Microbial

J Infect Dis, 2001 Feb 15, 183(4), 662 - 5 Epub 2001 Jan 11.
Altered expression of CD88 and associated impairment of complement 5a-induced neutrophil responses in human immunodeficiency virus type 1-infected patients with and without pulmonary tuberculosis; Meddows-Taylor S et al.; The effect of infection with human immunodeficiency virus type 1 (HIV patient group), infection with Mycobacterium tuberculosis (TB patient group), and coinfection with both of these organisms (HIV/TB patient group) on the expression of CD88 on polymorphonuclear leukocytes (PMNL) was determined by using a receptor-specific monoclonal antibody and flow cytometry . A significant reduction in the fluorescence intensity of CD88 on PMNL was observed in the HIV and HIV/TB groups, compared with both the healthy donor (HD) and TB groups . Furthermore, when degranulation of PMNL was induced by ligation of CD88 by complement 5a (C5a), a large proportion of patients in the HIV and the HIV/TB groups was found to have reciprocal degranulation responses . Patients in the 2 HIV groups also were found to have significantly reduced C5a-induced chemotactic responses and significantly elevated peripheral levels of C5a des Arg, compared with the HD and TB groups . These differences may contribute to the increased susceptibility of HIV-1-infected individuals to secondary microbial infections.

Eur J Immunol, 2000 Dec, 30(12), 3591 - 7
Bacterial CpG-DNA activates dendritic cells in vivo: T helper cell-independent cytotoxic T cell responses to soluble proteins; Sparwasser T et al.; Receptors for conserved molecular patterns associated with microbial pathogens induce synthesis of co-stimulatory molecules and cytokines in immature dendritic cells (DC), as do antigen-reactive CD4 T helper cells via CD40 signaling . Once activated, antigen-presenting DC may activate CD8 T cell responses in a T helper cell-independent fashion . Using immunostimulatory CpG-oligonucleotides (ODN) mimicking bacterial CpG-DNA, we tested whether CpG-DNA bypasses the need for T helper cells in CTL responses towards proteins by directly activating antigen-presenting DC to transit into professional APC . We describe that immature DC in situ constitutively process soluble proteins and generate CD8 T cell determinants yet CD8 T cell responses remain abortive . Induction of primary antigen-specific CD8 cytotoxic T lymphocyte (CTL)-mediated responses becomes initiated in wild-type as well as T helper cell-deficient mice, provided soluble protein and CpG-ODN are draining into the same lymph node . Specifically we show that CpG-ODN trigger antigen-presenting immature DC within the draining lymph node to acutely up-regulate co-stimulatory molecules and produce IL-12 . These results provide new insights for generating in vivo efficient CTL responses to soluble proteins which may influence vaccination strategies.

J Appl Microbiol, 2001 Feb, 90(2), 294 - 300
Epithelium-associated bacteria in the gastrointestinal tract of Arctic charr (Salvelinus alpinus L.) . An electron microscopical study; Ringo E et al.; AIMS: The primary aim was to use transmission electron microscopy (TEM) and scanning electron microscopy (SEM) to define the location of epithelium-associated bacteria in the digestive tract of the salmonid fish, Arctic charr (Salvelinus alpinus) . METHODS AND RESULTS: TEM and SEM examination of the gastrointestinal tract demonstrated substantial numbers of ovoid and rod-shaped bacterial cells associated with the microvillous brush borders of enterocytes . Bacteria were found at the tips of microvilli as well as between adjacent microvilli . Endocytosis of bacteria by epithelial cells was observed in two regions (pyloric caeca and midgut) . CONCLUSION: Electron microscope examination of the gut is an important tool for evaluating the microbial ecology of the fish digestive tract ecosystem . SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the current study clearly demonstrate that the intestine is involved in bacterial endocytosis.

Eur J Biochem, 2001 Jan, 268(2), 374 - 83
Mannosylerythritol lipid induces characteristics of neuronal differentiation in PC12 cells through an ERK-related signal cascade; Wakamatsu Y et al.; Rat pheochromocytoma PC12 cells undergo neuronal differentiation in response to nerve growth factor (NGF) . The differentiation involves protein kinase cascades that include the kinases MEK and ERK, as well as activation of the transcription factors c-Jun and c-Fos . We show here, that exposure of PC12 cells to mannosylerythritol lipid (MEL), a yeast extracellular glycolipid, enhances the activity of acetylcholinesterase and interrupts the cell cycle at the G1 phase, with resulting outgrowth of neurites and partial cellular differentiation . Treatment with MEL stimulates the phosphorylation of ERK to a similar extent as treatment with NGF, although, the appearance of phosphorylated ERK is somewhat delayed . Both the MEL-induced outgrowth of neurites and the increase in the activity of acetylcholinesterase are prevented by PD98059, a specific inhibitor of MEK . Northern blotting analysis of c-jun transcripts and analysis of transcription in PC12 cells of a c-jun/CAT reporter construct demonstrated a significant increase in the rate of transcription of the c-jun gene upon treatment with MEL . The sequence elements required for the MEL-mediated activation of transcription of the c-jun gene are located between nucleotides -126 and -79 in the 5' flanking region . Our results suggest that MEL induces characteristics of neuronal differentiation in PC12 cells, with transactivation of the c-jun gene, via an ERK-related signal cascade that is partially overlapping the pathways activated in response to NGF . These results might provide the groundwork for the use of microbial extracellular glycolipids as novel reagents for the treatment of cancer cells.

Clin Oral Implants Res, 2000, 11 Suppl 1, 33 - 58
The scientific basis for and clinical experiences with Straumann implants including the ITI Dental Implant System: a consensus report; Cochran DL; Successful endosseous implant therapy requires integration of the implant with bone, soft connective tissue and epithelium . This report from a consensus conference on Straumann dental implants including the ITI Dental Implant System documents the interaction of these nonsubmerged one-piece implants with the oral tissues and reviews clinical studies supporting the high success achievable with these implants in patients . Light and electron microscopy reveal that epithelial structures similar to teeth are found around the implants . A connective tissue zone exists between the apical extension of the junctional epithelium and the alveolar bone . This connective tissue comprises a dense circular avascular zone of connective tissue fibers surrounded by a loose vascular connective tissue . The histologic dimensions of the epithelium and connective tissue comprising the biologic width are similar to the same tissues around teeth . The nonsubmerged one-piece design of the Straumann implants, which have been used for over 20 years, has set a standard in implant dentistry, with other implants now being manufactured and placed using similar techniques . Straumann implants have an endosseous portion that is either coated with a well-characterized and well-documented titanium plasma-sprayed surface or is sandblasted and acid attacked . Both surfaces have been shown to have advantages for osseous integration compared to machined and other smoother implant surfaces . These advantages include greater amounts of bone-to-implant contact, more rapid integration with bone tissue, and higher removal torque values . The lack of component connection at or below the alveolar crest provides additional benefits . Component connection at the alveolar crest, as seen with submerged implants, results in microbial contamination, crestal bone loss and a more apical epithelial location . Numerous human clinical trials document the successful use of Straumann implants in a variety of indications and areas of the mouth . These include prospective long-term trials using strict criteria of success and life table analyses . Taken together, the clinical studies reveal that Straumann implants can be used predictably in partially edentulous and completely edentulous maxilla and mandibles with high success rates . Furthermore, the animal and microscopic studies reviewed provide a scientific basis for the integration of Straumann implants with bone, connective tissue and epithelium.

J Microbiol Methods, 2001 Feb 1, 44(1), 79 - 88
Ecological and physiological analyses of Pseudomonad species within a phenol remediation system; Whiteley AS et al.; A diverse collection of 700 bacteria obtained from an operational phenolic remediating industrial treatment plant was made to select potential strains as microbial biosensors . Pseudomonads were the most abundant group, of which 48 selected from the liquor or suspended solids were assessed for their physiological response to phenolic pollutant loading and niche specialisation . By FAME-MIS identification the Pseudomonads were clustered into six major species groups . Those isolates able to utilise phenol as a sole carbon source predominantly belonged to a non-clonal Pseudomonas pseudoalcaligenes cluster determined by REP-PCR genotyping . Rapid microtitre based respiration assays were developed to contrast activity in response to increasing concentrations of phenol . A considerable range in response for both phenol degrader and non-degrader strains was observed . This natural phenotypic and physiological heterogeneity could facilitate the selection of isolates for the development of a suite of ecologically relevant, custom designed sensors with predictable toxicity susceptibilities to monitor process efficacy.

Environ Exp Bot, 2001 Feb, 45(1), 1 - 9
UV-C irradiation reduces microbial populations and deterioration in Cucurbita pepo fruit tissue; Erkan M et al.; Tissue slices of zucchini squash (Cucurbita pepo L., cv . Tigress) fruit were exposed to ultraviolet-C (UV-C) radiation from germicidal lamps for 1, 10 or 20 min; however, only 10 and 20 min UV-C exposure significantly reduced microbial activity and deterioration during subsequent storage at 5 or 10 degrees C . UV-C treated slices had higher respiration rates than controls; however, the ethylene production of the slices was not affected by UV-C treatments . Slight UV-C irradiation damage (reddish brown discoloration) was detected on the surface of 10 and 20 min-treated slices after 12 days of storage at 10 degrees C . Slices stored at 5 degrees C did not show UV-C damage . Chilling injury was not observed until after 20 days of storage at 5 degrees C . The symptoms of chilling injury appeared as dried sunken brown spots on the surface of cortex tissue . UV-C treatments did not affect the degree of chilling injury during storage at 5 degrees C . UV-C treatment also had no consistent effect on sugar or malic acid concentrations . The most pronounced effect of UV-C irradiation was to retard microbial growth thereby providing a basis for the frequently observed delay in senescence and subsequent deterioration in fruit tissues.

Mol Immunol, 2000 Jun, 37(9), 545 - 52
Targeting early events in T cell activation to construct improved vaccines; Buchan GS et al.; Live, attenuated vaccines currently offer the best protection against virulent pathogens . Recent advances in Immunology and Molecular Biology provide an opportunity to design vaccines that will be more effective and safer than existing ones . Immunologists are rapidly developing the capacity to identify and construct the minimal immunogenic units from pathogens . The molecular signals required to fully activate antigen presenting cells (APCs) and responder T cells are becoming apparent . Improved vaccine delivery systems are being designed which will mimic the actions of pathogens in vivo . These vaccines will incorporate protective epitopes fused to immunoregulatory cytokines in chimeric proteins . They will be encapsulated in formulations which allow for the slow release of these chimeric proteins thereby inducing the memory T cells required for long-lived immunity . These vaccine formulations will target receptors present on the most active APCs . Here we discuss how these advances will allow us to rationally construct "virtual pathogens" which will provide improved protection against new and old microbial foes.

Hum Immunol, 2000 Dec, 61(12), 1270 - 3
Association of IL-10 genotype with sudden infant death syndrome; Summers AM et al.; Sudden infant death syndrome (SIDS) is a major cause of infant death of unknown etiology . We propose that SIDS results from a genetically determined imbalance in the production of inflammatory and anti-inflammatory cytokines in response to the infant's microbial flora . We were especially interested to know the relationship between SIDS and genetically determined higher or lower production of IL-10, an anti-inflammatory cytokine . Biallelic polymorphisms in the promoter region of the IL-10 gene associated with higher or lower production of IL-10 were determined in a SIDS and in a control group using a sequence-specific oligonucleotide approach . One particular allele of the IL-10 gene, the IL-10-592*A allele, was significantly associated with SIDS . Indeed, 70% of the SIDS babies carried the IL-10-592*A allele (p = 0.007 compared with control) . In addition, there was a significant reduction in the frequency of homozygosity for the allele IL-10-592*C (p = 0.001 compared with control) . Carrying the A allele (either A/A or A/C) had an odds ratio of 3.3 (95% confidence interval 1.4-8.0) . In the same patients there was no association with other IL-10 gene polymorphisms nor with other cytokine (TNF-alpha, TGF-beta 1) genotypes, emphasizing the particular relationship between SIDS and the IL-10-592*A allele.

Am J Ophthalmol, 2001 Jan, 131(1), 131 - 3
Increased incidence of corneal perforation after topical fluoroquinolone treatment for microbial keratitis; Mallari PL et al.; PURPOSE: To compare the incidence of corneal perforation in eyes treated with topical fluoroquinolone or fortified antibiotics for microbial keratitis . METHODS: A retrospective medical record review of patients hospitalized for bacterial keratitis from January 1991 through November 1999 . RESULTS: Two hundred seventy-seven cases of bacterial keratitis were identified . The incidence of corneal perforations was significantly higher in fluoroquinolone-treated eyes (18 out of 142, 12.7%) compared with eyes treated with fortified antibiotics (1 out of 135, 0.7%) (chi-square = 13.6, degrees of freedom (df) = 2, P <.001) . Within the fluoroquinolone-treated group, all corneal perforations occurred in those eyes treated with ofloxacin (18 out of 125, 14.4%) . Corneal perforations that occurred in the ofloxacin-treated eyes were not associated with any particular microbial isolate . CONCLUSION: Our data suggest an increased risk of corneal perforation after fluoroquinolone treatment for bacterial keratitis compared with treatment with fortified antibiotics . Further studies are warranted to verify this association and establish possible mechanisms by which topical fluoroquinolones may alter corneal collagen or keratocyte function.

Ecotoxicol Environ Saf, 2001 Jan, 48(1), 85 - 98
Comparative effects of a genetically engineered insect virus and a growth-regulating insecticide on microbial communities in aquatic microcosms; Kreutzweiser D et al.; The effects of a genetically engineered insect baculovirus on indigenous aquatic microbial communities were determined in closed, recirculating aquatic microcosms, and compared with the effects of a natural strain of the virus and of a growth-regulating insecticide, Dimilin . The recombinant virus was a nuclear polyhedrosis virus (NPV) of the spruce budworm (Choristoneura fumiferana (Cf)) with a lacZ marker gene inserted into the egt region of the CfNPV . The natural virus was Ireland strain CfNPV . Microbial measurement endpoints included decomposition activity (mass loss of organic material), respiration on two different substrates (O2 consumption), heterotrophic bacterial abundance (plate counts), and microbial community metabolic profiles (carbon source utilization patterns in Biolog GN microplates) . Viral DNA of both the natural strain and the recombinant viruses, detected by polymerase chain reaction techniques, settled out of the microcosm water and accumulated on bottom substrates within 3 days of the microcosm inoculations . The viral DNA persisted in bottom substrates for the duration of the 21-day experimental period, although there was some evidence that the recombinant virus was less stable than the natural strain in particulate organic matter . No significant changes in microbial decomposition or respiration activity, bacterial abundance, or average metabolic responses were detected by a time trend analysis in microcosms inoculated with either the lacZ recombinant virus or the natural Ireland strain CfNPV . Significant effects on microbial decomposition and respiration activity were detected in microcosms treated with the growth-regulating insecticide at, and above, the expected environmental concentrations . Despite significant effects on microbial community functional attributes in Dimilin-treated microcosms, there were no detectable changes in community structure in terms of metabolic profiles or bacterial abundance.

Occup Environ Med, 2001 Feb, 58(2), 113 - 8
Short term exposure to airborne microbial agents during farm work: exposure-response relations with eye and respiratory symptoms; Eduard W et al.; OBJECTIVES: Exposure to high levels of non-infectious microbial agents is recognised as a cause of respiratory disease in working populations, but except for endotoxins, little is known about exposure-response relations . As these effects do not depend on viability, exposure to non-viable microbial agents is important . Various methods not based on microbial cultures were explored to study the complex microbial exposure of farmers and associations with acute symptoms during work . METHODS: Airborne exposure was measured when farmers carried out specific tasks . Fungal spores, bacteria, endotoxins, beta(1-->3)-glucans, fungal antigens specific for Penicillium and Aspergillus species, and mites were measured by methods not based on microbial cultures . Also silica, inorganic and organic dust, ammonia, hydrogen sulphide, and nitrogen dioxide were measured . Respiratory, and nose and eye symptoms experienced during measurements were recorded by a short questionnaire . Both univariate and multivariate statistical analyses were applied to assess the relations between exposure and acute symptoms . RESULTS: 106 Farmers and their spouses participated in this study . Prevalences of work related symptoms were: wheezing 3%; chest tightness 7%; cough 14%; eye symptoms 18%; and nose symptoms 22% . Prevalence ratios for nose and eye symptoms were 4-8 after exposure to 20-500x10(3) fungal spores/m(3) and higher, and a prevalence ratio for cough was 4 after exposure to 500-17,000x10(3) fungal spores/m(3) . Nose symptoms were also associated with exposure to silica with prevalence ratios of 4-6 after exposure to 0.015-0.075 mg /m(3) and higher . CONCLUSIONS: Farmers had a high occurrence of symptoms of the nose and eyes as well as cough during work . These symptoms were associated in a dose dependent manner with exposure to fungal spores . Nose symptoms were also associated with exposure to silica.

J Immunol, 2001 Feb 15, 166(4), 2855 - 62
V beta 6+ and V beta 4+ T cells exert cooperative activity in clearance of secondary infection with Histoplasma capsulatum; Gomez FJ et al.; We previously studied the lung Vbeta TCR repertoire of C57BL/6 mice during primary infection with the pathogen Histoplasma capsulatum . We observed a consistent oligoclonal expansion of Vbeta4(+) T cells during the peak of infection and early stages of resolution . The Vbeta4(+) family played a role in protective immunity against the fungus . Depletion of this subpopulation of T cells hindered optimal clearance of infection from tissues . In this report we analyze the flux of the Vbeta TCR repertoire in the lungs of C57BL/6 mice with reinfection histoplasmosis . We observed a significant increase in Vbeta6(+) T cells on days 7, 10, and 14, the peak and early resolution phases of infection . This skewing was preceded by an increased number of memory T cells within Vbeta6(+) cells . The VDJ sequences of Vbeta6 chains were oligoclonal during the early stages of the infection, suggesting that the expansion was driven by a small number of Ags . More than 96% of the expanded Vbeta6(+) cells were CD4(+) . Depletion of Vbeta6(+) T cells but not Vbeta4(+) T cells induced a modest but significant delay in fungal clearance . Simultaneous depletion of Vbeta4(+) and Vbeta6(+) T cells induced a more pronounced impairment of host resistance . These studies illustrate the dynamic interactions between Vbeta families in the response to microbial challenge.

J Immunol, 2001 Feb 15, 166(4), 2444 - 50
Microbial lipopeptides stimulate dendritic cell maturation via Toll-like receptor 2; Hertz CJ et al.; The ability of dendritic cells (DC) to initiate immune responses in naive T cells is dependent upon a maturation process that allows the cells to develop their potent Ag-presenting capacity . Although immature DC can be derived in vitro by treatment of peripheral blood monocytes with GM-CSF and IL-4, additional signals such as those provided by TNF-alpha, CD40 ligand, or LPS are required for complete maturation and maximum APC function . Because we recently found that microbial lipoproteins can activate monocytes and DC through Toll-like receptor (TLR) 2, we also investigated whether lipoproteins can drive DC maturation . Immature DC were cultured with or without lipoproteins and were monitored for expression of cell surface markers indicative of maturation . Stimulation with lipopeptides increased expression of CD83, MHC class II, CD80, CD86, CD54, and CD58, and decreased CD32 expression and endocytic activity; these lipopeptide-matured DC also displayed enhanced T cell stimulatory capacity in MLR, as measured by T cell proliferation and IFN-gamma secretion . The lipid moiety of the lipopeptide was found to be essential for induction of maturation . Preincubation of maturing DC with an anti-TLR2 blocking Ab before addition of lipopeptide blocked the phenotypic and functional changes associated with DC maturation . These results demonstrate that lipopeptides can stimulate DC maturation via TLR2, providing a mechanism by which products of bacteria can participate in the initiation of an immune response.

J Immunol, 2001 Feb 15, 166(4), 2291 - 5
Distinct CpG DNA and polyinosinic-polycytidylic acid double-stranded RNA, respectively, stimulate CD11c- type 2 dendritic cell precursors and CD11c+ dendritic cells to produce type I IFN; Kadowaki N et al.; Two classes of nucleic acids, bacterial DNA containing unmethylated CpG motifs and dsRNA in viruses, induce the production of type I IFN that contributes to the immunostimulatory effects of these microbial molecules . Thus, it is important to determine which cells produce type I IFN in response to CpG DNA and dsRNA . CD4(+)CD11c(-) type 2 dendritic cell precursors (pre-DC2) were identified as the main producers of type I IFN in human blood in response to viruses . Here we asked whether pre-DC2 also produce type I IFN in response to CpG DNA and dsRNA . Oligodeoxynucleotides containing particular palindromic CpG motifs induced pre-DC2, but not CD11c(+) blood DC or monocytes, to produce IFN-alpha . In contrast, a synthetic dsRNA, polyinosinic polycytidylic-acid, induced CD11c(+) DC, but not pre-DC2 or monocytes, to produce IFN-alphabeta . These data indicate that CpG DNA and polyinosinic-polycytidylic acid stimulate different types of cells to produce type I IFN and that it is important to select oligodeoxynucleotides containing particular CpG motifs to induce pre-DC2 to produce type I IFN, which may play a key role in the strong adjuvant effects of CpG DNA.

Microbiology, 2001 Feb, 147(Pt 2), 373 - 82
Characterization of microbial consortia in a terephthalate-degrading anaerobic granular sludge system; Wu JH et al.; The microbial composition and spatial distribution in a terephthalate-degrading anaerobic granular sludge system were characterized using molecular techniques . 16S rDNA clone library and sequence analysis revealed that 78.5% of 106 bacterial clones belonged to the delta subclass of the class Proteobacteria; the remaining clones were assigned to the green non-sulfur bacteria (7.5%), Synergistes (0.9%) and unidentified divisions (13.1%) . Most of the bacterial clones in the delta-Proteobacteria formed a novel group containing no known bacterial isolates . For the domain Archaea, 81.7% and 18.3% of 72 archaeal clones were affiliated with Methanosaeta and Methanospirillum, respectively . Spatial localization of microbial populations inside granules was determined by transmission electron microscopy and fluorescent in situ hybridization with oligonucleotide probes targeting the novel delta-proteobacterial group, the acetoclastic Methanosaeta, and the hydrogenotrophic Methanospirillum and members of Methanobacteriaceae . The novel group included at least two different populations with identical rod-shape morphology, which made up more than 87% of the total bacterial cells, and were closely associated with methanogenic populations to form a nonlayered granular structure . This novel group was presumed to be the primary bacterial population involved in the terephthalate degradation in the methanogenic granular consortium.

J Antimicrob Chemother, 2001 Feb, 47(2), 199 - 202
Antimycobacterial action of B4128, a novel tetramethylpiperidyl-substituted phenazine; Matlola NM et al.; The effects of the novel tetramethylpiperidyl (TMP)-substituted phenazine, B4128 (0.6-2.5 mg/L), on growth, phospholipase A2(PLA2) activity, cation (K+, Ca2+) fluxes and energy metabolism (ATP) of Mycobacterium aurum A(+) and Mycobacterium tuberculosis (H37Ra) have been investigated in vitro . Growth, PLA2 and cation fluxes were measured radiometrically, while microbial ATP was assayed by means of a luciferin/luciferase chemiluminescence method . Exposure of the mycobacteria to B4128 resulted in immediate, dose-related enhancement of microbial PLA2 activity and inhibition of K+ influx, which preceded effects on microbial ATP, influx of Ca2+ and growth . These results demonstrate that B4128 possesses membrane-directed antimycobacterial properties that are similar to those of clofazimine.

Appl Environ Microbiol, 2001 Feb, 67(2), 702 - 12
Impact of dilution on microbial community structure and functional potential: comparison of numerical simulations and batch culture experiments; Franklin RB et al.; A series of microcosm experiments was performed using serial dilutions of a sewage microbial community to inoculate a set of batch cultures in sterile sewage . After inoculation, the dilution-defined communities were allowed to regrow for several days and a number of community attributes were measured in the regrown assemblages . Based upon a set of numerical simulations, community structure was expected to differ along the dilution gradient; the greatest differences in structure were anticipated between the undiluted-low-dilution communities and the communities regrown from the very dilute (more than 10(-4)) inocula . Furthermore, some differences were expected among the lower-dilution treatments (e.g., between undiluted and 10(-1)) depending upon the evenness of the original community . In general, each of the procedures used to examine the experimental community structures separated the communities into at least two, often three, distinct groups . The groupings were consistent with the simulated dilution of a mixture of organisms with a very uneven distribution . Significant differences in community structure were detected with genetic (amplified fragment length polymorphism and terminal restriction fragment length polymorphism), physiological (community level physiological profiling), and culture-based (colony morphology on R2A agar) measurements . Along with differences in community structure, differences in community size (acridine orange direct counting), composition (ratio of sewage medium counts to R2A counts, monitoring of each colony morphology across the treatments), and metabolic redundancy (i.e., generalist versus specialist) were also observed, suggesting that the differences in structure and diversity of communities maintained in the same environment can be manifested as differences in community organization and function.

Appl Environ Microbiol, 2001 Feb, 67(2), 696 - 701
Identification, purification, and characterization of iminodiacetate oxidase from the EDTA-degrading bacterium BNC1; Liu Y et al.; Microbial degradation of synthetic chelating agents, such as EDTA and nitrilotriacetate (NTA), may help immobilizing radionuclides and heavy metals in the environment . The EDTA- and NTA-degrading bacterium BNC1 uses EDTA monooxygenase to oxidize NTA to iminodiacetate (IDA) and EDTA to ethylenediaminediacetate (EDDA) . IDA- and EDDA-degrading enzymes have not been purified and characterized to date . In this report, an IDA oxidase was purified to apparent homogeneity from strain BNC1 by using a combination of eight purification steps . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single protein band of 40 kDa, and by using size exclusion chromatography, we estimated the native enzyme to be a homodimer . Flavin adenine dinucleotide was determined as its prosthetic group . The purified enzyme oxidized IDA to glycine and glyoxylate with the consumption of O2 . The temperature and pH optima for IDA oxidation were 35 degrees C and 8, respectively . The apparent Km for IDA was 4.0 mM with a kcat of 5.3 s(-1) . When the N-terminal amino acid sequence was determined, it matched exactly with that encoded by a previously sequenced hypothetical oxidase gene of BNC1 . The gene was expressed in Escherichia coli, and the gene product as a C-terminal fusion with a His tag was purified by a one-step nickel affinity chromatography . The purified fusion protein had essentially the same enzymatic activity and properties as the native IDA oxidase . IDA oxidase also oxidized EDDA to ethylenediamine and glyoxylate . Thus, IDA oxidase is likely the second enzyme in both NTA and EDTA degradation pathways in strain BNC1.

Bioresour Technol, 2001 Jan, 76(2), 125 - 9
Organic matter components and aggregate stability after the application of different amendments to a horticultural soil; Albiach R et al.; The effects of usual or recommended rates of application of five organic amendments (24 t/ha yr of MSW compost, sewage sludge, and ovine manure, 2.4 t/ha yr of commercial vermicompost, and 100 l/ha yr of a commercial humic acids solution) on the soil contents of organic matter, total humified substances, humic acids, carbohydrates and microbial gums, and the structural stability of aggregates were investigated . Four and five years after the beginning of the experiment, significant increments in most of the parameters studied were found after the application of organic residues, whereas the two commercial amendments did not produce any significant change, suggesting that rates recommended by the producers and imposed by their high prices are too low to be useful . MSW compost yielded the highest increases, even if the amount of organic matter applied as ovine manure was very similar . Organic matter and carbohydrates appeared to be the parameters most closely related to soil aggregate stability.

PDA J Pharm Sci Technol, 1999 Sep-Oct, 53(5), 235 - 9
Pharmaceutical container/closure integrity . V: An evaluation of the WILCO "LFC" method for leak testing pharmaceutical glass-stoppered vials; Kirsch LE et al.; The sensitivity, reliability, reproducibility and ease of use of the WILCO LFC package integrity test method was evaluated by preparing and testing a series of rubber-stoppered glass vials which were modified by affixing a glass micropipette through the vial side wall . The test units contained water, 50% aqueous ethanol, 20% lithium chloride or 20% aqueous glycerol . Leakage measurement obtained by LFC testing were compared to helium leak rate measurements . The LFC methods detected all leak > 0.0014 standard cubic centimeters per second (sccs), which represents a sensitivity about fourteen-fold greater than standard vacuum decay methods . The minimum detectable leak corresponded to a nominal micropipette internal diameter of between 1 and 2 microns . The effective detection range corresponded to a leak size associated with a 40 to 100% probability of microbial ingress based on a previously reported logistical regression model between helium leak rate and microbial immersion . The sensitivity did not vary with solvent or testing duration in range of 5 to 10 seconds . The coefficient of variation was about 3% . The LFC operation was rapid and without apparent mechanical or electronic problems over the two day testing period used in these studies.

Aquat Toxicol, 2000 Mar 1, 48(2-3), 135 - 150
Responses of aquatic communities to 25-6 alcohol ethoxylate in model stream ecosystems; Belanger SE et al.; A model stream ecosystem evaluation of the non-commercial alcohol ethoxylate 25-6 alcohol ethoxylate (AE) was performed in 1994 . Algal, heterotrophic microbial, protozoan, and invertebrate communities were assessed over an 8-week exposure period that followed an 8-week colonization period . Streams were exposed to nominal concentrations of 0, 12, 37, 111, 333 and 1000 microg AE/l . Confirmed concentrations (8-week means) were 7 (at the detection limit), 13, 36, 76, 259, and 760 microg AE/l as measured at the head of streams on a weekly basis . Microbial communities were initially enhanced by AE exposure (first 2 weeks of exposure), but by the conclusion of the study autotrophic and heterotrophic microbial communities were similar across treatments . In contrast, invertebrate populations and communities responded strongly to AE exposure with adverse effects indicated at 259-760 microg/l by 4 weeks and at 36-760 microg/l by 8 weeks . Key affected groups were the sensitive mayfly, stonefly, and caddisfly fauna . Species richness and abundance of selected populations such as Stenonema (mayfly), Chimarra (caddisfly), and Corbicula (Asiatic clam) were affected . These data indicated a model ecosystem no-observed effect concentration of 13 microg/l for 25-6 AE . In contrast to other published AE model ecosystem studies, 25-6 appears more toxic . Structure-activity-relationships at the model ecosystem level still demonstrate good relationships across a range of surfactants with calculated K(ow)s of 3-6 . Importantly, the collective information on fate and effects measured in laboratory and field systems confirms low risk to the environment.

J Agric Food Chem, 2000 Dec, 48(12), 6230 - 3
Substrate specificities of microbial transglutaminase for primary amines; Ohtsuka T et al.; Transglutaminase (epsilon-glutaminyl-peptide:amine gamma-glutaminyl-transferase, EC 2.3.2.13) (TGase) is an enzyme that catalyzes acyl transfer reactions between primary amines and Gln residues in proteins and peptides . The substrate specificity of TGase for primary amines was investigated to incorporate various functional groups into proteins and peptides . In this study, microbial transglutaminase and guinea pig liver transglutaminase were used . For the primary amines to be incorporated into benzyloxycarbonyl-L-Gln-Gly (Z-Gln-Gly), they were required to have more than four carbon chains without side chains between the functional groups . These results suggest that with appropriate primary amines as spacers, various functional groups, carboxyl groups, phosphate groups, saccharides, and so on, can be incorporated into proteins by using TGase.

Ann Rheum Dis, 2001 Feb, 60(2), 166 - 9
Effect of interleukin 2 on killer cell inhibitory receptors in patients with rheumatoid arthritis; Kogure T et al.; OBJECTIVE: The genes for killer cell inhibitory receptors (KIRs) have been cloned and their functions and responses to other molecules, including cytokines, have been partially clarified . However, the expression of KIRs has not been analysed in patients with autoimmune diseases, such as rheumatoid arthritis (RA), who are highly susceptible to microbial infection . Therefore, KIR expression on lymphocytes in patients with RA, and the regulation of KIR expression by interleukin 2 (IL2) in RA was investigated . METHODS: CD158a/b expression on peripheral blood mononuclear cells (PBMC) obtained from 25 patients with RA and 14 healthy subjects was analysed by flow cytometry . Additionally, PBMC from the two groups of subjects were cultured in RPMI 1640 medium with or without IL2 for 48 hours, and then their CD158a/b expression was analysed . RESULTS: The rate of CD158a expression on the CD8+ cells was lower in patients with RA than in healthy subjects, though there was no significant difference in the CD158a/b expression on the CD16+ cells between the two groups . The upregulation of CD16+CD158a/b+ cells in response to IL2 was significantly reduced in patients with RA compared with healthy subjects . CONCLUSION: The reduced induction of KIR expression in response to IL2 may provide insight into the reason for the high susceptibility of patients with RA to microbial infection.

Int J Syst Evol Microbiol, 2000 Nov, 50 Pt 6, 2043 - 55
New genus-specific primers for the PCR identification of members of the genus Saccharomonospora and evaluation of the microbial diversity of wild-type isolates of Saccharomonospora detected from soil DNAs; Salazar O et al.; Members of the genus Saccharomonospora are isolated infrequently, probably due to the low occurrence of these actinomycetes in the environment . Although members of this genus can easily be identified by micromorphological criteria, the extensive chemotaxonomic characterization of each new isolate is a time-consuming task which cannot always be undertaken when handling large numbers of strains as is the case in natural products screening programmes . In this work, the design of one set of genus-specific oligonucleotides which allows rapid detection of members of the genus Saccharomonospora by means of PCR-specific amplification is presented . The genus specificity of these primers was validated on a wide range of collection and wild-type strains, and subsequently applied to evaluate the presence of representatives of this taxon directly from soil DNAs . Partial 16S rDNA sequencing of representative wild-type strains was used to validate their genus assignment . Further analyses of PCR fingerprinting patterns and 16S-23S internal transcribed spacer sequences were used to determine the diversity of wild-type isolates obtained from soils . This study shows the usefulness of the application of these primers for the direct identification of members of this genus and in assessment of its occurrence within natural microbial habitats.

Dermatol Clin, 2001 Jan, 19(1), 79 - 86, viii
Dapsone and sulfapyridine; Paniker U et al.; Dapsone and sulfapyridine are structurally related compounds with anti-microbial and anti-inflammatory effects . Dapsone remains the most important drug for leprosy and is useful in the prophylaxis of Pneumocystis pneumonia in patients with HIV disease . The medical treatment of choice for dermatitis herpetiformis is dapsone; and sulfapyridine also can be used for those patients who are intolerant of dapsone . Other neutrophilic disorders also may respond to these drugs . Toxic side effects of both dapsone and sulfapyridine are mediated through the hydroxylamine metabolite . These include hemolysis, methemoglobinemia, and agranulocytosis . Careful monitoring for possible adverse reactions includes frequently performing complete blood counts and regular blood chemistry profile determinations.

Curr Opin Immunol, 2001 Feb, 13(1), 109 - 13
CD1 and lipid antigens: intracellular pathways for antigen presentation; Jayawardena-Wolf J et al.; Recently, different members of the CD1 family of MHC-like molecules have been shown to sample different intracellular compartments to present lipid and glycolipid antigens to T cells . Emerging models suggest that CD1 may have evolved to monitor the integrity of membrane lipids and/or to present microbial lipid antigens to both alpha beta and gamma delta T cells.

Curr Opin Immunol, 2001 Feb, 13(1), 55 - 62
Innate immunity in plants; Cohn J et al.; Studies of receptors and signal-transduction components that play a role in plant disease resistance have revealed remarkable similarities with innate immunity pathways in insects and mammals . In plants, specific receptors encoded by disease-resistance genes interact with products of microbial effector genes to activate defence responses . Resistance proteins have been found to have motifs in common with components of immune response pathways in mammals and invertebrates, and to rely on similar downstream signalling components . In the future, the sharing of ideas among plant and animal biologists is likely to broaden our understanding of defence responses in diverse organisms.

Curr Opin Immunol, 2001 Feb, 13(1), 19 - 25
Presentation of self and microbial lipids by CD1 molecules; Matsuda JL et al.; CD1 molecules present both self lipids and microbial lipids . Recent studies have elucidated novel antigenic structures that can be presented by CD1 for T cell stimulation, as well as new pathways for lipid-antigen presentation . Additionally, the development of lipid-CD1 tetramers now permits the tracking of CD1-reactive T cells during immune responses . Despite this, the roles of CD1-reactive T cells in both host defense and immune regulation remain to be unequivocally defined.

Clin Exp Pharmacol Physiol, 2001 Jan-Feb, 28(1-2), 1 - 8
Sunlight, immunosuppression and skin cancer: role of histamine and mast cells; Hart PH et al.; 1 . The development into tumours of skin cells transformed by ultraviolet (UV) B radiation of wavelengths 290-320 nm is enhanced by the ability of UVB to suppress an immune response that would otherwise destroy them . Ultraviolet B-induced immunomodulation may be by multiple mechanisms, but generally manifests in an antigen-presenting cell defect and an altered cytokine environment in the draining lymph nodes . 2 . Immune responses to microbial or self-antigens may be dysfunctional by similar mechanisms following UVB exposure . 3 . Earliest-acting intermediates in the initiation of UVB-induced immunosuppression are the UVB absorbers (photoreceptors) of the skin, notably DNA resulting in immunoregulatory cytokine production, and trans-urocanic acid (UCA), which, upon isomerization to its cis isomer, signals downstream immunosuppressive events . 4 . In mice, dermal mast cells are critical to UVB-induced systemic immunomodulation . In mice, there is a functional link as well as a linear relationship between the prevalence of histamine-staining dermal mast cells and the log of the dose of UVB required for 50% immunosuppression . Studies with histamine receptor antagonists support histamine as the main' product of mast cells involved . Histamine acts in large part via a prostanoid-dependent pathway . 5 . Approximately 50% of humans and greater than 90% of patients with non-melanoma skin cancer are UVB susceptible for suppression of a contact hypersensitivity response . Neither cytokine polymorphisms nor UVB-induced levels of cis-UCA in irradiated skin have been linked to UVB susceptibility . Patients with basal cell carcinomas (BCC) have an increased dermal mast cell prevalence in non-sun-exposed buttock skin . We propose that mast cells function in humans, as in mice, by initiating immunosuppression and, thereby, allowing a permissive environment for BCC development.

Bratisl Lek Listy, 2000, 101(8), 445 - 9
{Proteinases and antiproteinases: biomedical correlations}; Kotyza J; Both degradative and limited proteolysis are involved in physiological processes . Once activated, proteinases are controlled by endogenous inhibitors (antiproteinases) . A variety of genetic disorders and exogenous microbial proteinases disturb the balance between proteinases and cognate inhibitors . alpha 1-Antitrypsin deficiency is a model disorder resulting in an insufficient control of leukocyte elastase with a subsequent alveolar tissue damage . It appears that the manifold molar excess of inhibitor required to compensate the deficiency, is the consequence of both the local relative inaccessibility of the serpin, and the mode of proteinase-serpin interaction, in accord with the "branched pathway" mechanism . Slow-binding kinetics and the leak of native proteinase from the complex is illustrated by the peptidolytic action of porcine elastase in the presence of human alpha 1-antitrypsin and by gelatin zymography, respectively . (Tab . 2, Fig . 5, Ref . 44.)

Ann Agric Environ Med, 2000, 7(2), 111 - 8
Effects of exposure to flax dust in Polish farmers: work-related symptoms and immunologic response to microbial antigens associated with dust; Skorska C et al.; Medical examinations were performed in a group of 51 Polish farmers heavily exposed to flax dust during harvesting and scutching (threshing) and in a group of 50 healthy urban dwellers not exposed to organic dusts (controls) . The examinations included: interview concerning the occurrence of respiratory disorders and work-related symptoms, physical examination, X-ray examination of chest, lung function tests, oxymetric examinations, determination of the concentration of cytokines (IL-1alpha IL-6, TNFalpha) in blood serum and allergological tests with microbial antigens associated with organic dust, comprising: skin prick test with 4 antigens, agar-gel precipitation test with 12 antigens and test for specific inhibition of leukocyte migration with 4 antigens . As many as 32 farmers (62.7%) reported the occurrence of work-related symptoms during harvesting, transporting and scutching of flax . The most common complaint was general weakness reported by 15 farmers (29.4%), followed by headache reported by 14 (27.5%), blocking of the nose - by 11 (21.6%), dry cough, shivering, and eyes itching - each by 10 (19.8%), chest tightness and hoarseness - each by 9 (17.6%) . No control subjects reported these work-related symptoms . The mean spirometric values in the examined group of farmers were within a normal range and did not show a significant post-shift decline . In contrast, a significant post-shift decline of oxymetric values was found among flax farmers . The farmers showed a frequency of the positive early skin reactions to environmental allergens in the range of 0-19.6%, a frequency of positive precipitin reactions in a range of 0-56.9%, and frequency of positive reactions of leukocyte migration inhibition in a range of 7.8-21.6% . The members of the control group responded to the majority of allergens with a significantly lower frequency of positive results compared to the farmers . Elevated concentrations of IL-1alpha and IL-6, but not TNFalpha, were found in blood sera of flax farmers . In conclusion, farmers engaged in harvesting and scutching of flax represent a group of elevated professional risk because of high incidence of work-related symptoms and high frequency of allergic reactions to bacteria and fungi associated with organic dust.

Inorg Chem, 2000 Dec 11, 39(25), 5591 - 602
Kinetics and mechanism of iron(III) dissociation from the dihydroxamate siderophores alcaligin and rhodotorulic acid; Boukhalfa H et al.; The kinetics and mechanism of siderophore ligand dissociation from their fully chelated Fe(III) complexes is described for the highly preorganized cyclic tetradentate alcaligin and random linear tetradentate rhodotorulic acid in aqueous solution at 25 degrees C (Fe2L3 + 6H+ reversible 2 Fe3+ aq + 3 H2L) . At siderophore:Fe(III) ratios where Fe(III) is hexacoordinated, kinetic data for the H(+)-driven ligand dissociation from the Fe2L3 species is consistent with a singly ligand bridged structure for both the alcaligin and rhodotorulic acid complexes . Proton-driven ligand dissociation is found to proceed via parallel reaction paths for rhodotorulic acid, in contrast with the single path previously observed for the linear trihydroxamate siderophore ferrioxamine B . Parallel paths are also available for ligand dissociation from Fe2(alcaligin)3, although the efficiency of one path is greatly diminished and dissociation of the bis coordinated complex Fe(alcaligin)(OH2)2+ is extremely slow (k = 10(-5) M-1 s-1) due to the high degree of preorganization in the alcaligin siderophore . Mechanistic interpretations were further confirmed by investigating the kinetics of ligand dissociation from the ternary complexes Fe(alcaligin)(L) in aqueous acid where L = N-methylacetohydroxamic acid and glycine hydroxamic acid . The existence of multiple ligand dissociation paths is discussed in the context of siderophore mediated microbial iron transport.

J Allergy Clin Immunol, 2001 Jan, 107(1), 160 - 4
Antibody responses to bacteriophage phi X-174 in human subjects exposed to the antarctic winter-over model of spaceflight; Shearer WT et al.; BACKGROUND: It has been proposed that exposure to long-term spaceflight conditions (stress, isolation, sleep disruption, containment, microbial contamination, and solar radiation) or to ground-based models of spaceflight will alter human immune responses, but specific antibody responses have not been fully evaluated . OBJECTIVE: We sought to determine whether exposure to the 8-month Antarctic winter-over model of spaceflight would alter human antibody responses . METHODS: During the 1999 Australian National Antarctic Research Expeditions, 11 adult study subjects at Casey, Antarctica, and 7 control subjects at Macquarie Island, sub-Antarctica, received primary and secondary immunizations with the T cell-dependent neoantigen bacteriophage phi X-174 . Periodic plasma samples were analyzed for specific antibody function . RESULTS: All of the subjects from Casey, Antarctica, cleared bacteriophage phi X-174 normally by 1 week after primary immunization, and all had normal primary and secondary antibody responses, including immunologic memory amplification and switch from IgM to IgG antibody production . One subject showed a high normal pattern, and one subject had a low normal pattern . The control subjects from Macquarie Island also had normal immune responses to bacteriophage phi X-174 . CONCLUSIONS: These data do not support the hypothesis that de novo specific antibody responses of subjects become defective during the conditions of the Antarctic winter-over . Because the Antarctic winter-over model of spaceflight lacks the important factors of microgravity and solar radiation, caution must be used in interpreting these data to anticipate normal antibody responses in long-term spaceflight.

J Cell Biol, 2001 Jan 8, 152(1), 165 - 80
The phagosome proteome: insight into phagosome functions; Garin J et al.; Phagosomes are key organelles for the innate ability of macrophages to participate in tissue remodeling, clear apoptotic cells, and restrict the spread of intracellular pathogens . To understand the functions of phagosomes, we initiated the systematic identification of their proteins . Using a proteomic approach, we identified >140 proteins associated with latex bead-containing phagosomes . Among these were hydrolases, proton pump ATPase subunits, and proteins of the fusion machinery, validating our approach . A series of unexpected proteins not previously described along the endocytic/phagocytic pathways were also identified, including the apoptotic proteins galectin3, Alix, and TRAIL, the anti-apoptotic protein 14-3-3, the lipid raft-enriched flotillin-1, the anti-microbial molecule lactadherin, and the small GTPase rab14 . In addition, 24 spots from which the peptide masses could not be matched to entries in any database potentially represent new phagosomal proteins . The elaboration of a two-dimensional gel database of >160 identified spots allowed us to analyze how phagosome composition is modulated during phagolysosome biogenesis . Remarkably, during this process, hydrolases are not delivered in bulk to phagosomes, but are instead acquired sequentially . The systematic characterization of phagosome proteins provided new insights into phagosome functions and the protein or groups of proteins involved in and regulating these functions.

Diagn Microbiol Infect Dis, 2000 Dec, 38(4), 213 - 21
Discriminative power of fatty acid methyl ester (FAME) analysis using the microbial identification system (MIS) for Candida (Torulopsis) glabrata and Saccharomyces cerevisiae; Peltroche-Llacsahuanga H et al.; Candida (Torulopsis) glabrata is frequently isolated in cases of fungal infection and commonly shows acquired or innate fluconazole resistance . Saccharomyces cerevisiae, an emerging opportunistic yeast pathogen, causes serious systemic infections in immunocompromised, and vaginitis and superficial infections in immunocompetent patients . For both species reliable identification in the routine laboratory is mandatory, but species identification of strains, e.g . trehalose-negative C . glabrata, may be difficult . Therefore, gas-liquid chromatography (GLC) of whole cell fatty acid methyl ester (FAME) profiles, that is independent of assimilation profiles of strains and suitable for reliable and rapid identification of clinically important yeasts, was applied . However, frequent misidentification of C . glabrata as S . cerevisiae has been reported when using the Yeast Clinical Database of MIS . Accuracy of MIS identification may be strongly influenced by the amounts of cell mass analyzed . Therefore, the present study compared the MIS results of these two yeasts achieved with different cell masses . Primarily we optimized, especially with respect to cost-effectiveness, the recommended streaking technique yielding a maximal recovery of 90-130 mg of cell mass from one plate, enabling testing of poor growing strains of C . glabrata . For all C . glabrata strains tested (n = 10) the highest identification scores (SI {Similarity Index} range 0.525-0.963, median 0.832) were achieved with 30 to 45 mg of cell mass . Only 5 of 10 S . cerevisiae strains revealed good library comparisons (SI > or = 0.5) when using 30 mg of cell mass, whereas with 45 mg all strains but two revealed this SI-level . For S . cerevisiae a higher amount of cell mass processed (up to 90 mg) was correlated with better identification scores (SI range using 90 mg: 0.464-0.870, median, 0.737) . Several passages prior to FAME analysis of C . glabrata strains on recommended media revealed narrowing of SI ranges, but differences in SI values were not statistically significant.

Crit Rev Immunol, 2000, 20(5), 393 - 405
Critical roles of Toll-like receptors in host defense; Kaisho T et al.; Drosophila Toll is involved not only in dorsoventral patterning of embryos but also in immune responses to microbial infection . Several Toll-like receptors (TLRs) have also been identified in mammals . They are expressed on macrophages or dendritic cells (DCs), which are essential sentinels for innate immunity . These cells utilize TLRs as a recognition and signal transducing receptor for microbial molecular components . The most characterized mammalian TLR, TLR4, is a receptor for lipopolysaccharides (LPS) . TLR2 recognizes other components, such as peptideglycans (PGN) . This recognition, called pattern recognition, is essential for the establishment of innate immunity, which is the basis for host defense . In this article, we review recent findings about this expanding receptor family.

Mil Med, 2000 Nov, 165(11 Suppl), 1 - 21
Aeromedical evacuation of biological warfare casualties: a treatise on infectious diseases on aircraft; Withers MR et al.; A basic understanding of the transmission and isolation of infections would be essential to the safe and effective aeromedical evacuation (AE) of biological warfare (BW) casualties . First, the airframe as microbial environment is considered, and relevant preventive and disinfecting measures are discussed . A survey of past infectious disease transmission on civilian aircraft (including tuberculosis, influenza, measles, smallpox, and viral hemorrhagic fevers) is presented, and the communicability and stability of likely BW agents is described . A brief history of U.S . military aeromedical evacuation (as it relates to contagious diseases and U.S . Air Force BW doctrine) is also outlined . Special containment procedures (especially as used by the U.S . Army Aeromedical Isolation Team) are described . Finally, international legal and regulatory aspects of the AE of BW casualties are considered, and some unanswered questions and suggestions for future research are offered . It is concluded that, given adequate foresight, expertise, and resources, the AE of even contagious BW casualties could be safely and effectively accomplished.

Biochemistry, 2001 Jan 9, 40(1), 84 - 92
Characterization of BphF, a Rieske-type ferredoxin with a low reduction potential; Couture MM et al.; BphF is a small, soluble, Rieske-type ferredoxin involved in the microbial degradation of biphenyl . The rapid, anaerobic purification of a heterologously expressed, his-tagged BphF yielded 15 mg of highly homogeneous recombinant protein, rcBphF, per liter of cell culture . The reduction potential of rcBphF, determined using a highly oriented pyrolytic graphite (HOPG) electrode, was -157+/- 2 mV vs the standard hydrogen electrode (SHE) (20 mM MOPS, 80 mM KCl, and 1 mM dithiothreitol, pH 7.0, 22 degrees C) . The electron paramagnetic resonance spectrum of the reduced rcBphF is typical of a Rieske cluster while the close similarity of the circular dichroic (CD) spectra of rcBphF and BedB, a homologous protein from the benzene dioxygenase system, indicates that the environment of the cluster is highly conserved in these two proteins . The reduction potential and CD spectra of rcBphF were relatively independent of pH between 5 and 10, indicating that the pK(a)s of the cluster's histidinyl ligands are not within this range . Gel filtration studies demonstrated that rcBphF readily oligomerizes in solution . Crystals of rcBphF were obtained using sodium formate or poly(ethylene glycol) (PEG) as the major precipitant . Analysis of the intermolecular contacts in the crystal revealed a head-to-tail interaction that occludes the cluster, but is very unlikely to be found in solution . Oligomerization of rcBphF in solution was reversed by the addition of dithiothreitol and is unrelated to the noncovalent crystallographic interactions . Moreover, the oligomerization state of rcBphF did not influence the latter's reduction potential . These results indicate that the 450 mV spread in reduction potential of Rieske clusters of dioxygenase-associated ferredoxins and mitochondrial bc(1) complexes is not due to significant differences in their solvent exposure.

Clin Diagn Lab Immunol, 2001 Jan, 8(1), 21 - 30
Defective neutrophil degranulation induced by interleukin-8 and complement 5a and down-regulation of associated receptors in children vertically infected with human immunodeficiency virus type 1; Meddows-Taylor S et al.; The polymorphonuclear neutrophils (PMNs) of patients infected with human immunodeficiency virus type 1 (HIV-1) show impaired microbicidal responses . The present study assessed the functional integrity of PMN degranulation responses and the expression of specific receptors that mediate these responses in a group of children vertically infected with HIV-1 . PMN degranulation in response to interleukin-8 (IL-8) and complement 5a (C5a) was measured in a group of HIV-1-infected children with mild and severe clinical disease and in an uninfected control group . In addition, the expression of CXCR1, CXCR2, and CD88 on whole-blood PMNs was quantified by flow cytometry . Although CXCR1 expression was found to be largely unaltered in the HIV-1-infected children relative to that in the control children, the intensity of CXCR2 expression was significantly reduced in those with severe disease . Furthermore, there was a significant reduction in the percentage of cells expressing CD88 and in the intensity of CD88 fluorescence in the HIV-1-infected children compared to that in control children, with CD88 fluorescence intensity more significantly reduced in the presence of severe disease . PMNs from a large proportion of the HIV-1-infected children either showed reciprocal degranulation responses or were unresponsive to IL-8 and C5a, whereas the PMNs from the uninfected children showed positive responses . Inefficient agonist-induced degranulation may contribute to the increased susceptibility of HIV-1-infected children to secondary microbial infections . Furthermore, reduced expression of CXCR2 and CD88 may be suggestive of defects in other functions of PMNs from HIV-1-infected children.

Extremophiles, 2000 Dec, 4(6), 343 - 9
Diverse genes of cellulase homologues of glycosyl hydrolase family 45 from the symbiotic protists in the hindgut of the termite Reticulitermes speratus; Ohtoko K et al.; Diverse genes encoding cellulase homologues belonging to glycosyl hydrolase family 45 were identified from the symbiotic protists in the hindgut of the termite Reticulitermes speratus through the use of consensus PCR and the screening of a cDNA library . Fifteen full-length cDNA clones were isolated and sequenced, which encoded polypeptides consisting of 218-221 amino acid residues showing up to 63% identity to known family 45 cellulases . The cellulase sequences of the termite symbiotic protists were phylogenetically monophyletic, showing more than 75% amino acid identity with each other . These enzymes consist of a single catalytic domain, lacking the ancillary domains found in most microbial cellulases . By whole-cell in situ hybridization using oligonucleotide probes specific for regions conserved in some of the sequences, the origin of the genes was identified as symbiotic hypermastigote protists . The presence of diverse cellulase homologues suggests that symbiotic protists of termites may be rich reservoirs of novel cellulase sequences.

Int J Food Microbiol, 2000 Dec 5, 62(1-2), 37 - 45
Delimination of brewing yeast strains using different molecular techniques; Tornai-Lehoczki J et al.; In general, the genetic characteristics, the phenotype and the microbial purity of the production brewing yeast strains are among the most important factors in maintaining a consistently good quality of products . Analysis of restriction fragment length polymorphism (RFLP) patterns of 18S rRNA-coding DNA was investigated to group ale and lager strains . All production brewing yeast strains showed the same RFLP pattern as the type strain and synonym type strains of S . cerevisiae, and were quite different from the type and synonym type strains of S . pastorianus . Based on these data, all production brewing yeast strains investigated in this study appeared to belong to S . cerevisiae . Electrophoretic karyotyping and random amplified polymorphic DNA (RAPD) analysis appeared to be suitable methods for distinguishing not only the type and synonym type strain of S . cerevisiae and S . pastorianus, but also the ale and the lager strains.

Hum Immunol, 2000 Nov, 61(11), 1150 - 6
HLA-G and HLA-E polymorphisms in an indigenous African population . The ZVITAMBO Study Group; Matte C et al.; Studies of HLA-G and HLA-E polymorphisms in different populations from many industrialized countries have uniformly reported little sequence variation within these genes . To date, the polymorphism of these genes has not been characterized in populations from developing countries where more sequence variation would be anticipated due to greater exposure to microbial pathogens . In order to address this issue, we have recently investigated the polymorphism of these MHC genes in an indigenous African population (Shona ethnic group) . HLA-G and HLA-E alleles were determined by PCR-restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system (ARMS) analysis in 108 unrelated women recruited from maternity clinics and hospitals in Harare (Zimbabwe) . The genotyping analysis identified six different HLA-G alleles and three HLA-E alleles in this population . Of interest, the null-allele HLA-G*0105N was found with 11.1% of frequency, which is higher than in other populations tested so far . We observed the presence of 15 distinct HLA-G genotypes and 6 HLA-E genotypes . These data indicate that this African population contains low levels of allelic polymorphism similar to ethnic groups from industrialized countries . This is the first report describing HLA-G and HLA-E polymorphisms in an indigenous African population.

J Exp Biol, 2001 Jan, 204(Pt 2), 349 - 58
Quantifying nutrient production by the microbial symbionts in an aphid; Douglas AE et al.; The symbiotic bacteria Buchnera sp . provide aphids with essential amino acids, nutrients in short supply in the aphid diet of plant phloem sap . The contribution of Buchnera-derived amino acids to net protein growth of the aphid Aphis fabae was quantified from the protein growth of aphids reared on chemically defined diets lacking individual amino acids . The amino acid production rates varied among the nine essential amino acids over the range 8-156 pmol microg(-1)protein day(-1) (for tryptophan and leucine, respectively), equivalent to 0.02-0.33 fmol Buchnera(-1)day(-1) . In a complementary metabolic analysis, the aphids incorporated radioactivity from dietary {(14)C}glutamic acid into the essential amino acids isoleucine, lysine and threonine . Incorporation into isoleucine was significantly elevated by the omission of dietary isoleucine, indicating that dietary supply may affect the biosynthetic rates of certain amino acids by Buchnera . Aphids experimentally deprived of Buchnera did not synthesize essential amino acids from dietary glutamic acid . The mortality of aposymbionts was high over 7 days on the phenylalanine-free diet, and their assimilation of dietary leucine was depressed on the complete diet, suggesting that both the absence of bacteria-derived amino acids and the low rates of assimilation of certain dietary amino acids may contribute to the poor growth of these insects.

FEMS Microbiol Ecol, 2001 Jan, 34(3), 255 - 266
Microbial community dynamics in Mediterranean nutrient-enriched seawater mesocosms: changes in abundances, activity and composition; Lebaron P et al.; Quantitative and qualitative changes in bacterial communities from the Mediterranean Sea were compared in duplicate batch mesocosms with or without addition of inorganic nutrients . Methods including traditional microbial ecology techniques, molecular biology and flow cytometry were combined to determine abundances, production, cell size, activity, culturability and taxonomic diversity of bacterial cells . Addition of nutrients and confinement resulted in an increase of bacterial densities which were rapidly controlled by protozoan grazing . Changes in bacterial activity and morphology were observed during the growth phase of bacteria and under grazing pressure . The proportion of medium-size and culturable cells increased during the growth phase . These cells were preferentially consumed by grazers resulting in a strong limitation of bacterial production . As a consequence of the grazing pressure, large cells were produced and contributed to the remaining bacterial productivity after grazing . Grazing had an effect on the taxonomic composition of bacterial communities by preferentially eliminating gamma-Proteobacteria, alpha-Proteobacteria were preserved . It seems that some species from the genera Ruegeria and Cytophaga may have developed defence strategies to escape predation.

Plant J, 2000 Dec, 24(6), 837 - 47
Identification of genes encoding receptor-like protein kinases as possible targets of pathogen- and salicylic acid-induced WRKY DNA-binding proteins in Arabidopsis; Du L et al.; To understand how plant host genes are regulated during the activation of plant defence responses, we are studying a group of pathogen- and salicylic acid (SA)-induced DNA-binding proteins containing the novel WRKY domain . To identify downstream target genes of these WRKY proteins, we have searched the Arabidopsis genome and identified four closely linked genes on chromosome IV that contain an unusually large number of the W-box sequences {(T)TGAC(C/T)} recognized by WRKY proteins within a few hundred base pairs upstream of their coding regions . All four genes encode proteins characteristic of receptor-like protein kinases (RLK), each consisting of an N-terminal signal sequence, an extracellular receptor domain, a single transmembrane domain and a C-terminal cytoplasmic serine/threonine protein kinase domain . All four RLK genes were induced by treatment with SA or infection by a bacterial pathogen . Studies with one of the RLK genes (RLK4) indicated that a cluster of W-box elements in its promoter region were recognized by both purified WRKY proteins and SA-induced W-box binding activities from SA-treated Arabidopsis plants . Further analysis using the RLK4 gene promoter fused to a reporter gene in transgenic Arabidopsis indicated that the consensus WRKY protein-binding sites in the RLK4 gene promoter were important for the inducible expression of the reporter gene . These results indicate that pathogen- and SA-induced W-box binding proteins regulate not only genes encoding defence proteins with direct or indirect anti-microbial activities, but also genes encoding proteins with regulatory functions.

J Clin Invest, 2001 Jan, 107(1), 121 - 30
EBNA1-specific CD4+ T cells in healthy carriers of Epstein-Barr virus are primarily Th1 in function; Bickham K et al.; The Epstein-Barr virus (EBV) nuclear antigen-1 (EBNA1) maintains the viral episome in all host cells infected with EBV . Recently, EBNA1 was found to be the main EBV latency antigen for CD4+ T cells and could be recognized in cultures from all donors tested . We now identify a polarized Th1 phenotype and obtain evidence for its presence in vivo . When T cells were stimulated with dendritic cells infected with vaccinia vectors expressing EBNA1, 18 of 19 donors secreted IFN-gamma, whereas only two of 19 secreted IL-4 . Magnetic selection was then used to isolate cells from fresh blood based on EBNA1-induced cytokine production . Specific IFN-gamma CD4+ cell lines were established from six of six donors and IL-4 lines from three of six . Only the Th1 lines specifically lysed targets expressing three different sources of EBNA1 protein . When the IgG isotype of EBNA1 plasma Ab's was tested, most specific Ab's were IgG1 and of a high titer, confirming a Th1 response to EBNA1 in vivo . Ab's to other microbial antigens generally were not skewed toward IgG1 . Given emerging evidence that Th1 CD4+ T cells have several critical roles in host defense to viral infection and tumors, we propose that EBNA1-specific CD4+ Th1 cells contribute to resistance to EBV and EBV-associated malignancies.

Appl Environ Microbiol, 2001 Jan, 67(1), 100 - 9
Microbial activity in aquatic environments measured by dimethyl sulfoxide reduction and intercomparison with commonly used methods; Griebler C et al.; A new method to determine microbial (bacterial and fungal) activity in various freshwater habitats is described . Based on microbial reduction of dimethyl sulfoxide (DMSO) to dimethyl sulfide (DMS), our DMSO reduction method allows measurement of the respiratory activity in interstitial water, as well as in the water column . DMSO is added to water samples at a concentration (0.75% {vol/vol} or 106 mM) high enough to compete with other naturally occurring electron acceptors, as determined with oxygen and nitrate, without stimulating or inhibiting microbial activity . Addition of NaN(3), KCN, and formaldehyde, as well as autoclaving, inhibited the production of DMS, which proves that the reduction of DMSO is a biotic process . DMSO reduction is readily detectable via the formation of DMS even at low microbial activities . All water samples showed significant DMSO reduction over several hours . Microbially reduced DMSO is recovered in the form of DMS from water samples by a purge and trap system and is quantified by gas chromatography and detection with a flame photometric detector . The DMSO reduction method was compared with other methods commonly used for assessment of microbial activity . DMSO reduction activity correlated well with bacterial production in predator-free batch cultures . Cell-production-specific DMSO reduction rates did not differ significantly in batch cultures with different nutrient regimes but were different in different growth phases . Overall, a cell-production-specific DMSO reduction rate of 1.26 x 10(-17) +/- 0 . 12 x 10(-17) mol of DMS per produced cell (mean +/- standard error; R(2) = 0.78) was calculated . We suggest that the relationship of DMSO reduction rates to thymidine and leucine incorporation is linear (the R(2) values ranged from 0.783 to 0.944), whereas there is an exponential relationship between DMSO reduction rates and glucose uptake, as well as incorporation (the R(2) values ranged from 0.821 to 0.931) . Based on our results, we conclude that the DMSO reduction method is a nonradioactive alternative to other methods commonly used to assess microbial activity.

J Antibiot (Tokyo), 2000 Oct, 53(10), 1117 - 22
Fistupyrone, a novel inhibitor of the infection of Chinese cabbage by Alternaria brassicicola, from Streptomyces sp . TP-A0569; Igarashi Y et al.; A new microbial metabolite, designated fistupyrone, was isolated from the culture broth of a plant-associated Streptomyces sp . TP-A0569 . Fistupyrone inhibited the in vivo infection of the seedlings of Chinese cabbage by Alternaria brassicicola TP-F0423, the cause of Alternaria leaf spot, without any in vitro fungicidal activity.

Crit Rev Oral Biol Med, 2000, 11(4), 383 - 408
Epithelial structural proteins of the skin and oral cavity: function in health and disease; Presland RB et al.; Epithelial tissues function to protect the organism from physical, chemical, and microbial damage and are essential for survival . To perform this role, epithelial keratinocytes undergo a well-defined differentiation program that results in the expression of structural proteins which maintain the integrity of epithelial tissues and function as a protective barrier . This review focuses on structural proteins of the epidermis and oral mucosa . Keratin proteins comprise the predominant cytoskeletal component of these epithelia . Keratin filaments are attached to the plasma membrane via desmosomes, and together these structural components form a three-dimensional array within the cytoplasm of epithelial cells and tissues . Desmosomes contain two types of transmembrane proteins, the desmogleins and desmocollins, that are members of the cadherin family . The desmosomal cadherins are linked to the keratin cytoskeleton via several cytoplasmic plaque proteins, including desmoplakin and plakoglobin (gamma-catenin) . Epidermal and oral keratinocytes express additional differentiation markers, including filaggrin and trichohyalin, that associate with the keratin cytoskeleton during terminal differentiation, and proteins such as loricrin, small proline-rich proteins, and involucrin, that are cross-linked into the cornified envelope by transglutaminase enzymes . The importance of these cellular structures is highlighted by the large numbers of genetic and acquired (autoimmune) human disorders that involve mutations in, or autoantibodies to, keratins and desmosomal and cornified envelope proteins . While much progress has been made in the identification of the structural proteins and enzymes involved in epithelial differentiation, regulation of this process is less clear . Both calcium and retinoids influence epithelial differentiation by altering the transcription of target genes and by regulating activity of enzymes critical in epithelial differentiation, such as transglutaminases, proteinases, and protein kinases . These studies have furthered our understanding of how epithelial tissue and cell integrity is maintained and provide a basis for the future treatment of skin and oral disorders by gene therapy and other novel therapeutics.

J Hepatol, 2000 Dec, 33(6), 926 - 32
Covalent adducts of proteins with acetaldehyde in the liver as a result of acetaldehyde administration in drinking water; Jokelainen K et al.; BACKGROUND/AIMS: Acetaldehyde, the first metabolic product of ethanol, has been suggested to be responsible for several adverse effects of ethanol through its ability to form covalent adducts with proteins and cellular constituents . It has recently been suggested that acetaldehyde derived from microbial ethanol oxidation in the gut could also contribute to the effects of ethanol in the liver . The present work aimed to examine whether modification of proteins by acetaldehyde occurs in rat liver as a result of acetaldehyde administration in drinking water . METHODS: Rats were fed with either 0.7% acetaldehyde (n=10) or water (n=10) for 11 weeks . At the end of the feeding period, liver specimens were processed for immunohistochemistry for protein adducts with acetaldehyde and for hepatic cell type-specific protein markers . RESULTS: Mild fatty change was found in the liver of the acetaldehyde-treated animals but not in the control animals . Immunohistochemical stainings for acetaldehyde adducts revealed intensive positive staining for acetaldehyde adducts in eight (80%) of the animals fed with acetaldehyde . The adducts were predominantly perivenular, although positive staining also occurred along the sinusoids and in the periportal area . Double immunofluorescence staining experiments revealed that hepatocytes were the primary targets of acetaldehyde adduct deposition, although stellate cells and Kupffer cells also showed weak positive reactions . CONCLUSIONS: The present data indicate that acetaldehyde-protein adducts are formed in the liver of animals following acetaldehyde administration in drinking water, which may contribute to the hepatotoxicity of extrahepatic acetaldehyde . These findings should be implicated in studies on the extrahepatic pathways of ethanol oxidation.

Pediatr Clin North Am, 2000 Dec, 47(6), 1339 - 54
Complement deficiencies; Frank MM; The complement proteins play an important role in innate immunity, promoting inflammation and microbial killing . They play a role in the adaptive immune response, as well . Inherited total deficiencies of complement proteins are extremely rare . Table 1 lists more than 40 proteins that comprise the elements of the complement system . Deficiency of the proteins that promote lysis and opsonization is so rare that two papers are able to list all the observed cases to 1991 . The exception is mannan-binding lectin, in which deficiency may be commoner . Diseases of regulatory proteins, such as occurs in hereditary angioedema or paroxysmal nocturnal hemoglobinuria, are commoner but still are quite rare . As we learn more about complement proteins and their mechanism of action, we will understand more clearly how these proteins function . Polymorphisms of the proteins exists . Learning how these polymorphisms contribute to the development of disease will be the focus of complement studies in the next decade.

Phytochemistry, 2000 Nov, 55(6), 481 - 504
Advances in flavonoid research since 1992; Harborne JB et al.; Some of the recent advances in flavonoid research are reviewed . The role of anthocyanins and flavones in providing stable blue flower colours in the angiosperms is outlined . The contribution of leaf flavonoids to UV-B protection in plants is critically discussed . Advances in understanding the part played by flavonoids in warding off microbial infection and protecting plants from herbivory are described . The biological properties of flavonoids are considered in an evaluation of the medicinal and nutritional values of these compounds.

Int Rev Immunol, 2000, 19(6), 557 - 72
Pathogenic thyroglobulin peptides as model antigens: insights on the induction and maintenance of autoimmune thyroiditis; Carayanniotis G et al.; In recent years, the discovery of pathogenic thyroglobulin (Tg) peptides has given a new impetus to study, at the basic level, mechanisms of induction and immunoregulation of autoimmune thyroiditis . The genetic control of the immune response against defined Tg epitopes and the diversity of the T-cell receptor repertoire recruited for their recognition were among the first issues examined . Some of these epitopes contained hormonogenic sites, i.e . thyroxine residues, and thus offered an excellent opportunity to study how post-translational modifications such as iodination, can influence induction of thyroiditogenic cells . The delineation of pathogenic Tg determinants also enabled the search for "molecular mimics" i.e . peptides of microbial origin that may be involved in the pathogenesis of the disease . In addition, factors promoting the generation of pathogenic epitopes during Tg processing in antigen presenting cells could now be systematically investigated . This review summarizes recent findings in these areas.

Pharmazie, 2000 Nov, 55(11), 791 - 7
Vermiculine: a diolide with immunoregulatory activity; Proksa B et al.; Vermiculine (1), is a macrodiolide which displays a selective effect on certain reactions of cellular immunity . This paper summarizes data on its biological activity, isolation, microbial production, chemical properties and methods for its synthesis.

Curr Top Microbiol Immunol, 2000, 252, 189 - 200
Neo-self antigens and the expansion of B-1 cells: lessons from atherosclerosis-prone mice; Silverman GJ et al.; The pathogenesis of atherosclerosis involves an inflammatory process that is modulated by the immune system, and within these complex responses we have discerned a possible role for an archetypic B-1 clone . We speculate that due to their immunogenicity and in vivo distribution the "neo"-self determinants created in oxidatively modified LDL are highly stimulatory for certain B-1 cell clones . These neo-self determinants, which can be created chemically, by somatic processes, may in fact represent the molecular analogues of somatic maturation, or even aging . These changes, including those on non-protein antigens induced by oxidative metabolism, amongst others, create neo-determinants against which the host no doubt can not develop rigorous B-cell tolerance . The onset of expression of these oxidative neo-determinants relatively late in development may well serve a useful function for the highly evolved mammalian immune system, as targeting by evolutionarily selected B-1 clones may facilitate the amplification of other useful antibody-mediated physiologic functions . As in the case of the T15 clone, these antibodies may aid in protection against common microbial pathogens . Hence we postulate that during the evolution of the adaptive immune system the neo-self antigenic milieu may have been exploited for the natural selection of primordial clonal specificities . The T15 B-1 clone may then illustrate a common paradigm in which there has been natural selection based on utility for the defense of the individual from environmental threats, as well as for possible "housekeeping" role(s) and the maintenance of cellular homeostasis.

Plant J, 2000 Dec, 24(5), 569 - 82
Oxidative burst and cognate redox signalling reported by luciferase imaging: identification of a signal network that functions independently of ethylene, SA and Me-JA but is dependent on MAPKK activity; Grant JJ et al.; Recognition of avirulent microbial pathogens activates an oxidative burst leading to the accumulation of reactive oxygen intermediates (ROIs), which are thought to integrate a diverse set of defence mechanisms resulting in the establishment of plant disease resistance . A novel transgenic Arabidopsis line containing a gst1:luc transgene was developed and employed to report the temporal and spatial dynamics of ROI accumulation and cognate redox signalling in response to attempted infection by avirulent strains of Pseudomonas syringae pv . tomato (Pst) . Strong engagement of the oxidative burst was dependent on the presence of functional Pst hrpS and hrpA gene products . Experiments employing pharmacological agents suggested that at least two distinct sources, including an NADPH oxidase and a peroxidase-type enzyme, contributed to the generation of redox cues . The analysis of gst1 and pal1 gene expression in nahG, coi1 and etr1 plants suggested that engagement of the oxidative burst and cognate redox signalling functioned independently of salicylic acid, methyl jasmonate and ethylene . In contrast, studies using a panel of protein kinase and phosphatase inhibitors and in-gel kinase assays in these mutant backgrounds suggested that a 48 kDa mitogen-activated protein kinase (MAPK) activity was required for the activation of gst1 and pal1 in response to redox cues . Thus the engagement of a bifurcating redox signalling pathway possessing a MAPK module may contribute both to the establishment of plant disease resistance, and to the development of cellular protectant mechanisms.

J Immunol, 2001 Jan 1, 166(1), 633 - 41
Production of IL-12 by human monocyte-derived dendritic cells is optimal when the stimulus is given at the onset of maturation, and is further enhanced by IL-4; Ebner S et al.; Dendritic cells produce IL-12 both in response to microbial stimuli and to T cells, and can thus skew T cell reactivity toward a Th1 pattern . We investigated the capacity of dendritic cells to elaborate IL-12 with special regard to their state of maturation, different maturation stimuli, and its regulation by Th1/Th2-influencing cytokines . Monocyte-derived dendritic cells were generated with GM-CSF and IL-4 for 7 days, followed by another 3 days +/- monocyte-conditioned media, yielding mature (CD83(+)/dendritic cell-lysosome-associated membrane glycoprotein(+)) and immature (CD83(-)/dendritic cell-lysosome-associated membrane glycoprotein(-)) dendritic cells . These dendritic cells were stimulated for another 48 h, and IL-12 p70 was measured by ELISA . We found the following: 1) Immature dendritic cells stimulated with CD154/CD40 ligand or bacteria (both of which concurrently also induced maturation) secreted always more IL-12 than already mature dendritic cells . Mature CD154-stimulated dendritic cells still made significant levels (up to 4 ng/ml) . 2) Terminally mature skin-derived dendritic cells did not make any IL-12 in response to these stimuli . 3) Appropriate maturation stimuli are required for IL-12 production: CD40 ligation and bacteria are sufficient; monocyte-conditioned media are not . 4) Unexpectedly, IL-4 markedly increased the amount of IL-12 produced by both immature and mature dendritic cells, when present during stimulation . 5) IL-10 inhibited the production of IL-12 . Our results, employing a cell culture system that is now being widely used in immunotherapy, extend prior data that IL-12 is produced most abundantly by dendritic cells that are beginning to respond to maturation stimuli . Surprisingly, IL-12 is only elicited by select maturation stimuli, but can be markedly enhanced by the addition of the Th2 cytokine, IL-4.

Clin Exp Immunol, 2000 Dec, 122(3), 477 - 83
Parasite-mediated down-regulation of collagen-induced arthritis (CIA) in DA rats; Mattsson L et al.; Microbial infection can impact on the course of autoimmune disease, both in disease-inducing and disease-protecting capacities . Here we investigated if infection with Trypanosoma brucei brucei (Tbb), the protozoan causative agent of African Sleeping Sickness, could ameliorate the course of CIA in the Dark Agouti rat, an experimental model which shares many features with human rheumatoid arthritis . Infection of animals with living, but not inoculation with dead Tbb resulted in complete or significant reduction of clinical arthritic symptoms . Infection prior to collagen immunization was more effective than a later treatment, and this effect was related to the level of parasitaemia . Using reverse transcriptase-polymerase chain reaction we detected an increase in interferon-gamma mRNA in the draining lymph nodes of Tbb-treated animals relative to controls at day 28 after disease induction . Transforming growth factor-beta could be detected in the lymph nodes in four out of six animals that had received Tbb . In the joints, immunohistochemistry revealed reduced production of tumour necrosis factor-alpha in Tbb-treated animals relative to controls . The most striking difference between Tbb-infected and control groups, as measured by ELISA, was the down-regulation of anti-collagen II IgG antibody responses in parasite-infected animals . We conclude that live parasites can exert an immunomodulatory and protective effect in CIA in which several mechanisms may work in parallel, although the almost complete down-regulation of the anti-collagen antibody response may alone explain the protective effect in CIA . The described model may be useful in further attempts to use the mechanisms involved in parasite immune defence to prevent and treat certain autoimmune conditions.

Curr Opin Microbiol, 2000 Dec, 3(6), 625 - 30
Signalling molecules involved in cellular differentiation during Dictyostelium morphogenesis; Weeks G; GSK-3, Dd-STATa, PKA, rZIP and Ras all play important roles in cell type determination of Dictyostelium discoideum . The fact that homologs of these proteins also function in metazoan development emphasizes the importance of Dictyostelium as a model microbial organism for studying the molecular mechanisms that regulate development . The recent elaboration of the central role for GSK-3 in cell type determination has been of particular importance . The stimulatory effect of extracellular cAMP on GSK-3 activity has been shown to act through the cell surface receptor cAR3 and a tyrosine protein kinase ZAK1, which directly activates and phosphorylates GSK-3 . Several proteins, including Dd-STATa, have been identified as substrates for GSK-3, and are therefore potential transducers of the signals involved in cell type determination.

Antimicrob Agents Chemother, 2001 Jan, 45(1), 187 - 95
Efficacies of lipophilic inhibitors of dihydrofolate reductase against parasitic protozoa; Lau H et al.; Competitive inhibitors of dihydrofolate reductase (DHFR) are used in chemotherapy or prophylaxis of many microbial pathogens, including the eukaryotic parasites Plasmodium falciparum and Toxoplasma gondii . Unfortunately, point mutations in the DHFR gene can confer resistance to inhibitors specific to these pathogens . We have developed a rapid system for testing inhibitors of DHFRs from a variety of parasites . We replaced the DHFR gene from the budding yeast Saccharomyces cerevisiae with the DHFR-coding region from humans, P . falciparum, T . gondii, Pneumocystis carinii, and bovine or human-derived Cryptosporidium parvum . We studied 84 dicyclic and tricyclic 2,4-diaminopyrimidine derivatives in this heterologous system and identified those most effective against the DHFR enzymes from each of the pathogens . Among these compounds, six tetrahydroquinazolines were effective inhibitors of every strain tested, but they also inhibited the human DHFR and were not selective for the parasites . However, two quinazolines and four tetrahydroquinazolines were both potent and selective inhibitors of the P . falciparum DHFR . These compounds show promise for development as antimalarial drugs.

Infect Immun, 2001 Jan, 69(1), 456 - 62
Borrelia spirochetes upregulate release and activation of matrix metalloproteinase gelatinase B (MMP-9) and collagenase 1 (MMP-1) in human cells; Gebbia JA et al.; Borrelia burgdorferi, the spirochetal agent of Lyme disease, stimulated human peripheral blood monocytes to release pro-matrix metalloproteinase-9 (gelatinase B; pro-MMP-9) and active matrix metalloproteinase-1 (collagenase-1; MMP-1) . Human neutrophils also released pro-MMP-9 and a 130-kDa protein with gelatinolytic activity in response to live B . burgdorferi . In addition, U937 cells and human keratinocyte cells were also stimulated to release pro-MMP-9 under the same conditions . However, human umbilical vein endothelial cells (HUVECs) released pro-MMP-9 and pro-MMP-2 in a constitutive manner and were not influenced by live spirochetes . MMPs produced by human monocytes also enhanced the penetration of B . burgdorferi through extracellular matrix component barriers in vitro . Plasmin stabilized on the surface of the Lyme disease spirochete was shown to activate pro-MMP-9 to its active form . This active form was also observed in the plasma of mice infected with a relapsing fever borrelia . These results suggest that borreliae can upregulate MMPs and possibly mediate an activation cascade initiated by plasmin bound to the microbial surface . MMPs may play a role in dissemination of the Lyme disease spirochete and in the pathogenesis of Borrelia infection.

Scand J Immunol, 2000 Dec, 52(6), 609 - 17
Expression of CD1d in the duodenum of patients with cow's milk hypersensitivity; Ulanova M et al.; CD1 cell surface glycoproteins represent a family of non-major histocompatibility complex (MHC) encoded antigen-presenting molecules . All members of the CD1 family appear to mediate the recognition of microbial or endogenous lipid and glycolipid antigens . The recognition of CD1d by a unique subset of natural killer (NK) T cells that leads to rapid production of large amounts of both type 1 and type 2 cytokines can be augmented by some synthetic glycolipids . Because of the proposed role of such CD1d-restricted T cells in immunoregulation, we hypothesized that CD1d molecules participate in mucosal immune responses in patients with gastrointestinal symptoms owing to food hypersensitivity . Patients of that category represent a heterogeneous group in which poorly defined immunological mechanisms are believed to contribute to disease pathogenesis . The expression of CD1 in duodenal biopsy samples from six patients with verified intolerance to cow's milk and six healthy controls was studied by immunoperoxidase staining of cryostat sections using a panel of mouse monoclonal antibodies (MoAbs) specific for CD1a, b, c, and d . Large numbers of CD1d positive cells were found in the lamina propria of all the patients, both during the symptomatic and the asymptomatic periods, whereas healthy controls were virtually devoid of CD1d expression in the duodenum . The localization of CD1d positive cells corresponded to areas where B cells, plasma cells and dendritic cells (DC) were present . A positive correlation was found between the numbers of CD1d(+) and CD19(+) cells in the lamina propria . In contrast to previous reports, no CD1d expression was found on the epithelial cells . Although less numerous than CD1d(+) the CD1c(+) cells were also present in all the patients and in five out of six controls . No staining for CD1a or CD1b was detected in the duodenal biopsy samples from any of the subjects . The exclusive presence of CD1d in the duodenal lamina propria of the patients with cow's milk hypersensitivity might suggest the participation of these molecules in the pathogenesis of allergic reactions to food.

Virology, 2000 Dec 20, 278(2), 514 - 9
In vitro induction of HIV-1 replication in resting CD4(+) T cells derived from individuals with undetectable plasma viremia upon stimulation with human T-cell leukemia virus type I; Moriuchi H et al.; Microbial coinfections have been associated with transient bursts of human immunodeficiency virus (HIV) viremia in patients . In this study we investigated whether human T-cell leukemia virus type I (HTLV-I), another human retrovirus that is prevalent among certain HIV-infected populations, can induce HIV-1 replication in patients who had been successfully treated with highly active antiretroviral therapy . We demonstrate that supernatants from HTLV-I-producing MT-2 cells can induce in vitro replication of HIV-1 from highly purified, resting CD4(+) T cells obtained from individuals with undetectable plasma viremia . Depletion of proinflammatory cytokines from the supernatants reduced, but did not abrogate, the ability to induce HIV-1 replication, indicating that other factors such as HTLV-I Tax or Env also have a role . The HTLV-I-mediated effect does not require productive infection: exposure to heat-inactivated HTLV-I virions, purified Tax protein, or HTLV-I Env glycoprotein also induced expression of HIV-1 . Furthermore, we demonstrate that coculture of resting CD4(+) T cells with autologous CD8(+) T cells markedly inhibits the HTLV-I-induced virus replication . Our results suggest that coinfection with HTLV-I may induce viral replication in the latent viral reservoirs; however, CD8(+) T cells may play an important role in controlling the spread of virus upon microbial stimulation .

Nature, 2000 Nov 30, 408(6812), 580 - 3
Bacterial dehalorespiration with chlorinated benzenes; Adrian L et al.; Chlorobenzenes are toxic, highly persistent and ubiquitously distributed environmental contaminants that accumulate in the food chain . The only known microbial transformation of 1,2,3,5-tetrachlorobenzene (TeCB) and higher chlorinated benzenes is the reductive dechlorination to lower chlorinated benzenes under anaerobic conditions observed with mixed bacterial cultures . The lower chlorinated benzenes can subsequently be mineralized by aerobic bacteria . Here we describe the isolation of the oxygen-sensitive strain CBDB1, a pure culture capable of reductive dechlorination of chlorobenzenes . Strain CBDB1 is a highly specialized bacterium that stoichiometrically dechlorinates 1,2,3-trichlorobenzene (TCB), 1,2,4-TCB, 1,2,3,4-TeCB, 1,2,3,5-TeCB and 1,2,4,5-TeCB to dichlorobenzenes or 1,3,5-TCB . The presence of chlorobenzene as an electron acceptor and hydrogen as an electron donor is essential for growth, and indicates that strain CBDB1 meets its energy needs by a dehalorespiratory process . According to their 16S rRNA gene sequences, strain CBDB1, Dehalococcoides ethenogenes and several uncultivated bacteria form a new bacterial cluster, of which strain CBDB1 is the first, so far, to thrive on a purely synthetic medium.

Curr Microbiol, 2001 Jan, 42(1), 32 - 8
Evaluation of nitrogenous substrates such as peptones from fish:a new method based on Gompertz modeling of microbial growth; Dufosse L et al.; Fish peptones from tuna, cod, salmon, and unspecified fish were compared with a casein one by using a new method based on Gompertz modeling of microbial growth . Cumulative results obtained from six species of bacteria, yeasts, and fungi showed that, in most cases, these fish peptones are very effective . Nevertheless, this study raised some questions about the standardization of fish raw material, the enzymatic hydrolysis of fish proteins, and the composition of the culture medium used for testing the peptones.

Vaccine, 2000 Nov 22, 19(7-8), 840 - 3
Protection of cattle against rinderpest by intranasal immunisation with a dry powder tissue culture vaccine; Anderson J et al.; Dry powder tissue culture rinderpest vaccine containing 10(2.5) TCID(50) of virus per dose administered intranasally to cattle induced high titre circulating antibody responses and protection against challenge with a virulent strain of rinderpest virus . A reduction in the dose of virus to 10(1.1) TCID(50) resulted in a failure to elicit detectable antibody responses and a lack of protection . Intranasal powder vaccine offers several advantages over conventional needle-administered aqueous rinderpest vaccine, including greater stability in the absence of a cold chain, reduced risk of 'needle transfer' of other microbial agents present in the vaccinated herd and lower cost.

Clin Infect Dis, 2000 Oct, 31 Suppl 5, S190 - 201
Endogenous antipyretics; Tatro JB; Fever is the hallmark of the stereotyped host response to microbial infection, although it is just one of a number of high-risk strategies employed by the infected host to clear itself of invading pathogens . The febrile response is accompanied by activation of multiple endogenous antipyretic systems that serve to suppress its magnitude or duration . These include neuroactive substances of neural and humoral origin, some of which (e.g., glucocorticoids, melanocortins, and IL-10) have broad-ranging anti-inflammatory actions . Glucocorticoids, vasopressin, and melanocortins appear to exert their antipyretic effects by acting on receptors within the brain, but beyond this the mechanisms involved are unknown . It is hypothesized, but not proven, that endogenous antipyretic systems protect the host against the destructive consequences of unchecked fever . Importantly, pharmacological blockade of the actions of endogenous antipyretic systems increases fevers of even low to moderate intensity . Therefore, in addition to protecting against catastrophic consequences of high fever, endogenous antipyretic systems seem to play a fundamental physiological role in determining the normal course of fever . Elucidating the neural and biochemical mechanisms involved in suppression of fever by physiological antipyretic systems will yield a rich benefit, both by advancing the basic understanding of host defense strategies, and by permitting the design of novel antipyretic and anti-inflammatory strategies for therapeutic intervention in human disease.

Int J Med Microbiol, 2000 Oct, 290(4-5), 307 - 11
Toxin genes on pathogenicity islands: impact for microbial evolution; Dobrindt U et al.; Toxin-specific genes are often located on mobile genetic elements such as phages, plasmids and pathogenicity islands (PAIs) . The uropathogenic E . coli strain 536 carries two alpha-hemolysin gene clusters, which are part of the pathogenicity islands I536 and II536, respectively . Using different genetic techniques, two additional PAIs were identified in the genome of the E . coli strain 536, and it is likely that further PAIs are located on the genome of this strain . Pathogenicity islands are often associated with tRNA genes . In the case of the E . coli strain 536, the PAI-associated tRNA gene leuX, which encodes a minor leucyl-tRNA, affects the expression of various virulence traits including alpha-hemolysin production . The exact mode of action of the tRNA5Leu-dependent gene expression has to be identified in the future.

Clin Pediatr (Phila), 2000 Nov, 39(11), 627 - 33
Radiologic differentiation between bacterial and viral lower respiratory infection in children: a systematic literature review; Swingler GH; A systematic literature review was performed to quantify the accuracy of chest radiography in differentiating bacterial from viral lower respiratory infection in children . Relevant studies were identified in a systematic literature search and were included in the review according to predetermined criteria . Five of 13 relevant identified studies met the inclusion criteria . No clinically useful degree of accuracy was demonstrated, but great caution is needed in interpreting the findings because of the suboptimal nature of the reference standards, even in included studies . It is recommended that future surveys of the microbial etiology of pneumonia that employ a credible reference standard (such as lung aspiration) be used as opportunities to perform studies of diagnostic accuracy.

Proc Inst Mech Eng {H}, 2000, 214(5), 513 - 8
Influence of gelatin and bovine serum lubricants on ultra-high molecular weight polyethylene wear debris generated in in vitro simulations; Bell J et al.; Ultra-high molecular weight polyethylene (UHMWPE) wear debris induced osteolysis has a major role in the late aseptic loosening and ultimate failure of total hip replacements (THR) . Clinically relevant in vitro simulations of wear are essential to predict the osteolytic potential of bearing surfaces in artificial hip joints . Newborn calf or bovine serum has been accepted as a boundary lubricant for such in vitro tests, but its biological stability has been questioned . This study compared the wear factors, number of wear particles and levels of microbial contamination produced in bovine serum and a gelatin-based lubricant . The wear factors produced by the two lubricants were not significantly different, however the wear debris morphology produced was substantially different . The bovine serum became contaminated with micro-organisms within 28 h, whereas the protein-based lubricant remained uncontaminated . The results showed that bovine serum was not a stable boundary lubricant . They also showed that although the wear factors for the two solutions were not significantly different, the protein-based lubricant was not a suitable alternative to bovine serum because the wear debris produced was not clinically relevant.

Int J Hyg Environ Health, 2000 Oct, 203(2), 97 - 104
Exposure to airborne fungi, MVOC and mycotoxins in biowaste-handling facilities; Fischer G et al.; Health impacts due to fungi in indoor air can only be estimated reliably, if both fungal propagules and fungal secondary metabolites are qualified and quantified . In the present study, the fungal species composition in a compost facility is compared to the spectrum of microbial metabolites in the air with regard to the physiological properties of different fungal species . A number of relevant fungi was tested for the production of both volatile and non-volatile metabolites on different substrata . The profiles of mycotoxins and microbial volatile organic compounds (MVOC) turned out to be specific for certain species in pure culture . Consequently, the fungi may have different toxicological health impacts, though information on the relevance of microbial volatiles is still limited.

Diagn Microbiol Infect Dis, 2000 Nov, 38(3), 185 - 7
A new diagnostic problem: isolation of Escherichia coli O157:H7 strains with aberrant biochemical properties; Ware JM et al.; Over a five-year period (1995-1999) the Microbial Diseases Laboratory received 34 strains of E . coli O157:H7 each with a single aberrant biochemical property . In addition, 27 O157 strains with negative or delayed motility were noted during the same time period . These observations suggest that there may be an increased likelihood to misdiagnose O157:H7 infections using commercial systems in the future due to increasing phenotypic variability.

Int Microbiol, 2000 Sep, 3(3), 139 - 46
DNA methylation in ciliates: implications in differentiation processes; Gutierrez JC et al.; Much experimental evidence on the role of DNA methylation in gene expression has been reported . Here we review reports on DNA methylation in ciliated protozoa, emphasizing its implications in cell differentiation processes . Both types of methylated bases (adenine and cytosine) can be found in macronuclear DNA . The division cycle and conjugation have been studied with regard to adenine methylation, and several different functions have been assigned to the methylation changes detected in these processes . Cytosine methylation changes were analyzed during stomatogenesis of Paramecium and encystment of Colpoda inflata . A comparative analysis with other similar microbial eukaryotic differentiation processes is carried out.

Int Microbiol, 2000 Jun, 3(2), 97 - 101
Mineralogical composition and biomass studies of the microbial mats sediments from the Ebro Delta, Spain; Urmeneta J et al.; The mineral composition of the microbial mats at La Banya spit was studied . The spit is formed by a narrow sand bar and a peninsula and is located south of the main body of the Ebro Delta (Tarragona, Spain) . Although quartz was the predominant mineral component in all sampling sites, clay, feldspars, calcite, aragonite, halite, dolomite and gypsum were also found . An increase in both the fine material (clay) and the halite content was observed in the sites influenced by nearby salterns . The amount of each mineral did not differ significantly along a 55 cm deep profile, except for halite and aragonite, which reached a maximum in the surface and decreased with depth . Dolomite, which ranged from 0.5 to 5% (w/w), is a possible indicator of sulfate-reducing bacteria activity in the past . Organic carbon and total nitrogen were quantified for biomass assessment . Total nitrogen ranged from 0.1 to 0.56% in the uppermost layer, where the microbial mat is active, but was undetectable at deeper layers . Organic carbon ranged from 1 to 5.5% in the active microbial mat layers and decreased to 0.3% at deeper layers . During the summer, both organic carbon and total nitrogen contents (biomass) of the microbial mat samples from some sites increase, whereas other sites show constant concentrations throughout the year, and others have a fluctuant biomass content.

Int Microbiol, 1998 Mar, 1(1), 27 - 34
Cosmopolitan distribution of the large composite microbial mat spirochete, Spirosymplokos deltaeiberi; Margulis L et al.; Inocula from organic-rich black muds immediately underlying intertidal laminated microbial mats dominated by Microcoleus chthonoplastes yielded large, variable diameter spirochetes . These unusual spirochetes, previously reported only from the Alfacs Peninsula at the delta of the Ebro river in northeast Spain, contain striking arrays of cytoplasmic granules packed into their protoplasmic cylinders . On several occasions, both in summer and winter, the huge spirochetes were recognized in samples from mats growing in the Sippewissett salt marsh at Woods Hole Massachusetts . They were also seen in similar samples from microbial mats at North Pond, Laguna Figueroa, Baja California Norte, Mexico . The identity of these spirochetes was confirmed by electron microscopy: number and disposition of flagella, composite structure, measurements of their distinctive cytoplasmic granules . The granules, larger, more conspicuous and present in addition to ribosomes, are hypothesized to contain ATPases . As culture conditions worsen, these spirochetes retract into membrane-bounded round bodies in which they form refractile inclusions . From morphology and behavior we conclude the North American spirochetes from both Atlantic and Pacific intertidal microbial mats are indistinguishable from those at the delta of the Ebro river . We conclude a cosmopolitan distribution for Spirosymplokos deltaeiberi.

Braz J Infect Dis, 1997 Mar, 1(1), 17 - 26
Emerging Pathogens Associated with Tick-Borne Infections; Roberts R et al.; In recent years, several microbial agents have been identified that result in significant morbidity and mortality . The newly recognized tick borne infections, babesiosis and ehrlichiosis, may be transmitted by the same tick that transmits Borrelia burgdorferi and simultaneous infections may occur . Babesia are intraerythrocytic protozoa that may cause severe hemolytic anemia, whereas Ehrlichia, depending on the species, may infect either monocytes or granulocytes, with associated leukopenia, thrombocytopenia and anemia . Improved laboratory surveillance is urgently needed to assess the prevalence of these worldwide pathogens in order to institute appropriate infection control efforts.

J Small Anim Pract, 2000 Nov, 41(11), 501 - 7
Diagnostic features, confirmation and disease progression in 28 cases of lethal acrodermatitis of bull terriers; McEwan NA et al.; Lethal acrodermatitis (LAD) is a genetically determined metabolic disease of bull terriers first described in the USA in the 1980s . In this study, the largest so far reported, 28 bull terriers born in the UK were diagnosed as suffering from LAD, and the clinical findings and the progression of the disease with time are described . The main characteristics of LAD are stunting, splayed digits, eating difficulties, skin disease of the face and feet, and increased susceptibility to microbial infections . In older dogs, paronychia, nail disease and hyperkeratosis of the footpads develops, becoming severe in dogs over six months of age . A diagnosis of LAD can be strongly suspected in any bull terrier showing a combination of the aforementioned signs from an early age . Dermatohistopathological demonstration of marked parakeratotic hyperkeratosis is strongly supportive of the diagnosis of LAD and, in association with the typical clinical findings, is sufficient to confirm a diagnosis . Although many of the clinical signs and the pathology of this condition suggest zinc deficiency, the measurement of blood zinc levels as a diagnostic aid is of limited value.

Braz J Infect Dis, 1997 Aug, 1(4), 153 - 176
Emerging Pathogens Associated with Infectious Diarrhea; Roberts RB; In recent years, emerging microbial pathogens associated with infectious diarrhea have caused significant morbidity and mortality . Although sporadic cases of infectious diarrhea have occurred worldwide in the past, recent outbreaks in the United States traced to contaminated water or food have raised concerns about the safety of the water supply and the adequacy of surveillance of the food supply and foodborne diseases . In 1993, Cryptosporidium parvum, an important cause of unrelenting diarrhea and severe weight loss in AIDS patients, was associated with the largest outbreak of infectious diarrhea caused by contaminated municipal water that has ever been reported in the U.S . During the early summer of 1996, a major outbreak of Cyclospora cayetanensis that infected approximately 1,500 persons in 20 states, Washington, D.C . and two Canadian provinces was reported from North America . The suspected food vehicle in this outbreak was contaminated raspberries imported from Guatemala . In addition to these coccidian protozoa,Escherichia coli 0157:H7, first recognized in 1982 as a cause of hemorrhagic colitis, has recently been responsible for a multi-state outbreak in the U.S . due to contamination of commercial ground beef, and an outbreak in Japan that infected over 9,500 persons, two-thirds of whom were children . The contaminated food vehicle in the latter outbreak, although suspected to be radish sprouts, remains unknown . These recent massive outbreaks underscore the importance of a well-established public health infrastructure and an effective surveillance system for the early identification and reporting of infected patients that will lead to appropriate epidemiologic investigations and the rapid detection of contaminated vehicles.

Nat Struct Biol, 2000 Nov, 7 Suppl, 932 - 4
An overview of structural genomics; Burley SK; With access to sequences of entire human genomes plus those of various model organisms and many important microbial pathogens, structural biology is on the verge of a dramatic transformation . Our newfound wealth of sequence information will serve as the foundation for an important initiative in structural genomics . We are poised to embark on a systematic program of high-throughput X-ray crystallography and NMR spectroscopy aimed at developing a comprehensive view of the protein structure universe . Structural genomics will yield a large number of experimental protein structures (tens of thousands) and an even larger number of calculated comparative protein structure models (millions) . This enormous body of structural data will be freely available, and promises to accelerate scientific discovery in all areas of biological science, including biodiversity and evolution in natural ecosystems, agricultural plant genetics, breeding of farm and domestic animals, and human health and disease.

Curr Biol, 2000 Nov 16, 10(22), R823 - 5
Microbial pathogenesis: lipid rafts as pathogen portals; Rosenberger CM et al.; The route of initial entry influences how host cells respond to intracellular pathogens . Recent studies have demonstrated that a wide variety of pathogens target lipid microdomains in host cell membranes, known as lipid rafts, to enter host cells as an infectious strategy.

Curr Opin Biotechnol, 2000 Dec, 11(6), 547 - 53
Enzymatic hydroxylation reactions; Holland HL et al.; During the past 18 months, considerable progress has been made in the understanding of the key enzyme-substrate interactions that control the regioselectivity and stereoselectivity of the hydroxylation reaction performed by cytochrome-P450-dependent enzymes of mammalian origin . The manipulation of microbial hydroxylating enzymes, in both whole-cell and cell-free environments, has also been examined in the context of controlling the regioselectivity and stereoselectivity of the hydroxylation reaction . Several new applications for hydroxylating enzymes have been reported, and the construction of chimeric hydroxylating enzymes has been used both for mechanistic studies and for the production of enzymes with high hydroxylating activity for a defined substrate.

Curr Opin Biotechnol, 2000 Dec, 11(6), 520 - 6
Enzymatic asymmetric synthesis by decarboxylases; Ward OP et al.; Decarboxylation reactions using microbial cells or enzymes are increasingly being used for the synthesis of enantiomerically pure compounds because of their high degree of regio- and stereo-specificity . Pyruvate decarboxylase, benzoylformate decarboxylase and phenylpyruvate decarboxylase enzymes are capable of acyloin-type condensation reactions leading to formation of chiral alpha-hydroxy ketones, which are versatile building blocks in the pharmaceutical and chemical industries . Availability of three-dimensional structures of some decarboxylases in recent years has facilitated understanding of reaction mechanisms and the creation of mutants with enhanced activity and stability.

Baillieres Best Pract Res Clin Haematol, 2000 Dec, 13(4), 615 - 29
Will genome detection replace serology in blood screening for microbial agents?
Allain JP.
The residual risk of transfusion-transmitted viral infection in developed countries is considered minimal or negligible . However, zero risk remains a strong political objective . Genomic screening for HCV, HIV and HBV represents a major advance, eliminating infectious blood donations collected during the pre-seroconversion window period, rare cases of immunosilent infections and, possibly, a large spectrum of viral variants . In Western countries, HCV RNA genomic screening started on pools of 16-400 plasma samples from individual donations . Pooling may produce false-positive and false-negative results . Individual donation testing is more suitable to blood screening but requires multiplexing, automation, and affordable cost . Because donations from individuals who are HBV DNA-negative/serologically positive, or those apparently recovered from HCV infection, may remain infectious, it is unlikely that HBsAg, anti-HCV, and anti-HIV will be discontinued when genomic screening is extended to all three viruses . HIV-1 p24 antigen may prove redundant with HIV RNA screening . Anti-HTLV-I and HTLV-II will remain more effective than genomic testing .

FEMS Microbiol Ecol, 2000 Dec 1, 34(2), 173 - 180
Genetic diversity of carbofuran-degrading soil bacteria; Desaint S et al.; The genetic diversity of 128 carbofuran-degrading bacteria was determined by ARDRA (amplified ribosomal DNA restriction analysis) of 16S rDNA and restriction fragment length polymorphism analysis of the 16S-23S rDNA spacer region (IGS) using five endonucleases . The isolates were distributed in 26 distinct ARDRA groups and 45 IGS types revealing a high level of microbial diversity confirmed by ARDRA clustering and sequencing of 16S rDNA . The occurrence of a methylcarbamate-degrading gene (mcd) was monitored by polymerase chain reaction amplification using specific primers . The mcd gene was detected only in 58 bacteria and there was no clear relationship between the presence of this gene and the phylogenetic position of the strain.

FEMS Microbiol Ecol, 2000 Dec 1, 34(2), 161 - 171
Competition between methanogenesis and quinone respiration for ecologically important substrates in anaerobic consortia; Cervantes FJ et al.; Anaerobic consortia obtained from a wide variety of environments were tested for oxidizing several ecologically significant substrates with the humic model compound, anthraquinone-2,6-disulfonate (AQDS), as terminal electron acceptor . All the substrates, including hydrogen, acetate, propionate, methanol and lactate, were completely or partially converted to methane when bicarbonate was the only electron acceptor available . Addition of AQDS (20 mM) to the cultures prevented methanogenesis in most cases and AQDS reduction became the preferred pathway . AQDS was shown to be toxic for methanogenesis and this effect played an important role in enabling quinone-respiring bacteria to outcompete methanogens . Furthermore, AQDS respiration is thermodynamically more favorable than methanogenesis . All the consortia evaluated were capable of oxidizing hydrogen linked to the reduction of AQDS . Most inocula tested were also able to oxidize acetate and lactate in the same way . When methanol was provided as an electron donor competition between methanogenesis and acetogenesis occurred . Acetate accumulated from the latter process was responsible for quinone respiration . These results suggest that quinone-respiring bacteria are ubiquitous and that quinones in humus may significantly contribute to carbon cycling process by serving as a terminal electron acceptor for the anaerobic microbial oxidation of a wide variety of ecologically important substrates.

FEMS Microbiol Ecol, 2000 Dec 1, 34(2), 157 - 160
Carbon monoxide production is not enhanced by nitrogenase activity; Milligan P et al.; A diverse group of nitrogen-fixing bacteria and two heme degraders were grown with and without fixed nitrogen sources under oxic and suboxic conditions, with and without addition of heme-containing compounds . Several of the strains produced carbon monoxide (CO) under one or more of these conditions, but nitrogenase activity did not stimulate rates of production relative to controls . Although nitrogenase can reduce CO(2) to CO in vitro in the absence of N(2), this process likely contributes minimally to CO production in soils under in situ conditions . In contrast, myoglobin or hematin addition under oxic conditions significantly stimulated CO production by the heme degraders . However, estimates of CO production from microbial heme turnover suggest that this too is likely to be only a small source of CO in soils in situ.

J Gastroenterol Hepatol, 2000 Oct, 15 Suppl, G69 - 72
Disinfection of gastrointestinal endoscopes and accessories; Tandon RK; A worldwide concern has emerged with regard to endoscope disinfection and many gastrointestinal endoscopy associations have developed guidelines for proper disinfection of endoscopes and endoscopic accessories . A working party was convened to formulate guidelines for the Asia-Pacific region, pertaining to any setting in which gastrointestinal endoscopy is performed . Endoscope reprocessing that meets the established standard of practice helps to ensure a microbial-free endoscope for all patients, reduces the risk of disease transmission and helps to prolong the life of the endoscope . The recommendations included mechanical cleaning as the first and most important step followed by immersion in 2% glutaraldehyde for a minimum period of 10 min . Automated disinfectors have been recommended for busy endoscopy centres to ensure better compliance . Reuse of endoscopic accessories meant for 'single use' remains a controversial issue . Strict quality assurance programmes are a must to preclude lack of compliance with these guidelines.

Bone Marrow Transplant, 2000 Nov, 26(9), 1025 - 8
Successful treatment of invasive aspergillosis in chronic granulomatous disease by granulocyte transfusions followed by peripheral blood stem cell transplantation; Bielorai B et al.; Chronic granulomatous disease (CGD) is a primary immunodeficiency disorder characterized by impaired microbial killing and susceptibility to bacterial and fungal infections . Cure of the disease can be achieved by stem cell transplantation when performed early in its course, and before severe infections have developed . Invasive aspergillosis constitutes a very high risk for transplantation . We report a 4-year-old boy with X-linked CGD who underwent successful HLA-identical peripheral blood stem cell (PBSC) transplantation during invasive pulmonary aspergillosis and osteomyelitis of the left fourth rib, which was unresponsive to antifungal treatment . During the 2 months prior to the transplant he received G-CSF-mobilized granulocyte transfusions (GTX) from unrelated donors three times a week in addition to the antifungal treatment . This resulted in clinical improvement in his respiratory status . He also received GTX during the aplastic period after the conditioning regimen, until he had engrafted . Post-transplant superoxide generation test revealed that neutrophil function was within normal range . One year post transplant the CT scan showed almost complete clearance of the pulmonary infiltrates and a marked improvement in the osteomyelitic process . Based on other reports and our own experience, GTX can serve as important treatment in patients with CGD who have failed conventional anti-fungal treatment and for whom stem cell transplantation is the only chance for cure.

Microbes Infect . 2000 Oct;2(12):1537.
'Infectious web'; Kotra LP et al.; Exhaustive information on the Epstein-Barr virus, a member of the herpes family, is described at the International Herpes Management Forum web-site . Cervical cancer associations, AIDS treatment projects, and the Los Alamos National Laboratories provide useful information on papillomavirus infections, as well as hyperlinks to recent international papillomavirus conferences . A private pharmaceutical company, in collaboration with the National Institutes of Health, has launched a lively web-site covering different aspects of microbial infections for the general public.

Biochimie, 2000 Nov, 82(11), 1043 - 52
The His gap motif in microbial lipases: a determinant of stereoselectivity toward triacylglycerols and analogs; Pleiss J et al.; Lipases preferably hydrolyze the sn-1 and sn-3 acyl chain of triacylglycerols and sn-2 substituted analogs . Molecular modeling studies of the stereopreference of microbial lipases from Rhizopus oryzae, Rhizomucor miehei, Candida rugosa, and lipase B from Candida antarctica toward the hydrolysis of triacylglycerols and analogs revealed that sterical interactions occurring between the sn-2 substituent and the His gap affect substrate geometry, which can be monitored by a single torsion angle . This torsion angle correlates to the experimentally determined stereopreference and is, therefore, suitable to predict stereopreference by molecular modeling . For a given microbial lipase, stereopreference can be estimated by measuring the distance between the side chains of the His gap residues: a narrow His gap cleft implies sn-3 stereopreference for all investigated substrates; a medium-sized His gap discriminates by flexibility of the substrates: flexible substrates are hydrolyzed in sn-1, while rigid substrates are hydrolyzed in sn-3 . A wide open His gap implies sn-1 stereopreference for all substrates . This rule holds for all investigated microbial wild type lipases and mutants.

Appl Environ Microbiol, 2000 Dec, 66(12), 5488 - 91
Rapid method for coextraction of DNA and RNA from natural environments for analysis of ribosomal DNA- and rRNA-based microbial community composition; Griffiths RI et al.; A rapid protocol for the extraction of total nucleic acids from environmental samples is described . The method facilitates concomitant assessment of microbial 16S rRNA diversity by PCR and reverse transcription-PCR amplification from a single extraction . Denaturing gradient gel electrophoresis microbial community analysis differentiated the active component (rRNA derived) from the total bacterial diversity (ribosomal DNA derived) down the horizons of an established grassland soil.

FEMS Microbiol Ecol, 2000 Sep 1, 33(3), 197 - 208
Occurrence of sulfate-reducing bacteria under a wide range of physico-chemical conditions in Au and Cu-Zn mine tailings; Fortin D et al.; Sulfate-reducing bacteria (SRB) have been observed in mining environments, but their presence has not been linked to specific physico-chemical and mineralogical factors . The present study was undertaken to assess the presence of SRB in several Au and Cu-Zn mine tailings located near Timmins, Ont., Canada, and determine the factors responsible for their presence . Vegetated and non-vegetated mine tailings were sampled for SRB enumeration, pH, Eh, water content, total carbon content and sequential chemical extraction . Results first showed that SRB populations were present at all sites and that their distribution varied with depth . Populations were recovered from neutral pH and slightly anoxic tailings and from highly acidic (pH 2) and oxic tailings . The total carbon content of the tailings was generally low and not related to the presence of vegetation . In addition, the carbon content did not affect SRB population distribution and appeared to be more related to the type of tailings, i.e., oxidized and acidic Cu-Zn tailings contained on average more carbon than Au tailings . Results also indicated that the water content of the tailings varied greatly with depth and was not related to the presence of SRB populations . The sequential chemical extraction showed that the pyrite content of the tailings was lower in Au tailings than in Cu-Zn tailings, and that some oxidized Cu-Zn sites were depleted in pyrite due to microbial and chemical oxidation . Our results indicate that SRB could be cultured from a variety of sites and sample types, and that factors such as pH, Eh, water content and carbon content at the collection sites did not exert control on their presence.

FEMS Microbiol Ecol, 2000 Sep 1, 33(3), 181 - 189
Seasonal variation in rates of methane production from peat of various botanical origins: effects of temperature and substrate quality; Bergman I I et al.; The methane produced in peat soils can vary over the growing season due to variations in the supply of available substrate, the activity of the microbial community or changes in temperature . Our aim was to study how these factors regulate the methane production over the season from five different peat types of different botanical origin . Peat samples were collected on seven occasions between June and September . After each sampling, the peat soils were incubated at five different temperatures (7, 10, 15, 20 and 25 degrees C) without added substrate, or at 20 degrees C with added substrate (glucose, or H(2)/CO(2), or starch) . Rates of methane production averaged over the season differed significantly (P<0.05, R(2)=0.76) among the five peat types, the minerotrophic lawn producing the highest rates, and the hummock peat producing the lowest . The seasonal average Q(10) values for each plant community varied between 4.6 and 9.2, the highest value being associated with the ombrotrophic lawn and the lowest value with the mud-bottom plant community . For the unamended peat samples, the rates of methane production from each plant community varied significantly (P<0.05) over the season . This implies that the quality of organic matter, in combination with changes in temperature, explains the seasonal variation in methane production . However, addition of saturating amounts of glucose, H(2)/CO(2) or starch at 20 degrees C significantly reduced the seasonal variation (P<0.05) in methane production in peat from the minerotrophic lawn, wet carpet and mud-bottom plant communities . This suggests that substrate supply (e.g . root exudates) for the micro-organisms also varied over the season at these sites . Seasonal variation in methane production rates was apparent in peat from the hummock and ombrotrophic lawn plant communities even after addition of substrates, suggesting that the active biomass of the anaerobic microbial populations at these sites was regulated by other factors than the ones studied.

Acta Microbiol Pol, 2000, 49(2), 135 - 47
Oxidation of elemental sulfur by bacteria and fungi in soil; Czaban J et al.; Laboratory experiments were used to determine the effects of antibiotics, organic C and CaCO3 amendments of sterile reinoculated soil on S0 oxidation by bacteria and fungi . The rate of S0 oxidation in soil with nystatin added was higher than in soil amended with penicillin + streptomycin . This tells us that bacteria were more efficient than fungi in the S0 oxidation process . It was demonstrated that neutrophilic chemolithotrophs were more efficient in this process than heterotrophs . Glucose introduced to the soil had a negative effect and CaCO3 had a positive effect on S0 oxidation . In soil enriched with glucose the number of chemolithotrophs was very low in comparison with extremely numerous heterotrophic bacteria and fungi . It suggests that the role of heterotrophs in S0 oxidation could be important in habitats rich in organic C, e.g . rhizosphere . In soil containing S0, qualitative changes of fungal communities to genera with higher S0 oxidation ability was also noted . In the presented paper, after comparison of the own results with the data of others concerning the natural soils, the role of various microbial groups in S0 oxidation process in soils is discussed.

Eur J Immunol, 2000 Nov, 30(11), 3140 - 6
Expression of MHC class II molecules contributes to lipopolysaccharide responsiveness; Piani A et al.; Activation of phagocytes by lipopolysaccharide (LPS) causes synthesis and secretion of various mediators of inflammation . CD14, a glycosylphosphatidylinositol-anchored monocytic antigen serving as receptor for LPS, and members of the family of Toll-like receptors mediate cellular activation in response to LPS . Here we investigated whether expression of MHC class II molecules modified the response to LPS . Comparing LPS responsiveness of human and murine cells differing for expression of MHC class II molecules, we found that lack or a low level of expression of MHC class II molecules resulted in diminished secretion of pro-inflammatory cytokines following stimulation with LPS . Thus, expression of MHC class II molecules modifies LPS responsiveness, a finding suggesting that these molecules contribute to the pathogenesis not only of exotoxin-triggered toxic shock but also of endotoxin-triggered septic shock . Additionally to their role in antigen-specific immunity MHC class II molecules may influence the inflammatory response triggered by microbial constituents.

Acta Crystallogr D Biol Crystallogr, 2000 Dec, 56 Pt 12, 1694 - 5
Crystallization and preliminary crystallographic study of b0220, an 'ORFan' protein of unknown function from Escherichia coli; Abergel C et al.; Newly sequenced microbial genomes continue to reveal up to 50% functionally uncharacterized 'anonymous' genes . A significant fraction of these anonymous ORFs does not exhibit any sequence similarity to any protein in the databases and constitutes a set of unique sequences, denoted 'ORFans' . The structure determination of ORFan proteins is both of evolutionary and functional interest . Here, the first crystallization of an Escherichia coli ORFan gene product, the 157 amino-acid b0220 protein, is reported . The crystals belong to the trigonal space group P3 or P3(1), with unit-cell parameters a = b = 47.2, c = 88.4 A . There are two molecules in the asymetric unit . Frozen crystals diffract to 1.6 A resolution using synchrotron radiation . Phasing was performed using multiwavelength anomalous dispersion (MAD) on the selenomethionine-substituted b0220 protein.

Appl Microbiol Biotechnol, 2000 Oct, 54(4), 510 - 4
Enhanced stability of laccase in the presence of phenolic compounds; Mai C et al.; The storage stability of laccase (EC 1.10.3.2) from the white-rot basidomycete Trametes versicolor in potassium-citrate buffer was enhanced by various phenolic compounds as well as by lignin sulfonate . The highest storage stability was obtained with phenolics, e.g . phloroglucin and 3,5-dihydroxybenzoic acid; these represent substrates of laccase which are oxidized slowly because of their relatively high redox potential and which did not precipitate from the solution within the tested period of time . Sterilization enhanced the stability of laccase but additional stabilization by phenolics was observed both under sterile and non-sterile conditions . We thus concluded that stabilization occurred not only through prevention of microbial degradation.

Poult Sci, 2000 Nov, 79(11), 1517 - 23
Chemical preservation of whole broiler carcasses utilizing alkaline hydroxides; Shafer DJ et al.; The chemical preservation of whole broiler carcasses utilizing aqueous alkaline hydroxide solutions was examined as an alternative method of mortality management . Conversion of the preserved carcasses and solutions into an acceptable poultry by-product meal was examined . This research identified the basic parameters for effective preservative solutions that simultaneously hydrolyzed feathers and preserved the carcass . Euthanized, fully feathered, mature broilers were placed in potassium hydroxide (0.5 to 2.0 M) and sodium hydroxide (0.12 to 2.0 M) solutions for 5 and 10 d . Effectiveness was evaluated by visible feather degradation and carcass solubilization, odor production, inhibition of microbial growth, and solids content of the alkaline solutions . Sodium hydroxide at 1.9 and 2.0 M diffused throughout the carcass and produced adequate preservation without apparent putrefaction through 10 d . Aerobic bacteria were not recovered from sodium hydroxide solutions, carcass skin, or intestine samples at the 1.9 M concentration . Treatments of 2.0 M potassium hydroxide and a mixture of 1.5 M potassium hydroxide with 0.5 M sodium hydroxide produced the highest degree of carcass liquification at 10 d without visible putrefaction . Sodium hydroxide solution (2.0 M):carcass weight ratios ranging from 1:1 through 4:1 (wt:wt) were effective in preserving individual carcasses for more than 60 d without putrefaction.

Mol Cell, 2000 Oct, 6(4), 899 - 907
Structure of the TPR domain of p67phox in complex with Rac.GTP; Lapouge K et al.; p67phox is an essential part of the NADPH oxidase, a multiprotein enzyme complex that produces superoxide ions in response to microbial infection . Binding of the small GTPase Rac to p67phox is a key step in the assembly of the active enzyme complex . The structure of Rac.GTP bound to the N-terminal TPR (tetratricopeptide repeat) domain of p67phox reveals a novel mode of Rho family/effector interaction and explains the basis of GTPase specificity . Complex formation is largely mediated by an insertion between two TPR motifs, suggesting an unsuspected versatility of TPR domains in target recognition and in their more general role as scaffolds for the assembly of multiprotein complexes.

Curr Opin Genet Dev, 2000 Dec, 10(6), 606 - 11
Horizontal gene transfer among microbial genomes: new insights from complete genome analysis; Eisen JA; The determination and analysis of complete genome sequences has led to the suggestion that horizontal gene transfer may be much more extensive than previously appreciated . Many of these studies, however, rely on evidence that could be generated by forces other than gene transfer including selection, variable evolutionary rates, and biased sampling.

Ned Tijdschr Geneeskd, 2000 Nov 4, 144(45), 2152 - 6
{Tropical diseases and imported disorders in 1763 patients seen at the Outpatient Clinic for Tropical Diseases, Academic Medical Center, Amsterdam (1996-1997)}; Wetsteyn JC et al.; OBJECTIVE: To describe an analysis of the patients seen at the Outpatient Department (OPD) for Tropical Diseases in the Academic Medical Centre, Amsterdam, during 1996 and 1997 . DESIGN: Descriptive cross-sectional study . METHOD: From our database of OPD-patients the following data were analysed: age, country of birth, travel destination and most frequent complaints at presentation . These were further analysed in relation to travel destination, diagnosis and need of admission . RESULTS: In 1996 and 1997 1763 patients visited the OPD . Abdominal complaints, fever, general malaise and skin diseases were the main problems . Abdominal complaints were more often acquired in Asia, fever in sub-Saharan Africa and skin problems in South America . General malaise was not related to a specific travel destination . Abdominal complaints, fever and general malaise were more often caused by parasites, and skin problems by bacteria . Plasmodia were the most frequently encountered microbial cause . Malaria was found in 1 out of every 3 Dutch, and 9 out of every 10 Ghanaian patients with fever from Africa . CONCLUSION: The analysis of the database yielded useful information regarding patients with import diseases in the Netherlands and with respect to travellers to tropical areas.

J Immunol, 2000 Dec 1, 165(11), 6538 - 44
The N-terminal lipopeptide of a 44-kDa membrane-bound lipoprotein of Mycoplasma salivarium is responsible for the expression of intercellular adhesion molecule-1 on the cell surface of normal human gingival fibroblasts; Shibata K et al.; The activities to induce TNF-alpha production by a monocytic cell line, THP-1, and ICAM-1 expression and IL-6 production by human gingival fibroblasts were detected in plural membrane lipoproteins of Mycoplasma salivarium . Although SDS-PAGE of the lipoproteins digested by proteinase K did not reveal any protein bands with molecular masses higher than approximately10 kDa, these activities were detected in the front of the gel . A lipoprotein with a molecular mass of 44 kDa (Lp44) was purified . Proteinase K did not affect the ICAM-1 expression-inducing activity of Lp44, but lipoprotein lipase abrogated the activity . These results suggested that the proteinase K-resistant and low molecular mass entity, possibly the N-terminal lipid moiety, played a key role in the expression of the activity . The N-terminal lipid moiety of Lp44 was purified from Lp44 digested with proteinase K by HPLC . Judging from the structure of microbial lipopeptides as well as the amino acid sequence and infrared spectrum of Lp44, the structure of the N-terminal lipid moiety of Lp44 was speculated to be S-(2, 3-bisacyloxypropyl)-cysteine-GDPKHPKSFTEWV- . Its analogue, S-(2, 3-bispalmitoyloxypropyl)-cysteine-GDPKHPKSF, was synthesized . The lipopeptide was similar to the N-terminal lipid moiety of Lp44 in the infrared spectrum and the ICAM-1 expression-inducing activity . Thus, this study suggested that the active entity of Lp44 was its N-terminal lipopeptide moiety, the structure of which was very similar to S-(2, 3-bispalmitoyloxypropyl)-cysteine-GDPKHPKSF.

J Exp Med, 2000 Nov 20, 192(10), 1425 - 40
Lymph-borne chemokines and other low molecular weight molecules reach high endothelial venules via specialized conduits while a functional barrier limits access to the lymphocyte microenvironments in lymph node cortex; Gretz JE et al.; Lymph-borne, soluble factors (e.g., chemokines and others) influence lymphocyte recirculation and endothelial phenotype at high endothelial venules (HEVs) in lymph node cortex . Yet the route lymph-borne soluble molecules travel from the subcapsular sinus to the HEVs is unclear . Therefore, we injected subcutaneously into mice and rats a wide variety of fluorophore-labeled, soluble molecules and examined their distribution in the draining lymph nodes . Rather than percolating throughout the draining lymph node, all molecules, including microbial lipopolysaccharide, were very visible in the subcapsular and medullary sinuses but were largely excluded from the cortical lymphocyte microenvironments . Exclusion prevailed even during the acute lymph node enlargement accompanying viral infection . However, low molecular mass (MW) molecules, including chemokines, did gain entry into the cortex, but in a very defined manner . Low MW, fluorophore-labeled molecules highlighted the subcapsular sinus, the reticular fibers, and the abluminal and luminal surfaces of the associated HEVs . These low MW molecules were in the fibers of the reticular network, a meshwork of collagen fibers ensheathed by fibroblastic reticular cells that connects the subcapsular sinus floor and the HEVs by intertwining with their basement membranes . Thus, low MW, lymph-borne molecules, including chemokines, traveled rapidly from the subcapsular sinus to the HEVs using the reticular network as a conduit.

Drug Saf, 2000 Nov, 23(5), 351 - 62
Chinese proprietary medicine in Singapore: regulatory control of toxic heavy metals and undeclared drugs; Koh HL et al.; Traditional Chinese medicine (TCM) is gaining popularity as a form of complementary and alternative medicine . Reports of efficacy of TCM are increasing in numbers . TCM includes both crude Chinese medicinal materials (plants, animal parts and minerals) and Chinese proprietary medicine (CPM) {final dosage forms} . Despite the belief that CPM and herbal remedies are of natural origin, unlike Western medicine, and are hence safe and without many adverse effects, there have been numerous reports of adverse effects associated with herbal remedies . Factors affecting the safety of herbal medicines include intrinsic toxicity, adulteration, substitution, contamination, misidentification, lack of standardisation, incorrect preparation and/or dosage and inappropriate labelling and/or advertising . Hence, new regulations on the control of CPM were enforced in Singapore with effect from 1 September 1999 . These include licensing and labelling requirements, as well as control of microbial contamination . This article also reviews reports of excessive toxic heavy metals and undeclared drugs in CPM in Singapore between 1990 and 1997 . The names, uses, toxic heavy metal or drug detected and the year of detection are tabulated . Information on the brand or manufacturer's name are provided whenever available . The public and healthcare professionals should be better informed of the basic concept of TCM and its usefulness, as well as the potential adverse effects associated with its use . Greater control over the safety and quality of CPM could be achieved through good manufacturing practice, regulatory control, research, education, reporting usage of Chinese medicine (as in drug history) as well as reporting of adverse events.

Hum Gene Ther, 2000 Nov 1, 11(16), 2247 - 51
Virus-mediated gene transfer for cutaneous gene therapy; Ghazizadeh S et al.; Cutaneous gene therapy offers unique opportunities and limitations in the use of viral vectors for corrective gene transfer . Skin presents a formidable barrier to microbial invasion and is nourished by small blood vessels, thus ruling out the possibility of directed virus delivery through cannulated blood vessels . However, skin is physically accessible and its resident keratinocyte stem cell population is susceptible to direct in vivo transduction with retroviral vectors . Furthermore, keratinocyte stem cells transduced in culture have been shown to persist and to express the encoded transgene when grafted to immunocompromised mice . Cutaneous gene therapy trials are likely to involve virus-mediated transduction as a principal means of gene transfer.

Am J Obstet Gynecol, 2000 Nov, 183(5), 1138 - 43
Participation of the novel cytokine interleukin 18 in the host response to intra-amniotic infection; Pacora P et al.; OBJECTIVE: Interleukin 18 is a proinflammatory pleiotropic cytokine that has been implicated in the host defense against infection . This study was undertaken to determine whether interleukin 18 concentrations change in the maternal, fetal, and amniotic fluid compartments with labor (term and preterm) and microbial invasion of the amniotic cavity . STUDY DESIGN: Amniotic fluid was assayed for interleukin 18 in samples obtained from 285 patients in the following groups: (1) term not in labor (n = 22), in labor (n = 19), and with microbial invasion of the amniotic cavity (n = 16); (2) preterm labor who delivered at term (n = 38), who delivered preterm but without microbial invasion of the amniotic cavity (n = 41), and preterm labor with microbial invasion of the amniotic cavity (n = 24); (3) preterm premature rupture of membranes without microbial invasion of the amniotic cavity (n = 30) and with microbial invasion of the amniotic cavity (n = 34); (4) term premature rupture of membranes not in labor (n = 20) and term premature rupture of membranes in labor (n = 19); and (5) midtrimester (n = 22) . In addition, cord and maternal plasma samples from women at term not in labor (n = 20) and in labor (n = 20) were assayed for interleukin 18 . RESULTS: (1) Interleukin 18 was detectable in all amniotic fluid samples and maternal and umbilical cord blood samples . (2) Interleukin 18 concentrations increased with advancing gestational age (r = 0.47; P <.0001) . (3) Microbial invasion of the amniotic cavity in either preterm or term parturition was associated with a significant increase in the amniotic fluid concentration of interleukin 18 (preterm labor without microbial invasion of the amniotic cavity: median, 14.95 pg/mL; range, 3.9-277.0 pg/mL; vs preterm labor with microbial invasion of the amniotic cavity: median, 20.75 pg/mL; range, 5.53-160.21 pg/mL; P <.02; term labor without microbial invasion of the amniotic cavity: median, 18.73 pg/mL; range, 5.09-95.44 pg/mL; vs term labor with microbial invasion of the amniotic cavity: median, 24.35 pg/mL; range, 10.07-144.42 pg/mL; P<.004) . (4) Both term and preterm parturition were associated with a modest increase in amniotic fluid interleukin 18 concentrations, although this trend did not reach statistical significance . (5) Rupture of membranes at term was associated with a significant decrease in amniotic fluid interleukin 18 concentrations (intact membranes: median, 14.96 pg/mL; range, <3.89-26.07 pg/mL; vs rupture of membranes: median, 10.1 pg/mL; range, 4.29-21.44 pg/mL; P <.001) . CONCLUSION: (1) Interleukin 18 is increased in cases of microbial invasion of the amniotic cavity . (2) Interleukin 18 is detectable in the amniotic, maternal, and fetal compartments . (3) We propose that this novel cytokine plays a role in the host defense against infection.

J Microbiol Methods, 2000 Dec 1, 43(1), 3 - 31
Artificial neural networks: fundamentals, computing, design, and application; Basheer IA et al.; Artificial neural networks (ANNs) are relatively new computational tools that have found extensive utilization in solving many complex real-world problems . The attractiveness of ANNs comes from their remarkable information processing characteristics pertinent mainly to nonlinearity, high parallelism, fault and noise tolerance, and learning and generalization capabilities . This paper aims to familiarize the reader with ANN-based computing (neurocomputing) and to serve as a useful companion practical guide and toolkit for the ANNs modeler along the course of ANN project development . The history of the evolution of neurocomputing and its relation to the field of neurobiology is briefly discussed . ANNs are compared to both expert systems and statistical regression and their advantages and limitations are outlined . A bird's eye review of the various types of ANNs and the related learning rules is presented, with special emphasis on backpropagation (BP) ANNs theory and design . A generalized methodology for developing successful ANNs projects from conceptualization, to design, to implementation, is described . The most common problems that BPANNs developers face during training are summarized in conjunction with possible causes and remedies . Finally, as a practical application, BPANNs were used to model the microbial growth curves of S . flexneri . The developed model was reasonably accurate in simulating both training and test time-dependent growth curves as affected by temperature and pH.

Arthritis Rheum, 2000 Nov, 43(11), 2583 - 9
Oral antibiotics as a novel therapy for arthritis: evidence for a beneficial effect of intestinal Escherichia coli; Nieuwenhuis EE et al.; OBJECTIVE: The intestinal flora is thought to play an important role in regulation of immune responses . We investigated the effects of changing the intestinal flora on the course of adjuvant-induced arthritis (AIA) and on experimental autoimmune encephalomyelitis (EAE) by the use of oral antibiotics . METHODS: Oral treatment with either vancomycin or vancomycin, tobramycin, and colistin was started after AIA and EAE induction . Clinical symptoms of AIA and EAE were monitored, and microbial analysis of ileal samples was performed . RESULTS: Oral vancomycin treatment after disease induction significantly decreased clinical symptoms of AIA . Simultaneously, increased concentrations of Escherichia coli were detected in the distal ileum of vancomycin-treated rats . Ileal concentrations of E coli were inversely related to disease scores in rats with AIA . Coadministration of colistin/tobramycin to prevent the increase in E coli abrogated the beneficial effect of vancomycin on AIA . Vancomycin treatment also reduced the clinical symptoms of EAE . CONCLUSION: We propose oral vancomycin as a novel therapeutic strategy in autoimmune diseases.

Arthritis Rheum, 2000 Nov, 43(11), 2578 - 82
Unmethylated oligo-DNA containing CpG motifs aggravates collagen-induced arthritis in mice; Miyata M et al.; OBJECTIVE: To investigate the effects of an intradermal injection of an unmethylated oligodeoxynucleotide (ODN) containing CpG motifs on the severity of collagen-induced arthritis (CIA) . METHODS: CIA was induced in DBA/1 LacJ mice by immunization with bovine type II collagen (CII) in Freund's complete adjuvant followed 3 weeks later by immunization with CII in Freund's incomplete adjuvant (yielding CIA mice) . Unmethylated ODN containing a CpG motif was injected intradermally into DBA/1 LacJ mice at a dosage of 20 microg (yielding CpG-CIA mice) 1 week prior to the first immunization with CII . Unmethylated ODN containing a GpC motif instead of a CpG motif and ODN containing a methylated CpG motif were used to produce controls (GpC-CIA mice and mCpG-CIA mice, respectively) . After the second immunization with CII, arthritis scores were measured weekly up to the eighth week . At the eighth week, the mice were killed, histopathologic changes in the ankle joints were examined, and titers of interferon-gamma (IFNgamma) in the supernatants of splenocytes (1 x 10(7)) stimulated in culture by CII for 3 days were determined by enzyme-linked immunosorbent assay . RESULTS: CpG-CIA mice had significantly higher arthritis scores than CIA mice . CpG-CIA mice had more severe histopathologic changes than CIA mice and mCpG-CIA mice . Moreover, splenocytes in CpG-CIA mice produced higher IFNgamma titers in response to CII than did splenocytes in CIA mice and mCpG-CIA mice . CONCLUSION: Injection of unmethylated oligo-DNA containing CpG motifs aggravated CIA through activation of the Th1-type immune response, suggesting that microbial infection could be one of the mechanisms for aggravation or exacerbation of arthritis or, alternatively, that such infection could be an adjuvant in the induction of arthritis in rheumatoid arthritis.

Yakugaku Zasshi, 2000 Oct, 120(10), 891 - 8
{Nostalgic review of one student of Prof . Tsuda about natural product, carbanion, thiazoline and beta-lactam chemistry}; Hirai K; This article describes the works of one of Tsuda's students who owes their performance to him through the long-term, direct and indirect influence of the Tsuda school, especially supervised by him during his very last period as a Professor at the University of Tokyo . They consist of i) natural product chemistry, i.e . Siccanin and related congeners, ii) carbanion chemistry, i.e . iodomethylation (C1) and trans-iodopropenylation (C3) reactions using thiazoline derivatives and their synthetic application, and iii) beta-lactam chemistry, i.e . 1,3-dipoler cycloaddition reaction of iminochloride of penicillin, new beta-lactam ring synthesis, synthesis of carbapenem derivatives including microbial hydration and stereo-specific 1 beta-methyl carbapenem synthesis.

Eur J Biochem, 2000 Dec, 267(23), 6740 - 52
A modular esterase from Penicillium funiculosum which releases ferulic acid from plant cell walls and binds crystalline cellulose contains a carbohydrate binding module; Kroon PA et al.; An esterase was isolated from cultures of the filamentous fungus Penicillium funiculosum grown on sugar beet pulp as the sole carbon source . The enzyme (ferulic acid esterase B, FAEB) was shown to be a cinnamoyl esterase (CE), efficiently releasing hydroxycinnamic acids from synthetic ester substrates and plant cell walls, and bound strongly to microcrystalline cellulose . A gene fragment was obtained by PCR using partial amino-acid sequences obtained from the pure enzyme and used to a probe a P . funiculosum genomic DNA library . A clone containing a 1120-bp ORF, faeB, was obtained which encoded a putative 353-residue preprotein including an 18-residue signal peptide, which when expressed in Eschericia coli produced CE activity . Northern analysis showed that transcription of faeB was tightly regulated, being stimulated by growth of the fungus on sugar beet pulp but inhibited by free glucose . The faeB promoter sequence contains putative motifs for binding an activator protein, XLNR, and a carbon catabolite repressor protein, CREA . FAEB was comprised of two distinct domains separated by a 20 residue Thr/Ser/Pro linker region . The N-terminal domain comprised 276 amino acids, contained a G-X-S-X-G motif typical of serine esterases, and was shown to be a member of a family comprising serine esterases, including microbial acetyl xylan esterases, poly (3-hydroxyalkanoate) depolymerases and CEs, and proteins of unknown function from Mycobacterium spp . and plants . The C-terminal domain comprised 39 amino acids and closely resembled the family 1 cellulose binding carbohydrate-binding modules (CBM) of fungal glycosyl hydrolases . This is the first report of a fungal CE with a CBM.

Plant Physiol, 2000 Nov, 124(3), 1373 - 80
How alfalfa root hairs discriminate between Nod factors and oligochitin elicitors; Felle HH et al.; Using ion-selective microelectrodes, the problem of how signals coming from symbiotic partners or from potential microbial intruders are distinguished was investigated on root hairs of alfalfa (Medicago sativa) . The Nod factor, NodRm-IV(C16:2,S), was used to trigger the symbiotic signal and (GlcNAc)(8) was selected from (GlcNAc)(4-8), to elicit defense-related reactions . To both compounds, root hairs responded with initial transient depolarizations and alkalinizations, which were followed by a hyperpolarization and external acidification in the presence of (GlcNAc)(8) . We propose that alfalfa recognizes tetrameric Nod factors and N-acetylchitooligosaccharides (n = 4-8) with separate perception sites: (a) (GlcNAc)(4) and (GlcNAc)(6) reduced the depolarization response to (GlcNAc)(8), but not to NodRm-IV(C16:2, S); and (b) depolarization and external alkalization were enhanced when NodRm-IV(C16:2,S) and (GlcNAc)(8) were added jointly without preincubation . We suggest further that changes in cytosolic pH and Ca(2+) are key events in the transduction, as well as in the discrimination of signals leading to symbiotic responses or defense-related reactions . To (GlcNAc)(8), cells responded with a cytosolic acidification, and they responded to NodRm-IV(C16:2,S) with a sustained alkalinization . When both agents were added jointly, the cytosol first alkalized and then acidified . (GlcNAc)(8) and NodRm-IV(C16:2,S) transiently increased cytosolic Ca(2+) activity, whereby the response to (GlcNAc)(8) exceeded the one to NodRm-IV(C16:2,S) by at least a factor of two.

Plant Physiol, 2000 Nov, 124(3), 1169 - 80
Sensing of osmotic pressure changes in tomato cells; Felix G et al.; Cells of tomato (Lycopersicon esculentum) growing in suspension gradually depleted their culture medium and caused a steady decrease in its osmolality . When confronted with a sudden change in medium osmolality (a hypo-osmotic or hyperosmotic shock), respectively, these cells responded with volume changes and stress symptoms such as rapid extracellular alkalinization, efflux of K(+)-ions, and induction of 1-aminocyclopropane-1-carboxylate synthase acid, the key enzyme of ethylene biosynthesis . This array of stress symptoms is well known from cultured plant cells treated with microbial elicitors . Compared with elicitor treatment, induction of responses by hyperosmotic shock was slow and occurred only after increases of approximately 200,000 Pa in osmotic pressure . In contrast, hypo-osmotic shock induced responses without measurable lag and faster than elicitor treatments . Measurable medium alkalinization was induced when medium osmolality was reduced by as little as approximately 10 mosmol, a change corresponding to only approximately 0.2 bar in osmotic pressure . Like treatment with elicitors, hypo-osmotic shock induced specific changes in protein phosphorylations as demonstrated by in vivo labeling with {(33)P}orthophosphate . Exposure of cells to consecutive up- and down-shifts in medium osmolality showed that sensing of osmotic changes occurred within seconds, whereas adaptation to new osmotic conditions proceeded over hours . In conclusion, suspension-cultured plant cells display rapid, easily measurable macroscopic responses to osmotic shock and provide an interesting model system to study osmoregulation, a key process in plant growth and development.

Microb Ecol, 2000 Aug, 40(3), 238 - 249
Effect of Supplemental Electron Donors on the Microbial Reduction of Fe(III), Sulfate, and CO(2) in Coal Mining-Impacted Freshwater Lake Sediments; Kusel K et al.; In acidic mining-impacted lake sediments, the microbial reduction of Fe(III) is the dominant electron-accepting process, whereas the reduction of sulfate seems to be restricted to a narrow sediment zone of elevated pH and lower amounts of total and reactive iron . To evaluate the microbial heterogeneity and the commensal interactions of the microbial community, the flow of supplemental carbon and reductant was evaluated in four different zones of the sediment in anoxic microcosms at the in situ temperature of 12 degrees C . Substrate consumption, product formation, and the potential to reduce Fe(III) and sulfate were similar with both upper and lower sediment zones . In the upper acidic iron-rich sediment zone, the rate of Fe(II) formation 204 nmol ml(-1) d(-1) was enhanced to 833 nmol ml(-1) d(-1) and 462 nmol ml(-1) d(-1) by supplemental glucose and H(2), respectively . Supplemental lactate and acetate were not consumed under acidic conditions and decreased the rate of Fe(II) formation to 130 nmol ml(-1) d(-1) and 52 nmol ml(-1) d(-1), respectively . When the pH of the upper sediment increased above pH 5, acetate-dependent reduction of sulfate was initiated even though the pool of Fe(III) was not depleted . In deeper sediment zones with elevated pH, the rapid consumption of acetate was always coincident to a decrease in the concentration of sulfate and soluble Fe(II), indicating the formation of Fe(II) sulfides . Although the reduction of Fe(III) was still an ongoing process in deeper sediment zones, the formation of Fe(II) was only slightly enhanced by the consumption of glucose or cellobiose, but not by H(2) or acetate . H(2)-utilizing acetogens seemed to be involved in the consumption of H(2) . These collective results indicated (i) that the reduction of Fe(III) predominated over the reduction of sulfate as long as the sediment remained acidic and carbon-limited, and (ii) that the sulfate-reducing microbiota in this heterogeneous sediment were better adapted to the geochemical gradients present than were other neutrophilic dissimilatory Fe(III) reducers.

Microb Ecol, 2000 Aug, 40(3), 209 - 222
Succession of Protists on Estuarine Aggregates; Worner U et al.; Colonization by and succession of bacteria and bacterivorous protists on laboratory-made aggregates were determined over a period of 14 days during winter and spring in 1997 . Aggregates were generated from natural water from the limnetic zone of the Elbe Estuary using a tilting tube roller system . Within 1 h after the beginning of the experiments, macroaggregates started to form . Aggregates reached a maximum size of 1 mm with a tendency toward large sizes at the end of the experiment after the 10th day . On the first day, high bacterial densities of more than 10(9) cells ml(-1) were detected within the aggregates . The abundances of flagellates and ciliates within aggregates were also two or three orders of magnitude higher than in the surrounding water . Densities of aggregate associated organisms are comparable to those occuring in sediments . The first protistan colonizers on the aggregates were small heterotrophic flagellates, such as choanoflagellates and small euglenids . Later, beginning on the 4th day, small sarcodines and ciliates became abundant . The most abundant ciliates associated with aggregates were small species of the Hypotrichia, Cyrtophorida, and Hymenostomata . After 9 days, large omnivorous and carnivorous ciliates, such as large members of the Hypotrichia and the Pleurostomatida, occurred . In spring, large heterotrophic flagellates and amebae also appeared at this time . These findings indicated the existence of a succession of protists on newly formed aggregates and a microbial food net within the aggregates based on bacterial production . Additionally, most of the species observed during this study were adapted for living on surfaces . In natural environments they are more common in benthic than in pelagic environments . For them, aggregates are havens in the water column comparable to sediment communities.

Steroids, 2000 Dec, 65(12), 863 - 70
Microbial transformation of 3-hydroxy-5,6-cyclopropanocholestanes--an alternative route to 6-methylsteroids; Yan JL et al.; Incubation of 3beta-hydroxy-5,6alpha-cyclopropano-5alpha-cholestane (4), 3beta-hydroxy-5,6beta-cyclopropano-5beta-cholestane (5), and 3beta-hydroxy-5,6alpha-cyclopropano-5alpha-cholest-7-e ne (6) with Mycobacterium sp . (NRRL B-3805) gave a mixture of side chain cleaved 17-keto steroids as the major products in 52, 57, and 69% yields, respectively . Among these 17-keto steroids, the cyclopropyl ring eliminated product, androst-4-ene-3,17-dione (9), was isolated in 6, 4, and 8% yields, respectively . A cyclopropyl ring migration product, 6alpha,7alpha-cyclopropanoandrost-4-ene-3,17-dione (16), was isolated from the incubation mixture of 6 in 4% yield, also 10% yield of 16 was obtained when 5, 6alpha-cyclopropano-5alpha-androst-7-ene-3,17-dione (12) was incubated . The cyclopropyl ring opening and subsequent reduction followed by oxidation of the two major biotransformation products, 5, 6beta-cyclopropano-5beta-androsta-3,17-dione (10) and 5, 6alpha-cyclopropano-5alpha-androsta-3,17-dione (7), gave 6beta- and 6alpha-methylandrost-4-ene-3,17-dione in 60, and 45% yields, respectively.

J Nat Prod, 2000 Oct, 63(10), 1400 - 5
Available pathways database (APD): an essential resource for combinatorial biology; Pirrung MC et al.; A relational database, the Available Pathways Database (APD), has been constructed of microbial natural products, their producing strains, and their biosynthetic pathways . The database allows the ready selection of donor strains for combinatorial biology experiments . It provides the same type of resource for combinatorial biology as the Available Chemicals Directory (ACD) does for combinatorial chemical library generation . Its cataloging ability can also provide insight into novel aspects of biosynthetic routes . In particular, no 10-unit Type I polyketides were found in the compilation of this edition of the APD (Version I).

J Hosp Infect, 2000 Nov, 46(3), 210 - 5
Endoscope washers--a protocol for their use; Marchetti MG et al.; A protocol for the disinfection of gastroduodenoscopes, retrograde cholangiopancreatography endoscopes and colonoscopes using endoscope washers is described . The process recommends initial manual washing with a disinfectant containing didecyldimethylammonium chloride, surfactants and enzymes, a second washing in the endoscope washer using a detergent associated with a bacteriostatic, fungistatic substance (benzoisothiazolone) and finally the use of a 2% glutaraldehyde product buffered at pH6 . After treatment with 2% Steranios added to the washer, less than 1 micro-organism/ml liquid was found in the following units: 83% of the colonoscopes, 83% of the oesophagogastroduodenoscopes, 83% of the main channels of the retrograde cholangiopancreatography endoscopes and 75% of the auxiliary channels of the latter instruments . In 14% of the colonoscopes, 42% of the gastroduodenoscopes, 42% of the main and 50% of auxiliary channels in the retrograde cholangiopancreatography endoscopes there were no signs of microbial growth in the wash liquid . The results obtained indicated that this protocol allowed adequate disinfection of the endoscope channels, structurally the most difficult part of the instrument to disinfect . Emphasis is given to the degree by which instrument contamination can increase during overnight storage, suggesting that endoscopes need to be submitted to further disinfection after overnight storage . Moreover, the water flowing into the washers can also cause instrument recontamination, particularly during the final rinses . Therefore, to better safeguard the health of patients undergoing endoscopy, special care must be taken to maintain the filters and disinfect the washers themselves, no matter how effective the disinfection protocol used may be .

Org Lett, 2000 Nov 16, 2(23), 3583 - 6
Chemoenzymatic synthesis of (+)-aspicilin from chlorobenzene; Banwell MG et al.; The enantiomerically pure cis-1,2-dihydrocatechol 2, which is obtained by microbial oxidation of chlorobenzene, has been converted, via intermediate 3, into the natural product (+)-aspicilin (1).

Eur J Haematol, 2000 Oct, 65(4), 215 - 20
Circulating endothelial cells in vascular disorders: new insights into an old concept; Dignat-George F et al.; The endothelial contribution to vascular disorders has been widely documented in experimental models . However, its implication in human pathology is difficult to investigate, owing to the paucity of noninvasive methods and of specific endothelial markers . The enumeration of circulating endothelial cells (CEC) released in peripheral blood after vascular injury represents a direct exploration of the endothelium . For this purpose, we have produced a monoclonal antibody (S-Endo 1), which recognizes CD 146, a molecule expressed on all types of human endothelial cells but absent from haemopoietic cells . Using this antibody, we have designed a specific and sensitive immunocapture test, which allowed us to detect high numbers of CEC in thrombotic, infectious or immunological disorders, while CEC were found to be very rare ( < 3/ml) in normal subjects . This quantitative approach using CEC might prove useful as a marker of vascular wall injury . Their enumeration is of interest in the clinical follow-up of vascular disorders, in the evaluation of therapeutic effectiveness or in the direct diagnosis of infectious diseases involving intra-endothelial microbial agents . Furthermore, an immunological and/or functional study of CEC could allow one to assess their procoagulant and proadhesive properties, as well as their viability, opening new perspectives for CEC investigation in vascular pathology.

Presse Med, 2000 Oct 7, 29(29), 1630 - 3
{Infection and cancer: general information and specific questions}; Viot M et al.; The main risk factors of infectious complications in cancer patients result from immune deficiency more or less related to cancer . Prognosis is related to the type and grade of the underlying disease . Prospective studies should be conducted to update data on the frequency of infections, morbidity and mortality (expert agreement) . Prospective studies are needed to follow the epidemiology in cancer patients, particularly in neutropenic patients (expert agreement) . Prospective studies should be conducted to determine prognosis factors allowing precise recognition of "low-risk" neutropenic patients with fever who could benefit from home care (expert agreement) . When infection is suspected, the first criterion determining the therapeutic attitude concern signs of gravity requiring emergency care (septic shock) . Beyond this situation, the first criterion determining the therapeutic attitude is the severity of the neutropenia . Microbial diagnosis is essential for initiating and later adapting anti-infectious treatment as well as for assessing efficacy.

Semin Arthritis Rheum, 2000 Oct, 30(2), 111 - 20
Genetic susceptibility and the link between cat exposure and rheumatoid arthritis; Penglis PS et al.; OBJECTIVES: Rheumatoid arthritis (RA) is a chronic inflammatory disease for which immunogenetic susceptibility factors have been defined . In a recent case control study, it was shown that a prior intimate relationship with pet cats or budgerigars confers risk for subsequent development of RA after a period of latency . Pets are a potential reservoir for putative microbial agents that could be a stimulus for chronic inflammation subject to the influence of immunogenetic factors . Therefore, a study was undertaken to determine whether the presence of HLA-DRB1 alleles bearing the RA susceptibility motif influenced risk for RA associated with prior exposure to pets . METHODS: Blood samples were obtained from available RA patients and their case controls who had participated in the prior epidemiologic study . DR and DQ genotypes were determined by sequence analysis of oligonucleotides amplified from the DRB1 and DQB1 genes by polymerase chain reactions (PCR) . Subjects were segregated according to pet exposure (as determined previously) and genotype for statistical analyses . RESULTS: The odds ratio (OR) for prepubertal exposure to cats and RA in available subjects irrespective of DRB1 genotype was 4.2 (CI, 2.1 to 8.5; P<.00002) . The OR between prior exposure to cats and RA in subjects with the RA susceptibility genotype DRB1 *0401 and *0404 was 5.8 (CI, 1.4 to 26; P<.02) and >24 (CI, 1.6 to 813; P<.01), respectively . In subjects with the genotype DRB1 *1501, the association between RA and prior cat exposure was OR 8.4 (CI, 1.7 to 45; P<.01) . No significant association between RA and pet exposure was found in patients selected according to other genotypes . The association between RA and the recognized HLA-DR susceptibility motif was slightly stronger in subjects with a history of intimate cat exposure (OR 4.7 {CI, 1.5 to 14.8}, P<.005) than subjects without prior intimate exposure (OR 3.3 {CI; 1.2 to 9.3}, P<.02) . In the small number of subjects who had reported an intimate association with pet birds, no influence of DR genotype on risk for RA was discerned . CONCLUSIONS: Risk for RA associated with prior intimate exposure to cats is concentrated in subjects with the RA-susceptibility conferring genotypes DRB1 *0401 and *0404 . The findings suggest an interaction between an environmen-tal agent associated with pet cats and certain RA susceptibility-conferring DR genotypes . The risk for RA associated with intimate cat exposure also was significant in subjects with DRB1*1501, a genotype not otherwise associated with RA, but which shares with known RA susceptibility-bearing alleles the presence of an electropositive pocket (Pocket 4) in the DR peptide binding groove.

Digestion, 2000, 62(4), 255 - 61
Obstructive jaundice, bacterial translocation and interdigestive small-bowel motility in rats; Nieuwenhuijs VB et al.; BACKGROUND/AIMS: Translocation of gut bacteria occurs in obstructive jaundice, the underlying mechanisms are unclear . We designed this experimental study to investigate the association between interdigestive motility and the pathogenesis of bacterial translocation during biliary obstruction . METHODS: Rats were fitted with jejunal myoelectrodes for the measurement of the interdigestive migrating motor complex (MMC) and with two cannulas in the proximal common bile duct (CBD) for exteriorization of biliary flow . This allowed measurement of MMCs under control conditions with an intact enterohepatic circulation and during 3 days of CBD obstruction without surgical intervention . Mesenteric lymph nodes, liver, spleen and segments of the duodenum, the jejunum and the caecum were removed for microbial culturing . RESULTS: The MMC cycle length increased from 17.3 min before CBD obstruction to 31.9, 34.1, and 25.3 min on days 1, 2 and 3, respectively, after CBD obstruction (p < 0.05 for all days) . Bacterial levels in the jejunum were significantly higher in CBD-obstructed rats than in control rats . The translocation incidence was significantly higher in rats with CBD obstruction (6/8) than in control rats (1/8) . The bacterial levels in the jejunum correlated significantly with the MMC cycle length (r = 0.60, p <0.05) . CONCLUSION: Experimental biliary obstruction is associated with disturbance of MMCs, small-bowel bacterial overgrowth and increased bacterial translocation .

Immunity, 2000 Oct, 13(4), 453 - 62
CD40 triggering of heterodimeric IL-12 p70 production by dendritic cells in vivo requires a microbial priming signal; Schulz O et al.; CD40 ligation triggers IL-12 production by dendritic cells (DC) in vitro . Here, we demonstrate that CD40 cross-linking alone is not sufficient to induce IL-12 production by DC in vivo . Indeed, resting DC make neither the IL-12 p35 nor IL-12 p40 subunits and express only low levels of CD40 . Nevertheless, after DC activation by microbial stimuli that primarily upregulate IL-12 p40 and augment CD40 expression, CD40 ligation induces a significant increase in IL-12 p35 and IL-12 p70 heterodimer production . Similarly, IL-12 p70 is produced during T cell activation in the presence but not in the absence of microbial stimuli . Thus, production of bioactive IL-12 by DC can be amplified by T cell-derived signals but must be initiated by innate signals.

Eur J Immunol, 2000 Oct, 30(10), 2881 - 7
Enhanced immunogenicity of aldehyde-bearing antigens: a possible link between innate and adaptive immunity; Allison ME et al.; Innate immunity directs the adaptive immune response by identifying antigens that are associated with infectious agents . Although some microbial antigens can be recognized by innate immune receptors, most cannot, and these require identification by some other means . The introduction of aldehydes into antigens by glycolaldehyde, which can be produced by activated neutrophils reacting with serine, or by the oxidation of an N-linked oligosaccharide with NaIO4, enhances by several orders of magnitude their immunogenicity in mice . The augmented immunogenicity requires the presence of an aldehyde on the antigen, and is not dependent on protein aggregation . An in vitro correlate of augmented immunogenicity is the enhanced presentation of glycolaldehyde-modified antigen to T cells by macrophages and bone marrow-derived dendritic cells . The potential clinical importance of this form of antigen modification is twofold: glycolaldehyde renders a model self antigen immunogenic, and it converts a relatively non-immunogenic malaria antigen, merozoite surface protein-1, into an effective immunogen . Thus, the tagging of antigens by the addition of aldehydes, which may be an innate immune mechanism to facilitate their recognition by the adaptive immune system, may have a role in the genesis of autoimmunity and the development of vaccines.

Lett Appl Microbiol, 2000 Oct, 31(4), 270 - 3
Selection of Saccharomyces cerevisiae strains for use as a microbial feed additive; Agarwal N et al.; Saccharomyces cerevisiae ITCCF 2094, NCIM 3052, 1031, 1032, NCDC 42, 45, 47, 49 and 50 were screened for their tolerance to pH 2.0-7.0, various concentrations (0.00, 0.10, 0.25 0.50 and 1.0%) of a mixture of acetic, propionic and butyric acids (70:20:10), and bile salts (0.00, 0.30, 0.60 and 0.90%) . Low pH (2.0-4.0) and addition of organic acids or bile salts in the medium inhibited the growth of all the strains tested, but the percentage of inhibition was variable in the different strains of yeast . Two of the strains showing maximum tolerance, 42 and 49, were further tested for in vitro dry matter degradability (IVDMD) using green berseem, wheat straw and oat hay as substrates . Saccharomyces cerevisiae 49 enhanced the IVDMD of berseem and wheat straw whereas S . cerevisiae 42 was ineffective . Based on the results of the present experiment, S . cerevisiae NCDC 49 can be considered as the best strain which might tolerate the adverse conditions in the gastrointestinal tract when used as a live microbial feed supplement in the diet of the animals.

Biodegradation, 1999, 10(6), 443 - 7
Removal of mercury from mercury-contaminated sediments using a combined method of chemical leaching and volatilization of mercury by bacteria; Nakamura K et al.; A method for the removal of mercury sulfide from mercury-contaminated sediments was developed, which consists of chemical leaching and volatilization of mercury by bacteria . More than 85% of the mercury in sediment containing 0.11-37.4 mg/kg of mercury was efficiently extracted with 3 M HCl and 74 mM FeCl3 . Subsequent volatilization by bacteria resulted in the removal of 62.9-75.1% of mercury from mercury-contaminated Minamata Bay sediments . Methylmercury was also eliminated from soil at a high efficiency . Thus, this combined method of chemical and microbial treatments could be used for efficient removal of both organic and inorganic mercurials from natural sediments.

Biodegradation, 1999, 10(6), 405 - 14
Behavior of chemical contaminants under controlled redox conditions in an artificial sequential soil column system and in batch cultures; Nay M et al.; A leachate pollution plume was simulated in a sequential soil column system (SSCS), defined by a longitudinal redox cascade from methanogenic, sulfate-, nitrate-reducing to aerobic conditions . A mixture of contaminants, including compounds such as perchloroethene (PER), 1,1-dichloroethene (DCE), 1,4-dichlorobenzene (DCB), 2-nitrophenol (NP), and benzene (BEN) was supplied to the methanogenic column; their conversion was followed during passage through the SSCS . After establishment of a steady state, parameters were changed to simulate changes in the environment and to follow the reactions of the system . Acetate, initially introduced as an anaerobically easy degradable carbon source, was first omitted and later added again to the system . Furthermore, the medium flow rate and by that the load of the test compounds was increased 4 fold . Changes transiently increased the residual concentrations of most of the xenobiotics . Within seven months after acetate omission, turnover of contaminants had almost returned to the original steady state . Flow and load increase resulted in increased transformation of compounds such as PER, NP, and BEN . After readdition of acetate, the turnover of most of the xenobiotics increased within the next five weeks . Microbial populations for the anaerobic batch cultures, obtained from the specific columns, were used to investigate metabolites formed and conversion kinetics of the xenobiotics . Almost the same conversion pattern was found in batches and columns, except no transformation of DCE and DCB was detected in the SSCS, though both compounds were converted in batch cultures.

FEBS Lett, 2000 Nov 3, 484(2), 65 - 8
Kinetic mechanism of NADH-enoyl-ACP reductase from Brassica napus; Fawcett T et al.; Enoyl-ACP reductase, a component of fatty acid synthase, is a target for anti-microbial agents and herbicides . Here we demonstrate the kinetic mechanism to be a compulsory-order ternary complex with NADH binding before the acyl substrate . Matrix-assisted laser desorption ionisation mass spectrometry analysis of enzymatically and synthesised crotonyl-ACP substrate showed the former to contain a single acyl group, whereas the latter contained up to four additional crotonylations . The use of authentic crotonyl-ACP will be important in future kinetic and crystallographic studies.

J Immunol, 2000 Nov 15, 165(10), 5637 - 45
Priming by microbial antigens from the intestinal flora determines the ability of CD4+ T cells to rapidly secrete IL-4 in BALB/c mice infected with Leishmania major; Julia V et al.; Infection of BALB/c mice with Leishmania major results in the rapid accumulation of IL-4 transcripts within CD4(+) T cells that react to the parasite Leishmania homologue of mammalian RACK1 (LACK) Ag . Because memory/effector cells secrete IL-4 more rapidly than naive cells, we sought to analyze the phenotype of these lymphocytes before infection . Indeed, a fraction of LACK-specific CD4(+) T cells expressed a typical CD62 ligand(low)CD44(high)CD45RB(low) phenotype in uninfected mice . LACK-specific T cells were primed in gut-associated lymphoid tissues by cross-reactive microbial Ags as demonstrated by their reactivity with bacterial extracts and by the ability of APCs from the mesenteric LN of BALB/c mice to induce their proliferation . Also, mice in which the digestive tract has been decontaminated exhibited a reduced proportion of LACK-specific T cells expressing a memory/effector phenotype and did not exhibit the early accumulation of IL-4 transcripts induced by L . major . Thus, LACK-specific T cells represent a subset of CD4(+) T cells which have acquired the ability to rapidly secrete IL-4 as the result of their priming by cross-reactive microbial Ags . Tracking the fate of these cells may provide information about the regulation of cell-mediated immune responses to gut Ags in physiological and pathological situations.

Anal Biochem, 2000 Nov 15, 286(2), 198 - 205
Stable expression of varied levels of inducible nitric oxide synthase in primary cultures of endothelial cells; Zhang B et al.; Nitric oxide (NO*), generated by nitric oxide synthase (NOS II) from immunostimulated cells during infection, plays an important role in host immune defense against microbial invasion . The impact of different rates of NO* production on host cell function has not been defined . Herein, we describe the development of a method to express varied levels of murine NOS II in bovine pulmonary artery endothelial cells . A retroviral vector (pMFGSNOS) encoding NOS II was used to transduce primary cultures of endothelial cells . Bovine endothelial cells were susceptible to this transduction and up to 18% of the cells expressed immunodetectable murine NOS II . The NOS II-transduced endothelial cells were cultured on the three-dimensional matrix, Gelfoam, for 8-10 days . Stable expression of NOS II was assessed by measuring nitrite accumulation in media every 2 days . By day 10, endothelial cells on Gelfoam were found to secrete NO* at a rate exceeding 1.0 microM/h/10(6) cells, concomitant with an enhanced level of NOS II activity . Argininosuccinate synthetase, a key enzyme in the metabolism of l-citrulline to l-arginine, increased as well, perhaps in response to dimunition of the intracellular arginine pool corresponding to the observed high output of NO* . In spite of the continuous flux of NO*, endothelial cell viability was not effected . This system provides the opportunity to assess the impact of different levels of sustained NO* production on endothelial cell physiology .

J Endotoxin Res, 2000, 6(1), 51 - 6
Limits of a deletion spanning Tlr4 in C57BL/10ScCr mice; Poltorak A et al.; Proceeding from our observation that LPS-unresponsive mice of the strain C57BL/10ScCr mice fail to express the Tlr4 gene {Poltorak A, He X . Smirnova I et al . Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene . Science 1998; 282: 2085}, we have defined the exact limits of a deletion encompassing Tlr4 in the C57BL/10ScCr genome . The deletion removes 74723 bp of DNA, with reference to the control strain 129/J (from which the complete sequence of the Tlr4 locus was obtained) . There is no inserted element, and no re-arrangement of the chromosome (e.g . inversion or translocation) in the immediate region of Tlr4; the deletion removes only one recognizable gene . Hence, other immunological anomalies that have been identified in C57BL/10ScCr mice (a non-healing phenotype in Leishmania inoculation and failure to produce interferon-gamma in response to numerous microbial infections) must be ascribed to one of two causes . Mutation(s) at other loci may be responsible for these defects . Alternatively, Tlr4 locus deletion may have phenotypic consequences that exceed the well known blockade of LPS signal transduction.

Surg Today, 2000, 30(10), 875 - 8
Changes in T-cell receptor subsets after cardiac surgery in children; Yamaguchi T et al.; T cells are divided into two subsets, alphabeta and gammadelta, according to the T-cell receptor (TCR) expressed . Recent findings indicate that gammadelta T cells serve as the first defense against microbial pathogens, and represent a link between innate and acquired immunity . We conducted a study to investigate the changes in circulating TCR subsets after cardiac surgery in children . Blood samples from 24 children who underwent cardiac surgery with cardiopulmonary bypass (CPB) were collected serially to analyze TCR subsets by flow cytometry . The alphabeta T cells reached a nadir on postoperative day (POD) 1, but recovered to pre-CPB levels on POD 3 . On the other hand, the gammadelta T cells decreased after CPB and did not recover to pre-CPB levels even after POD 7 . The alphabeta/gammadelta T-cell ratio was increased after POD 3 . In children, gammadelta T cells recover more slowly than alphabeta T cells after cardiac surgery . These changes in TCR subsets may contribute to postoperative immunosuppression.

Sheng Wu Gong Cheng Xue Bao, 2000 May, 16(3), 301 - 3
{Molecular modification and artificial evolution of microbial enzymes}; Wang ZX et al.; There exists a remarkably progress in molecular modification and artificial evolution of microbial enzymes in recent years . Error-prone polymerase chain reaction and DNA shuffling have been developed and successfully applied and a lot of startling achievements have been obtained in artificial evolution of enzymes.

J Biol Chem, 2001 Jan 26, 276(4), 2551 - 4 Epub 2000 Oct 31.
Human Nod1 confers responsiveness to bacterial lipopolysaccharides; Inohara N et al.; The immune response to microbial pathogens is initiated by recognition of specific pathogen components by host cells both at the cell surface and in the cytosol . While the response triggered by pathogen products at the surface of immune cells is well characterized, that initiated in the cytosol is poorly understood . Nod1 is a member of a growing family of intracellular proteins with structural homology to apoptosis regulators Apaf-1/Ced-4 and a class of plant disease-resistant gene products . Here we show that bacterial lipopolysaccharides, but not other pathogen components tested, induced TLR4- and MyD88-independent NF-kappaB activation in human embryonic kidney 293T cells expressing trace amounts of Nod1 . Nod2, another Nod family member, also conferred responsiveness to bacterial components but with a response pattern different from that observed with Nod1 . As it was reported for plant disease-resistant R proteins, the leucine-rich repeats of Nod1 and Nod2 were required for lipopolysaccharide-induced NF-kappaB activation . A lipopolysaccharide binding activity could be specifically coimmunopurified with Nod1 from cytosolic extracts . These observations suggest that Nod1 and Nod2 are mammalian counterparts of plant disease-resistant gene products that may function as cytosolic receptors for pathogen components derived from invading bacteria.

Extremophiles, 2000 Oct, 4(5), 259 - 65
Comparisons of the polar lipid and pigment profiles of two solar salterns located in Newark, California, USA, and Eilat, Israel; Litchfield CD et al.; The whole community pigments and lipids have been examined during a 5-year period in two commercial solar salterns located in the United States and in Israel . There were significant differences in the complexity of the lipid and pigment patterns within the California saltern system, and these differences were not consistent over the sampling period despite examination of ponds with the same salinity . The solar saltern system in Eilat, Israel, showed greater consistency during this sampling period and compared directly with previous studies . The complexity of the saltern in Newark, California, could be explained on the basis of the prevailing weather conditions (cooler and more rainfall) and the nutrient-enriched source water . The Eilat saltern, however, has an oligotrophic water source and has a considerably warmer and drier climate . This difference resulted in more diverse and more complex pigment and lipid patterns and presumably microbial populations in the Newark, California, plant than in the saltern in Eilat, Israel.

Chemosphere, 2000 Aug, 41(3), 437 - 46
Volatile metabolites from mold growth on building materials and synthetic media; Wilkins K et al.; Mold species which were isolated from damp buildings were grown on sterile building materials and some synthetic media in order to study the microbial volatile organic compounds produced . Patterns of the microbial volatile organic compounds (MVOC) were very media dependent but media which favor terpene biosynthesis may give patterns unique enough for identification of dominant indoor molds.

Chemosphere, 2000 Nov, 41(9), 1457 - 62
Anaerobic transformation of chlorophenols in methanogenic sludge unexposed to chlorophenols; Takeuchi R et al.; Transformation of all 19 chlorophenol (CP) isomers was investigated in a laboratory anaerobic methanogenic sludge that had not been exposed to synthetic chemicals . Concentration of CP was analyzed over time to calculate disappearance rate constants using first-order reaction kinetics and all possible CP degradation pathways were estimated . The rate constants ranged between 0.46 x 10(-3) and 0.161 day(-1) . CPs were transformed via dechlorination . The chlorine atom at the ortho-position was the most easily dechlorinated, whereas dechlorination rate at the para-position was lowest . The overall pathways of CP transformation were much less diverse than that we previously found for contaminated sediment . The Dolfing hypothesis of microbial selection of the most thermodynamically favorable pathways was not applicable for CP transformation in this study as well as previous study performed by our group.

Proc West Pharmacol Soc, 2000, 43, 29 - 30
Fungi test for evaluation of new antiandrogenic molecule; Cabeza M et al.; Antiandrogens have high impact in medicine as they are compounds capable of decreasing the effect of benign prostatic hyperplasia, cancer, and other androgen-dependent diseases that affect a large percentage of the male population in the world . {figure: see text} Dihydrotestosterone, a 5 alpha-reductase metabolite of testosterone, has been implicated as the responsible factor of these androgen abnormalities . This fact indicates that the logical step in the treatment of these diseases is the inhibition of this enzyme activity using molecules that compete selectively with its natural substrates . The study of the pharmacological effect of antiandrogens requires specific animal models using a large number of laboratory animals . On basis of this concept, presently there is a tendency to eliminate studies with animals . In this paper we report a new method for antiandrogenic evaluation using microbial transformations.

Appl Environ Microbiol, 2000 Nov, 66(11), 4962 - 71
Comparison of acid mine drainage microbial communities in physically and geochemically distinct ecosystems; Bond PL et al.; This study presents population analyses of microbial communities inhabiting a site of extreme acid mine drainage (AMD) production . The site is the inactive underground Richmond mine at Iron Mountain, Calif., where the weathering of a massive sulfide ore body (mostly pyrite) produces solutions with pHs of approximately 0.5 to approximately 1.0 . Here we used a suite of oligonucleotide probes, designed from molecular data recently acquired from the site, to analyze a number of microbial environments by fluorescent in situ hybridization . Microbial-community analyses were correlated with geochemical and mineralogical data from those environments . The environments investigated were within the ore body and thus at the site of pyrite dissolution, as opposed to environments that occur downstream of the dissolution . Few organism types, as defined by the specificities of the oligonucleotide probes, dominated the microbial communities . The majority of the dominant organisms detected were newly discovered or organisms only recently associated with acid-leaching environments . "Ferroplasma" spp . were detected in many of the communities and were particularly dominant in environments of lowest pH and highest ionic strength . Leptospirillum spp . were also detected in many slime and pyrite-dominated environments . In samples of an unusual subaerial slime, a new uncultured Leptospirillum sp . dominated . Sulfobacillus spp . were detected as a prominent inhabitant in warmer ( approximately 43 degrees C) environments . The information gathered here is critical for determining organisms important to AMD production at Iron Mountain and for directing future studies of this process . The findings presented here also have relevance to the microbiology of industrial bioleaching and to the understanding of geochemical iron and sulfur cycles.

Appl Environ Microbiol, 2000 Nov, 66(11), 4945 - 53
PCR bias in ecological analysis: a case study for quantitative Taq nuclease assays in analyses of microbial communities; Becker S et al.; Succession of ecotypes, physiologically diverse strains with negligible rRNA sequence divergence, may explain the dominance of small, red-pigmented (phycoerythrin-rich) cyanobacteria in the autotrophic picoplankton of deep lakes (C . Postius and A . Ernst, Arch . Microbiol . 172:69-75, 1999) . In order to test this hypothesis, it is necessary to determine the abundance of specific ecotypes or genotypes in a mixed background of phylogenetically similar organisms . In this study, we examined the performance of Taq nuclease assays (TNAs), PCR-based assays in which the amount of an amplicon is monitored by hydrolysis of a labeled oligonucleotide (TaqMan probe) when hybridized to the amplicon . High accuracy and a 7-order detection range made the real-time TNA superior to the corresponding end point technique . However, in samples containing mixtures of homologous target sequences, quantification can be biased due to limited specificity of PCR primers and probe oligonucleotides and due to accumulation of amplicons that are not detected by the TaqMan probe . A decrease in reaction efficiency, which can be recognized by direct monitoring of amplification, provides experimental evidence for the presence of such a problem and emphasizes the need for real-time technology in quantitative PCR . Use of specific primers and probes and control of amplification efficiency allow correct quantification of target DNA in the presence of an up to 10(4)-fold excess of phylogenetically similar DNA and of an up to 10(7)-fold excess of dissimilar DNA.

Appl Environ Microbiol, 2000 Nov, 66(11), 4849 - 53
Fate of selenate and selenite metabolized by Rhodobacter sphaeroides; Van Fleet-Stalder V et al.; Cultures of a purple nonsulfur bacterium, Rhodobacter sphaeroides, amended with approximately 1 or approximately 100 ppm selenate or selenite, were grown phototrophically to stationary phase . Analyses of culture headspace, separated cells, and filtered culture supernatant were carried out using gas chromatography, X-ray absorption spectroscopy, and inductively coupled plasma spectroscopy-mass spectrometry, respectively . While selenium-amended cultures showed much higher amounts of SeO(3)(2-) bioconversion than did analogous selenate experiments (94% uptake for SeO(3)(2-) as compared to 9.6% for SeO(4)(2-)-amended cultures from 100-ppm solutions), the chemical forms of selenium in the microbial cells were not very different except at exposure to high concentrations of selenite . Volatilization accounted for only a very small portion of the accumulated selenium; most was present in organic forms and the red elemental form.

Appl Environ Microbiol, 2000 Nov, 66(11), 4790 - 7
Effect of temperature on carbon and electron flow and on the archaeal community in methanogenic rice field soil; Fey A et al.; Temperature is an important factor controlling CH(4) production in anoxic rice soils . Soil slurries, prepared from Italian rice field soil, were incubated anaerobically in the dark at six temperatures of between 10 to 37 degrees C or in a temperature gradient block covering the same temperature range at intervals of 1 degrees C . Methane production reached quasi-steady state after 60 to 90 days . Steady-state CH(4) production rates increased with temperature, with an apparent activation energy of 61 kJ mol(-1) . Steady-state partial pressures of the methanogenic precursor H(2) also increased with increasing temperature from <0.5 to 3.5 Pa, so that the Gibbs free energy change of H(2) plus CO(2)-dependent methanogenesis was kept at -20 to -25 kJ mol of CH(4)(-1) over the whole temperature range . Steady-state concentrations of the methanogenic precursor acetate, on the other hand, increased with decreasing temperature from <5 to 50 microM . Simultaneously, the relative contribution of H(2) as methanogenic precursor decreased, as determined by the conversion of radioactive bicarbonate to (14)CH(4), so that the carbon and electron flow to CH(4) was increasingly dominated by acetate, indicating that psychrotolerant homoacetogenesis was important . The relative composition of the archaeal community was determined by terminal restriction fragment length polymorphism (T-RFLP) analysis of the 16S rRNA genes (16S rDNA) . T-RFLP analysis differentiated the archaeal Methanobacteriaceae, Methanomicrobiaceae, Methanosaetaceae, Methanosarcinaceae, and Rice clusters I, III, IV, V, and VI, which were all present in the rice field soil incubated at different temperatures . The 16S rRNA genes of Rice cluster I and Methanosaetaceae were the most frequent methanogenic groups . The relative abundance of Rice cluster I decreased with temperature . The substrates used by this microbial cluster, and thus its function in the microbial community, are unknown . The relative abundance of acetoclastic methanogens, on the other hand, was consistent with their physiology and the acetate concentrations observed at the different temperatures, i.e., the high-acetate-requiring Methanosarcinaceae decreased and the more modest Methanosaetaceae increased with increasing temperature . Our results demonstrate that temperature not only affected the activity but also changed the structure and the function (carbon and electron flow) of a complex methanogenic system.

Poult Sci, 2000 Oct, 79(10), 1485 - 90
The effect of varying mix uniformity (simulated) of phytase on growth performance, mineral retention, and bone mineralization in chicks; Johnston SL et al.; An 18-d experiment was conducted to determine the effect of varying mix uniformity of phytase on growth performance, mineral retention, and bone mineralization in chicks . Chicks (initial and final weights were 74.5 and 803.3 g) were allotted to seven treatments with six (Treatment 1) or seven (Treatments 2 to 7) replicates of seven chicks per replicate in a completely randomized design . Varying mix uniformity of phytase was simulated by alternately providing two diets with two different concentrations of microbial phytase; the diets were switched every 24 h . Treatments were: 1) positive control (CON) (Ca, 1.0%; available phosphorus (aP), 0.45%), 2) negative control (NEG) (Ca, 0.9%; aP, 0.35%), 3) NEG + 600 phytase units (FTU) daily (CV0), 4) NEG + 500 or 700 FTU (CV17), 5) NEG + 400 or 800 FTU (CV34), 6) NEG + 200 or 1,000 FTU (CV69), or 7) NEG + 0 or 1,200 FTU (CV103) . Gain, feed intake, and bone breaking strength were similar (P > 0.15) in the CON and CV0 treatments, but these response variables were decreased in the NEG treatment (P < 0.01) . Gain:feed was not affected by treatment (P = 0.15) . Bone ash was decreased (P < 0.02) by the NEG and CV0 treatments compared with the CON diet, but chicks fed the CV0 diet had greater bone ash than those fed the NEG (P < 0.01) diet . Increasing FTU CV decreased bone breaking strength and bone ash (P < 0.01) . Calcium and phosphorus retention (P < 0.08) and gain (P < 0.09) were numerically decreased, and phosphorus excretion was numerically increased (P < 0.07) as FTU CV increased . The difference between the CV0 and CV103 treatments was significant only for bone breaking strength and ash (P < 0.01) . In conclusion, increasing phytase CV had little effect on growth performance, whereas bone ash and breaking strength and calcium and phosphorus retention and excretion decreased only at the most extreme CV.

Nutrition, 2000 Oct, 16(10), 996 - 1005
Anorexia of infection: current prospects; Langhans W; The anorexia of infection is part of the host's acute phase response (APR) . Despite being beneficial in the beginning, long lasting anorexia delays recovery and is ultimately deleterious . Microbial products such as bacterial cell wall compounds (e.g., lipopolysaccharides and peptidoglycans), microbial nucleic acids (e . g., bacterial DNA and viral double-stranded RNA), and viral glycoproteins trigger the APR and presumably also the anorexia during infections . Microbial products stimulate the production of proinflammatory cytokines (e.g., interleukins {ILs}, tumor necrosis factor-alpha, interferons), which serve as endogenous mediators . Several microbial products and cytokines reduce food intake after parenteral administration, suggesting a role of these substances in the anorexia during infection . Microbial products are mainly released and cytokines are produced in the periphery during most infections; they might inhibit feeding through neural and humoral pathways activated by their peripheral actions . Activation of peripheral afferents by locally produced cytokines is involved in several cytokine effects, but is not crucial for the anorectic effect of microbial products and IL-1beta . Cytokines increase leptin expression in the adipose tissue, and leptin may contribute to, but is also not essential for, the anorectic effects of microbial products and cytokines . In addition, a direct action of cytokines and microbial products on the central nervous system (CNS) is presumably involved in the anorexia during infection . Cytokines can reach CNS receptors through circumventricular organs and through active or passive transport mechanisms or they can act through receptors on endothelial cells of the brain vasculature and stimulate the release of subsequent mediators such as eicosanoids . De novo CNS cytokine synthesis occurs in response to peripheral infections, but its role in the accompanying anorexia is still open to discussion . Central mediators of the anorexia during infection appear to be neurochemicals involved in the normal control of feeding, such as serotonin, dopamine, histamine, corticotropin releasing factor, neuropeptide Y, and alpha-melanocyte-stimulating hormone . Reciprocal, synergistic, and antagonistic interactions between various pleiotropic cytokines, and between cytokines and neurochemicals, form a complex network that mediates the anorexia during infection . Current knowledge on the mechanisms involved suggests some therapeutic options for treatment . Substances that block common key steps in cytokine synthesis or cytokine action, or inhibitors of eicosanoid synthesis, may hold more promise than attempts to antagonize specific cytokines . To target the neurochemical mediation of the anorexia during infection may be even more efficient . Future research should address these neurochemical mechanisms and the cytokine actions at the blood-brain barrier . Further unanswered questions concern the modulation of the anorexia during infection by gender and nutritional state.

Clin Sci (Lond), 2000 Nov, 99(5), 433 - 41
Pharmacokinetics of retinyl palmitate and retinol after intramuscular retinyl palmitate administration in severe malaria; Davis TM et al.; Retinol (vitamin A alcohol) is an accepted adjunctive treatment in infections such as measles . There is also indirect evidence from in vitro, animal and human studies that retinol supplementation may be beneficial in severe malaria . There have, however, been no studies that have examined the pharmacokinetics of acute retinol supplementation in severe illness . To establish whether mobilization of intramuscular retinyl palmitate (RP) and its availability as retinol are adequate in complicated falciparum malaria, we administered a single dose of 400000 i.u . of RP to six Vietnamese adults with severe malaria . Another 28 patients were not given RP . All patients had blood samples taken over 96 h for RP and retinol assay using HPLC, and received conventional anti-malarial and supportive therapy . Admission serum retinol concentrations were below the lower limit of the reference range (<1.0 micromol/l) in 74% of the 34 patients . In supplemented patients, analysis of serum RP between 0 and 96 h using a multi-compartmental model revealed a median (range) delay in mobilization of 6.9 h (0.7-15.1 h), a bioavailability of 55% (19-100%) and an elimination half-life of 13.5 h (4.2-23.7 h) . The area under the serum retinol curve expressed as an absolute or percentage change from baseline was greater in supplemented than in unsupplemented patients (P<0.05) . The separation in median serum retinol concentrations in the two groups was maximal at 48 h . The model-derived retinol half-life {1.5 (0.7-15.8) h} suggested rapid uptake, metabolism and/or excretion . In conclusion, there is variable RP bioavailability in severe malaria, but a significant if delayed increase in serum retinol over that associated with recovery from the infection . In severe infections, RP supplementation appears simple, well tolerated and of potential benefit once anti-microbial and supportive therapy have been established.

J Agric Food Chem, 2000 Oct, 48(10), 4826 - 9
Biotransformation of (-)-dihydromyrcenyl acetate using the plant parasitic fungus Glomerella cingulata as a biocatalyst; Miyazawa M et al.; The microbial transformation of (-)-dihydromyrcenyl acetate was investigated using the plant parasitic fungus Glomerella cingulata . As a result, (-)-dihydromyrcenyl acetate was converted to dihydromyrcenol, 3,7-dihydroxy-3,7-dimethyl-1-octene-7-carboxylate, 3,7-dihydroxy-3,7-dimethyl-1-octene, 3,7-dimethyloctane-1,2, 7-triol-7-carboxylate, and 3,7-dimethyloctane-1,2,7-triol . In addition, microbial transformation of dihydromyrcenol by G . cingulata was carried out . The metabolic pathway of (-)-dihydromyrcenyl acetate is discussed.

J Inorg Biochem, 2000 Aug 31, 81(3), 183 - 90
Microsomal cytochrome P450 2C5: comparison to microbial P450s and unique features; Williams PA et al.; Although microsomal P450s represent the majority of P450s, only microbial P450s have been amenable to crystal structure solution . We have recently solved the first crystal structure of a microsomal P450, 2C5, a progesterone hydroxylase from rabbit . We discuss the features of the protein in common with existing structures of microbial P450s and limitations of homology modeling mammalian P450s based on the microbial structures . Unique features involving membrane, substrate and cytochrome P450 reductase interactions are also discussed.

Immunol Cell Biol, 2000 Oct, 78(5), 545 - 53
Special feature for the Olympics: effects of exercise on the immune system: exercise-induced modulation of macrophage function; Woods J et al.; Macrophages are important effector cells involved in phagocytosis, microbial killing and antitumour activity . Macrophages also display accessory cell function, in that they can present antigen to foster the development of T lymphocyte-mediated immunity . Recent work, including studies from this group, has demonstrated that acute and chronic exercise can affect many facets of macrophage biology . Manifestation of these effects depends on exercise intensity and duration, the function measured, the timing of measurement in relation to exercise and the concentration of the macrophage-activating stimulus . Exercise has potent stimulatory effects on phagocytosis, antitumour activity, reactive oxygen and nitrogen metabolism, and chemotaxis . Indeed, it has been shown that exercise training can increase macrophage antitumour activity in mice of different ages . However, not all functions are enhanced by exercise . Exercise-induced reductions in macrophage MHC II expression and antigen-presentation capacity have been documented . These findings bring up the possibility that exercise, and perhaps other stressors, activate macrophages for effector functions while downregulating accessory cell functions . To a large extent, the mechanisms responsible for the exercise-induced changes in macrophage function remain unknown, but may depend on exercise-induced changes in neuroendocrine factors . Future studies need to explore the effects in a mechanistic way and provide documentation as to their physiological significance.

Trends Genet, 2000 Oct, 16(10), 442 - 9
Genes that fight infection: what the Drosophila genome says about animal immunity; Khush RS et al.; From deciphering the principles of heredity to identifying the genes that control development, the fruit fly Drosophila melanogaster is being used to deconstruct an increasing number of biological processes . Genetic studies of Drosophila responses to microbial infection have identified regulators of innate immunity that are functionally conserved in mammals . These recent findings highlight the ancient origins of animal immune responses and demonstrate the potential of Drosophila for dissecting host-pathogen interactions . The sequencing of the Drosophila genome both enhances genetic approaches and provides new clues for the identification of key components of innate immunity . This article summarizes how information gained from genomic analysis contributes to our understanding of how animals cope with infectious disease.

Hepatology, 2000 Nov, 32(5), 901 - 9
Fine specificity of T cells reactive to human PDC-E2 163-176 peptide, the immunodominant autoantigen in primary biliary cirrhosis: implications for molecular mimicry and cross-recognition among mitochondrial autoantigens; Shigematsu H et al.; The anti-mitochondrial antibody response in primary biliary cirrhosis (PBC) is primarily directed at E2 components of PDC, OGDC, and BCOADC, and E3BP . Previous work has shown that the immunodominant autoreactive T- cell epitope is the PDC-E2 163-176 peptide, restricted by HLA DR53 . To address molecular mimicry and cross-recognition among mitochondrial autoantigens, we analyzed reactivity, including agonism and antagonism assays, to a series of single amino acid-substituted peptides using cloned T-cell lines in PBC and controls . Interestingly, fine specificities were unique for every single T-cell clone, but the clones could be categorized into two distinct groups based on recognition motifs of the T-cell receptor (TCR) ligand: group A (170)ExDK(173) and group B (168)EIExD(172) . (170)E is the most critical TCR contact residue for both groups of cloned T-cell lines, whereas (173)K and (168)E are the critical TCR contact residues for group A and group B cloned T-cell lines, respectively . More importantly, some group A-cloned T-cell lines cross-reacted to human E3BP 34-47, human OGDC-E2 100-113, and several peptides derived from various microbial proteins carrying an ExDK motif, whereas group B-cloned T-cell lines reacted only to E3BP 34-47 carrying an EIExD motif . Furthermore, an RGxG motif was exclusively found in the complementarity-determining region (CDR3) of the TCR Vbeta in the group B-cloned T-cell lines, while G, S, and/or R were frequently found in the CDR3 of the TCR Vbeta in the group A-cloned T-cell lines . These data provide a framework for understanding molecular mimicry among mitochondrial antigens.

Yale J Biol Med, 1999 Sep-Oct, 72(5), 313 - 20
Welch, Sedgwick, and the Hopkins model of hygiene; Benson KR; William H . Welch and William T . Sedgwick, two of the founding fathers of American public health, were both early generation "Hopkins Men." Sedgwick was part of the first group of graduate students to attend Johns Hopkins University, and Welch was part of the initial faculty at the University's medical school . While they never worked together as colleagues at Hopkins, both became interested in the exciting new discoveries of the microbial nature of human disease and developed similar public health programs based on this information . Sedgwick expanded upon these investigations in the new sanitary science program at MIT, where academic public health first emerged in the United States following Sedgwick's appointment in 1883 . Welch, who had been exposed to European research in microbiology, promoted microbial research in pathology in Baltimore in 1884 . His laboratory-based investigations expanded until they led to the formation of the country's first school of public health in 1916 . Thus, a "Hopkins Model" for hygiene and public health emerged from the efforts of both Welch and Sedgwick.

J Oral Pathol Med, 2000 Nov, 29(10), 500 - 6
Vascular remodelling in chronic inflammatory periodontal disease; Chapple CC et al.; Periodontitis is a chronic inflammatory disease of the highly vascularised supporting tissues of the teeth . Little is known about the vascular changes in untreated advanced periodontitis . Using confocal immunofluorescence microscopy and morphometry, we defined and quantified vascular remodelling in this lesion . In the connective tissue subjacent to the altered epithelium lining of the periodontal pocket, there was a significant increase in the numerical density of vascular profiles, primarily accounted for by vessels > or = 25 microm in diameter . In addition, vascular basement membranes were thickened and there was accumulation of non-vascular basement membrane remnants . We investigated the distribution of major angiogenic growth factors in periodontitis using immunohistochemistry . Basic fibroblast growth factor, although consistently associated with blood vessels, showed no regional variation in its distribution . In contrast, there was a marked regional variation in the intensity of immunostaining for vascular endothelial growth factor, with significantly reduced staining of the pocket epithelium . The changes in the vascularity of the periodontal connective tissues in untreated advanced periodontitis may be, in part, a consequence of altered expression of angiogenic activity by the epithelium . In turn, this may reflect the epithelial response to microbial flora in the microenvironment of the periodontal pocket.

Pediatr Allergy Immunol, 2000, 11 Suppl 13, 19 - 22
Allergic diseases in farmers' children; Braun-Fahrlander C; Several studies have reported lower rates of allergic sensitization and allergies in children living in rural as compared to urban communities . This has been attributed to the lower levels of air pollution in rural areas . The question arises whether other factors in the rural environment could explain the lower prevalence rates of allergic sensitization and hay fever . A first report from rural South Bavaria in Germany demonstrated that children living in a home where coal and wood were used for heating had a significantly lower risk of suffering from hay fever (odds ratio 0.57 (0.34-0.98)), of being sensitized to common allergens (OR 0.67 (0.49-0.93)) and of having bronchial hyperresponsiveness (OR 0.55 (0.34-0.90)) than their peers living in homes with other heating systems . Subsequently, the Swiss Study on Childhood Allergy and Respiratory Symptoms with Respect to Air Pollution (SCARPOL) tested the hypothesis that farming as parental occupation was associated with a lower risk of hay fever and atopy . A total of 1620 (86.0%) 6-15-year-old schoolchildren living in three rural communities of Switzerland were examined using a standardized questionnaire completed by the parents and IgE antibodies against six common aeroallergens in serum samples of 404 (69.3.0%) of the 13-15-year-olds . Farming as parental occupation was significantly associated with lower rates of reported hay fever symptoms and allergic sensitization . Comparing children from farming with those from non-farming environments, the adjusted OR was 0.34 (95% CI: 0.12-0.89) for sneezing attacks during the pollen season, and 0.31 (95% CI: 0.13-0.73) for a sensitization to allergens . These results have recently been confirmed in a new and much larger survey in rural South Bavaria . Several alternative explanations have to be considered when interpreting these findings, namely, selection bias, the development of tolerance, increased microbial stimulation and a more traditional lifestyle (diet and housing conditions) . Based on present knowledge, the underlying environmental factor explaining the protective effect of the farming environment has not yet been identified.

J Immunol, 2000 Nov 1, 165(9), 5027 - 34
Microbial and T cell-derived stimuli regulate antigen presentation by dendritic cells in vivo; Manickasingham S et al.; B cells and dendritic cells (DC) internalize and degrade exogenous Ags and present them as peptides bound to MHC class II molecules for scrutiny by CD4(+) T cells . Here we use an Ab specific for a processed form of the model Ag, hen egg lysozyme (HEL), to demonstrate that this protein is not efficiently presented by lymph node DC following s.c . immunization . HEL presentation by the DC can be dramatically enhanced upon coinjection of a microbial adjuvant, which appears to act by enhancing peptide loading onto MHC class II . CD40 cross-linking or the presence of a high frequency of T cells specific for HEL can similarly improve presentation by DC in vivo . For any of these activating stimuli, CD8alpha(+) DC consistently display the highest proportion of HEL-loaded MHC class II molecules . These data indicate that exogenous Ags can be displayed to T cells in lymphoid tissues by a large cohort of resident DC whose presentation is regulated by innate and adaptive stimuli . Our data further reveal the existence of a feedback mechanism that augments Ag presentation during cognate APC-T cell interactions.

J Immunol, 2000 Nov 1, 165(9), 4797 - 801
Cutting edge: CD1d deficiency impairs murine host defense against the spirochete, Borrelia burgdorferi; Kumar H et al.; CD1 molecules can present microbial lipid Ag to T cells, suggesting that they participate in host defense against pathogens . In this study, we examined the role of CD1d in resistance to infection with the Lyme disease spirochete, Borrelia burgdorferi (Bb), an organism with proinflammatory lipid Ag . Bb infection of CD1d-deficient (CD1d(-/-)) mouse strains normally resistant to this pathogen resulted in arthritis . Pathology correlated with an increased prevalence of spirochete DNA in tissues and enhanced production of Bb-specific IgG, including IgG to Ag rapidly down-modulated on spirochetes in vivo . CD1d(-/-) mice exhibited high-titer Bb-specific IgG2a, an isotype commonly induced in disease-susceptible mice but not in the disease-resistant control mice in this study . These results show that CD1d deficiency impairs host resistance to a spirochete pathogen, and are the first example of a mutation that imparts Bb-resistant mice with the Ab and disease profile of a susceptible mouse strain.

Hum Reprod Update, 2000 Sep-Oct, 6(5), 432 - 41
Infections in IVF: review and guidelines; Steyaert SR et al.; Since the inception of in-vitro fertilization (IVF), questions about contamination and the transmission of infection have been raised . In this review, screening for Chlamydia trachomatis, as well as medical and ethical considerations on IVF in couples infected with hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus (HIV), are discussed . IVF is not contraindicated in case of HBV or HCV infection, but the decision is far more delicate in case of HIV infection . If donor gametes or embryos are used, prevention of infectious disease transmission resides in accurate donor selection, including screening for C . trachomatis, HIV, HBV, cytomegalovirus and Treponema pallidum . In the embryology laboratory, microbial contamination of the IVF system deserves attention, and can be prevented by using sterile technique and supplementing culture media with screened sera or serum substitutes and antibiotics . Persons whose biological material is to be cryopreserved should be screened for HBV, HCV and HIV, and separate containers should be used for infected and non-infected material . Finally, transmission of infectious diseases to laboratory personnel can be prevented by adherence to strict safety guidelines, wearing of protective clothing, HBV vaccination, prohibition of mouth pipetting, and developing a plan for the disposal of bio-hazardous material.

Adv Perit Dial, 2000, 16, 208 - 12
In vitro study of the efficacy of a two-way connection with disinfectant in the prevention of peritonitis; Buoncristiani U et al.; A new connection system for continuous ambulatory peritoneal dialysis (CAPD) has been established, and its efficacy in preventing microbial contamination of the peritoneal cavity has been tested in vitro . The system consists of a Y-shaped channel formed in the bottom of a Plexiglas cup . The Luer-lock shaped ends of the Y-shaped channel are designed to host the connectors from the drainage bag, the catheter transfer set, and the bag of fresh dialysate . Because the connectors from the catheter transfer set and the fresh bag are located at the inner surface of the cup bed, and because the cup is filled with disinfectant during the entire exchange procedure, all at-risk steps are continuously protected by disinfectant (that is, removal of the caps from the connectors, connection and disconnection, replacement of the caps) . Still, because the patient could inadvertently extract and contaminate one of the two connectors (although such a possibility is unlikely), the disinfecting efficacy of the system was tested in vitro . Despite contamination with various micro-organisms at the highest possible concentrations, all tests showed negative bacterial growth, thus confirming the absolute efficacy of the system in preventing exogenous transluminal peritonitis.

Presse Med, 2000 Sep 23, 29(27), 1532 - 4
{Standards, options and recommendations for good practice in hemoculture in cancerology}; Bussy Malgrange V et al.; Excepting emergency and aplasia: two to three blood samples should be draw for culture an hour apart within a 24 period (standard) . For emergency or aplasia: two to three blood samples should be drawn for culture before initiating early antibiotic therapy . The delay between samples drawn from different sites should be less than one hour (standard) . For patients on antibiotics: four to six blood samples should be drawn for culture within 48 hours, outside ongoing antibiotic administration . If the patient is given corticosteroids, it is recommended to draw two or three blood samples in case of deterioration (agreement of the experts) . Rigorous aseptic techniques must be used (standard) . Culture media are chosen according to the institution's microbial ecology (standard) . The volume of blood drawn should be adapted to the system used (standard) . Culture positivity is determined at 24 to 48 hours.

Biol Neonate, 2000 Oct, 78(3), 186 - 90
Determination of secretory IgA and albumin in saliva of newborn infants; Seidel BM et al.; As a first line of defence against microbial invasion, secretory IgA (sIgA) is the dominant immunoglobulin on all mucosal surfaces . In this study sIgA was determined by radial immunodiffusion in saliva samples of 63 newborn infants divided into the following age groups: (1) 1 day and younger, (2) 2-10 days . Concentrations of sIgA and albumin as well as their relation to age, postprandial time, gestational age and birth weight were analysed . sIgA could be detected in 75.0% (group 1) and 77.1% (group 2) of the saliva samples with a mean concentration of 190.2 mg/l (group 1) and 216.4 mg/l (group 2) . Differences failed to reach significance . Concentration of sIgA was found to be independent of age but positively related to the concentration of albumin in the same saliva sample . In conclusion, this study provides evidence that high levels of sIgA are found in saliva of newborn infants, indicating the existence of a competent oral mucosal immune system as early as within the first 10 days of life .

Trends Microbiol, 2000 Oct, 8(10), 452 - 6
The Toll receptor family and microbial recognition; Medzhitov R et al.; The survival of multicellular organisms is dependent on their ability to recognize invading microbial pathogens and to induce a variety of defense reactions . Recent evidence suggests that an evolutionarily ancient family of Toll-like receptors plays a crucial role in the detection of microbial infection and the induction of immune and inflammatory responses.

Int J Dermatol, 2000 Sep, 39(9), 672 - 7
Necrotizing eosinophilic folliculitis as a manifestation of the atopic diathesis; Magro CM et al.; BACKGROUND: Eosinophilic folliculitis (EF) is an idiopathic eruption of sterile pustules and papules involving the trunk, face, and extremities, associated in many cases with human immunodeficiency virus (HIV) infection . The classic histopathology is one of follicular-based, eosinophilic spongiosis with variable microabscess formation . We describe nine HIV-negative patients who manifested a novel form of pustular EF in the setting of atopy . MATERIALS AND METHODS: Paraffin sections of skin biopsies from ten patients, stained with hematoxylin and eosin and special preparations to evaluate for microbial pathogens, were examined . Detailed clinical histories and serologic studies were obtained . RESULTS: Among the clinical presentations in seven men, two women, and one girl (age range, 11-62 years) were ulcerative and/or nodular plaques mainly on the face and/or extremities, sometimes in an annular configuration . The clinical considerations included deep mycotic infection, ulcerative herpes, systemic vasculitis, Mucha Haberman disease, and pyoderma gangrenosum . All patients had a personal and/or family history of atopy . Co-existent medical illnesses included psoriasis, lupus erythematosus, and lymphoproliferative disease . One patient was on a calcium channel blocker, one on multiple antidepressants, and two on antihistamines, all of which are associated with immune dysregulation . All skin biopsies showed variable intra- follicular eosinophilic microabscesses, follicular necrosis, folliculocentric necrotizing eosinophilic vasculitis, marked degeneration of connective tissue fiber elements, and striking tissue eosinophilia, including flame figure formation and dermal eosinophilic abscesses . Apart from commensals, such as Pityrosporum and Demodex, microbial pathogens were not identified . CONCLUSIONS: The presentations differed from conventional EF by virtue of a strong association with atopy and by the presence of ulceration, nodule formation, follicular and dermal necrosis, and eosinophilic vasculitis . We propose the term "necrotizing eosinophilic folliculitis," and suggest that the basis of this novel form of EF is an unrepressed T-helper lymphocyte type 2 (Th2)-dominant response to various epicutaneous stimuli in patients with atopy, the prototypic immune dysregulatory state associated with a Th2-dominant cytokine milieu.

J Virol, 2000 Nov, 74(22), 10600 - 11
Strategy for systematic assembly of large RNA and DNA genomes: transmissible gastroenteritis virus model; Yount B et al.; A systematic method was developed to assemble functional full-length genomes of large RNA and DNA viruses . Coronaviruses contain the largest single-stranded positive-polarity RNA genome in nature . The approximately 30-kb genome, coupled with regions of genomic instability, has hindered the development of a full-length infectious cDNA construct . We have assembled a full-length infectious construct of transmissible gastroenteritis virus (TGEV), an important pathogen in swine . Using a novel approach, six adjoining cDNA subclones that span the entire TGEV genome were isolated . Each clone was engineered with unique flanking interconnecting junctions which determine a precise systematic assembly with only the adjacent cDNA subclones, resulting in an intact TGEV cDNA construct of approximately 28.5 kb in length . Transcripts derived from the full-length TGEV construct were infectious, and progeny virions were serially passaged in permissive host cells . Viral antigen production and subgenomic mRNA synthesis were evident during infection and throughout passage . Plaque-purified virus derived from the infectious construct replicated efficiently and displayed similar plaque morphology in permissive host cells . Host range phenotypes of the molecularly cloned and wild-type viruses were similar in cells of swine and feline origin . The recombinant viruses were sequenced across the unique interconnecting junctions, conclusively demonstrating the marker mutations and restriction sites that were engineered into the component clones . Full-length infectious constructs of TGEV will permit the precise genetic modification of the coronavirus genome . The method that we have designed to generate an infectious cDNA construct of TGEV could theoretically be used to precisely reconstruct microbial or eukaryotic genomes approaching several million base pairs in length.

Int J Med Microbiol, 2000 Mar, 290(1), 27 - 35
Bacterial adhesins: function and structure; Klemm P et al.; Specific adhesion to host tissue cells is an essential virulence factor of most bacterial pathogens . The fundamental processes that determine bacterial attachment to host tissue surfaces are mediated by microbial adhesins . Host specificity and tissue tropism are characteristics exhibited by different bacteria and are determined (at least in part) by the interaction between adhesins and their complementary receptors on host cell surfaces . A detailed picture of how bacteria are able to target to various receptors is emerging . A large number of bacterial adhesins with individual receptor specificities have been identified . Furthermore, recent research has shown that individual adhesins are prone to rapid microevolution that results in changes in the receptor specificity of individual adhesins . Microbial adhesins are often assembled into complex polymeric organelle structures, however non-organelle adhesins linked to the cell surface as monomers or simple oligomers also exist . This review gives an overview of bacterial adhesins and focuses on some general aspects of their biogenesis and role in bacterial colonization of host cell surfaces and as virulence factors.

Br J Perioper Nurs, 2000 Feb, 10(2), 102 - 7
Two gloves or not two gloves that is the question; Bernthal L; Surgical gloves were first introduced in 1889, when Halstead wanted to protect his scrub nurse from dermatitis caused by contact with mercuric chloride (Dodds et al 1990) . The use of gloves then became more widespread, principally to protect surgical teams whilst operating on infected patients (Dodds et al 1990) . Later, focus changed to protecting the wound from microbial contamination.

Biotechnol Bioeng, 2000 Dec 5, 70(5), 518 - 24
Enhanced bioaccumulation of heavy metals by bacterial cells displaying synthetic phytochelatins; Bae W et al.; A novel strategy using synthetic phytochelatins is described for the purpose of developing microbial agents for enhanced bioaccumulation of toxic metals . Synthetic genes encoding for several metal-chelating phytochelatin analogs (Glu-Cys)(n)Gly (EC8 (n = 8), EC11 (n = 11), and EC20 (n = 20)) were synthesized, linked to a lpp-ompA fusion gene, and displayed on the surface of E . coli . For comparison, EC20 was also expressed periplasmically as a fusion with the maltose-binding protein (MBP-EC20) . Purified MBP-EC20 was shown to accumulate more Cd(2+) per peptide than typical mammalian metallothioneins with a stoichiometry of 10 Cd(2+)/peptide . Cells displaying synthetic phytochelatins exhibited chain-length dependent increase in metal accumulation . For example, 18 nmoles of Cd(2+)/mg dry cells were accumulated by cells displaying EC8, whereas cells exhibiting EC20 accumulated a maximum of 60 nmoles of Cd(2+)/mg dry cells . Moreover, cells with surface-expressed EC20 accumulated twice the amount of Cd(2+) as cells expressing EC20 periplasmically . The ability to genetically engineer ECs with precisely defined chain length could provide an attractive strategy for developing high-affinity bioadsorbents suitable for heavy metal removal .

Curr Opin Struct Biol, 2000 Oct, 10(5), 548 - 56
Chaperone-assisted pilus assembly and bacterial attachment; Sauer FG et al.; Bacterial pili assembled by the chaperone-usher pathway can mediate microbial attachment, an early step in the establishment of an infection, by binding specifically to sugars present in host tissues . Recent work has begun to reveal the structural basis both of chaperone function in the biogenesis of these pili and of bacterial attachment.

Genome Res, 2000 Oct, 10(10), 1643 - 7
Links from genome proteins to known 3-D structures; Wang Y et al.; We describe a genome annotation service provided by the Entrez browser, All protein products identified in fully sequenced microbial genomes have been compared with proteins with known 3-D structure by use of the BLAST sequence comparison algorithm . For the approximately 20% of genome proteins in which unambiguous sequence similarity is detected, Entrez provides a link from the gene product to its predicted structure . The service uses the Cn3D molecular graphics viewer to present a 3-D view of the known structure, together with an alignment display mapping conserved residues from the genome protein onto the known structure . Using an example from Aeropyrum pernix, we illustrate how mapping to a 3-D structure can confirm predictions of biological function.

Biochem J, 2000 Nov 1, 351 Pt 3, 687 - 96
Regulation of Nramp1 mRNA stability by oxidants and protein kinase C in RAW264.7 macrophages expressing Nramp1(Gly169); Lafuse WP et al.; The murine Nramp1 (natural-resistance-associated macrophage protein) locus confers innate resistance against intracellular macrophage pathogens . The gene encodes a transporter molecule, which is rapidly recruited to the phagosome . Nramp1 functions as an iron transporter by transporting iron into the phagosome . Within the phagosome iron mediates anti-microbial killing by hydroxyl radical formation through the iron-catalysed Fenton/Haber-Weiss reaction . In addition to its effects on the growth of intracellular pathogens, Nramp1 exerts a wide range of pleiotropic effects in activated macrophages . One of these pleiotropic effects is the increased stability of a variety of mRNA species, including Nramp1 mRNA . In the present study, the stability of Nramp1 mRNA in Mycobacterium avium infected RAW264 . 7 macrophages expressing either the Nramp1(Gly169) resistant allele or the Nramp1(Asp169) susceptible allele was examined . Nramp1 mRNA stability was greater in Nramp1(Gly169) macrophages than in Nramp1(Asp169) macrophages . The increase in Nramp1 mRNA stability in resistant macrophages was inhibited by antioxidants and protein kinase C (PKC) inhibitors, suggesting that Nramp1 mRNA stability is regulated by an oxidant-generated signalling pathway that requires PKC activity . This was corroborated by treating Nramp1(Asp169) macrophages with menadione, which generates reactive oxygen species within cells . Menadione increased Nramp1 mRNA stability to the level observed in resistant macrophages; this increase was also inhibited by a PKC inhibitor . Further, PKC activity was found to be greater in M . avium-infected Nramp1(Gly169) macrophages than in infected Nramp1(Asp169) macrophages and inhibited by treatment with an antioxidant.

Bone Marrow Transplant, 2000 Sep, 26(6), 701 - 3
Fatal invasive cerebral Absidia corymbifera infection following bone marrow transplantation; Paterson PJ et al.; A 56-year-old dairy farmer received a fully HLA matched unrelated donor marrow transplant for high risk CML in chronic phase . His early post-transplant course was complicated by a series of massive intracerebral bleeds and by sepsis related to a malignant otitis externa . The microbial pathogen isolated from ear swabs was found to be Absidia corymbifera, but CT scan at the time showed no intracerebral extension . Despite neutrophil engraftment and aggressive antifungal management he succumbed . Autopsy revealed invasion of Absidia into the brain from the ear . We speculate that colonisation by Absidia resulted from occupational exposure.

J Food Prot, 2000 Oct, 63(10), 1404 - 9
Comparison of capillary and test tube procedures for analysis of thermal inactivation kinetics of mold spores; Fujikawa H et al.; Characteristics of capillary and test tube procedures for thermal inactivation kinetic analysis of microbial cells were studied for mold spores . During heating, capillaries were submerged in a water bath and test tubes were held with their caps positioned above the level of the heating medium . Thermal inactivation curves of Aspergillus niger spores in capillaries at around 60 degrees C consisted of a shoulder and a fast linear decline, whereas curves in test tubes consisted of a shoulder, a fast linear decline, and a horizontal tail . There were no significant differences in values of the rate and the delay of fast declines in curves between the procedures . Some experiments were done to clarify the cause for tailing with test tubes . There were no tails with test tubes whose inner walls were not contaminated by A . niger spores, suggesting that tails arise from A . niger spores contaminating the inner walls of test tubes . Temperature of the inner wall at the level of a heating medium was lower than that of the medium . Further, there were no tails for test tubes submerged in the heating medium . These results showed that the reason for survival of contaminants on the upper wall of test tubes was that cells were not subjected to sufficient inactivation temperature . Finally, thermal inactivation curves of A . niger spores in capillaries at various constant temperatures were studied . Curves consisted of a shoulder and a fast linear decline at 57 degrees C and above, whereas curves at below 57 degrees C consisted of a shoulder, a fast linear decline, and a sloping tail.

J Dent Res, 1988 Jan, 67(1), 46 - 50
The effect of long-term use of a dentifrice containing zinc citrate and a non-ionic agent on the oral flora; Jones CL et al.; The effect on the oral ecology of daily use, for seven months, of a dentifrice containing 0.5% (w/w) zinc citrate and a non-ionic agent, i.e., 0.2% (w/w) Triclosan (2,4,4'-trichloro-2'-hydroxydiphenyl ether), has been monitored on 13 adult volunteers . Plaque and saliva were sampled monthly and the bacterial flora analyzed . Twenty-six volunteers used a placebo dentifrice as part of their normal oral hygiene for four months to establish the baseline microbial flora . The volunteers were then split into two equal groups: One group continued to use the placebo dentifrice; the other used the dentifrice containing zinc citrate and Triclosan . There was no evidence that seven months' use of a dentifrice containing 0.5% zinc citrate and 0.2% Triclosan caused shifts in the oral microbial ecology, nor was there any evidence of developing bacterial resistance to Triclosan.

Luminescence, 2000 Sep-Oct, 15(5), 321 - 7
Effects of zinc on the reactive oxygen species generating capacity of human neutrophils and on the serum opsonic activity in vitro; Hasegawa H et al.; To investigate the effects of zinc on non-specific immune functions, we used the chemiluminescence method to examine the capacity of human neutrophils to produce reactive oxygen species and accompanying serum opsonic activity . When neutrophils were stimulated with both opsonized zymosan and phorbol myristate acetate in the presence of 1-10(-3) mmol/L zinc lucigenin-dependent CL responses were stable or declined, whereas luminol-dependent CL responses were significantly enhanced . The results suggest that zinc activates protein kinase C and promotes MPO degranulation and ROS metabolism, especially in hypochlorous acid production, which have the direct action of causing microbial death . Further, the lucigen-dependent CL response stimulated with OZ was strongly enhanced by anti-MPO antibodies, whereas the enhancement was less in the presence of zinc, suggesting that zinc may suppress the receptor-mediated signal transduction process . Both responses were inhibited at 10 mmol/L . Serum opsonic activity was enhanced by zinc at 10(-4) and 10(-3) mmol/L but reduced at 10 mmol/L . These data indicate that addition of zinc around and above normal physiological concentrations facilitates neutrophil functional activity and serum opsonic activity, whereas these are inhibited by a lack of zinc or an excessive amount, suggesting that zinc is essential for optimal functioning of non-specific immunity .

Adv Biochem Eng Biotechnol, 2000, 69, 87 - 124
Development of biotechnology in India; Ghose TK et al.; India has embarked upon a very ambitious program in biotechnology with a view to harnessing its available human and unlimited biodiversity resources . It has mainly been a government sponsored effort with very little private industry participation in investment . The Department of Biotechnology (DBT) established under the Ministry of Science and Technology in 1986 was the major instrument of action to bring together most talents, material resources, and budgetary provisions . It began sponsoring research in molecular biology, agricultural and medical sciences, plant and animal tissue culture, biofertilizers and biopesticides, environment, human genetics, microbial technology, and bioprocess engineering, etc . The establishment of a number of world class bioscience research institutes and provision of large research grants to some existing universities helped in developing specialized centres of biotechnology . Besides DBT, the Department of Science & Technology (DST), also under the Ministry of S&T, sponsors research at universities working in the basic areas of life sciences . Ministry of Education's most pioneering effort was instrumental in the creation of Biochemical Engineering Research Centre at IIT Delhi with substantial assistance from the Swiss Federal Institute of Technology, Zurich, Switzerland to make available state-of-the-art infrastructure for education, training, and research in biochemical engineering and biotechnology in 1974 . This initiative catalysed biotechnology training and research at many institutions a few years later . With a brief introduction, the major thrust areas of biotechnology development in India have been reviewed in this India Paper which include education and training, agricultural biotechnology, biofertilizers and biopesticides, tissue culture for tree and woody species, medicinal and aromatic plants, biodiversity conservation and environment, vaccine development, animal, aquaculture, seri and food biotechnology, microbial technology, industrial biotechnology, biochemical engineering and associated activities such as creation of biotechnology information system and national repositories . Current status of intellectual property rights has also been discussed . Contribution to the India's advances in biotechnology by the industry, excepting a limited few, has been far below expectations . The review concludes with some cautious notes.

Adv Biochem Eng Biotechnol, 2000, 69, 71 - 85
Microbial production of amino acids in Japan; Kumagai H; The microbial biotechnology of amino acids production which was developed and industrialized in Japan have been summarized . The amino acids include L-glutamic acid, L-lysine, L-threonine, L-aspartic acid, L-alanine, L-cysteine, L-dihydroxyphenylalanine, D-p-hydroxyphenyl-glycine, and hydroxy-L-proline.

Am J Obstet Gynecol, 2000 Oct, 183(4), 887 - 94
Evidence of in vivo differential bioavailability of the active forms of matrix metalloproteinases 9 and 2 in parturition, spontaneous rupture of membranes, and intra-amniotic infection; Maymon E et al.; OBJECTIVE: Matrix metalloproteinases (MMP-9 and MMP-2) have been implicated in the digestion of fetal membranes . The purpose of this study was to determine the amniotic fluid concentrations of active forms of MMP-2 and MMP-9 and to explore the participation of these enzymes in labor (term and preterm), rupture of membranes (term and preterm), and microbial invasion of the amniotic cavity . STUDY DESIGN: A cross-sectional study was conducted with 291 women in the following categories: (1) term not in labor, (2) term in labor, (3) preterm labor and intact membranes who delivered at term, (4) preterm labor who delivered preterm, (5) preterm labor with microbial invasion of the amniotic cavity, (6) preterm premature rupture of membranes without microbial invasion of the amniotic cavity, (7) preterm premature rupture of membranes with microbial invasion of the amniotic cavity, (8) term premature rupture of membranes not in labor, and (9) mid trimester . Active forms of MMP-2 and MMP-9 were measured by a novel assay that uses a substrate developed by protein engineering . RESULTS: (1) MMP-2 and MMP-9 were detected in 88% and 96% of amniotic fluid samples, respectively (255/291 and 279/291) . (2) The concentrations of active forms of MMP-2 and MMP-9 changed with advancing gestational age . (3) Spontaneous term parturition was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) and a significant decrease in the median concentration of the active forms of MMP-2 (P <.003) . (4) Preterm labor with intact membranes leading to preterm delivery in the absence of infection was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.2) . (5) Rupture of membranes (either term or preterm) was associated with a significant increase in the concentration of the active forms of MMP-9 and with a significant decrease in the concentration of the active forms of MMP-2 (P <.005 for term and P <.03 and P <.003 for preterm, respectively) . (6) Microbial invasion of the amniotic cavity in women with preterm premature rupture of membranes was also associated with a significant increase in the concentration of the active forms of MMP-9 (P <.03) and a decrease in the concentration of the active forms of MMP-2 (P <.05) . (7) Microbial invasion of the amniotic cavity in patients with preterm labor was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.6) . CONCLUSION: Spontaneous rupture of membranes (either term or preterm), parturition (either term or preterm), and microbial invasion of the amniotic cavity were associated with significant increases in the amniotic fluid concentration of the active forms of MMP-9 . In contrast, the concentration of the active forms of MMP-2 either decreased or remained the same in these conditions . Our observations provide evidence for a novel regulation of gelatinolytic activity in vivo.

J Immunol, 2000 Oct 15, 165(8), 4515 - 21
Rapid recruitment of neutrophils containing prestored IL-12 during microbial infection; Bliss SK et al.; Neutrophils are well known to rapidly migrate to foci of infection, where they exert microbicidal functions . We sought to determine whether neutrophils responding to in vivo infection with the protozoan pathogen Toxoplasma gondii were capable of IL-12 production as suggested by recent in vitro studies . Intraperitoneal infection induced a neutrophil influx by 4 h, accompanied by ex vivo IL-12 p40 and p70 release . Approximately 85% of the neutrophils displayed intracellular stores of IL-12, as determined by flow cytometry and confocal fluorescence microscopy . Neutrophils from IFN-gamma knockout mice also expressed IL-12, ruling out an IFN-gamma-priming requirement . Neither infected nor uninfected peritoneal macrophages displayed intracellular IL-12, but these cells were strongly IL-10(+) . Infection per se was unnecessary for IL-12 production because peritoneal and peripheral blood neutrophils from uninfected animals contained IL-12(+) populations . Expression of the granulocyte maturation marker Gr-1 (Ly-6G) was correlated with IL-12 production . Mice depleted of their granulocytes by mAb administration at the time of infection had decreased serum levels of IL-12 p40 . These results suggest a model in which neutrophils with prestored IL-12 are rapidly mobilized to an infection site where they are triggered by the parasite to release cytokine . Our findings place neutrophils prominently in the cascade of early events leading to IL-12-dependent immunity to T . gondii.

J Immunol, 2000 Oct 1, 165(7), 3804 - 10
Activation of toll-like receptor 2 on human dendritic cells triggers induction of IL-12, but not IL-10; Thoma-Uszynski S et al.; Mammalian Toll-like receptors (TLRs) are required for cell activation by bacterial lipoproteins (bLP) and LPS . Stimulation of monocytes with bLP and LPS results in a TLR-dependent induction of immunomodulatory genes leading to the production of pro-inflammatory cytokines . In this paper, we compared the expression and response of TLRs on monocytes and dendritic cells (DC) . TLR2, but not TLR4, was detected on peripheral blood monocytes and DC, in lymphoid tissue CD1alpha+ DC as well as on in vitro monocyte-derived DC . Upon stimulation with bLP or LPS, monocytes produced IL-12 and IL-10 at similar levels, whereas monocyte-derived DC produced comparable levels of IL-12, but little IL-10 . Greater than 90% of the bLP-induced production of IL-12 was blocked by anti-TLR2 mAb . Thus, DC express TLR2 and activation of this receptor by bLP provides an innate mechanism by which microbial pathogens preferentially activate cell-mediated immunity.

J Am Acad Nurse Pract, 2000 Jan, 12(1), 3 - 10
Nurse practitioner student prescriptive patterns; Fontana SA et al.; As employment of nurse practitioners (NPs) increases in health care systems, there is a need to have current data on their prescribing practices and patterns, and to implement a system for updating such data . This study reports prescriptive data based upon 10,421 primary care visits conducted by 55 family NP students over a 15-month period in 1997 and 1998 . Numbers of over-the-counter drugs taken regularly, prescription drugs currently prescribed and prescription drugs prescribed or refilled at the visit were recorded in addition to types of drugs, compliance issues, diagnoses rendered and sociodemographic information . Individual student data were aggregated and analyzed using Epi Info (Epidemiology Program Office of the Centers for Disease Control) and SPSS-PC . Results identified that: 1) the majority of patient visits involved the prescription of 1-2 drugs (88%); 2) major compliance issues included financial concerns, knowledge deficits, and complexity/demands of treatment; 3) commonly rendered diagnoses at drug visits for chronic conditions were hypertension and diabetes; for acute conditions, otitis sinusitis and upper respiratory infections; 4) anti-microbial agents, drugs used for relief of pain, and cardiovascular drugs account for 60% of drug mentions; and 5) the numbers of drugs prescribed or refilled at visits were similar by type of preceptor, except fewer single drugs were prescribed or refilled at visits supervised by nurse preceptors . Findings are discussed relative to deepening the understanding of advanced practice nursing education and the prescribing practices of NP students and their preceptors.

Sci Total Environ, 2000 Oct 9, 260(1-3), 125 - 33
Potential effects of forest fire and storm flow on total mercury and methylmercury in sediments of an arid-lands reservoir; Caldwell CA et al.; A study was conducted from July 1995 to June 1996 to examine the spatial and temporal changes of mercury concentrations in sediments of an arid-lands reservoir . Prior to the first sample collection in July, a forest fire burned 2930 ha of mixed conifer and ponderosa pine in the watershed of Caballo Reservoir in south-central New Mexico . The fire was eventually extinguished by summer rains and storm runoff resulting in the mobilization and transport of charred vegetative material into an intermittent tributary (Palomas Creek) that drains the watershed into Caballo Reservoir . Concentrations of total mercury (THg), monomethlymercury (MMHg), and total organic carbon (TOC) in surficial sediments revealed fire, followed by storm runoff, enhanced the transport of mercury and organic matter to the reservoir . Concentrations of THg in sediments increased from 7.5 etag/g in July to 46.1 etag/g by November 1995 at one site (Palomas) nearest the outflow of Palomas Creek . No other spatial or temporal trends were observed for THg at other sites throughout the remainder of the study . Concentrations of MMHg in sediments at the Palomas site increased from 0.428 etag/g in July to 12.46 etag/g by October 1995 compared to concentrations in sediments at the remaining sites which ranged from 0.11 to 1.50 etag/g throughout the study . The ratio of MMHg to THg (a gross index of methylation activity) was greatest in sediments from the Palomas site (5.4-33.8%) compared to the remaining sites (0.01-3.60%) . The ratio was mirrored by elevated TOC in sediments at the Palomas site (2.5-11.8%) that remained elevated throughout the study . Fire and subsequent late-summer rains may have had a twofold effect on mercury concentrations in Caballo Reservoir . The storm-driven runoff following the forest fire carried mercury complexed to organic matter which resulted in elevated levels of mercury as well as providing a carbon source for microbial methylation processes in sediment.

Cell, 2000 Sep 15, 102(6), 827 - 37
The polymeric immunoglobulin receptor translocates pneumococci across human nasopharyngeal epithelial cells; Zhang JR et al.; The polymeric immunoglobulin receptor (pIgR) plays a crucial role in mucosal immunity against microbial infection by transporting polymeric immunoglobulins (pIg) across the mucosal epithelium . We report here that the human pIgR (hpIgR) can bind to a major pneumococcal adhesin, CbpA . Expression of hpIgR in human nasopharyngeal cells and MDCK cells greatly enhanced pneumococcal adherence and invasion . The hpIgR-mediated bacterial adherence and invasion were abolished by either insertional knockout of cbpA or antibodies against either hpIgR or CbpA . In contrast, rabbit pIgR (rpIgR) did not bind to CbpA and its expression in MDCK cells did not enhance pneumococcal adherence and invasion . These results suggest that pneumococci are a novel example of a pathogen co-opting the pIg transcytosis machinery to promote translocation across a mucosal barrier.

Dig Liver Dis, 2000 Jun-Jul, 32(5), 440 - 6
Virus, liver and autoimmunity; Bogdanos DP et al.; The immune system's ability to distinguish self from nonself is essential for both host defence against microbial antigens and protection of self-antigens from autoimmune destruction . Such discrimination is complicated by extensive structural homology shared between micro-organisms and self-antigens, a condition known as molecular mimicry . Molecular mimicry provides the foundation for an immune response directed against an exogenous agent such as a virus to cross-react with mimicked host self sequences, leading to autoimmunity, and in some cases, tissue injury and autoimmune disease . In this review we analyse studies investigating the role of molecular mimicry and cross-reactive immunity in liver-related autoimmunity.

Eur J Anaesthesiol, 2000 Nov, 17(11), 680 - 7
A central venous catheter coated with benzalkonium chloride for the prevention of catheter-related microbial colonization; Moss HA et al.; In an attempt to overcome infections associated with central venous catheters, a new antiseptic central venous catheter coated with benzalkonium chloride on the internal and external surfaces has been developed and evaluated in a clinical trial . Patients (235) randomly received either a triple-lumen central venous catheter coated with benzalkonium chloride (117) or a polyurethane non-antiseptic catheter (118) . The incidence of microbial colonization of both catheters and retained antiseptic activity of the benzalkonium chloride device following removal were determined . The benzalkonium chloride resulted in a significant reduction of the incidence of microbial colonization on both the internal and external catheter surfaces . The reduction in colonization was detected at both the intradermal (21 benzalkonium chloride catheters vs . 38 controls, P = 0.0016) and distal segments of the antiseptic-coated catheters . Following catheter removal retained activity was demonstrated in benzalkonium chloride catheters which had been in place for up to 12 days . No patients developed adverse reactions to the benzalkonium chloride catheters . The findings demonstrate that the benzalkonium chloride catheter significantly reduced the incidence of catheter-associated colonization.

J Bacteriol, 2000 Nov, 182(21), 6042 - 8
Energy taxis is the dominant behavior in Azospirillum brasilense; Alexandre G et al.; Energy taxis encompasses aerotaxis, phototaxis, redox taxis, taxis to alternative electron acceptors, and chemotaxis to oxidizable substrates . The signal for this type of behavior is originated within the electron transport system . Energy taxis was demonstrated, as a part of an overall behavior, in several microbial species, but it did not appear as the dominant determinant in any of them . In this study, we show that most behavioral responses proceed through this mechanism in the alpha-proteobacterium Azospirillum brasilense . First, chemotaxis to most chemoeffectors typical of the azospirilla habitat was found to be metabolism dependent and required a functional electron transport system . Second, other energy-related responses, such as aerotaxis, redox taxis, and taxis to alternative electron acceptors, were found in A . brasilense . Finally, a mutant lacking a cytochrome c oxidase of the cbb(3) type was affected in chemotaxis, redox taxis, and aerotaxis . Altogether, the results indicate that behavioral responses to most stimuli in A . brasilense are triggered by changes in the electron transport system.

Am J Respir Crit Care Med, 2000 Oct, 162(4 Pt 1), 1348 - 54
(1-->3)-beta-D-glucan and endotoxin in house dust and peak flow variability in children; Douwes J et al.; House dust-associated bacterial endotoxins have been shown to be associated with asthma severity, and a similar role has been suggested for fungal (1-->3)-beta-D-glucans . In this study the relation between these agents and peak expiratory flow (PEF) variability was investigated in 148 children 7 to 11 yr of age of whom 50% had self- or parent-reported chronic respiratory symptoms . All children self-monitored twice daily their PEF for a period of 16 wk . Dust samples were collected from mattresses and from living room and bedroom floors, and endotoxin and (1-->3)-beta-D-glucan were measured in dust extracts . The relations with mean daily PEF variability (Ampl%mean) were investigated by linear regression analysis, adjusting for dust mite allergen levels, presence of pets, and type of floor cover . In unadjusted analyses the levels of both endotoxin and (1--> 3)-beta-D-glucan per square meter of living room floor were significantly associated with PEF-variability (but not when expressed per gram of sampled dust), particularly in atopic children with asthma symptoms . Adjusted analyses showed the same association for (1--> 3)-beta-D-glucan but not for endotoxin . Although no associations were found with microbial agent levels in bedroom floor or mattress dust, high levels of (1-->3)-beta-D-glucan in living room floor dust apparently increase PEF variability in asthmatic children.

Regul Toxicol Pharmacol, 2000 Aug, 32(1), 68 - 72
Incorporating model uncertainties along with data uncertainties in microbial risk assessment; Kang SH et al.; Much research on food safety has been conducted since the National Food Safety Initiative of 1997 . Risk assessment plays an important role in food safety practices and programs, and various dose-response models for estimating microbial risks have been investigated . Several dose-response models can provide reasonably good fits to the data in the experimental dose range, but yield risk estimates that differ by orders of magnitude in the low-dose range . Hence, model uncertainty can be just important as data uncertainty (experimental variation) in risk assessment . Although it is common in risk assessment to account for data uncertainty, it is uncommon to account for model uncertainties . In this paper we incorporate data uncertainties with confidence limits and model uncertainties with a weighted average of an estimate from each of various models . A numerical tool to compute the maximum likelihood estimates and confidence limits is addressed . The proposed method for incorporating model uncertainties is illustrated with real data sets .

Onderstepoort J Vet Res, 2000 Jun, 67(2), 105 - 14
Morphological changes associated with the development of the rumino-reticulum in growing lambs fed different rations; Swan GE et al.; Morphological changes associated with rumino-reticular development was compared in two groups of SA Mutton Merino lambs (n = 12) fed different diets at 3-5 weeks, 17-19 weeks and 31-33 weeks of age . Diet groups were identified as MMH or MHH according to the sequence at which the full-milk replacement (M) or hay (H) diet were fed to the lambs over the three study phases (phases I, II and III) preceding each age period . Prominent differences in the morphology (size and volume) and function (pH, proteolytic activity and microbial population) of the rumino-reticulum were observed in indicator lambs (n = 2) killed at every age period and also between milk-fed and hay-fed lambs . The size of the rumino-reticulum was rudimentary at 3-5 weeks of age and remained underdeveloped in lambs at 17-19 weeks of age which had received a full-milk replacement diet during phase II . One lamb, slaughtered at 3 weeks of age, showed a large distended rumen with severe sloughing of the surface cells of the stratum corneum.The size of the rumino-reticulum increased in size (2 x) in lambs which were fed hay relative to the milk-fed lambs during phase II and reached adult proportions in all lambs at 31-33 weeks of age . Ultrastructural examination showed that rumen papillae were more developed in lambs fed hay during phase II when compared to those of milk-fed lambs . Rumen papillae were best developed in phase III lambs.

Eur Respir J, 2000 Sep, 16(3), 464 - 71
Allergic bronchopulmonary aspergillosis in cystic fibrosis . A European epidemiological study . Epidemiologic Registry of Cystic Fibrosis; Mastella G et al.; Allergic bronchopulmonary aspergillosis (ABPA) is a disease resulting from a hypersensitivity response to Aspergillus fumigatus, although the pathogenesis of the disease is unknown and its prevalence in cystic fibrosis (CF) is still poorly defined . Data from the Epidemiologic Registry of Cystic Fibrosis (ERCF) on 12,447 CF patients gathered from 224 CF centres in nine European countries were analysed . The ERCF definition of ABPA diagnosis is a positive skin test and serum precipitins to A . fumigatus, together with serum immunoglobulin (Ig)E levels >1,000 U x mL(-1) and additional clinical or laboratory parameters . The overall prevalence of ABPA in the ERCF population was 7.8% (range: 2.1% in Sweden to 13.6% in Belgium) . Prevalence was low <6 yrs of age but was almost constant approximately 10% thereafter . No sex differences were observed . ABPA affected 8.0% of patients with a deltaF508/deltaF508 genotype and 5-6% with deltaF508/G551D, deltaF508/G542X and deltaF508/N1303K genotypes . ABPA patients presented a lower forced expiratory volume in one second (FEV1) than those without ABPA at any age and the prevalence ranged from 6.6% in patients with FEV1 > or =20-12.9% in those with FEV1 <40% . ABPA was associated with higher rates of microbial colonization, pneumothorax and massive haemoptysis, and with higher IgG serum levels and poorer nutritional status . A mixed model regression analysis of lung function showed that FEVI decline during the follow-up period was not substantially different in ABPA patients compared with non-ABPA patients for any subgroups based on age or disease severity at enrollment . To conclude, allergic bronchopulmonary aspergillosis is a frequent complication in cystic fibrosis patients, particularly after the age of 6 yrs, and it is generally associated with a poorer clinical condition . However, any clear independent influence of allergic bronchopulmonary aspergillosis on the rate of lung function decline in the short term was not shown.

Psychopharmacology (Berl), 2000 Sep, 151(4), 299 - 305
Endomorphin-1 induces antinociception without immunomodulatory effects in the rat; Carrigan KA et al.; RATIONALE: Although there is evidence that central opioid receptors are involved in immunomodulation, it has been only recently that an endogenous agonist, designated endomorphin-1, possessing high selectivity and affinity for the mu opioid receptor has been identified . OBJECTIVE: The present study assesses the immunomodulatory effects of endomorphin- in the rat and provides further evaluation of the antinociceptive effects of endomorphin-1 . METHODS: Rats were surgically implanted with cannulae directed at the lateral cerebral ventricle . Animals received vehicle or endomorphin-1 at doses of 31.63 or 56.23 microg (ICV) and were tested for antinociception in two different assays, the warm water tail withdrawal procedure and the hotplate assay . Additional studies assessed the effect of naltrexone on the antinociception produced by endomorphin-1 in both antinociceptive assessments . Assessments of immune status following endomorphin-1 treatment included measurements of splenic natural killer cell activity, production of interferon-y, and lymphocyte proliferative responses to mitogenic stimulation by Con-A, LPS, and the microbial superantigen, TSST-1 . RESULTS: Endomorphin-1 induced significant and naltrexone reversible antinociception 30 and 60 min following drug administration, as measured by the hotplate assay and warm water tail withdrawal procedure . In marked contrast, endomorphin-1 did not produce immunomodulatory effects up to 120 min following ICV administration . CONCLUSIONS: Endomorphin-1 produces antinociception but does not induce immunomodulatory effects in the rat . These findings suggest that it is possible to develop therapeutic strategies for separating antinociception and immunomodulatory properties through the mu opioid receptor.

Comp Biochem Physiol B Biochem Mol Biol, 2000 Aug, 126(4), 599 - 608
Purification and properties of alanine racemase from crayfish Procambarus clarkii; Shibata K et al.; Fresh water crayfish Procambarus clarkii is known to accumulate D-alanine remarkably in muscle after seawater acclimation, accompanied by an increase in alanine racemase activity . We have purified alanine racemase from crayfish muscle to homogeneity . The enzyme is a monomeric protein with a molecular mass of 58 kDa . It is highly specific to alanine and does not racemize L-serine, L-aspartate, L-glutamate, L-valine and L-arginine . The enzyme shows the highest activity at pH 9.0 in the conversion of L- to D-alanine and at pH 8.5 in the reverse conversion . Properties such as amino acid sequence, quaternary structure, pyridoxal 5'-phosphate (PLP)-dependency, pH-dependency and kinetic parameters seem to be distinct from those of the microbial alanine racemases . Various salts including NaCl at concentrations around seawater level were potently inhibitory for the activity in both of L- to -D and D- to -L direction.

Curr Opin Biotechnol, 2000 Oct, 11(5), 478 - 83
Filamentous fungi as microbial cell factories for food use; Archer DB; The knowledge base that will underpin the more efficient use of filamentous fungi as cell factories in food has increased during the past year in the areas of gene regulation, protein secretion, safety and synthesis of ingredients such as long-chain polyunsaturated fatty acids.

Curr Opin Biotechnol, 2000 Oct, 11(5), 434 - 9
Detection of genes essential in specific niches by signature-tagged mutagenesis; Lehoux DE et al.; Variations of the signature-tagged mutagenesis (STM) technique are now possible and the method can be applied to most pathogens that have an STM-selectable phenotype in a host system . STM screening of 15,040 mutants from 11 bacterial species identified 323 in vivo attenuated mutants . As a genome-scanning tool, STM will yield information about genes with unknown functions as well as information crucial for understanding microbial pathogenesis.

Contraception, 2000 Jul, 62(1), 39 - 43
Anti-nidatory effect of vaginally administered (Ala8,13, 18)-magainin II amide in the rhesus monkey; Dhawan L et al.; The hypothesis that timed application of a potent anti-microbial, anti-tumor agent like magainin peptides can inhibit blastocyst implantation was examined in the present study using the rhesus monkey as the primate model . Incidence of pregnancy, vaginal bleeding patterns, serum levels of progesterone, estrogen and monkey chorionic gonadotropin were examined following vaginal administration of (Ala8,13,18)-magainin II amide, a synthetic analogue of magainin 2, via tampon during days 20 to 26 of mated cycles . Implantation occurred in two out of three animals following administration of 0.25 mg magainin, while administration of 0.5 mg (Ala8,13,18)-magainin II amide resulted in inhibition of implantation in all females with no change in lengths of treatment cycles, and subsequent cycles . It appears from the present study that, besides being a local microbicidal agent, intravaginal administration of (Ala(8,13,18))-magainin II amide is a potential anti-implantation strategy for intercepting pregnancy.

Nucleic Acids Res, 2000 Oct 15, 28(20), 4029 - 36
Automatic detection of conserved gene clusters in multiple genomes by graph comparison and P-quasi grouping; Fujibuchi W et al.; We previously reported two graph algorithms for analysis of genomic information: a graph comparison algorithm to detect locally similar regions called correlated clusters and an algorithm to find a graph feature called P-quasi complete linkage . Based on these algorithms we have developed an automatic procedure to detect conserved gene clusters and align orthologous gene orders in multiple genomes . In the first step, the graph comparison is applied to pairwise genome comparisons, where the genome is considered as a one-dimensionally connected graph with genes as its nodes, and correlated clusters of genes that share sequence similarities are identified . In the next step, the P-quasi complete linkage analysis is applied to grouping of related clusters and conserved gene clusters in multiple genomes are identified . In the last step, orthologous relations of genes are established among each conserved cluster . We analyzed 17 completely sequenced microbial genomes and obtained 2313 clusters when the completeness parameter P: was 40% . About one quarter contained at least two genes that appeared in the metabolic and regulatory pathways in the KEGG database . This collection of conserved gene clusters is used to refine and augment ortholog group tables in KEGG and also to define ortholog identifiers as an extension of EC numbers.

Free Radic Res, 2000 Oct, 33(4), 341 - 8
Hemoglobin-based blood substitutes and the hazards of blood radicals; Alayash AI; Cell-free hemoglobins, chemically altered or genetically expressed in microbial host systems, have been developed as oxygen-carrying therapeutics . Site-directed modifications are introduced and serve to stabilize the protein molecules in a tetrameric and/or a polymeric functional form . Animal studies, as well as recent clinical studies, have suggested these proteins probably deliver oxygen to tissues . However, concerns still persist regarding the interference of hemoglobin and its oxidation products with the vascular redox balance, potentially impeding its clinical usefulness . This article reviews our current understanding of heme-mediated toxicities and some of the emerging protective strategies used to overcome hemoglobin side reactions.

Nippon Geka Gakkai Zasshi, 2000 Sep, 101(9), 588 - 92
{Infection and regulation of leukocytes by autonomic nervous system}; Abo T; The distribution of leukocyte pattern (i.e., the granulocyte to lymphocyte ratio) is changed by microbial infections . We found that the autonomic nervous system is also intimately associated with the inflammatory response . Even in the absence of such infections, the distribution of leukocyte pattern is changed by stimulation of the autonomic nervous system itself . When these changes become significant, people are susceptible to diseases such as purulent inflammation or allergic inflammation.

Isotopes Environ Health Stud, 2000, 36(1), 35 - 47
13C and 15N distributions in three spodic dystric cambisols under beech and spruce; Heil B et al.; The study of natural isotopic abundance signatures is useful to gain further insights in the processes resulting in depthwise changes in the composition of soil organic matter (SOM) . Objectives were to describe the delta 13C and delta 15N abundances of SOM with depth in soils from a 153-year old beech (B1), a 119-year old spruce (F1) and a 61-year old spruce (F2) stand at Solling, north-west Germany, and to study, how podzolisation affects the isotopic abundances of 13C and 15N in the SOM . The degree of podzolisation decreased in the order F1 > B1 > F2 . At the surface of the humus layer of all three sites, delta 13C values are approximately 1 to 4/1000 higher than in the leaves and needles, probably mainly due to the discrimination of 13C by microbial decomposition . 13C abundances in the organic layers of B1 and F2 increased only slightly from -27.6/1000 PDB (B1, L) to -27.2/1000 PDB (B1, Oh) and from -26.3/1000 PDB (F2, L) to -25.9/1000 PDB (F2, Oh), suggesting that biotic activity resulted in mixing of organic matter . At F1, however, 13C abundance increased from -27.5/1000 PDB (L) to -26.0/1000 PDB (Oh) which reflects the lack of mixing by animals . In the upper 2-4 cm of the mineral soil, i.e., in the eluvial horizons Aeh, 13C values showed a minimum at the spruce sites which was presumably related to a translocation of 13C enriched fulvic acids . Depthwise changes in delta 15N values were not related to podzolisation processes . At all three sites, a 13N enrichment with depth occurred in the mineral soil which is the result of the discrimination of 15N by microbial decomposition.

Crit Rev Oral Biol Med, 2000, 11(3), 366 - 80
Food starches and dental caries; Lingstrom P et al.; Sucrose and starches are the predominant dietary carbohydrates in modern societies . While the causal relationship between sucrose and dental caries development is indisputable, the relationship between food starch and dental caries continues to be debated and is the topic of this review . The current view of dental caries etiology suggests that in-depth evaluation of the starch-caries relationship requires the consideration of several critical cariogenic determinants: (1) the intensity (i.e., the amount and frequency) of exposure of tooth surfaces to both sugars and starches, (2) the bioavailability of the starches, (3) the nature of the microbial flora of dental plaque, (4) the pH-lowering capacity of dental plaque, and (5) the flow rate of saliva . Studies of caries in animals, human plaque pH response, and enamel/dentin demineralization leave no doubt that processed food starches in modern human diets possess a significant cariogenic potential . However, the available studies with humans do not provide unequivocal data on their actual cariogenicity . In this regard, we found it helpful to distinguish between two types of situations . The first, exemplified by our forebears, people in developing countries, and special subject groups in more modern countries, is characterized by starch consumption in combination with a low sugar intake, an eating frequency which is essentially limited to two or three meals per day, and a low-to-negligible caries activity . The second, exemplified by people in the more modern societies, e.g., urban populations, is characterized by starch consumption in combination with significantly increased sugar consumption, an eating frequency of three or more times per day, and a significantly elevated caries activity . It is in the first situation that food starches do not appear to be particularly caries-inducive . However, their contribution to caries development in the second situation is uncertain and requires further clarification . Although food starches do not appear to be particularly caries inducive in the first situation, the possibility cannot be excluded that they contribute significantly to caries activity in modern human populations . The commonly used term "dietary starch content" is misleading, since it represents a large array of single manufactured and processed foods of widely varying composition and potential cariogenicity . Hence, increased focus on the cariogenicity of single starchy foods is warranted . Other aspects of starchy foods consumption, deserving greater attention, include the bioavailability of starches in processed foods, their retentive properties, also in relation to sugars present (starches as co-cariogens), their consumption frequency, the effect of hyposalivation on their cariogenicity, and their impact on root caries . The starch-caries issue is a very complex problem, and much remains uncertain . More focused studies are needed . At present, it appears premature to consider or promote food starches in modern diets as safe for teeth.

Crit Rev Oral Biol Med, 2000, 11(3), 333 - 55
Biocompatibility of resin-modified filling materials; Geurtsen W; Increasing numbers of resin-based dental restorations have been placed over the past decade . During this same period, the public interest in the local and especially systemic adverse effects caused by dental materials has increased significantly . It has been found that each resin-based material releases several components into the oral environment . In particular, the comonomer, triethyleneglycol di-methacrylate (TEGDMA), and the 'hydrophilic' monomer, 2-hydroxy-ethyl-methacrylate (HEMA), are leached out from various composite resins and 'adhesive' materials (e.g., resin-modified glass-ionomer cements {GICs} and dentin adhesives) in considerable amounts during the first 24 hours after polymerization . Numerous unbound resin components may leach into saliva during the initial phase after polymerization, and later, due to degradation or erosion of the resinous restoration . Those substances may be systemically distributed and could potentially cause adverse systemic effects in patients . In addition, absorption of organic substances from unpolymerized material, through unprotected skin, due to manual contact may pose a special risk for dental personnel . This is borne out by the increasing numbers of dental nurses, technicians, and dentists who present with allergic reactions to one or more resin components, like HEMA, glutaraldehyde, ethyleneglycol di-methacrylate (EGDMA), and dibenzoyl peroxide (DPO) . However, it must be emphasized that, except for conventional composite resins, data reported on the release of substances from resin-based materials are scarce . There is very little reliable information with respect to the biological interactions between resin components and various tissues . Those interactions may be either protective, like absorption to dentin, or detrimental, e.g., inflammatory reactions of soft tissues . Microbial effects have also been observed which may contribute indirectly to caries and irritation of the pulp . Therefore, it is critical, both for our patients and for the profession, that the biological effects of resin-based filling materials be clarified in the near future.

APMIS Suppl, 2000, 100, 1 - 59
Collectins and collectin receptors in innate immunity; Holmskov UL; This thesis is based on nine papers and a review on the collectins and collectin receptors in innate immunity . The collectins are a family of proteins in which the individual chains consist of a C-type lectin domain attached to a collagen domain via an alpha-coiled neck region . The chains are organized into a triple collagen helix and oligomerized through N-terminally located cysteines . The collectins have a dual function: one is to bind specifically to carbohydrate structures on the surface of a pathogen; the other is subsequently to recruit other cells and molecules to destroy the pathogen . The C-type lectin domains contain 110-130 amino-acid residues arranged in a conserved sequence pattern which allows the domain to fold into a well-defined tertiary structure . Five collectins have been described . Lung surfactant proteins A and D (SP-A and SP-D) are mainly found in the surfactant coating the luminal surface of the pulmonary epithelial cells, but are also produced by cells lining the gastrointestinal tract . Mannan-binding lectin (MBL), conglutinin and collectin-43 (CL-43) are serum proteins produced by the liver . Conglutinin and CL-43 have so far only been found in Bovidae . The collectins are involved in innate, nonadaptive immune defense . They bind to microbial surface carbohydrates, inducing aggregation and thereby impeding infectivity or mediating phagocytosis through specific receptors on the phagocytes . After binding microbial carbohydrate, MBL can activate the complement system through a newly discovered pathway which makes use of two serine proteases (MASP-1 and MASP-2) to activate the complement factors C4 and C2 . In man, low serum MBL concentrations resulting from mutations in the collagen region are associated with a common opsonic defect . CL-43 was identified as a new collectin by its calcium-dependent binding to mannan and by its M(r) of 43 kDa in the reduced state on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) . The lectin was isolated by affinity chromatography on mannan-Sepharose, absorption with rabbit anti-bovine Ig coupled to Sepharose-4B and ion-exchange chromatography . CL-43 shows an apparent molecular mass of 120 kDa in the unreduced state on SDS-PAGE and elutes with an apparent molecular mass of 750 kDa on gel chromatography under nondissociating conditions . Amino-acid analysis and susceptibility to collagenase digestion indicated that CL-43 was a collectin . Electron microscopy of purified CL-43 revealed only rod-like monomer subunits 37.4 nm long . Two-dimensional gel electrophoresis showed that CL-43 has two isoforms of pI 4.9 and 5.3 respectively, corresponding to the native form of CL-43 and a truncated form which lacks the first 9 amino-acid residues . The N-terminal amino-acid sequence of CL-43 was used to design primers for PCR with a bovine liver cDNA as template . The cDNA of CL-43 was cloned and the open reading frame was found to encode a protein of 301 amino-acid residues, including an N-terminal region of 28 residues, a collagen region of 114 residues and a neck-CRD region of 159 residues . The amino-acid sequence of CL-43 shows 74% identity with bovine conglutinin and 70% identity with bovine SP-D, but the collagen region is considerably shorter than those of conglutinin and SP-D . Northern blot analysis showed that CL-43 was only synthesized in bovine liver, no signal being detected in a variety of other bovine tissues, including lung . No cross-hybridizing signals were detected in mRNA from ovine, human, rat or mouse liver . Since CL-43 and conglutinin have only been detected in members of the Bovidae, it is probable that an ancestral gene of these two proteins was first derived from a SP-D-like gene and that this ancestral gene underwent duplication during evolution . The carbohydrate binding profile of CL-43 was analyzed by an inhibition assay with biotinylated CL-43, using solid-phase mannan as the ligand . (ABSTRACT TRUNCATED)

Microbes Infect, 2000 Sep, 2(11), 1343 - 51
Structure of the intestinal flora responsible for development of the gut immune system in a rodent model; Umesaki Y et al.; The intestinal flora comprising indigenous, autochthonous bacteria is constantly present in the alimentary tract of host animals, including humans . The indigenous bacteria greatly affect the structure and functions of the intestinal mucosa . Studies involving gnotobiotic mice or rats have shown that the presence of limited kinds of intestinal bacteria is responsible for the development of the gut immune system, such as secretory IgA, major histocompatibility complex molecules and intraepithelial lymphocytes . Understanding of the structure of the intestinal flora or the organization of the microbial population in the intestine, based on evaluation of the immunological responses, may clarify its functions in the host animal.

J Microbiol Methods, 2000 Oct, 42(2), 203 - 7
The addition of low numbers of 3' thymine bases can be used to improve the hybridization signal of oligonucleotides for use within arrays on nylon supports; Brown TJ et al.; Oligonucleotide arrays can be used for the analysis of microbial nucleic acid . The addition of high numbers of dTTP to the 3' ends of oligonucleotides using terminal transferase has been shown to facilitate membrane binding . This paper demonstrates low numbers of thymine bases added to the 3' end of oligonucleotides during synthesis can improve hybridisation signal intensity where the signal seen with the unmodified oligonucleotides is poor . Thus, the addition of variable numbers of thymine bases to different oligonucleotides allows the production of oligonucleotide arrays producing strong interpretable hybridisation signals.

J Microbiol Methods, 2000 Oct, 42(2), 167 - 74
Discrimination among iron sulfide species formed in microbial cultures; Popa R et al.; A quantitative method for the study of iron sulfides precipitated in liquid cultures of bacteria is described . This method can be used to quantify and discriminate among amorphous iron sulfide (FeS(amorph)), iron monosulfide minerals such as mackinawite or greigite (FeS(min)), and iron disulfide minerals such as pyrite or marcasite (FeS(2min)) formed in liquid cultures . Degradation of iron sulfides is performed using a modified Cr(2+) reduction method with reflux distillation . The basic steps of the method are: first, separation of FeS(amorph); second, elimination of interfering species of S such as colloidal sulfur (S(c) degrees ), thiosulphate (S(2)O(3)(2-)) and polysulfides (S(x)(2-)); third, separation of FeS(min); and fourth, separation of FeS(2min) . The final product is H(2)S which is determined after trapping . The efficiency of recovery is 96-99% for FeS(amorph), 76-88% for FeS(min), and >97% for FeS(2min) . This method has a high reproducibility if the experimental conditions are rigorously applied and only glass conduits are used . A well ventilated fume hood must be used because of the toxicity and volatility of several reagents and products . The advantage relative to previously described methods are better resolution for iron sulfide species and use of the same bottles for both incubation of cultures and acid degradation . The method can also be used for Fe/S stoichiometry with sub-sampling and Fe analysis.

Mol Biol Evol, 2000 Oct, 17(10), 1467 - 82
Microsatellite markers reveal a spectrum of population structures in the malaria parasite Plasmodium falciparum; Anderson TJ et al.; Multilocus genotyping of microbial pathogens has revealed a range of population structures, with some bacteria showing extensive recombination and others showing almost complete clonality . The population structure of the protozoan parasite Plasmodium falciparum has been harder to evaluate, since most studies have used a limited number of antigen-encoding loci that are known to be under strong selection . We describe length variation at 12 microsatellite loci in 465 infections collected from 9 locations worldwide . These data reveal dramatic differences in parasite population structure in different locations . Strong linkage disequilibrium (LD) was observed in six of nine populations . Significant LD occurred in all locations with prevalence <1% and in only two of five of the populations from regions with higher transmission intensities . Where present, LD results largely from the presence of identical multilocus genotypes within populations, suggesting high levels of self-fertilization in populations with low levels of transmission . We also observed dramatic variation in diversity and geographical differentiation in different regions . Mean heterozygosities in South American countries (0.3-0.4) were less than half those observed in African locations (0 . 76-0.8), with intermediate heterozygosities in the Southeast Asia/Pacific samples (0.51-0.65) . Furthermore, variation was distributed among locations in South America (F:(ST) = 0.364) and within locations in Africa (F:(ST) = 0.007) . The intraspecific patterns of diversity and genetic differentiation observed in P . falciparum are strikingly similar to those seen in interspecific comparisons of plants and animals with differing levels of outcrossing, suggesting that similar processes may be involved . The differences observed may also reflect the recent colonization of non-African populations from an African source, and the relative influences of epidemiology and population history are difficult to disentangle . These data reveal a range of population structures within a single pathogen species and suggest intimate links between patterns of epidemiology and genetic structure in this organism.

Anal Biochem, 2000 Oct 15, 285(2), 225 - 9
Selective and sensitive biosensor for theophylline based on xanthine oxidase electrode; Stredansky M et al.; Milk and microbial xanthine oxidases (XOs) were used for the construction of amperometric enzyme electrodes . Substrate specificity differences of these enzymes were studied . Of the two enzymes, only the microbial XO was found to oxidize theophylline, but not theobromine and caffeine . The substrate specificity of microbial XO was affected by pH, where the optimum for xanthine was 5.5, while for theophylline it was in the range from 6.5 to 8.5 . The theophylline biosensor showed a low detection limit of 2 x 10(-7) M and signal linearity up to 5 x 10(-5) M . The sensitivity of the microbial XO electrode to theophylline could be selectively eliminated by immersion in alkaline phosphate solution, thus allowing for the construction of a blank electrode for differential measurements . The feasibility of this approach has been demonstrated by the determination of free (unbound) and total theophylline in blood samples . The biosensor exhibited good operational (>6 h) and shelf (>3 months) stability when trehalose was used as a stabilizer of the biocatalytic layer .

Nat Med, 2000 Oct, 6(10), 1147 - 53
Secretory leukocyte protease inhibitor mediates non-redundant functions necessary for normal wound healing; Ashcroft GS et al.; Secretory leukocyte protease inhibitor (SLPI) is a serine protease inhibitor with anti-microbial properties found in mucosal fluids . It is expressed during cutaneous wound healing . Impaired healing states are characterized by excessive proteolysis and often bacterial infection, leading to the hypothesis that SLPI may have a role in this process . We have generated mice null for the gene encoding SLPI (Slpi), which show impaired cutaneous wound healing with increased inflammation and elastase activity . The altered inflammatory profile involves enhanced activation of local TGF-beta in Slpi-null mice . We propose that SLPI is a pivotal endogenous factor necessary for optimal wound healing.

J Exp Med, 2000 Oct 2, 192(7), 965 - 76
CD1b-mediated T cell recognition of a glycolipid antigen generated from mycobacterial lipid and host carbohydrate during infection; Moody DB et al.; T cells recognize microbial glycolipids presented by CD1 proteins, but there is no information regarding the generation of natural glycolipid antigens within infected tissues . Therefore, we determined the molecular basis of CD1b-restricted T cell recognition of mycobacterial glycosylated mycolates, including those produced during tissue infection in vivo . Transfection of the T cell receptor (TCR) alpha and beta chains from a glucose monomycolate (GMM)-specific T cell line reconstituted GMM recognition in TCR-deficient T lymphoblastoma cells . This TCR-mediated response was highly specific for natural mycobacterial glucose-6-O-(2R, 3R) monomycolate, including the precise structure of the glucose moiety, the stereochemistry of the mycolate lipid, and the linkage between the carbohydrate and the lipid . Mycobacterial production of antigenic GMM absolutely required a nonmycobacterial source of glucose that could be supplied by adding glucose to media at concentrations found in mammalian tissues or by infecting tissue in vivo . These results indicate that mycobacteria synthesized antigenic GMM by coupling mycobacterial mycolates to host-derived glucose . Specific T cell recognition of an epitope formed by interaction of host and pathogen biosynthetic pathways provides a mechanism for immune response to those pathogenic mycobacteria that have productively infected tissues, as distinguished from ubiquitous, but innocuous, environmental mycobacteria.

Contact Dermatitis, 2000 Oct, 43(4), 200 - 5
Dyshidrotic eczema and occupation: a descriptive study; Lehucher-Michel MP et al.; Dyshidrotic eczema (DE) is a chronic and/or recurrent vesicular eczema of the palmar sides of the hands and sometimes the soles of the feet . The major aetiologies to be considered are atopic DE, contact DE, systemic contact DE and DE due to fungal or microbial allergy . To clarify the role of occupational contact allergens, 59 workers with DE were examined to determine whether they had atopy, environmental contact sensitization and/or contact allergy coupled with work activity . For all, patch testing was carried out using the European standard series and the material brought in by the patients . No significant difference was found between the rate (42.4%) of history of atopy among the 59 workers with DE and that (30.6%) observed among 160 workers with other forms of hand eczema (HE) . Positive patch test reactions were found in 72.8% of dyshidrotic patients . A relationship to occupational activity was strong for 18 (30.5%) of them but this rate was significantly lower than that (55%) observed among the 160 with other types of eczema . Results of the present study showed that contact sensitization is as much involved in workers with DE as in workers with other form of HE, even though the relationship to work is significantly less frequent for DE . It also suggests that atopic histories are no more involved in DE than in other forms of HE.

Appl Environ Microbiol, 2000 Oct, 66(10), 4279 - 91
Characterization, seasonal occurrence, and diel fluctuation of poly(hydroxyalkanoate) in photosynthetic microbial mats; Rothermich MM et al.; In situ poly(hydroxyalkanoate) (PHA) levels and repeating-unit compositions were examined in stratified photosynthetic microbial mats from Great Sippewissett Salt Marsh, Mass., and Ebro Delta, Spain . Unlike what has been observed in pure cultures of phototrophic bacteria, the prevalence of hydroxyvalerate (HV) repeating units relative to hydroxybutyrate (HB) repeating units was striking . In the cyanobacteria-dominated green material of Sippewissett mats, the mole percent ratio of repeating units was generally 1HB:1HV . In the purple sulfur bacteria-dominated pink material the relationship was typically 1HB:2HV . In Sippewissett mats, PHA contributed about 0.5 to 1% of the organic carbon in the green layer and up to 6% in the pink layer . In Ebro Delta mats, PHA of approximately 1HB:2HV-repeating-unit distribution contributed about 2% of the organic carbon of the composite photosynthetic layers (the green and pink layers were not separated) . Great Sippewissett Salt Marsh mats were utilized for more extensive investigation of seasonal, diel, and exogenous carbon effects . When the total PHA content was normalized to organic carbon, there was little seasonal variation in PHA levels . However, routine daily variation was evident at all sites and seasons . In every case, PHA levels increased during the night and decreased during the day . This phenomenon was conspicuous in the pink layer, where PHA levels doubled overnight . The daytime declines could be inhibited by artificial shading . Addition of exogenous acetate, lactate, and propionate induced two- to fivefold increases in the total PHA levels when applied in the daylight but had no effect when applied at night . The distinct diel pattern of in situ PHA accumulation at night appears to be related, in some phototrophs, to routine dark energy metabolism and is not influenced by the availability of organic nutrients.

Appl Environ Microbiol, 2000 Oct, 66(10), 4205 - 11
Rapid mineralization of benzo{a}pyrene by a microbial consortium growing on diesel fuel; Kanaly RA et al.; A microbial consortium which rapidly mineralized the environmentally persistent pollutant benzo{a}pyrene was recovered from soil . The consortium cometabolically converted {7-(14)C}benzo{a}pyrene to (14)CO(2) when it was grown on diesel fuel, and the extent of benzo{a}pyrene mineralization was dependent on both diesel fuel and benzo{a}pyrene concentrations . Addition of diesel fuel at concentrations ranging from 0.007 to 0.2% (wt/vol) stimulated the mineralization of 10 mg of benzo{a}pyrene per liter 33 to 65% during a 2-week incubation period . When the benzo{a}pyrene concentration was 10 to 100 mg liter(-1) and the diesel fuel concentration was 0.1% (wt/vol), an inoculum containing 1 mg of cell protein per liter (small inoculum) resulted in mineralization of up to 17.2 mg of benzo{a}pyrene per liter in 16 days . This corresponded to 35% of the added radiolabel when the concentration of benzo{a}pyrene was 50 mg liter(-1) . A radiocarbon mass balance analysis recovered 25% of the added benzo{a}pyrene solubilized in the culture suspension prior to mineralization . Populations growing on diesel fuel most likely promoted emulsification of benzo{a}pyrene through the production of surface-active compounds . The consortium was also analyzed by PCR-denaturing gradient gel electrophoresis of 16S rRNA gene fragments, and 12 dominant bands, representing different sequence types, were detected during a 19-day incubation period . The onset of benzo{a}pyrene mineralization was compared to changes in the consortium community structure and was found to correlate with the emergence of at least four sequence types . DNA from 10 sequence types were successfully purified and sequenced, and that data revealed that eight of the consortium members were related to the class Proteobacteria but that the consortium also included members which were related to the genera Mycobacterium and Sphingobacterium.

Cornea, 2000 Sep, 19(5), 650 - 8
Changing concepts in the diagnosis and management of blepharitis; McCulley JP et al.; PURPOSE: Chronic blepharitis is one of the most common conditions seen in the ophthalmologist's office; but, it is difficult to treat effectively . Nevertheless, much progress has been made over the last quarter century, not only in diagnosis but also in treatment of blepharitis . METHODS: Perhaps the most important progress has been made as the result of extensive and detailed clinical evaluations of patients, sometimes over extended periods of time with diverse patient populations . RESULTS: The availability of sophisticated instrumentation and laboratory techniques, as well as an array of antibiotics, has enhanced the whole picture for effective blepharitis treatment . CONCLUSION: Although much progress has been made, it is important to build on the present understanding . Most important is the continued development of targeted treatment protocols that address specific signs, as is now possible with microbial abnormalities . Progress in the understanding and treatment of relevant inflammatory processes will benefit from continuing biomedical discoveries.

Cornea, 2000 Sep, 19(5), 625 - 43
Twenty-five-year panorama of corneal immunology: emerging concepts in the immunopathogenesis of microbial keratitis, peripheral ulcerative keratitis, and corneal transplant rejection; Dana MR et al.; PURPOSE: To describe the most recent advances in our understanding of the cellular and molecular mechanisms involved in the immunopathogenesis of corneal immunoinflammatory disorders including microbial keratitis, peripheral ulcerative keratitis . and allograft rejection . METHODS: Review of the published peer-reviewed literature that has contributed significantly to our modern understanding of corneal immunology . In addition, the authors have summarized the information in conceptual diagrams that highlight the critical cellular and molecular pathways that lead to corneal immune responses in the two most thoroughly studied corneal immune disorders, herpes simplex keratitis (HSK) and transplant rejection . RESULTS: In spite of the wide array of molecular and cellular factors that mediate corneal immunity, critical mechanistic facets are shared by the various corneal immunoinflammatory disorders . These include activation and migration of local antigen-presenting cells (APCs), including Langerhans cells (LCs), upregulation in pleiotropic proinflammatory cytokines such as interleukin-1 (IL-1) and tumor necrosis factor-alfa (TNF-alpha) that can mediate a wide array of immune functions in addition to up-regulating protease expression . and chemokines that play a critical role on the one hand in attracting nonantigen-specific inflammatory cells such as neutrophils and on the other in attracting CD4+ T helper type 1 (Th1) cells that mediate most of the destruction in the cornea . CONCLUSIONS: In the last 25 years, we have seen our field develop from a descriptive stage into a new phase where the fundamental processes that mediate and effect corneal immunity are being accurately deciphered . It is anticipated that this new knowledge will allow development of specific molecular and genetic therapeutic strategies that could target critical steps in the immunopathogenesis of disease without the untoward side-effects of nonspecific generalized immune suppression that still remains the standard of care today.

Mikrobiologiia, 2000 Jul-Aug, 69(4), 453 - 64
{Structure and function of microbial communities in soil from the southern Taiga}; Golovchenko AV et al.; General regularities in the structure of the microbial communities of southern taiga soil ecosystems and taxonomic differences between the microbial communities of soils with different hydrothermal characteristics are discussed with reference to the main types of soils of the Central State Forest Biosphere Reserve.

Crit Care Med, 2000 Sep, 28(9 Suppl), S4 - 8
Pathophysiologic basis of sepsis: considerations for future strategies of intervention; Glauser MP; OBJECTIVE: To analyze the implications for therapeutic advances in the treatment of sepsis and septic shock based on the mechanisms underlying the response to microbial infection . DATA SOURCES: Clinical trials and experimental models designed to determine the key mechanisms involved in cellular and molecular processes of inflammatory reactions . STUDY SELECTION: Analyses of normal immune reactions to microbial infection, processes involved in the development of sepsis, and reasons for frequent failure of regimens based on current therapeutic rationales . DATA EXTRACTION/SYNTHESIS: Review of the data suggests that the dysregulation of the immune system resulting in sepsis/septic shock is ineffectually blocked by interfering with the action of inflammatory mediators or cascades, as these processes may be too complex to be easily antagonized . Rather, blockade of the action of microbial products or of the intracellular processes activated by receptor interaction with these products may provide a more promising therapeutic alternative . CONCLUSIONS: The sepsis syndrome induced by microbial pathogens reflects excessive stimulation of the processes of innate immunity . Bacterial components reacting with specific receptors activate intracellular processes, resulting in the release of excessive amounts of mediators of inflammation as well as triggering multiple complex proteolytic cascades . Blockade or antagonism of the actions of individual intermediary messenger molecules has proved therapeutically unsuccessful, because blockade of mediators alone does not block the direct activation of processes such as coagulation and complement . The dysregulation that characterizes sepsis may be amenable to blockade of the bacterial components or to the intracellular pathways triggered by these products.

Am J Ophthalmol, 2000 Jul, 130(1), 33 - 41
Treatment of persistent corneal epithelial defect with extended wear of a fluid-ventilated gas-permeable scleral contact lens; Rosenthal P et al.; PURPOSE: To report treatment of persistent corneal epithelial defects unresponsive to other therapies by extended wear of a fluid-ventilated gas-permeable scleral contact lens.METHODS: In this retrospective study, 14 eyes of 13 consecutive patients referred for the treatment of persistent corneal epithelial defects that failed to heal with conventional therapies or developed epithelial defects after penetrating keratoplasty for persistent corneal epithelial defects were fitted with an extended-wear gas-permeable scleral lens . These included seven eyes of six patients with Stevens-Johnson syndrome and seven eyes of seven patients who did not have Stevens-Johnson syndrome . Twelve eyes had undergone recent penetrating keratoplasty . All 14 eyes were fitted with a gas-permeable scleral contact lens designed to avoid the intrusion of air bubbles under its optic . An antibiotic and corticosteroid were added to the lens fluid reservoir or instilled before each lens insertion in 12 of 14 eyes . The lenses were worn continuously except for brief periods of removal for purposes of cleaning, replacement of the lens fluid reservoir, and examination and photography of the cornea.RESULTS: Five of the seven persistent corneal epithelial defects associated with Stevens-Johnson syndrome healed . The persistent corneal epithelial defects of four of these eyes re-epithelialized within 7 days, and a fifth healed in 27 days of gas-permeable scleral lens extended wear . A sixth persistent corneal epithelial defect that failed to heal initially re-epithelialized after a subsequent penetrating keratoplasty and gas-permeable scleral lens extended wear . The seventh eye healed after 3 days of gas-permeable scleral lens extended wear, but the persistent corneal epithelial defect subsequently recurred . Three of seven non-Stevens-Johnson syndrome persistent corneal epithelial defects re-epithelialized within 36 hours, 6 days, and 36 days, respectively . Of the six (six of 14) persistent corneal epithelial defects that failed to heal with a gas-permeable scleral lens extended wear, one subsequently healed after multiple amniotic membrane grafts . Microbial keratitis occurred in four eyes (four of 14) and graft failure in one eye, all of which required repeat penetrating keratoplasty.CONCLUSION: Extended wear of an appropriately designed gas-permeable scleral contact lens was effective in promoting the healing of persistent corneal epithelial defects in some eyes that failed to heal after other therapeutic measures . Re-epithelialization appears to be aided by a combination of oxygenation, moisture, and protection of the fragile epithelium afforded by the scleral lens . However, microbial keratitis represents a significant risk.

J Accid Emerg Med, 2000 Sep, 17(5), 341 - 3
Biological tissue adhesive for multiple use in the accident and emergency department; Gerrard C et al.; OBJECTIVE: To assess the strength of the glue and microbial contamination over 28 days from opening a vial of tissue adhesive in the accident and emergency setting, and to quantify cost savings of repeated use of the vials . METHOD: (1) Strips of reinforced nylon and a specially constructed piece of apparatus designed to measure the force at which the glue gave way were used to measure the strength of the tissue adhesive at various times after the glue was opened to assess if the glue strength deteriorated over time . (2) Microbial contamination of the glue was assessed . RESULTS: There was no deterioration in the glue strength over time . There was no evidence of microbial contamination of the glue . CONCLUSION: Cyanoacrylate tissue adhesive can safely be reused for a period of 28 days after opening with no risk of degradation of glue strength or contamination with micro-organisms . In our department this represents a potential saving of l5400 per year.

J Microbiol Methods, 2000 Sep, 42(1), 87 - 96
Glucose uptake rates of single E . coli cells grown in glucose-limited chemostat cultures; Natarajan A et al.; To evaluate the extent to which single-cell glucose uptake rates determine the overall specific growth rate of a culture, dilute chemostat cultures of Escherichia coli BL21 were grown in defined medium under glucose limitation . The glucose uptake dynamics of the cell population was examined at the single-cell level using the fluorescent glucose analog, 2-NBDG . Between dilution rates of 0.12 h(-1) and 0.40 h(-1), mean cellular protein content and steady-state, extracellular glucose concentrations increased with increasing dilution rate . However, the distribution of 2-NBDG uptake rates in the population remained constant over the range of dilution rates studied . This indicates that the growth of cells in continuous culture is not limited by the maximum rate of uptake of glucose but by the availability of glucose for transport . The work represents an example of how quantitative flow cytometry can be applied to gain detailed insight into microbial growth physiology.

Mol Immunol, 2000 Apr, 37(6), 311 - 20
A novel cellular interaction involving antigen-pulsed macrophage and antigen-specific B-lymphocytes; Weaver DJ Jr et al.; Extensive documentation shows that macrophage efficiently present antigen to CD 4(+) T-cells in conjunction with the MHC II molecule . Previously, a novel fluorescent probe, FITC-BSA, was developed to analyze intracellular antigen processing and presentation pathways within viable peritoneal murine macrophage . The studies revealed fluorescein's accessibility to antibody binding when associated with peptides bound within the MHC II cleft . To determine if MHC II-fluoresceinated-peptide complexes on the surface of macrophage were also sufficient to stimulate antigen-specific B-cells, nylon wool-purified splenic B-cells from FITC-KLH injected BALB/c mice (H-2(d)) were co-cultured with antigen-pulsed macrophage . B-cell stimulation and antibody production was observed in the presence of FITC-BSA-pulsed macrophage, whereas, macrophage incubated in the presence of unlabeled BSA were not stimulated . Compared with control cells, similar levels of stimulation were detected following depletion of Thy 1.2(+) cells from nylon wool-based spleen cell preparations . Stimulation was inhibited upon preincubation with anti-fluorescein IgG antibodies . Stimulation was not measurable using B-cells derived from the naive mice . The interaction was inhibited upon addition of MHC II specific antibodies and leupeptin, a microbial product that inhibits MHC II-peptide complex formation . Importantly, antibody production was not observed in the presence of antigen-pulsed macrophage from H-2(b) mice . Moreover, B-cell stimulation via this pathway was dependent upon antigen concentration as well as the cell to cell ratio.

Bioorg Med Chem Lett, 2000 Sep 18, 10(18), 2087 - 90
Synthesis and PKC isozyme surrogate binding of indothiolactam-V, a new thioamide analogue of tumor promoting indolactam-V; Nakagawa Y et al.; To investigate the role of the amide group of (-)-indolactam-V (1) on PKC binding, we synthesized (-)-indothiolactam-V (2), a new thioamide analogue of 1, by microbial conversion using Streptomyces blastmyceticum . Compounds 2 and 1 showed similar binding affinities to conventional PKCs but 2 had lower affinities to novel PKCs, suggesting that novel PKCs recognize amide modifications more effectively than conventional PKCs.

Emerg Infect Dis, 2000 Sep-Oct, 6(5), 505 - 12
Comparative genomics and understanding of microbial biology; Fraser CM et al.; The sequences of close to 30 microbial genomes have been completed during the past 5 years, and the sequences of more than 100 genomes should be completed in the next 2 to 4 years . Soon, completed microbial genome sequences will represent a collection of >200,000 predicted coding sequences . While analysis of a single genome provides tremendous biological insights on any given organism, comparative analysis of multiple genomes provides substantially more information on the physiology and evolution of microbial species and expands our ability to better assign putative function to predicted coding sequences.

Plant J, 2000 Sep, 23(6), 817 - 24
In vivo imaging of an elicitor-induced nitric oxide burst in tobacco; Foissner I et al.; A growing body of evidence suggests that nitric oxide (NO), an important signalling and defence molecule in mammals, plays a key role in activating disease resistance in plants, acting as signalling molecule and possibly as direct anti-microbial agent . Recently, a novel fluorophore (diaminofluorescein diacetate, DAF-2 DA) has been developed which allows bio-imaging of NO in vivo . Here we use the cell-permeable DAF-2 DA, in conjunction with confocal laser scanning microscopy, for real-time imaging of NO in living plant cells . Epidermal tobacco cells treated with cryptogein, a fungal elicitor from Phytophthora cryptogea, respond to the elicitor with a strong increase of intracellular NO . NO-induced fluorescence was found in several cellular compartments, and could be inhibited by a NO scavenger and an inhibitor of nitric oxide synthase . The NO burst was triggered within minutes, reminiscent of the oxidative burst during hypersensitive response reactions . These results reveal additional similarities between plant and animal host responses to infection.

Eur J Haematol, 2000 Feb, 64(2), 114 - 20
Flow cytometric analysis of defensins in blood and marrow neutrophils; Klut ME et al.; Polymorphonuclear neutrophils (PMN) are vital in host defense against microbial infections . This study provides a flow cytometric method for the quantitative analysis of microbicidal peptides (defensins) in cells of PMN lineage . Rabbit neutrophil peptides, NP-2 and NP-5, were measured in all PMN and in subpopulations of PMN expressing 1-selectin . PMN lineage counts were made on Wright's-stained blood smears and marrow cytospins . Immunoreactivity for NP-2, and NP-5 was detected by using the alkaline phosphatase anti-alkaline phosphatase technique . The results show that marrow PMN express higher levels of NP-2 and NP-5 than blood PMN, p < 0.001 and that these levels are associated with elevated numbers of myeloid precursors . In both blood and marrow, NP-2 occurs in two PMN subpopulations and the mean fluorescence intensity of NP-2 is consistently higher than that of NP-5 . Increased levels of defensins are observed in circulating PMN depicting the most 1-selectin p < 0.05 . Immunocytochemical results indicate that PMN defensins reside in cytoplasmic granules and are not constitutively expressed on the cell surface . Furthermore, defensins are not detected in monocytes, eosinophils, lymphocytes and erythrocytes . The flow cytometric method described here provides a novel means of quantitating host natural defenses, allows the characterization of PMN subpopulations and has clinical applications.

Toxicology, 2000 Sep 7, 150(1-3), 191 - 206
The immune system as a potential target for environmental estrogens (endocrine disrupters): a new emerging field; Ahmed SA; It is now well known that natural (17beta-estradiol) and synthetic (e.g . diethylstilbestrol) estrogens not only affect the reproductive system, but also markedly influence the immune system . Recently, a new class of estrogens that is abundant in the environment (in industrial chemicals, pesticides, and surfactants) has been recognized . Some of these estrogenic chemicals (which are a large subgroup of endocrine disrupters) have also been shown to influence the immune system . This review assimilates growing evidence in wildlife, laboratory animals and to a limited extent in humans, which suggests that environmental chemicals may also affect the immune system . Further studies are needed to ascertain the immunological consequences of exposure to environmental estrogens, especially in humans . At the present time, it is not known whether the human immune system responds to a low dose of environmental estrogens or if environmental estrogens influence certain subsets of human populations, rather than the general population . Conceivably, an alteration of the immune system by environmental estrogens could affect the individuals' ability to mount well-regulated immune responses to microbial and vaccine antigens, allergens, self and tumor antigens . Possible changes in the immune system must be investigated routinely in toxicity studies . A comprehensive mechanistic understanding of potential immunomodulatory chemicals is needed . In this regard, relevant laboratory animals may be especially useful in identifying susceptible periods of life, whether both genders are equally affected, in analysis of changes in target lymphoid organs, and to determine the immunological effects of mixtures of chemicals.

J Agric Food Chem, 2000 Sep, 48(9), 4341 - 51
Isolation and acclimation of a microbial consortium for improved aerobic degradation of alpha-hexachlorocyclohexane; Manonmani HK et al.; A microbial consortium that can utilize alpha-hexachlorocyclohexane (alpha-HCH) as a sole source of carbon and energy was isolated from soil and sewage through a novel technique involving an initial enrichment in a glass column reactor followed by a shake flask enrichment . This consortium took 14 days to completely mineralize 5 and 10 microg mL(-)(1) alpha-HCH in mineral salts medium in shake flasks . The degradative ability of this consortium improved very markedly on acclimation by successive and repeated passages through media containing increasing concentrations of alpha-HCH . The acclimated consortium could degrade 100 microg mL(-)(1) of alpha-HCH within 72 h at a degradation rate of 58 microg mL(-)(1) day(-)(1) with concomitant release of stoichiometric amounts of chloride . Accumulation of any intermediary metabolites was not detected in the culture broth as tested by TLC and GC, implying complete mineralization of the substrate . The acclimated consortium contained eight bacterial strains and a fungus . The individual strains and the different permutations and combinations of them, however, were able to utilize only 10 microg mL(-)(1) of alpha-HCH . Mesophilic temperatures (20-30 degrees C) and near-neutral pH (6.0-8.0) were most favorable for alpha-HCH degradation . Among the auxiliary carbon sources tested, ethanol, benzoate, and glucose (at higher concentrations) retarded the degradation of alpha-HCH, whereas the addition of cellulose, sawdust, and low concentrations of glucose (<200 microg mL(-)(1)) and acetone enhanced the rate of degradation.

J Agric Food Chem, 2000 Sep, 48(9), 4204 - 8
Maintaining quality of fresh-cut mangoes using antibrowning agents and modified atmosphere packaging; Gonzalez-Aguilar GA et al.; Treatments to inhibit browning and decay and prolong shelf life of fresh-cut mangoes were investigated . Combinations of antibrowning agents and modified atmosphere packaging (MAP) resulted in a reduction of browning and deterioration of fresh-cut mangoes stored at 10 degrees C . Combinations of several browning inhibitors were more effective than those applied individually . Among these treatments, solutions containing 4-hexylresorcinol (0.001 M) (HR) plus potassium sorbate (0.05 M) (KS) and HR plus KS plus D-isoascorbic acid (0.5 M) (ER) reduced changes in color (L, a, and b) and microbial growth and did not affect sensory characteristics of fresh-cut mangoes . In general, these treatments did not affect significantly the changes in organic acids and sugar content of slices during the 14 days of storage at 10 degrees C . High humidity created in the in-package atmosphere alleviated tissue dryness and was an important factor in the ability of the antibrowning solutions to prevent browning and decay . It appears that the maintenance of quality of fresh-cut mangoes is more related to particular combinations of the antibrowning agents used rather than the modified atmosphere created inside the package . HR + ER + KS treatment in combination with MAP could be used to inhibit browning, decay, and deterioration of fresh-cut mangoes.

Acc Chem Res, 2000 Sep, 33(9), 579 - 89
Covalent inhibition of digestive lipases by chiral phosphonates; Cavalier JF et al.; Designing and synthesizing specific inhibitors is of fundamental value for understanding the molecular mechanisms involved in the interfacial adsorption step as well as the catalytic activity of lipases . In this Account, we will review and discuss results obtained mostly at our laboratory concerning the covalent inhibition of human gastric and human pancreatic lipases by chiral phosphonates . Rather than presenting an exhaustive list of compounds tested so far with lipases of animal and microbial origin, we selected recent experimental data illustrating well the specific problems encountered during the covalent inhibition of these digestive lipases.

J Exp Med, 2000 Sep 18, 192(6), 823 - 33
Interleukin (IL)-4 is a major regulatory cytokine governing bioactive IL-12 production by mouse and human dendritic cells; Hochrein H et al.; Interleukin (IL)-12 may be secreted as a bioactive T helper type 1 (Th1) cell-inducing heterodimer, as a monomer, or as an antagonistic homodimer . We analyzed the IL-12 produced by mouse splenic dendritic cells (DCs), human thymic DCs, and cultured human monocyte-derived DCs . IL-12 production required both a microbial or T cell-derived stimulus and an appropriate cytokine milieu . The different IL-12 forms were differentially regulated by the cytokines present rather than the stimulus used . IL-4 alone or together with granulocyte/macrophage colony-stimulating factor or interferon gamma effectively enhanced the production of the bioactive heterodimer and selectively reduced the antagonistic homodimer of IL-12 . Therefore, IL-4, the major Th2-driving cytokine, provides a negative feedback causing DCs to produce the major Th1-inducing cytokine, bioactive IL-12.

Ecotoxicol Environ Saf, 2000 Sep, 47(1), 96 - 103
An in situ respirometric technique to measure pollution-induced microbial community tolerance in soils contaminated with 2,4, 6-trinitrotoluene; Gong P et al.; Long-term exposure to 2,4,6-trinitrotoluene (TNT) can induce changes in the structure and activities of soil microbial communities . Such changes may be associated with an elevated microbial tolerance . An in situ respirometry technique based on the analysis of the substrate-induced respiration response to freshly added TNT was used to examine soil microbial tolerance to TNT at the community level . The specific growth rate derived by fitting an exponential equation to respiration data was taken as the measurement endpoint . Microbial tolerance was evaluated using a tolerance index defined as the ratio of the specific growth rate at a spiking dose of 2000 microg TNT/g soil to that of the control with no spiked TNT . Three soils with long-term exposure histories (TNT level in soil: 1.5, 32, and 620 microg TNT/g, respectively) exhibited significantly higher microbial community tolerance to TNT than two uncontaminated control soils . A soil containing 29,000 microg TNT/g exhibited the highest tolerance . Findings from this study support the hypothesis that pollution-induced community tolerance can be used as a means of identifying those compounds that have exerted selective pressure on the community .

Ecotoxicol Environ Saf, 2000 Sep, 47(1), 65 - 73
A stepwise procedure for assessment of the microbial respiratory activity of soil samples contaminated with organic compounds; Eisentraeger A et al.; Soil respiration measurements are used frequently for the characterization of soil samples . Identical methods are used for the ecotoxicological characterization of contaminated soil samples as well as for quantification of the active microbial biomass in agriculturally used soils . In this study four soil samples contaminated with large amounts of volatile organic compounds, polyaromatic hydrocarbons, or nitroaromatic compounds are characterized after stepwise addition of carbon, nitrogen, and phosphorus . The respiration kinetics are assessed over a period of 5 days . By means of qualitative evaluation of the results, it is demonstrated that this stepwise addition allows one to distinguish between growth-promoting effects of biodegradable organic compounds on the one hand and the toxic influence of these compounds on the other hand . Finally it is stated that a comprehensive ecotoxicological characterization cannot be performed routinely using only one or several parameters of respiration curves . There is need for further research and validation if soil respiration measurements are to be performed quantitatively in the future .

Free Radic Res, 2000 Sep, 33(3), 261 - 5
Antioxidant binding of caeruloplasmin to myeloperoxidase: myeloperoxidase is inhibited, but oxidase, peroxidase and immunoreactive properties of caeruloplasmin remain intact; Park YS et al.; The neutrophil enzyme myeloperoxidase (MPO) purposefully makes hypochlorous acid (HOCl) as part of the cells defence against microbial infections . During cell lysis, however, MPO will be released into the extracellular environment where production of HOCl, a powerful oxidant, will lead to molecular damage . Extracellular MPO binds to the copper-containing protein caeruloplasmin (Cp) and prevents MPO making HOCl . Cp has several important antioxidant functions in extracellular fluids associated with its ability to catalyse oxidation of ferrous ions and to remove peroxides . The binding of MPO to Cp did not inhibit these important extracellular antioxidant activities of Cp, but in so doing it provided additional antioxidant protection against formation of HOCl.

Br J Pharmacol, 2000 Sep, 131(2), 335 - 41
Methyl p-hydroxybenzoate (E-218) a preservative for drugs and food is an activator of the ryanodine receptor Ca(2+) release channel; Cavagna D et al.; 1 . Haloperidol is a drug used in the management of several psychotic disorders and its use has been linked to Neuroleptic Malignant Syndrome . In the present study we have investigated the effect of a commercial preparation of haloperidol, Serenase, on skeletal muscle sarcoplasmic reticulum . 2 . Addition of Serenase to isolated terminal cisternae caused a rapid release of calcium . We tested whether the active Ca(2+)-releasing substance was haloperidol or another compound present in the preparation . 3 . Our results show that methyl p-hydroxybenzoate, one of the preservatives and a commonly used anti-microbial agent (E-218) is an activator of Ca(2+) release (E.C . 50=2.0 mM), mediated by a ruthenium red-sensitive Ca(2+) release channel present in skeletal muscle terminal cisternae.

Rev Epidemiol Sante Publique, 2000 Aug, 48(## Suppl 2), 11 - 22
{Quantitative risk assessment related to microbial food contamination} {In Process Citation}; Sanaa M et al.; The four keystones of quantitative risk assessment are hazard identification, exposure assessment, dose-response assessment and risk characterization . This paper considers the different steps of risk assessment and their application to food microbiology . Traditionally, quantitative risk assessment has been viewed simply as a method to estimate risk . However, when we conduct a complete risk assessment including different factors from "farm to fork" it can serve to understand the risk process . Quantitative risk assessment can also provide valuable insights as how to best manage the risk.

Theriogenology, 2000 Jul 1, 54(1), 57 - 67
Fertility results using bovine semen cryopreserved with extenders based on egg yolk and soy bean extract; van Wagtendonk-de Leeuw AM et al.; Semen extenders containing components such as egg yolk and skim milk are difficult to standardize and they introduce the risk of microbial contamination . A well-defined extender not originating from animal tissues would present a valuable contribution to the AI industry . We evaluated the fertility of bovine semen cryopreserved with 3 different extenders: 1) TRIS-Standard, prepared at 2 local AI laboratories, containing 20% (v/v) pasteurized egg yolk, 2) TRIS-Concentrate, prepared by adding 20% (v/v) pasteurized egg yolk and 1:5 (v/v) nonpyrogenic water, and 3) Biociphos Plus, a soybean extract containing extender, prepared by adding 1:5 nonpyrogenic water . Ejaculates of 4 Holstein bulls were split into 3 aliquots and cryopreserved with the 3 extenders . Prior to this study, the semen dose-response curve for each of the 4 bulls was developed in a field trial by freezing the semen and randomly distributing the straws throughout the Netherlands for insemination . Optimal semen doses were thus established to detect the effect of extenders on fertility, evaluated by 56-day non-return rate (NR56), and by the estimated conception rate and the calving rate, given a conception . We used the multiphasic model developed by Grossman et al . (7) . A total of 22,246 first and second inseminations were recorded . The NR56 ranged among bulls from 67.0 to 70.1% for Tris-Standard, from 67.5 to 69.9% for Tris-Concentrate and from 60.2 to 66.7% for Biociphos Plus . No significant differences in NR56 were detected between Tris-Standard and Tris-Concentrate (P=0.54), whereas Biociphos Plus resulted in a significantly lower NR56 than Tris-Standard and Tris-Concentrate (P<0.05) . Estimated conception rate was 72.1, 73.6 and 69.6% and estimated calving rate, given a conception was 80.6, 78.3 and 77.1 for Tris-Standard, Tris-Concentrate and Biociphos Plus, respectively . These results indicate that 1) semen extended with a custom made TRIS-Concentrate can be succesfully used in the field resulting in comparable fertility with Tris-Standard; 2) semen extended with Biociphos Plus results in a significant reduction in the NR56; 3) extender source may affect both conception rate and calving rate, given a conception, i.e., extrinsic and intrinsic sperm factors (4).

Bull Acad Natl Med, 2000, 184(2), 313 - 23; discussion 323-4
{Intracellular iron metabolism}; Beaumont C; Iron is essential to life but it is poorly soluble in biological fluids and toxic in excess . Organisms have developed multiple proteins to insure iron transport and storage, and some of these have only been discovered recently . Among them, members of the Nramp family are transmembrane proteins which transport several divalent cations, including Fe2+ . These proteins participate in intestinal iron absorption, erythopoiesis and microbial defence . Frataxin, which is deficient in Friedreich's ataxia, regulates iron flux through mitochondria . The intracellular labile iron pool, an intermediate form of iron between different subcellular compartments, regulates ferritin synthesis and stability of mRNA coding for proteins of iron transport . Subunit composition of ferritin plays an important role in the control of the size of this labile iron pool.

Trends Microbiol, 2000 Sep, 8(9), 419 - 25
CD1 and CD1-restricted T cells in infections with intracellular bacteria; Schaible UE et al.; Glycolipid-specific, CD1a-, b- and c-dependent cytotoxic T cells have recently been shown to be involved in the host response against tuberculosis . These CD1 molecules 'sample' mycobacterial glycolipids from different intracellular sites in the infected cell . Additionally, upon microbial encounter, CD1d-dependent natural killer T cells promptly produce cytokines and perform regulatory activities . Here, we discuss the intracellular localization of CD1 molecules and mycobacterial lipids and the role of CD1-mediated T-cell responses in mycobacterial infections.

Pharmacol Res, 2000 Oct, 42(4), 373 - 81
Molecular mechanism of staurosporine-induced apoptosis in osteoblasts; Chae HJ et al.; Staurosporine, a microbial alkaloid, is a strong inhibitor of protein kinases . We induced apoptosis in murine osteoblast MC3T3E-1 cells by exposure to the staurosporine . Staurosporine transiently increased the phosphotransferase activity of c-Jun N-terminal kinase-1 (JNK1), which in turn may activate the transcriptional activity of activating protein-1 (AP-1) . We then prepared extracts from staurosporine-treated MC3T3E-1 cells and monitored the cleavage of acetyl-YVAD-AMC and acetyl-DEVD-AMC, fluorogenic substrates of caspase-1-like and caspase-3-like proteases, respectively . Staurosporine caused a significant increase in the proteolytic activity of caspase-3-like proteases, but not in the activity of caspase-1-like proteases . Furthermore, staurosporine increased the transcriptional activity of nuclear factor- kappa B (NF- kappa B) . These data suggest that staurosporine-induced apoptosis in osteoblasts may occur via activation of JNK1, caspase-3-like proteases, and transcriptional factors including AP-1 and NF- kappa B .

Org Lett, 2000 Sep 21, 2(19), 2921 - 2
A short formal synthesis of squalamine from a microbial metabolite; Kinney WA et al.; A short formal synthesis of squalamine is described, utilizing the biotransformation product 2, which is available in one step from commercially available 3-keto-23,24-bisnorchol-4-en-22-ol (1) . Regioselective C-22 oxidation and C-24 sulfation of the corresponding alcohols in the presence of a free C-7 alcohol make for an efficient preparation of squalamine intermediate 11.

J Chromatogr A, 2000 Aug 11, 889(1-2), 245 - 52
Unknown bisethylisooctanollactone isomers in industrial waste water . Isolation, identification and occurrence in surface water; Knepper TP et al.; Unknown bisethylisooctanollactone isomers (BIOL isomers) which are chemical by-products of butyraldehyde synthesis, were isolated from industrial waste water applying various purification methods with subsequent semi-preparative high-performance liquid chromatography . Through interpretation of mass spectra after gas chromatographic separation the individual BIOL isomers were identified as stereoisomers of 2,4-diethyl-3-n-propyl-delta-valerolactone . Thus, it was possible for the first time to quantify the BIOL isomers in the river Rhine, Germany, with a mean sum concentration of 1.6 microg l(-1) . A regular analysis performed over a period of almost two years of the river Rhine always gave the same ratio among the individual isomers . Drinking water production out of such water was studied, revealing that activated carbon filtration led to a 95% reduction of the BIOL concentration . Additional subsoil passage and a subsequent slowsand filtration led to a total elimination due to microbial degradation . Even if the BIOL isomers proved not to be relevant to drinking water, their behavior in the aquatic environment needs to be more thoroughly investigated since these compounds have been discharged for many years in high amounts into the river Rhine.

J Anim Sci, 2000 Sep, 78(9), 2421 - 30
Compartmental modeling with nitrogen-15 to determine effects of degree of fat saturation on intraruminal N recycling; Oldick BS et al.; Two- and three-compartment models were developed to describe N kinetics within the rumen using three Holstein heifers and one nonlactating Holstein cow fitted with ruminal and duodenal cannulas . A 4 x 4 Latin square design included a control diet containing no supplemental fat and diets containing 4.85% of diet dry matter as partially hydrogenated tallow (iodine value = 13), tallow (iodine value = 51), or animal-vegetable fat (iodine value = 110) . Effects of fat on intraruminal N recycling and relationships between intraruminal N recycling and ruminal protozoa concentration or the efficiency of microbial protein synthesis were determined . A pulse dose of 15(NH4)2SO4 was introduced into the ruminal NH3 N pool, and samples were taken over time from the ruminal NH3 N and nonammonia N pools . For the three-compartment model, precipitates of nonammonia N after trichloroacetic acid and ethanol extraction were defined as slowly turning over nonammonia N; rapidly turning over nonammonia N was determined by difference . Curves of 15N enrichment were fit to models with two (NH3 N and nonammonia N) or three (NH3 N, rapidly turning over nonammonia N, and slowly turning over nonammonia N) compartments using the software SAAM II . Because the three-compartment model did not remove a small systematic bias or improve the fit of the data, the two-compartment model was used to provide measurements of intraruminal N recycling . Intraruminal NH3 N recycling (45% for control) decreased linearly as fat unsaturation increased (50.2, 43.0, and 41.7% for partially hydrogenated tallow, tallow, and animal-vegetable fat, respectively) . Intraruminal nitrogen recycling was not correlated with efficiency of microbial protein synthesis or ruminal protozoa counts.

J Anim Sci, 2000 Sep, 78(9), 2412 - 20
Effects of degree of fat saturation on fiber digestion and microbial protein synthesis when diets are fed twelve times daily; Oldick BS et al.; Three Holstein heifers and one nonlactating cow, fitted with ruminal and duodenal cannulas, were arranged in a 4 x 4 Latin square design to determine the effects of degree of fat saturation on ruminal neutral detergent fiber digestion and microbial protein synthesis and to determine whether changes in the efficiency of microbial protein synthesis were related to protozoal populations in the rumen . Corn silage-based diets contained no added fat or 4.85% of diet dry matter as partially hydrogenated tallow, tallow, or animal-vegetable fat . Iodine values of fat sources were 12.8, 50.6, and 109.7 for partially hydrogenated tallow, tallow, and animal-vegetable fat, respectively . Cattle were fed every 2 h and consumed 1.5% of body weight as dry matter daily . Ruminal neutral detergent fiber digestibility was decreased by added fat but was not affected by increasing iodine value . Flows of microbial N and non-NH3-nonmicrobial N to the duodenum were not affected by treatment . Ruminal protozoa concentration decreased linearly as the iodine value of fats increased . The efficiency of microbial protein synthesis was increased and protozoa concentrations tended to decrease when fat was fed . Decreased ruminal protozoa concentration may have decreased intraruminal N recycling . Biohydrogenation of added fat may result in a low ruminal concentration of unsaturated fatty acids when cows are fed frequently, reducing the negative effects of unsaturated fat sources on ruminal neutral detergent fiber digestibility . Protozoa were inhibited by unsaturated fat, but it is not clear if biohydrogenation and frequent feeding lessened inhibition.

J Leukoc Biol, 2000 Sep, 68(3), 400 - 4
Macrophage-derived chemokine (MDC); Mantovani A et al.; Macrophage-derived chemokine (MDC) is a CC chemokine paradigmatic of emerging aspects of chemokine immunobiology . It is constitutively expressed, yet microbial products and cytokines regulate its expression with divergent effects of type II (IL-4 and IL-13) and type I (interferon) cytokines . Processing of the mature protein by dipeptidyl peptidase IV/CD26 provides a further level of regulation . It acts on diverse cellular targets including dendritic cells (DC), NK cells, and T cell subsets . Among these, MDC is a potent attractant for CCR4 expressing polarized Th2 and Tc2 cells, and evidence is consistent with a role of this chemokine as an amplification loop of polarized type II responses . Emerging indications on the involvement of MDC in diverse pathologies, ranging from allergic reactions to HIV infection and neoplasia, are discussed.

J Leukoc Biol, 2000 Sep, 68(3), 383 - 90
Proinflammatory response and IL-12 expression in HIV-1 infection; Ma X et al.; HIV-1 infection elicits a broad range of host responses, many of which interfere with the regulatory pathways of gene expression of interleukin-12 (IL-12), a heterodimeric cytokine essential for cell-mediated immunity against microbial infection . The inhibition of IL-12 production by accessory cells after HIV-1 infection has been identified as a potential factor responsible for impaired innate and Th1 cell-mediated responses observed in AIDS patients . The mechanism by which HIV-1 infection suppresses IL-12 gene expression is largely uncharacterized . Here we review all pathways identified that could potentially mediate HIV-induced impairment of IL-12 gene expression, such as IL-10, transforming growth factor beta, interferon-alpha/beta, tumor necrosis factor alpha, Fc receptors, complement regulatory proteins, and receptors . Also discussed is the decreased CD40 ligand induction in CD4 T cells during HIV infection, which may have a strong impact on T cell-dependent IL-12 production that is critical for the establishment and maintenance of a Th1 response.

J Dairy Sci, 2000 Aug, 83(8), 1782 - 94
Effects of altering energy and protein supply to dairy cows during the dry period . 1 . Intake, body condition, and milk production; Dewhurst RJ et al.; We used 48 Holstein-Friesian cows to investigate the effects of altering energy and protein supply to dry cows . Cows were fed one of three diets for 6 wk prior to parturition: (a) a 60:40 (DM basis) mixture of grass silage with barley straw ad libitum; (b) grass silage ad libitum; or (c) 0.5 kg/d of prairie meal with grass silage ad libitum . The standard lactation diet was a flat-rate allocation of concentrates and grass silage ad libitum . We evaluated dry-period diets using four dry fistulated cows; rumen pH remained high (mean = 6.6) and ammonia concentrations followed N intake . The inclusion of straw reduced apparent ruminal digestion of OM, N, and NDF as well as microbial protein yield, though microbial yield per unit of OM apparently digested in the rumen remained unchanged . Voluntary intake of forage was reduced by the inclusion of straw, while the inclusion of prairie meal had little effect . The decline in intake as calving approached was lower with the silage and straw mix diet . There were large differences in the BW change over the final 5 wk of the dry period, although the opposite effect was seen in early lactation, and differences in BW and body condition score were small by lactation wk 22 . Despite the substantial differences in nutrient supply and effects on body reserves, there was little effect of dry-period diet on subsequent performance . Lower forage intakes and yields of protein and lactose were confined to the first month of lactation for cows previously offered straw.

J Food Prot, 2000 Sep, 63(9), 1262 - 7
Effect of high hydrostatic pressure on Cryptosporidium parvum infectivity; Slifko TR et al.; The incidence of foodborne disease outbreaks caused by contaminated low-pH fruit juices is increasing . With recent mandatory pasteurization of apple juice and the industry's concerns of food safety, fruit juice processors are showing more interest in alternative nonthermal technologies that can kill >99.99% of microbial pathogens present in foods . The association of the coccidian protozoan, Cryptosporidium, with diarrheal disease outbreaks from contaminated tap water and fruit juice raises a safety concern in the food and beverage industries . The objective of this study was to evaluate the effects of high hydrostatic pressure (HHP) on C . parvum oocysts . Oocysts were suspended in apple and orange juice and HHP treated at 5.5 x 10(8) Pa (80,000 psi) for 0, 30, 45, 60, 90, and 120 s . Oocyst viability was assessed by excystation using bile salts and trypsin while the cell culture foci detection method was used to assess infectivity . Results indicated that HHP inactivated C . parvum oocysts by at least 3.4 log10 after 30 s of treatment . No infectivity was detected in samples exposed to > or =60 s of HHP and >99.995% inactivation was observed . This study demonstrated that HHP efficiently rendered the oocysts nonviable and noninfectious after treatment at 5.5 x 10(8) Pa.

J Food Prot, 2000 Sep, 63(9), 1240 - 7
Analysis of the fluctuating microbial counts in commercial raw milk--a case study; Nussinovitch A et al.; Microbial counts of raw milk from eight commercial dairy farms recorded over a period of about 2 years were obtained from a processing plant . All the counts formed irregular fluctuating series whose statistical properties were examined . In all eight cases the autocorrelation function indicated that the counts were random and had no underlying periodicity . This suggested that the fluctuations, at least to some extent, reflected the combined effect of numerous factors, some unknown, and that these factors operate randomly and independently . In five out of the eight cases the fluctuating pattern of the record could be described by a probabilistic model based on the assumption that the counts have a lognormal distribution . The validity of the model was confirmed by comparing the calculated frequencies of counts exceeding preselected levels with those actually observed in fresh data . The model enabled ranking the five sources on the basis of anticipated frequencies of excessive counts rather than on the basis of mean performance . Two sequences had clearly identified regions with different statistical characteristics . They were interpreted as a reflection of changes in the sanitary conditions at the corresponding farms . In one case, both the overall magnitudes and the fluctuation amplitude of the counts progressively decreased, a situation for which the described model as formulated was inapplicable.

Vestn Khir Im I I Grek, 2000, 159(3), 55 - 8
{Programmed irrigation and drainage in the treatment of patients with suppurative arthritis and intramedullary phlegmons}; Iusupov IuN et al.; Programmed irrigation and drainage were used in the process of treatment of 17 patients with purulent arthritis of the knee joint and talocrural articulation and of 13 patients with intramedullary phlegmons of the femoral, tibial and humeral bones . The programmed irrigation and drainage were proved to decrease the microbial dissemination of the wash liquid to 10 colony-forming units per 1 ml during 1-3 days . The method allowed to avoid synovectomy and resection of the joint in patients with purulent gonitis, to decrease the risk of appearance of secondary infections . The programmed irrigation and drainage for intramedullary phlegmons provide complete sanitation of the purulent focus during 7-10 days, create favorable conditions for consolidation of fractures and prevent the development of chronic osteomyelitis.

Ocul Immunol Inflamm, 2000 Jun, 8(2), 101 - 9
Clinical features and keratoplasty results in keratoconus complicated by acute hydrops; Akova YA et al.; PURPOSE: Acute hydrops develops when Descemet's membrane and the endothelium of the ectatic cornea separates to allow aqueous humor to enter the stroma . The current study was done to determine if penetrating keratoplasty (PK) in keratoconus patients with resolved hydrops has a poor prognosis . METHODS: A retrospective chart review of 35 eyes of 35 patients with keratoconus who underwent PK after the resolution of acute hydrops (Group 1) and of 74 eyes of 69 patients who had PK without a previous history of acute hydrops (Group 2) was performed . The chi-square test was used for statistical analysis . RESULTS: The mean age in Group 1 was 21.1+/-11 . 0 years . The mean age in Group 2 was 23.2+/-14.4 years . Vernal keratoconjunctivitis was present in 11 patients in Group 1 (31%) and in 12 (17.4%) patients in Group 2 . Loose sutures were present in two patients (6%) in Group 1; both resulted in loose suture-related corneal neovascularization . Loose sutures developed in five patients (7%) in Group 2 . In Group 1, three patients (9%) had microbial keratitis . Microbial keratitis was seen in five patients (7%) in Group 2 . Two patients (6%) in Group 1 developed late endothelial graft failure . In Group 2, one eye (1%) developed early and one eye (1%) late epithelial graft rejection; five eyes (7%) developed late endothelial graft rejection . There was no statistically significant difference between the two groups with respect to the incidence of graft rejection . At the last examination, 34 eyes in Group 1 had clear grafts; in the second group, graft clarity was achieved in 71 eyes . CONCLUSION: Penetrating keratoplasty in hydrops is successful in terms of graft clarity and visual outcome in patients with keratoconus after the resolution of hydrops . The patients should have preoperative and postoperative anti-allergic treatment and close follow-up for possible complications.

Curr Opin Genet Dev, 2000 Oct, 10(5), 497 - 502
Toll signaling: the enigma variations; Wasserman SA; Experiments reported in the past year have revealed considerable diversity in Toll-mediated pathways for signal transduction in development and innate immunity . Rather than function as a well conserved signaling cassette, Toll receptors and associated factors have apparently evolved as a diverse set of configurations to defend against microbial infection in species ranging from plants to humans.

Matrix Biol, 2000 Sep, 19(5), 443 - 53
Identification and localization of lysozyme as a component of eggshell membranes and eggshell matrix; Hincke MT et al.; The avian eggshell is a composite biomaterial composed of non-calcifying eggshell membranes and the overlying calcified shell matrix . The calcified shell forms in a uterine fluid where the concentration of different protein species varies between the initial, rapid calcification and terminal phases of eggshell deposition . The role of these avian eggshell matrix proteins during shell formation is poorly understood . The properties of the individual components must be determined in order to gain insight into their function during eggshell mineralization . In this study, we have identified lysozyme as a component of the uterine fluid by microsequencing, and used western blotting, immunofluorescence and colloidal-gold immunocytochemistry to document its localization in the eggshell membranes and the shell matrix . Furthermore, Northern blotting and RT-PCR indicates that there is a gradient to the expression of lysozyme message by different regions of the oviduct, with significant albeit low levels expressed in the isthmus and uterus . Lysozyme protein is abundant in the limiting membrane that circumscribes the egg white and forms the innermost layer of the shell membranes . It is also present in the shell membranes, and in the matrix of the calcified shell . Calcite crystals grown in the presence of purified hen lysozyme exhibited altered crystal morphology . Therefore, in addition to its well-known anti-microbial properties that could add to the protective function of the eggshell during embryonic development, shell matrix lysozyme may also be a structural protein which in soluble form influences calcium carbonate deposition during calcification.

Nutrition, 2000 Sep, 16(9), 719 - 22
Microbial contamination of enteral feeding formulas and diarrhea; Okuma T et al.; Twenty-five medical and surgical patients receiving liquid ready-to-use sterile enteral formulas were evaluated prospectively to investigate the relation of diarrhea to serum albumin level, total lymphocyte count, delayed hypersensitivity to purified protein derivative, antibiotic therapy, administration rate and site of enteral formula, and microbial contamination of enteral feeds . Formulas were administered to 6 patients with hang times of up to 6 h by pump-assisted continuous drip and to 19 patients with hang times of up to 3 h as a bolus feeding . Samples of formulas for microbial culture were obtained aseptically before and after feeding on the first and eighth day of the study period . The incidence of microbial contamination of the formula before and after feeding was 1 of 49 samples (2.0%) and 10 of 48 samples (20.8%), respectively . There were 2 patients with diarrhea, which occurred on the second day . Formula samples from 2 patients (100%) with diarrhea and 2 samples from 23 patients (8.7%) without diarrhea were contaminated with 10(4) cfu/mL or more, respectively . A significant difference (P = 0.04) was detected between the two groups . The other factors studied showed no significant association with the incidence of diarrhea . In conclusion, contaminated formula appears to play a significant role in the etiology of diarrhea in patients receiving enteral feeding.

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