of the linamarase and amygdalase activities of Penicillium aurantiogriseum P35; Petruccioli M et al.; The effects of medium composition on the production of beta-glucosidase (amygdalase and linamarase) by Penicillium aurantiogriseum P35 were studied and the medium optimized as follows (g/l of deionized water): pectin, 10.0; (NH4)2SO4, 8.0; KH2PO4, 8.0; Na2HPO4, 2.8; MgSO4.7H2O, 0.5; yeast extract, 4.0; initial pH 6.0 . When grown in a bench fermenter on this medium, the fungus produced 50.5 mU of amygdalase and 9.4 mU of linamarase per ml of culture broth . Two beta-glucosidases (PGI and PGII), each having amygdalase and linamarase activities, were recovered from the culture broth and purified; their relative molecular weights, as native enzymes, were estimated to be about 247,000 and 147,000, respectively . Both enzymes showed the same optimum pH (6.0) but different optimum temperatures (55 and 60 degrees C for PGI and PGII, respectively) . Thermostability (10 min at 60 degrees C) and half-life of enzyme activity (7 hours at 60 degrees C) of PGII were higher than those of PGI (10 min at 50 degrees C and 2 hours at 55 degrees C, respectively) . A wide range of cyanogenic glycosides (such as tetraphyllin B, epivolkenin, gynocardin, passibiflorin, prunasin, taxiphyllin, amygdalin, lucumin, sambunigrin, dhurrin, linamarin and cardiospermin sulfate) were hydrolyzed by both enzymes.

Bioelectrochem Bioenerg, 1999 May, 48(2), 389 - 96
Genetic programming as an analytical tool for non-linear dielectric spectroscopy; Woodward AM et al.; By modelling the non-linear effects of membranous enzymes on an applied oscillating electromagnetic field using supervised multivariate analysis methods, Non-Linear Dielectric Spectroscopy (NLDS) has previously been shown to produce quantitative information that is indicative of the metabolic state of various organisms . The use of Genetic Programming (GP) for the multivariate analysis of NLDS data recorded from yeast fermentations is discussed, and GPs are compared with previous results using Partial Least Squares (PLS) and Artificial Neural Nets (NN) . GP considerably outperforms these methods, both in terms of the precision of the predictions and their interpretability.

Acta Microbiol Immunol Hung, 1999, 46(2-3), 289 - 95
Impact of yeast genetics and molecular biology on traditional and new biotechnology; Maraz A; Developments in yeast genetics, biochemistry, physiology and process engineering provided bases of rapid development in modern biotechnology . Elaboration of the recombinant DNA technique is far the most important milestone in this field . Other molecular genetic techniques, as molecular genotyping of yeast strains proved also very beneficial in yeast fermentation technologies . Saccharomyces cerevisiae is the most exploited eukaryotic microorganism in biotechnology but non-Saccharomyces species are becoming more and more important in the production of perfectly translated heterologous proteins.

Infect Immun, 1999 Jul, 67(7), 3390 - 8
Mycoplasmal lipopeptide MALP-2 induces the chemoattractant proteins macrophage inflammatory protein 1alpha (MIP-1alpha), monocyte chemoattractant protein 1, and MIP-2 and promotes leukocyte infiltration in mice; Deiters U et al.; Natural as well as experimental infections with pathogenic mycoplasmas lead to cellular responses characterized by early polymorphonuclear leukocyte influx, which in turn is followed by infiltration of macrophages . Since some of the most potent leukocyte chemoattractants are macrophage products, we investigated whether the 2-kDa macrophage-activating lipopeptide (MALP-2) from Mycoplasma fermentans was capable of inducing chemoattractant chemokines and initiating an in vivo inflammatory effect . MALP-2 was a potent in vitro inducer of the chemokines macrophage inflammatory protein 1alpha (MIP-1alpha), monocyte chemoattractant protein 1 (MCP-1), and MIP-2, yielding a maximal response at 0.1 ng/ml (5 x 10(-11) M) . Leukocyte infiltration was determined after intraperitoneal injection of MALP-2, liposome-encapsulated MALP-2, and heat-killed mycoplasmas . There was a steady increase in the number of peritoneal cells over 72 h in response to these agents . Polymorph counts were maximal by 24 to 48 h, decreasing thereafter . Monocytes/macrophages had significantly increased after 3 days . MIP-1alpha, MCP-1, and MIP-2 levels in serum or peritoneal lavage fluid were determined . MIP-1alpha and MCP-1 levels were elevated by 2 to 6 h after injection and were still above control values after 24 h . In contrast, MIP-2 levels reached their maximum at 2 h, dropping to control values after 24 h . We conclude that macrophage-stimulating mycoplasmal lipoproteins, exemplified by MALP-2, play an important role in the late phase of phagocyte recruitment at sites of infection and that this is affected by leukoattractive chemokines.

Infect Immun, 1999 Jul, 67(7), 3188 - 92
Effects of Mycoplasma fermentans incognitus on differentiation of THP-1 cells; Reyes L et al.; Mycoplasma fermentans incognitus has been isolated from human tissue in patients both with and without AIDS who died of systemic infection . M . fermentans incognitus and other strains of M . fermentans have been associated with rheumatoid arthritis . While cell extracts of M . fermentans incognitus can induce changes in murine and human cells of the monocytic lineage, little is known about interactions of viable organisms with such cells . Because of the central role of macrophages in chronic inflammation, we examined the effects of M . fermentans incognitus on surface markers and functions of THP-1 cells, a well-characterized human monocytic cell line . This cell line has been used extensively in studies of macrophage differentiation, especially following exposure to phorbol esters . Changes in cell morphology, phagocytosis, rate of cell division, and selected surface markers were evaluated in cultures of THP-1 cells exposed to phorbol myristate acetate (PMA), M . fermentans incognitus, or both . As reported by other investigators, PMA induced THP-1 cells to differentiate into cells resembling tissue macrophages . M . fermentans incognitus only minimally affected changes induced by PMA, slightly increasing the percentage of cells positive for FCgammaRI and major histocompatibility complex (MHC) class II antigens . M . fermentans incognitus alone induced an incomplete arrest in the cell cycle at G0 phase, increased phagocytic ability, and enhanced expression of FCgammaRI, CR3, CR4, and MHC class II antigens.

Microbiology, 1999 May, 145 ( Pt 5), 1005 - 14
The large plasmids of Shiga-toxin-producing Escherichia coli (STEC) are highly variable genetic elements; Brunder W et al.; Shiga-toxin-producing Escherichia coli (STEC) of different serotypes are known to harbour large plasmids . The aim of this study was to investigate, using the example of the plasmid-encoded serine protease EspP, whether these plasmids are a uniform genetic element present in STEC . Examination of 201 diarrhoeagenic E . coli strains using a newly developed espP-specific PCR showed that espP is specific for STEC and present in 57% of STEC belonging to 16 different serotypes . The espP genes of the 16 STEC serotypes varied to a certain extent, as shown by nucleotide sequence and restriction enzyme analyses, but the DNA regions adjacent to the espP gene were completely different . When two further STEC-plasmid markers, the catalase-peroxidase gene katP and the enterohaemorrhagic E . coli-haemolysin gene EHEC-hlyA were included, many combinations of the three markers were found, depending in part on the serotype . In addition, strains possessing none of the three markers still harboured large plasmids . In the most prevalent STEC serogroup, O157, it was observed that the plasmid of sorbitol-fermenting STEC O157:H- lacks the espP and katP genes although both genes are present in the plasmid of the non-sorbitol-fermenting STEC O157:H7 . The EHEC-hlyA gene, however, is present in both . In conclusion, this study shows that the large plasmids of STEC are not uniform genetic elements but heterogeneous in both their gene composition and arrangement.

Biol Pharm Bull, 1999 May, 22(5), 546 - 8
A new cytotoxic 2H-pyran compound produced by acid treatment of the fermentation broth of Streptomyces sp; Nosaka C et al.; A new cytotoxic substance was isolated following acid treatment of fermented broth of a Streptomyces strain numbered MJ915-WF12 . The structure was elucidated by spectroscopic analyses to be 5-dihydro-4-formyl-6-hydroxy-2-hydroxymethyl-6-methyl-2H-pyran . It was thought to be generated by dehydration and ring reformation of the precursor material in cultured broth by acid.

Mol Cell Biol, 1999 Jul, 19(7), 4561 - 71
Std1 and Mth1 proteins interact with the glucose sensors to control glucose-regulated gene expression in Saccharomyces cerevisiae; Schmidt MC et al.; The Std1 protein modulates the expression of glucose-regulated genes, but its exact molecular role in this process is unclear . A two-hybrid screen for Std1-interacting proteins identified the hydrophilic C-terminal domains of the glucose sensors, Snf3 and Rgt2 . The homologue of Std1, Mth1, behaves differently from Std1 in this assay by interacting with Snf3 but not Rgt2 . Genetic interactions between STD1, MTH1, SNF3, and RGT2 suggest that the glucose signaling is mediated, at least in part, through interactions of the products of these four genes . Mutations in MTH1 can suppress the raffinose growth defect of a snf3 mutant as well as the glucose fermentation defect present in cells lacking both glucose sensors (snf3 rgt2) . Genetic suppression by mutations in MTH1 is likely to be due to the increased and unregulated expression of hexose transporter genes . In media lacking glucose or with low levels of glucose, the hexose transporter genes are subject to repression by a mechanism that requires the Std1 and Mth1 proteins . An additional mechanism for glucose sensing must exist since a strain lacking all four genes (snf3 rgt2 std1 mth1) is still able to regulate SUC2 gene expression in response to changes in glucose concentration . Finally, studies with green fluorescent protein fusions indicate that Std1 is localized to the cell periphery and the cell nucleus, supporting the idea that it may transduce signals from the plasma membrane to the nucleus.

Mol Cell Probes, 1999 Jun, 13(3), 183 - 9
Mycoplasma orale has a sequence similar to the insertion-like sequence of M . fermentans; Ditty SE et al.; Polymerase chain reaction (PCR) using the Mycoplasma fermentans insertion sequence-like element (ISLE) RW primer set amplifies DNA from Mycoplasma orale when more than 1 ng is present in the reaction tube . In this study, amplified products from 11 different clinical isolates and the ATCC prototype of M . orale were sequenced and compared to 206 bp amplicons from eight isolates and the ATCC strain of M . fermentans . The nucleotide sequences of the amplified M . orale products had high sequence homology (88-92%) to those from M . fermentans, but differed at several key positions . The M . orale products contained a DraI restriction enzyme site not found in any of the M . fermentans amplified products . Consistent with this finding, the PCR products from M . orale were digested by DraI while the PCR products from M . fermentans were resistant to DraI digestion . The results suggest that M . orale may carry a similar IS-like element that complicates but does not negate using the ISLE PCR assay designed to detect M . fermentans . It appears possible for the RW primers to amplify M . orale if the mycoplasmas are present at higher concentrations . The amplified products can be differentiated from those from M . fermentans by a rapid DraI restriction endonuclease digestion or by Southern blot analysis using the RW006 internal probe under highly stringent conditions .

J AOAC Int, 1999 May-Jun, 82(3), 657 - 62
Fusarium mycotoxins in corn and corn products in a high-risk area for gastric cancer in Shandong Province, China; Groves FD et al.; Consumption of fermented, but not unfermented, corn pancakes has been linked with elevated stomach cancer mortality rates in rural Linqu County in Shandong Province, China . Previous surveys of fungal contamination of corn in China have detected fumonisins, which are mycotoxins produced by Fusarium moniliforme . To determine whether mycotoxins might account for the increased risk of cancer among those consuming fermented pancakes, we obtained specimens of corn, cornmeal, unfermented and fermented pancake batter, and cooked fermented pancakes from each of 16 households in Linqu County for analysis by the U.S . Department of Agriculture . Fumonisins B1, B2, and B3 were detected (> or = 0.5 microgram/g) in 19, 25, and 6% of the corn specimens, respectively, as well as in various corn products . No type A trichothecenes were detected; however, the type B trichothecenes deoxynivalenol and 15-acetyldeoxynivalenol were detected (> or = 0.5 microgram/g) in 58 and 17% of the corn specimens, respectively, and zearalenone was detected (> or = 0.5 microgram/g) in 15% of the cornmeal specimens . The mycotoxins were detected only at low levels (< 10 micrograms/g), which did not increase with fermentation . These findings do not support the hypothesis that mycotoxin contamination increases the risk of gastric cancer among those who consume fermented Chinese pancakes.

J Bacteriol, 1999 Jun, 181(12), 3716 - 20
Functioning of DcuC as the C4-dicarboxylate carrier during glucose fermentation by Escherichia coli; Zientz E et al.; The dcuC gene of Escherichia coli encodes an alternative C4-dicarboxylate carrier (DcuC) with low transport activity . The expression of dcuC was investigated . dcuC was expressed only under anaerobic conditions; nitrate and fumarate caused slight repression and stimulation of expression, respectively . Anaerobic induction depended mainly on the transcriptional regulator FNR . Fumarate stimulation was independent of the fumarate response regulator DcuR . The expression of dcuC was not significantly inhibited by glucose, assigning a role to DcuC during glucose fermentation . The inactivation of dcuC increased fumarate-succinate exchange and fumarate uptake by DcuA and DcuB, suggesting a preferential function of DcuC in succinate efflux during glucose fermentation . Upon overexpression in a dcuC promoter mutant (dcuC*), DcuC was able to compensate for DcuA and DcuB in fumarate-succinate exchange and fumarate uptake.

Br J Biomed Sci, 1998 Sep, 55(3), 184 - 91
Differential effects of sodium butyrate on the transcription of the human TIS11 family of early-response genes in colorectal cancer cells; Maclean KN et al.; The tetradecanoyl phorbol acetate (TPA)-inducible sequence 11 (TIS11) family of early-response proteins consists of at least five members . They share a highly conserved Cys3His zinc-binding motif, but otherwise have little sequence similarity . Their function remains unknown, but all are induced rapidly and transiently in response to extracellular hormone and growth factor signals . Sodium butyrate, a fermentation product of dietary fibre, effects colorectal cancer cell proliferation by inducing growth arrest, differentiation and apoptosis . In this communication, we report that butyrate has differential effects on the transcription of the three human TIS11 family members identified so far in T84 and HT-29 human colorectal cancer cell lines . Butyrate response factor 1 (BRF1) transcription is repressed, butyrate response factor 2 (BRF2) transcription is activated and there is no apparent effect on the transcription of human TIS11 (HTIS11) . Induction and repression occur rapidly, with altered mRNA levels detectable within 15 min of butyrate addition . Two other short-chain fatty acids, propionate and acetate, have no detectable effects on BRF1 or BRF2 transcription.

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