Ecotoxicol Environ Saf, 1997 Dec, 38(3), 200 - 4 Microbial toxicity of Cd and Hg in different soils related to total and water-soluble contents; Welp G et al.; Microbial toxicity tests with Cd and Hg and up to 18 soil samples were combined with sorption and solubility measurements . The data ought to indicate to what extent toxic effects of the metals are influenced by sorption and the other factors that may alter their biocidal action . The microbial reduction of Fe(III) oxides to Fe2+ ions was used as a parameter of microbial activity to calculate the total amounts of Cd and Hg that cause 10, 50, and 90% inhibition {effective doses (ED)10, ED50, ED90} . The corresponding solution concentrations {effective concentrations: (EC)10, EC50, EC90} were derived from Freundlich adsorption isotherms . The very large variability in ED values in different soils (ED10: Cd, 5-95 mg/kg; Hg, 0.125-125 mg/kg) can be related to the sorption and solubility behavior of the metals . Nevertheless, the EC values of both metals also indicate a considerable variation (EC10: Cd, 0.01-1.13 mg/liter; Hg, < 0.001-0.041 mg/liter) . The influence of soluble soilborne substances on metal speciation is the main reason for their varying toxic potential in different soil solutions . The strong and complex influence of soil properties on the toxicity of heavy metals in soils indicates that both chemical and biological methods of soil analysis are needed to assess the risks of soil contamination adequately. Acta Ophthalmol Scand, 1997 Oct, 75(5), 483 - 6 Corneoscleral discs excised from enucleated and non-enucleated eyes are equally suitable for transplantation; Garweg J et al.; PURPOSE: To assess whether the biological quality of corneoscleral tissue dissected in situ is, after organ culture, comparable to that harvested after enucleation . METHODS: Corneoscleral discs were prepared from 23 donor eyes, either after enucleation, under laminar flow conditions (right eyes; group 1) or by direct excision in situ (left eyes; group 2) . Endothelial cell counts were made and the degree of tissue contamination assessed both prior to and upon termination of organ culture . RESULTS: Microbial growth was found in 12/22 conjunctival swabs collected from group 1 eyes and in 14/22 of those obtained from group 2 globes (p = 0.76) . Bacterial growth was detected in four primary culture media, two from each group, at low colony densities . No significant difference in endothelial cell counts were encountered between the two groups, either immediately after dissection {group 1: 2940 +/- 308 (2100-3500) c/mm2; group 2: 2947 +/- 345 (2200-3700) c/mm2; p = 0.945} or upon termination of organ culture {group 1: 2646 +/- 321 (1895-3200); group 2: 2723 +/- 312 (2100-3650); p = 0.413} . CONCLUSION: Dissection of corneoscleral discs in situ may serve as an alternative to the conventional technique if consent is obtained to remove only the cornea . The risk of contamination is no higher and endothelial cell viability no lower than in tissue derived from enucleated globes, provided that the excision is performed by a skilled surgeon and a rigorous disinfection protocol is instigated. Aliment Pharmacol Ther, 1997 Dec, 11 Suppl 3, 40 - 4; discussion 44-6 Review article: Epithelial disposition of antigen; Lamm ME; Exogenous antigens can impinge upon the luminal surface of mucous membranes and to a limited degree, can even penetrate into the lamina propria . As a result of infection, microbial antigens can be present in mucosal epithelium or lamina propria . At all three levels (lumen, epithelium, lamina propria), locally produced antibodies, principally immunoglobulin A (IgA), can combine with antigens . As a result, antigens can be prevented from attaching to the epithelium, viruses can be neutralized inside epithelial cells and immune complexes in the lamina propria can be excreted into the lumen . As the Fc portion of IgA is specialized to facilitate epithelial transport of IgA rather than to activate inflammatory mediator systems (as is the case with other classes of immunoglobulin), the predominance of IgA among mucosal antibodies serves to limit the degree of local inflammation following antigen-antibody reactions. Microbios, 1997, 91(366), 55 - 65 Measurement of the dielectrophoretic enrichment of yeast on grid electrodes using image analysis; Brown AP et al.; Dielectrophoresis using grid electrode structures offers many advantages for the abstraction and enrichment of cells in analytical microbiology . Modifications to the quantification method of a grid-based dielectrophoresis system are described, utilizing a more rapid and efficient image analysis technique . The grid electrode arrangement enabled dielectrophoretic enrichment of Saccharomyces cerevisiae, resulting in between 36.2 and 46.1% concentration of cells (by a single pass), relative to an identical control sample suspension analysed without an applied electric field . These enrichment factors compare favourably with previously obtained data using alternative measurement techniques . In addition, the image analysis technique is considerably less labour intensive and enables the continuous detection of cellular concentration and dielectrophoretic behaviour . Utilizing differences in frequency dependent cell responses, the system described could be suitable for highly selective pre-enrichment or cell separation on a large scale . Dielectrophoretic techniques, including enrichment, viable cell analysis and assessment of low microbial concentrations, could enhance or replace many existing methods for the analysis of micro-organisms and other cells. Analyst, 1997 Oct, 122(10), 1125 - 8 Microbial detection by a glucose biosensor coupled to a microdialysis fibre; Palmisano F et al.; The use of a glucose biosensor coupled to microdialysis sampling in a flow injection analysis system is described to follow the growth of Escherichia coli in a glucose-containing liquid culture medium . The experimental set-up permitted a throughput rate of 25 samples h-1 . Growth curves were modelled by a modified Gompertz equation, which permitted the determination of lag time and maximum specific growth rate . The time required to produce an appreciable variation in the biosensor response (minimum detection time, MDT) was determined . A plot of MDT versus microbial concentration was found to be linear in the range 10(6)-10(10) colony forming units (cfu) ml-1 . A microbial concentration of 10(6) cfu ml-1 can be detected after about 5 h. Trans R Soc Trop Med Hyg, 1997 Sep-Oct, 91(5), 618 - 22 Higher proportion of CD8+ T cells in the blood in healthy adults from Ethiopia and Bangladesh compared with Sweden; Worku S et al.; The phenotypic composition of peripheral blood lymphocytes in 45 healthy adults (15 each from Bangladesh, Ethiopia and Sweden) was analysed as an indicator of the influence of environment and/or ethnic background on the human immune response . The possible interference of technical factors was minimized by highly standardized handling of samples and by use of a similar simultaneous 3-colour flow cytometry analysis technique for all samples . The percentage of CD4+ cells was lower, and the percentage of CD8+ cells was higher, in Bangladeshi and Ethiopian subjects than in those from Sweden . A higher percentage of CD57+/CD8+ T cells was also found in these 2 groups than in Swedish subjects . The percentage of gamma delta T cells was higher in Bangladeshi subjects and a difference in T cell receptor V beta expression was also noted between Bangladeshi and Swedish subjects . The data suggest that environmental or genetic factors are important bias factors to be considered in immunophenotyping studies . Possibly differences in the pattern or level of microbial challenge, as well as nutritional factors, may lead to different adaptive changes in the immune response . The potential influence of such immune adaptation on the response to vaccination or pharmaceutical therapy may be important in the development of new strategies of medical intervention in different geographical regions or ethnic groupsPIP: The phenotypic composition of peripheral blood lymphocytes in 45 healthy adult blood donors (15 each from Bangladesh, Ethiopia, and Sweden) was examined as an indicator of the influence of environment and/or ethnic background on the human immune response . A highly standardized 3-color flow cytometry analysis was performed on all samples . The percentage of CD4+ cells was lower and the percentage of CD8+ cells was higher in Bangladeshi and Ethiopian subjects than in those from Sweden . A higher percentage of CD57+/CD8+ T cells was likewise found in these 2 groups compared to Swedish subjects . The percentage of gammadelta T cells was higher in Bangladeshi subjects and a difference in T cell receptor Vbeta expression was also noted between Bangladeshi and Swedish subjects . The findings suggest that environmental or genetic factors are important bias factors to be taken into account in phenotyping studies . Possible variations in the patterns or level of microbial challenge, as well as nutritional factors, may lead to different adaptive changes in the immune response . The potential influence of such immune adaptation on the response to vaccination or pharmaceutical therapy may be essential in the development of new medical intervention approaches in different geographical regions or ethnic groups . Biochim Biophys Acta, 1998 Jan 15, 1389(2), 123 - 31 Inhibition of lipases from Chromobacterium viscosum and Rhizopus oryzae by tetrahydrolipstatin; Potthoff AP et al.; Tetrahydrolipstatin is known as an inhibitor for pancreatic lipase but not for microbial lipases . In this paper we demonstrate that in the presence of water-insoluble substrates like tributyrin or olive oil, tetrahydrolipstatin inhibits the lipases of Chromobacterium viscosum and Rhizopus oryzae, although with different potency . In contrast to porcine pancreatic lipase, which forms an irreversible and covalent enzyme-inhibitor complex with tetrahydrolipstatin, the inhibition of the microbial lipases is reversible as the inhibitor can be removed from the enzyme-inhibitor complex by solvent extraction . Moreover, after inhibition of Chromobacterium viscosum lipase tetrahydrolipstatin remains chemically unchanged. Zh Mikrobiol Epidemiol Immunobiol, 1997 Nov-Dec, (6), 59 - 61 {The characteristics of the changes in the submicroscopic structure of the immunocompetent cells in the development of experimental mycobacterioses}; Sibirnaia RI et al.; The electron-microscopic study of the ultrastructure of peritoneal exudate cells of laboratory animals during the development of infectious processes caused by Mycobacterium tuberculosis and atypical mycobacteria was carried out . The study revealed that in all pathological processes under study typical manifestation of cell-mediated immunity took place, characterized by microbial phagocytosis on the level of cell ultrastructure, an increase in the lysosomal activity and the exhaustion of cells involved in the process up to their destruction . These changes in processes caused by the action of the agents of different mycobacterial infections were manifested with different intensity. Bioorg Med Chem, 1997 Dec, 5(12), 2165 - 71 O-methylasparvenone, a nitrogen-free serotonin antagonist; Bos M et al.; O-Methylasparvenone (1) and asparvenone (2) were isolated from an Aspergillus parvulus Smith broth in a microbial screening for 5-HT2C ligands and found to be 5-HT2C antagonists . They represent the first nitrogen-free serotonin ligands . The absolute configuration of 1 was determined to be S by X-ray analysis of the corresponding Mosher-ester . A short and efficient synthesis of rac-1 was developed . This protocol was applied to the synthesis of derivatives of 1 and a structure-affinity relationship was established. Proc Natl Acad Sci U S A, 1997 Dec 9, 94(25), 13414 - 9 Catalytic mechanism of the adenylyl and guanylyl cyclases: modeling and mutational analysis; Liu Y et al.; The adenylyl and guanylyl cyclases catalyze the formation of 3', 5'-cyclic adenosine or guanosine monophosphate from the corresponding nucleoside 5'-triphosphate . The guanylyl cyclases, the mammalian adenylyl cyclases, and their microbial homologues function as pairs of homologous catalytic domains . The crystal structure of the rat type II adenylyl cyclase C2 catalytic domain was used to model by homology a mammalian adenylyl cyclase C1-C2 domain pair, a homodimeric adenylyl cyclase of Dictyostelium discoideum, a heterodimeric soluble guanylyl cyclase, and a homodimeric membrane guanylyl cyclase . Mg2+ATP or Mg2+GTP were docked into the active sites based on known stereochemical constraints on their conformation . The models are consistent with the activities of seven active-site mutants . Asp-310 and Glu-432 of type I adenylyl cyclase coordinate a Mg2+ ion . The D310S and D310A mutants have 10-fold reduced Vmax and altered {Mg2+} dependence . The NTP purine moieties bind in mostly hydrophobic pockets . Specificity is conferred by a Lys and an Asp in adenylyl cyclase, and a Glu, an Arg, and a Cys in guanylyl cyclase . The models predict that an Asp from one domain is a general base in the reaction, and that the transition state is stabilized by a conserved Asn-Arg pair on the other domain. Z Gastroenterol, 1997 Aug, 35(8), 637 - 49 {Nutrition in the etiopathogenesis of chronic inflammatory bowel diseases}; Weinand I et al.; It is currently held that the pathogenesis of inflammatory bowel disease (IBD) involves a complex interaction of host responses, some immunologic and genetically determined, and external influences including microbial and dietary factors . Nutritional surveys and studies testing stepwise exclusion dietary compounds have suggested that dietary factors might be linked to the occurrence of IBD . Food allergy was suggested early as a possible trigger for the inflammatory response; however, no firm evidence has been gathered over the years to substantiate this possibility . The possibility that patients with Crohn's disease may have an unusual premorbid pattern of dietary intake has also been examined since the early seventies . Several groups of investigators report a high intake of refined sugars (i.e . sucrose), recycled cooking oil, a more frequent consumption of fast foods in patients with Crohn's disease and ulcerative colitis respectively . However, a critical comparison of incidence and prevalence studies published from 1976 until 1994 clearly showed no pre-existing nutritional abnormality that has been identified consistently in patients who develop inflammatory bowel disease. Prikl Biokhim Mikrobiol, 1997 Jul-Aug, 33(4), 455 - 60 {A bioluminescent method of determining antibiotic sensitivity of microbial cells in septic blood}; Frundzhian VG et al.; A bioluminescence assay for rapid (5-7 h) determination of susceptibility to antibiotics was applied to samples of septic blood and optimized . The method comprises hemolysis of blood, reduction of osmolarity by adding concentrated nutritive media, and further incubation of the samples in the presence or absence of therapeutic doses of the antibiotic examined . Growth of bacteria is estimated by the level of bacterial ATP in the sample, which is determined by a bioluminescence assay . Hemolysis and further incubation of samples in nutrition media reduced the concentration of nonbacterial ATP to a level that did not interfere with the determination of bacterial ATP . There was a positive correlation between the levels of resistance to antibiotics determined by the bioluminescence assay and standard plate counts. G Ital Med Lav Ergon, 1997 Jan-Mar, 19(1), 33 - 5 {Reduction of the lung cancer risk among dairy ranchers: dose response relationship with the traditional indicators of professional exposure}; Mastrangelo G et al.; An epidemiological study of mortality was carried out in the Province of Padua in a cohort of family farmers . The subjects, identified from the SCAU files, were enrolled in the cohort if informations were available on complete registry data, date of beginning and of termination of farm work, size of farm, and number of dairy cattle . The 2,415 farmers selected were followed-up for mortality through the registry offices of their Communes of residence . The cause of death was obtained for the 541 decedents . The standardised mortality ratio (SMR) was the ratio between observed and expected mortality, calculated on the basis of the mortality rates in the regional general population . The lung cancer SMR was 0.54 (0.36-0.79; observed = 27) among the 1641 dairy farmers, and 0.78 (0.46-1.25; observed = 17) among the 774 arable farmers . Among dairy farmers, moreover, lung cancer SMRs showed a significant trend across the quartiles of length of work, number of cattle, area of farm, age at beginning farm work, and age at work termination . Dairy farmers are known to be exposed to higher airborne endotoxin concentrations, and it is reasonable to assume that this cumulative exposure further increases with years of work, number of cattle, and area of farm . Microbial endotoxin might have protected dairy farmers against lung cancer through a host factor, the Tumor Necrosis Factor, produced by alveolar macrophages. Plant Mol Biol, 1997 Nov, 35(5), 673 - 9 In vivo characterization of transcriptional regulatory sequences involved in the defence-associated expression of the tobacco retrotransposon Tnt1; Vernhettes S et al.; The expression of the tobacco retrotransposon Tnt1 is induced by wounding, pathogen infections as well as microbial elicitors and abiotic factors known to induce the plant defence response . We report here that the LTR U3 region is sufficient to mediate transcriptional activation by biotic and abiotic elicitors in stable transgenic conditions . We have used in vivo footprinting techniques in order to analyse the cis-regulatory elements of the LTR U3 region that mediate the induction of Tnt1 expression . Our results indicate that a tandemly repeated short element, named BII box, is involved in the transcriptional activation of the tobacco retrotransposon Tnt1 in association with the plant defence signaling cascade. Plant Mol Biol, 1997 Nov, 35(5), 551 - 60 Isolation of the cDNAs encoding (+)6a-hydroxymaackiain 3-O-methyltransferase, the terminal step for the synthesis of the phytoalexin pisatin in Pisum sativum; Wu Q et al.; Pisatin is the major phytoalexin produced by pea upon microbial infection . The enzyme that catalyzes the terminal step in the pisatin biosynthetic pathway is (+)6a-hydroxymaackiain 3-O-methyltransferase (HMM) . We report here the isolation and characterization of two HMM cDNA clones (pHMM1 and pHMM2) made from RNA obtained from Nectria haematococca-infected pea tissue . The two clones were confirmed to encode HMM activity by heterologous expression in Escherichia coli . The substrate specificity of the methyltransferases in E . coli was similar to the activity detected in CuCl2-treated pea tissue . Nucleotide sequence analysis of Hmm1 and Hmm2 revealed an open reading frame of 1080 bp and 360 amino acid residues which would encode 40.36 kda and 40.41 kDa polypeptides, respectively . The deduced amino acid sequence of HMM1 has 95.8% identity to HMM2, 40.6% identity to Zrp4, a putative O-methyltransferase (OMT) in maize root, and 39.1% to pBH72-F1, a putative OMT induced in barley by fungal pathogens or UV light . Comparison of the deduced amino acid sequences of the cDNA clones to OMTs from other higher plants identified the binding sites of S-adenosylmethionine (AdoMet) . Southern blot analysis showed two closely linked genes with strong homology to Hmm in the pea genome. J Hosp Infect, 1997 Dec, 37(4), 263 - 72 How useful are microbial filters in respiratory apparatus? Das I, Fraise AP. Following an outbreak of hepatitis C in surgical patients in Australia, it has been suggested that transmission can take place as a result of contaminated anaesthetic circuits . It has therefore been recommended that filters should be placed between patients and breathing systems with a new filter being used for each patient . Although nosocomial pneumonia is a major manifestation of hospital-acquired infection, it is unclear whether contamination of ventilator circuits is implicated in the aetiology of this condition . Some data suggest that bacteria cannot survive well in anaesthetic circuits and several studies have failed to demonstrate significant contamination of circuits in clinical situation . Several outbreaks of pneumonia related to contaminated anaesthetic equipment have been described, but many of these were controlled by appropriate decontamination of the respiratory equipment . Although ventilator filters are used by the majority of intensive care units and filters do have the ability to filter bacteria and viruses, there are few data suggesting that the use of filters reduce the rate of pulmonary infections in long-term ventilated patients . Furthermore, to change filters between operations would have significant financial implications, and there is no conclusive evidence that they would reduce cross infection . Until more data are available on the role of filters in both long-term ventilated patients and operations, standard hygienic measures such as appropriate disinfection protocols are still the most effective way of reducing ventilator-associated infections. Rev Med Chir Soc Med Nat Iasi, 1996 Jul-Dec, 100(3-4), 193 - 7 The patients with cariogenic high-risk after the cervico-facial radiotherapy and their management; Lacatusu S et al.; Studying through clinical and laboratory means the bucco-dental status of the 28 patients who followed radiotherapeutical cures in the cervico-facial territory, the authors establish the apparition, at different periods of time, of the rampant caries . The rampant caries have been found in 46.42 percent of the cases, especially in patients exposed to prolonged or repeated cures of cervico-facial radiotherapy (88.88 percent) and concern the cervical zones, incisal edges and cusps, zones that are usually resistant at caries . The appearance and the severity of these types of caries are correlated with: a) the degree of reduction of the repose and stimulated salivary flow; b) the maintaining time of the low salivary flow that is dependent on the received X-ray dose and the number of cures; c) the association with sweet-drinks intake; d) the manner of effectuation of the bucco-dental hygiene; e) the setting-up or not of the some prevention measures . There are presented some preventive measures that must be set-up in this category of patients and treatment-protocols, that can be applied in the clinical situations of appearance of rampant caries after the irradiant treatment of the head and neck . The authors recommend the compulsory use of the prophylactic treatment and suggest a suitable plan of treatment that includes the reduction of microbial flora, the change of the alimentary diet, and salivary stimulates and substitutes . These measures can avoid the apparition of rampant caries, which will require ample restoration . The conclusions emphasize the fact that exists real possibilities of protection and treatment for these types of caries, depending on the clinical status of patient and the disease prognosis, when exists a tight collaboration between the radiotherapist, patients, and dentist. Avian Dis, 1997 Oct-Dec, 41(4), 977 - 80 Transmission electron microscopic demonstration of abnormalities on the tracheal cilia of chicks exposed to formaldehyde during hatching; Sander JE et al.; Formaldehyde gas used to fumigate hatcheries for control of microbial contamination has an adverse effect on tracheal cilia function and morphology . Evaluation of the changes revealed alterations in the ultrastructure of the axoneme with the absence of B subfibers and the production of additional A subfibers . Spikes and vesicular blebs in the cilia walls were evident in formaldehyde-exposed cilia . These changes could result in ciliastasis and cilia loss. Biochem Pharmacol, 1998 Jan 15, 55(2), 105 - 11 Multiple controls in inflammation . Extracellular and intracellular phospholipase A2, inducible and constitutive cyclooxygenase, and inducible nitric oxide synthase; Cirino G; Inflammation occurs as a defensive response to invasion of the host by foreign material, often of microbial nature . This response is normally a localized protective response that at the microscopic level involves a complex series of events including dilatation of arterioles, venules, and capillaries with increased vascular permeability, exudation of fluids including plasma proteins, and leukocyte migration into the inflammatory area . Since disease characterized by inflammation is an important cause of morbidity and mortality in humans, the processes involved in the host defense in inflammation have been and continue to be the object of several experimental studies . The role of several mediators such as histamine, serotonin, bradykinin, prostaglandins, and, more recently, cytokines and nitric oxide has been evaluated, and a contribution for each one of these mediators has been proposed . With the development of powerful molecular biology tools, it has become possible to study enzymes involved in this complex phenomenon by measuring the expression or evaluating the signaling pathways following a specific stimulus . These techniques have generated a proliferation of studies on the role of several enzymes and cytokines in inflammation . Most of these studies have been conducted in vitro on cell lines, and not many of the results have been confirmed by in vivo studies . This commentary does not pretend to analyze all of the studies and their possible in congruences, but endeavors to provoke in the reader a critical review of dogmas and current beliefs that most of the time are built on unilateral interpretation of the data. Leukemia, 1997 Dec, 11(12), 2131 - 6 Expression of NADPH oxidase is induced by all-trans retinoic acid but not by phorbol myristate acetate and 1,25 dihydroxyvitamin D3 in the human promyelocytic cell line NB4; N'Diaye EN et al.; Human promyelocytic cells, NB4, differentiate into neutrophils in response to all-trans retinoic acid (ATRA) . It has recently been proposed that NB4 cells have bilineage potential because these cells are also able to differentiate into monocyte/macrophages when exposed to a combination of 1,25-dihydroxyvitamin D3 (VD3) and phorbol myristate acetate (PMA) . Differentiation of myeloid cells into neutrophils or monocytes is associated with the acquisition of the O2- producing enzyme, NADPH oxidase, which plays a critical role in microbial killing . In this study, the expression of the components of the NADPH oxidase complex during the differentiation of NB4 cells into neutrophils or macrophages has been investigated . Whereas cells exposed to ATRA were able to produce O2- after 2 days of differentiation, they remain unable to generate O2- when exposed to PMA or PMA + VD3 . With the exception of p21rac, none of the other oxidase components was expressed in non-differentiated cells . Addition of ATRA induced the progressive expression and accumulation of p22phox, p91phox, p47phox and p67phox . Compared to the other components, p67phox was expressed late and its expression appeared to correlate most closely with the generation of O2- in the differentiation process . In PMA or PMA + VD3-differentiated NB4 cells, expression of the NADPH oxidase components was incomplete . Therefore, ATRA induced the expression of a functional NADPH oxidase complex in neutrophil-like NB4 cells . In contrast, when NB4 cells are exposed to monocytic differentiating agents, they acquire only part of the phenotypic characteristics of monocytes and lack one of the major phagocytic functionalities, the respiratory burst oxidase. Arch Oral Biol, 1997 Oct-Nov, 42(10-11), 735 - 42 Scanning electron microscopy of microbial colonization of 'rapid' and 'slow' dental-plaque formers in vivo; Zee KY et al.; The aim was to investigate the morphological features of supragingival plaque development in 'rapid' and 'slow' plaque formers using scanning electron microscopy (SEM) . Forty-nine healthy volunteers were screened for their plaque-formation rate after 3 days of oral hygiene abstinence using the Plaque Index (PI) . Five with the highest and six with the lowest mean PI were selected as 'rapid' and 'slow' plaque formers, respectively . Six enamel blocks measuring 2 x 2 x 1 mm were bonded onto the buccal surfaces of the upper left canine, premolars and first molar of each selected participant after a series of prophylaxes and oral hygiene instruction to ensure clinical gingival health . A 14-day period with no oral hygiene began thereafter . An enamel block was removed at 3 hr, 6 hr, 1 day, 3 days, 7 days and 14 days from each individual, fixed, and processed for SEM . Quantitatively, more bacteria were observed on the 3-hr to 1-day enamel blocks of the 'rapid' plaque formers . The day-1 specimens of the 'rapid' group showed a more complex supragingival plaque structure than those of the 'slow' group . From days 3 to 14, during the maturation period of supragingival plaque, there were no discernible differences between the two groups except that intermicrobial matrix was more prominent in the 'rapid' group . In general, the development of supragingival plaque followed known patterns . These observations indicate that morphological features of supragingival plaque in 'slow' and 'rapid' plaque formers differ, especially in the early developmental phases. Med Parazitol (Mosk), 1997 Oct-Dec, (4), 32 - 6 {The isolation and results of the study of the protein structure of spirochetes isolated from I . ricinus ticks in Byelarus}; Trofimov NM et al.; Four hundred and eighty Ixodes ricinus (278 female and 202 male) samples collected in the natural biotopes of 13 administrative districts in southwest Byelarus were studied via DSK-H medium (Sigma) inoculation . Twenty-four spirochetes isolates (3, 2, 1, and 18 in the Brest, Gomel, Mogilev, and Grodno regions, respectively) were obtained on the territory of Byelarus . After adapted to the medium, most isolates (as many as 7 x 10(6)-5 x 10(7) microbial cells per ml) in the stationary phase . All the obtained isolates were cryoconserved at the level of 2-6 passages and after -70 degrees C storage during 4-6 months (a followup period) they were able to recover their initial reproductive activity in the fresh BSK-H medium . Proceeding from preidentification using a comparative electrophoretic analysis of the molecular mass of polypeptides, the pattern of their specific reactivity with polyclonal serum antibodies from the rabbit immunized with cultured Borrelia afzelii (Ip21 strain), in immunoblotting and indirect immunofluorescence assay, I . ricinus spirochetes were referred to Borrelia burgdorferi sensu lato isolates. Regul Toxicol Pharmacol, 1997 Dec, 26(3), 307 - 21 Health risk assessment practices in the U.S . Food and Drug Administration; Gaylor DW et al.; The U.S . Food and Drug Administration (FDA) regulates a wide variety of consumer products . Safety issues involve chemical and microbial contaminants in food, biologies, and medical devices; side effects from prescription and nonprescription drugs; residues of animal drugs in food; and radiation from electronic devices . Because of this wide diversity, the legal standards, rules, and policies governing the regulation of these products differ considerably . Hence, risk assessment and risk management practices within the FDA are of necessity quite diverse . This paper presents a summary of risk assessment practices at each of the product centers of the FDA (Center for Food Safety and Applied Nutrition, Center for Drug Evaluation and Research, Center for Biologics Evaluation and Research, Center for Devices and Radiological Health, and Center for Veterinary Medicine) and of the development of risk assessment procedures at the National Center for Toxicological Research. J Bacteriol, 1998 Jan, 180(2), 400 - 2 Detoxification of protoanemonin by dienelactone hydrolase; Bruckmann M et al.; Protoanemonin is a toxic metabolite which may be formed during the degradation of some chloroaromatic compounds, such as polychlorinated biphenyls, by natural microbial consortia . We show here that protoanemonin can be transformed by dienelactone hydrolase of Pseudomonas sp . strain B13 to cis-acetylacrylate . Although similar Km values were observed for cis-dienelactone and protoanemonin, the turnover rate of protoanemonin was only 1% that of cis-dienelactone . This indicates that at least this percentage of the enzyme is in the active state, even in the absence of activation . The trans-dienelactone hydrolase of Pseudomonas sp . strain RW10 did not detectably transform protoanemonin . Obviously, Pseudomonas sp . strain B13 possesses at least two mechanisms to avoid protoanemonin toxicity, namely a highly active chloromuconate cycloisomerase, which routes most of the 3-chloro-cis,cis-muconate to the cis-dienelactone, thereby largely preventing protoanemonin formation, and dienelactone hydrolase, which detoxifies any small amount of protoanemonin that might nevertheless be formed. Epidemiol Infect, 1997 Dec, 119(3), 319 - 25 Residual moisture determines the level of touch-contact-associated bacterial transfer following hand washing; Patrick DR et al.; We report here a new and critical determinant of the effectiveness of hand hygiene procedures, namely the amount of residual moisture left on the hands after washing and drying . When samples of skin, food and utilities were touched with wet, undried hands, microbial numbers in the order of 68000, 31000 and 1900 respectively translocated to these representative surfaces . Bacterial numbers translocating on touch contact decreased progressively as drying with an air or cloth towel system removed residual moisture from the hands . A 10 s cloth towel-20 s air towel protocol reduced the bacterial numbers translocating to skin, food and utilities on touch contact to 140, 655 and 28 respectively and achieved a 99.8, 94 and 99% reduction in the level of bacterial translocation associated with wet hands . Careful hand drying is a critical factor determining the level of touch-contact-associated bacterial transfer after hand washing and its recognition could make a significant contribution towards improving handcare practices in clinical and public health sectors. Genetica, 1997, 100(1-3), 241 - 52 The expression of the tobacco Tnt1 retrotransposon is linked to plant defense responses; Grandbastien MA et al.; Activation of retrotransposons by stresses and external changes is common in all eukaryotic systems, including plants . The transcription of the tobacco Tnt1 retrotransposon was studied in its natural host as well as in Arabidopsis and tomato . It is activated by factors of microbial origin, by external stresses, and by viral, bacterial, and fungal attacks . Tnt1 expression is linked with the biological responses of the plant to the elicitor or to the pathogen attack and in particular with the early steps of the metabolic pathways leading to the activation of plant defense genes . In most cases, the basic features of Tnt1 regulation in tobacco are maintained in tomato and Arabidopsis, but some host-specific regulations were shown . The U3 region of the Tnt1 LTR contains the major cis-acting components of Tnt1 transcriptional activation in association with the plant defense responses . Furthermore, the Tnt1 U3 region, and especially the tandemly repeated BII boxes, contains several sequences similar to well-characterized motifs involved in the activation of several plant defense genes . The possible origin of Tnt1 regulatory sequences as well as the biological implications of Tnt1 activation by pathogen attacks are discussed. Arch Biochem Biophys, 1998 Jan 1, 349(1), 161 - 6 Site-directed mutagenesis of bacterial hemoglobin: the role of glutamine (E7) in oxygen-binding in the distal heme pocket; Dikshit KL et al.; The bacterial and yeast hemoglobins have a glutamine instead of histidine in the E7 position of the distal heme pocket . The recently determined crystal structure of Vitreoscilla hemoglobin (VHb) indicates that this residue is oriented out of the heme pocket and may not ligand the bound oxygen . This is in contrast to elephant myoglobin which also has a Gln(E7) but which does ligand the bound oxygen . This residue was changed in VHb using site-directed mutagenesis to leucine (VHbL) or to histidine (VHbH) . Spectral and kinetic studies of the binding of oxygen and CO to VHbL showed that this substitution had little effect on the ligand-binding properties of this protein, evidence that Gln(E7) does not H-bond the bound ligand, in agreement with the findings of the crystallographic study of VHb . In contrast, the functional properties of VHbH were drastically altered in a way suggesting that the E7His may itself be liganded to the heme iron . These studies are further evidence that the distal heme pocket in VHb and related microbial hemoglobins differs from that in mammalian hemoglobins and may resemble in some ways the heme pocket in cytochrome b5. Am J Ophthalmol, 1997 Sep, 124(3), 394 - 5 Corneal ulcer associated with contamination of aerosol saline spray tip; Donzis PB; PURPOSE: To report a complication of aerosol saline use in a contact lens wearer . METHODS: Case report . A 57-year-old woman who used soft contact lenses on an extended-wear basis developed a unilateral Pseudomonas corneal ulcer associated with the use of preservative-free aerosol saline . RESULTS: The solution inside the aerosol can was free of microbial contamination . The spray tip, however, was contaminated with P aeruginosa . CONCLUSIONS: Aerosol spray cans may minimize contamination of the solution inside the can . The spray tip is still susceptible to microbial contamination, especially with continued use of the solution over an extended period of time. J Dairy Sci, 1997 Dec, 80(12), 3283 - 92 Effects of dietary supplementation on nutrient digestion and the milk yield of intensively grazed lactating dairy cows; Jones-Endsley JM et al.; Fourteen midlactation Holstein cows were used in an 80-d study to examine supplementation strategies during intensive rotational grazing . Factors examined were the concentration of protein in the supplement {12 or 16% crude protein (CP) on a dry matter basis} and the amount of supplement offered (6.4 or 9.6 kg/d per cow) . The supplement was offered in equal portions three times daily after milking . Pasture, composed of alfalfa (Medicago sativa) and orchardgrass (Dactylis glomerata), was divided into 27 0.16-ha paddocks that were grazed for 12 to 24 h . Intake of forage tended to increase as CP in the supplement increased but was unaffected by the amount of supplement offered . Intakes of organic matter and dry matter and the digestion of these nutrients in the total tract increased as CP in the supplement increased and as the amount of supplement increased . Ruminal pH and concentrations of volatile fatty acids were unaffected by treatments, but concentrations of NH3 N increased as CP in the supplement increased . An increase in CP resulted in a greater intake and flow of total N to the duodenum . The flow of microbial N to the duodenum and the efficiency of microbial protein synthesis were unaffected by treatment . Flows of total amino acids and essential amino acids to the duodenum tended to increase as CP in the supplement increased . Yield of fat-corrected milk and concentrations of fat and protein in milk were unaffected by treatment . These results suggested that the supply and digestion of nutrients in grazing dairy cows may be improved through an increase in the CP concentration of the supplement or the amount of supplement offered . However, effects on the yield of milk and milk components may be small. Adv Biochem Eng Biotechnol, 1998, 59, 73 - 121 Novel screen methodologies for identification of new microbial metabolites with pharmacological activity; Hill DC et al.; Micro-organisms continue to provide an important source of chemical diversity for the discovery of compounds with new biological activities . Microbial metabolites discovered recently using assays to detect compounds with potential pharmacological utility are surveyed and found to represent an extensive range of structural types produced by a wide variety of organisms . Assays used for screening samples produced by microbial processes must be robust, sensitive and specific and able to operate above a background of potential interferences from a number of sources . Discovery assays currently in use fall into three main categories cell-based, receptor-ligand interaction and enzyme inhibition assays . Trends in the use of these assays and new developments in assay technology applicable to the screening of microbial samples are examined with particular reference to the high throughput screening environment . For microbial screening to be a competitive route to new drug leads, the disciplines involved must be engineered into a seamlessly integrated process to deliver novel compounds with the required biological properties rapidly. Appl Environ Microbiol, 1998 Jan, 64(1), 185 - 91 Use of subtractive hybridization to design habitat-based oligonucleotide probes for investigation of natural bacterial communities; Mau M et al.; We describe a rapid oligonucleotide probe design strategy based on subtractive hybridization which yields probes for 16S rRNA or rRNA genes of individual members of microbial communities that are specific within the context of those communities . This strategy circumvents the need to sequence many similar or identical clones of dominant members of a community . Radioactively labeled subfragments of a cloned 16S rRNA gene sequence for which a probe is required (target) were hybridized with biotinylated total 16S ribosomal DNA (rDNA) amplified from the microbial community, and the hybrids formed were subsequently discarded . The remaining enriched fragments were used to screen a library consisting of cloned subfragments of the target sequence by colony hybridization in order to identify the variable regions of the 16S rRNA gene with the required specificity . The sequencing of random clones in one 16S rDNA library demonstrated that only those clones with 100% sequence identity with the probe fragment were detected by it . Moreover, sequencing of other, randomly selected, probe-positive clones revealed 100% sequence identity with the probe . Probes developed in this way tended to correspond to more variable regions of the 16S rRNA if the target sequences were similar to the sequences of other clones in the library and to less variable regions if the target sequences were phylogenetically isolated within the clone library . Although the absolute specificity of the latter probes, as assessed by comparison with available database sequences, was lower than the absolute specificity of the probes from the more variable regions, they were specific within the context of the environmental samples from which they were derived. Ophthal Plast Reconstr Surg, 1997 Dec, 13(4), 289 - 92 Conjunctival cilia entrapment: an unrecognized cause of ocular irritation; Hunts JH et al.; Conjunctival entrapment of cilia is form of ocular irritation that to our knowledge has not yet been described . Scarring predisposes the conjunctiva to this phenomenon by creating blind recesses in which loose cilia can become entrapped . Once caught, the rigid cilia can act as a mechanical irritant or act as a nidus for microbial growth . We present herein three patients with entrapped cilia and discuss their pathophysiology and management . The symptoms in all three patients resolved with conjunctivoplasty and removal of the cilia. Phytochemistry, 1997 Dec, 46(7), 1193 - 5 Glucose-conjugation of the flavones of Psiadia arabica by Cunninghamella elegans; Ibrahim AR et al.; Microbial transformation of psiadiarabin and its 6-desmethoxy analogue 5,3' dihydroxy-7,2',4'5'-tetramethoxyflavone by Cunninghamella elegans NRRL 1392 gave the 3'-glucoside conjugates of the two flavones . Structural elucidation of these two new metabolites was achieved using 1D and 2D NMR spectroscopy and CIMS. Shock, 1997 Dec, 8(6), 391 - 401 Wound healing in sepsis and trauma; Thornton FJ et al.; Wound healing represents a dynamic and immediate response of the body to tissue injury with the purpose of restoring anatomical continuity, structure and function . Success or failure of this complex cascade of events is determined largely by competence of the host's immune system . Sepsis represents one of the most formidable threats to successful wound healing . It can present as a local bacterial colonization of the injury site with minimal systemic reaction or the "systemic inflammatory response syndrome," a primary cause of mortality among critically ill patients . Trauma also predisposes patients to wound complications especially as a result of post-traumatic immunosuppression . This phenomenon exposes the patient to the risk of microbial infection and ultimately the sepsis syndrome . The immune system, therefore, represents a vulnerable gateway through which trauma and sepsis exert their deleterious effect on the wound healing process resulting in increased morbidity and mortality for the surgical patient. Electrophoresis, 1997 Oct, 18(11), 2019 - 23 Sodium dodecyl sulfate-polymer capillary electrophoresis for the analysis of cell culture proteins; Klyushnichenko V et al.; The application of sodium dodecyl sulfate (SDS)-polymer capillary electrophoresis (CE) to the analysis of proteins of hybridoma cell culture supernatant is demonstrated . All steps of the developed analysis are shown and discussed . Following optimization of sample preparation and concentration, as well as of the mode of injection all main protein components were separated with high resolution and selectivity . Proteolytic degradation of IgG, due to microbial contamination, was observed on SDS-gel CE and capillary zone electrophoresis (CZE) . The technique was applied to monitor the cell cultivation process and concentrations of main components are demonstrated . IgG was found to increase from initially 5 mg/L to 20 mg/L after 4 days and to > 130 mg/L after 14 days of cultivation . The techniques may be applied to other biotechnological processes for the production or purification of proteins. Crit Rev Immunol, 1997, 17(5-6), 469 - 80 Initiation and regulation of CNS autoimmunity; Goverman J et al.; Our studies addressed the questions of how self-reactive T cells escape tolerance and what stimuli cause these T cells to initiate autoimmune responses . We employed experimental allergic encephalomyelitis (EAE) as an animal model of multiple sclerosis (MS) . Endogenous expression of myelin basic protein (MBP) induces tolerance in T cells that recognize one region of MBP, whereas T cells specific for a different region escape tolerance . Triggers of disease induction were investigated in a T-cell receptor (TCR) transgenic model in which the majority of T cells recognize the MBP epitope that does not induce tolerance . EAE occurs spontaneously in this model and the incidence of disease depends on microbial exposure . EAE can also be actively induced by immunization with MBP peptide accompanied by injection of pertussis toxin as well as by administration of pertussis toxin alone . Immunization with MBP peptide without pertussis toxin, however, stimulates the transgenic T cells, but the activated T cells do not accumulate in the central nervous system (CNS) or induce EAE . Our studies suggest that initiation of autoimmune disease involves complex interactions between the neuroendocrine system as well as the innate and specific immune systems. J Infect Dis, 1998 Jan, 177(1), 182 - 7 Pneumocystis carinii glycoprotein A inhibits surfactant phospholipid secretion by rat alveolar type II cells; Lipschik GY et al.; Pneumocystis carinii pneumonia (PCP) remains a major cause of morbidity in AIDS . The pathogenesis of PCP is poorly understood, but evidence of surfactant abnormalities is mounting . The role of the major surface glycoprotein of P . carinii, gpA, in producing surfactant abnormalities was investigated . Rat type II pneumocytes were incubated with {3H}choline, purified gpA, and modulators . Lipid was extracted, and {3H}dipalmitoyl phosphatidylcholine (DPPC) secretion was calculated . Contaminating endotoxin had no effect on DPPC secretion . gpA inhibited basal and ATP-stimulated DPPC secretion in a dose- and time-dependent manner . An anti-gpA monoclonal antibody attenuated inhibition of DPPC secretion . Unglycosylated recombinant gpA inhibited secretion, suggesting that functional activity resides in the protein moiety of gpA . These results suggest that gpA is a specific trigger for abnormalities of surfactant lipids in PCP . This is a unique role for a microbial product in disease pathogenesis and a potentially exploitable therapeutic target. Trends Biotechnol, 1997 Dec, 15(12), 523 - 30 Directed evolution of enzyme catalysts; Kuchner O et al.; Directed enzyme evolution has emerged in the past few years as a powerful alternative to rational approaches for engineering biocatalysts . Prerequisites for successful directed evolution are functional expression in a suitable microbial host, a rapid screen for the desired feature(s) and a well-thought-out working strategy for navigating protein landscapes . The rapidly growing body of literature on enzyme evolution in vitro includes techniques for creating and searching combinatorial enzyme libraries, as well as several successful examples of different evolutionary strategies being used. J Exp Med, 1997 Dec 1, 186(11), 1819 - 29 In vivo microbial stimulation induces rapid CD40 ligand-independent production of interleukin 12 by dendritic cells and their redistribution to T cell areas; Reis e Sousa C et al.; The early induction of interleukin (IL)-12 is a critical event in determining the development of both innate resistance and adaptive immunity to many intracellular pathogens . Previous in vitro studies have suggested that the macrophage (MPhi) is a major source of the initial IL-12 produced upon microbial stimulation and that this response promotes the differentiation of protective T helper cell 1 (Th1) CD4+ lymphocytes from precursors that are primed on antigen-bearing dendritic cells (DC) . Here, we demonstrate by immunolocalization experiments and flow cytometric analysis that, contrary to expectation, DC and not MPhi are the initial cells to synthesize IL-12 in the spleens of mice exposed in vivo to an extract of Toxoplasma gondii or to lipopolysaccharide, two well characterized microbial stimulants of the cytokine . Importantly, this production of IL-12 occurs very rapidly and is independent of interferon gamma priming or of signals from T cells, such as CD40 ligand . IL-12 production by splenic DC is accompanied by an increase in number of DCs, as well as a redistribution to the T cell areas and the acquisition of markers characteristic of interdigitating dendritic cells . The capacity of splenic DC but not MPhi to synthesize de novo high levels of IL-12 within hours of exposure to microbial products in vivo, as well as the ability of the same stimuli to induce migration of DC to the T cell areas, argues that DC function simultaneously as both antigen-presenting cells and IL-12 producing accessory cells in the initiation of cell-mediated immunity to intracellular pathogens . This model avoids the need to invoke a three-cell interaction for Th1 differentiation and points to the DC as both a sentinel for innate recognition and the dictator of class selection in the subsequent adaptive response. Plant Physiol, 1997 Dec, 115(4), 1619 - 27 Identification and characterization of a novel arabinoxylanase from wheat flour; Cleemput G et al.; An endogenous wheat (Triticum aestivum) flour endoxylanase was purified to homogeneity from a crude wheat flour extract by ammonium sulfate precipitation and cation-exchange chromatography . The 30-kD protein had an isoelectric point of 9.3 or higher . A sequence of 19 amino acids at the NH2 terminus showed 84.2% identity with an internal sequence of 15-kD grain-softness protein, friabilin . High-performance anion-exchange chromatography and gel-permeation analysis of the hydrolysis products indicated the preferential hydrolysis of highly branched structures by the enzyme; wheat arabinoxylan and rye (Secale cereale) arabinoxylan (high arabinose to xylose ratios) were hydrolyzed more efficiently by this enzyme than oat (Avena sativa) spelt xylan (low arabinose to xylose ratios) . The release of the hydrolysis products as a function of time suggested that the endoxylanolytic activity was associated with the release of arabinose units from the polysaccharides, suggesting that the enzyme action is similar to that by endoxylanases from Ceratocystis paradoxa, Aspergillus niger, and Neurospora crassa . Although the enzyme released arabinose from arabinoxylan, it did not hydrolyze p-nitrophenyl-alpha-L-arabinofuranoside . From the above, it follows that the enzyme, called arabinoxylanase, differs from most microbial endoxylanases and from an endoxylanase purified earlier from wheat flour. Clin Chest Med, 1997 Dec, 18(4), 695 - 706 Etiology of sarcoidosis; Moller DR; Since sarcoidosis was first recognized as a distinct clinical entity, investigators have speculated that a transmissible agent may cause sarcoidosis . Recent attempts at directly isolating infectious organisms or indirectly detecting microbial DNA or RNA from sarcoid tissue have led to inconclusive results . Studies on the immunopathogenic origins of sarcoidosis have provided evidence of persistent antigenic stimulation at sites of inflammation that are associated with dysregulated cytokine production . To date, however, the challenge of defining the cause of sarcoidosis remains unmet. Semin Liver Dis, 1997, 17(4), 297 - 310 Complement and complement deficiencies; Whaley K et al.; The complement system provides a first line of defense and mediates a large variety of cellular and humoral interactions within the immune response, including chemotaxis, phagocytosis, cell adhesion, and B-cell differentiation . The system involves more than 30 serum components and numerous cell surface regulators and receptors . Similar to the blood clotting system, complement activation is initiated through a series of complex activation cascades involving enzymatic cleavage . Three independent complement activation cascades, the classical, the alternative, and the lectin pathway, have been described . The liver is the main site of biosynthesis for most of the serum components of complement and diseases of the liver can lead to alterations of the normally stable plasma levels of complement . Deficiencies of single components can lead to a broad variety of secondary diseases, caused by either imbalanced activation or defects in the humoral or cellular response to microbial infections. J Capillary Electrophor, 1996 Jul-Aug, 3(4), 209 - 13 Capillary electrophoretic separation of TNT and its transformation products; Pucik L et al.; 2,4,6-Trinitrotoluene (TNT), 2-aminodinitrotoluene (2ADNT), 4-aminodinitrotoluene (4ADNT), and diamino-6-nitrotoluene (DA6NT) in a variety of matrices (including chemical, microbial, and plant extract) were successfully separated and quantified using micellar electrokinetic capillary electrophoresis (MEKC) . The method used a buffer solution of 20 mM in borax, 50 mM in sodium dodecyl sulfate (SDS), and of pH 8.5 . Voltage applied across the capillary was 20 kV and temperature was maintained at 25 degrees C . Compounds eluted within approximately 6-11 min and were identified by both their migration times and spectra. Ophthalmology, 1997 Nov, 104(11), 1902 - 9 Ofloxacin monotherapy for the primary treatment of microbial keratitis: a double-masked, randomized, controlled trial with conventional dual therapy . The Ofloxacin Study Group; Crystal structure of methyl-coenzyme M reductase: the key enzyme of biological methane formation; Max-Planck-Institut fur Biophysik, Heinrich-Hoffmann-Strabetae 7, 60528 Frankfurt, GermanyMethyl-coenzyme M reductase (MCR), the enzyme responsible for the microbial formation of methane, is a 300-kilodalton protein organized as a hexamer in an alpha2beta2gamma2 arrangement . The crystal structure of the enzyme from Methanobacterium thermoautotrophicum, determined at 1.45 angstrom resolution for the inactive enzyme state MCRox1-silent, reveals that two molecules of the nickel porphinoid coenzyme F430 are embedded between the subunits alpha, alpha', beta, and gamma and alpha', alpha, beta', and gamma', forming two identical active sites . Each site is accessible for the substrate methyl-coenzyme M through a narrow channel locked after binding of the second substrate coenzyme B . Together with a second structurally characterized enzyme state (MCRsilent) containing the heterodisulfide of coenzymes M and B, a reaction mechanism is proposed that uses a radical intermediate and a nickel organic compound. Appl Environ Microbiol, 1997 Dec, 63(12), 4645 - 50 Frequency of formation of chimeric molecules as a consequence of PCR coamplification of 16S rRNA genes from mixed bacterial genomes; Wang GC et al.; PCR is routinely used in amplification and cloning of rRNA genes from environmental DNA samples for studies of microbial community structure and identification of novel organisms . There have been concerns about generation of chimeric sequences as a consequence of PCR coamplification of highly conserved genes, because such sequences may lead to reports of nonexistent organisms . To quantify the frequency of chimeric molecule formation, mixed genomic DNAs from eight actinomycete species whose 16S rRNA sequences had been determined were used for PCR coamplification of 16S rRNA genes . A large number of cloned 16S ribosomal DNAs were examined by sequence analysis, and chimeric molecules were identified by multiple-sequence alignment with reference species . Here, we report that the level of occurrence of chimeric sequences after 30 cycles of PCR amplification was 32% . We also show that PCR-induced chimeras were formed between different rRNA gene copies from the same organism . Because of the wide use of PCR for direct isolation of 16S rRNA sequences from environmental DNA to assess microbial diversity, the extent of chimeric molecule formation deserves serious attention. J Dairy Sci, 1997 Nov, 80(11), 2901 - 6 Substitution of neutral detergent fiber from forage with neutral detergent fiber from by-products in the diets of lactating cows; Zhu JS et al.; Four lactating dairy cows that were ruminally and duodenally cannulated were used in an experiment with a 4 x 4 Latin square design to determine the effects of the substitution of neutral detergent fiber (NDF) from forage with NDF from wheat middlings, corn gluten feed, or a blend of distillers dried grains and hominy . Dietary crude protein and NDF averaged 18 and 31%, respectively, for the diet with 71.2% of the NDF from forage (control diet) and for diets with 55% of the NDF from forage (by-product diets) . The substitution of NDF from these by-products for forage NDF did not affect dry matter intake (20.1 kg/d) or digestibility of organic matter . Total tract digestibility of acid detergent fiber was lower for cows fed the diet containing a blend of distillers dried grains and hominy than for cows fed the diet containing corn gluten feed . Microbial crude protein synthesis, milk production (23.9 kg/d), and milk fat percentage were similar for all cows, regardless of diet . Cows fed the diets containing wheat middlings or a blend of distillers dried grains and hominy had reduced ruminal pH compared with that of cows fed the diet containing corn gluten feed or the control diet . Diets containing 55% of total NDF from forage with 31% of total NDF from corn gluten feed, wheat middlings, or a blend of distillers dried grains and hominy can supply sufficient effective fiber to maintain normal ruminal function. Aust Vet J, 1997 Nov, 75(11), 796 - 9 Use of a multifenestrated indwelling lavage system for treatment of septic digital tenosynovitis in cattle; Anderson DE et al.; Septic tenosynovitis was diagnosed in seven cattle on the basis of history, physical examination, radiographs, cytological examination of tendon sheath fluids, and microbial culture . A commercially available indwelling multifenestrated silicone rubber drain was used to perform frequent lavage of the flexor tendon sheaths . The sepsis resolved in all cattle . Five of six cattle for which long-term (> 1 year) follow-up information was available were clinically sound on the affected limb and had remained productive members of the herd. Anaesthesia, 1997 Nov, 52(11), 1073 - 6 Convection warmers--not just hot air; Avidan MS et al.; We sought to determine whether the forced air convection warmers (nine Bair Huggers, Augustine Medical, and one Warm Touch, Mallinkrodt Medical) used in our operating theatres could be a source of microbial pathogens . Agar plates were placed directly in the air stream of the warmers . Four of these grew potentially pathogenic organisms . When the warmers were set to blow through perforated blankets, no growth occurred . Three of the warmers were swabbed and sites of colonisation were found in their hoses . After fixing a microbial filter to the end of the hose, organisms were no longer detectable . We conclude that these warming devices are a potential source of nosocomial infection . They should only be used in conjunction with perforated blankets, should have their microbial filters changed regularly and their hoses sterilised . The inclusion of a microbial filter into the nozzle of the hose could be incorporated into the design of the warmer. J Periodontal Res, 1997 Oct, 32(7), 608 - 13 Polyclonal B cell activators and in vitro induction of auto-antibody reactive with collagen; Hahn CL et al.; Cells producing autoantibodies are known to be present in chronically inflamed periodontal tissues . In sites of chronic inflammation, polyclonal B cell activators (PBA) are known to exhibit adjuvant activity when combined with foreign antigens . These results prompted an examination of PBA in eliciting an antibody response to an autoantigen (i.e . collagen type I) . Rat lymphocytes were stimulated with rat collagen (type I), microbial PBA (LPS) or the combination of LPS plus rat collagen in vitro . Anti-collagen antibody-forming cells (AFC) were enumerated using an ELISPOT assay . Collagen or LPS alone elicited few anti-collagen AFC but the addition of LPS to collagen resulted in a substantial adjuvant effect and yielded maximal responses to collagen . Comparisons of anti-collagen AFC from short-term immunized (2-6 wk after booster), non-immunized and long-term immunized (3-4 months after booster) animals were performed . It revealed that cells from recently immunized rats were significantly easier to activate than the other 2 groups . The adjuvant effect of microbial PBA may be important in anti-collagen antibody production and thus the localization of PBA in periodontal pockets may explain why anti-collagen AFC are restricted to the chronically inflamed periodontal tissues. J Leukoc Biol, 1997 Dec, 62(6), 693 - 701 Natural killer cells: endothelial interactions, migration, and target cell recognition; Timonen T; Natural killer (NK) cells form a unique third group of lymphocytes that differs from T and B cells in surface phenotype, target recognition, and function . By producing cytokines and exerting cytotoxicity, NK cells participate in the resistance against microbial infections and malignant disease . The research on the molecular mechanisms of migration and target cell recognition by NK cells has recently developed rapidly . NK cells express a number of adhesion molecules common to hematopoietic lineage, bind to endothelium, extravasate, and respond to chemotactic stimuli, much resembling T cells in those respects . However, NK cells are probably capable of transmigration and infiltration merely through activation by cytokines and chemokines, as opposed to the requirement of antigen presentation in the initial activation of T cells . Target cell recognition and ensuing cytotoxicity of NK cells is a sum effect of a delicate balance between the effects of inhibitory and activating NK cell receptors . NK cells express several well-defined MHC I-recognizing receptors that inactivate their functions . In pathological alterations of MHC I expression, the inhibitory receptors do not engage and thus permit the lysis of the target cell . The receptors that trigger the cytolytic machinery of NK cells are less well known . Some candidate triggering receptors have been identified and it seems that NK cell triggering is mediated by multiple receptors, as is the inhibition of cytotoxicity . For example, NK cells clearly detect target cell-bound antibodies and thus mediate antibody-dependent cytotoxicity . They may also detect carbohydrate moieties, normal but pathologically distributed adhesion molecules, as well as ligands for a number of co-stimulatory receptors. J Biomater Sci Polym Ed, 1997, 8(12), 947 - 61 Rifampicin carrying polyhydroxybutyrate microspheres as a potential chemoembolization agent; Kassab AC et al.; In this study, we attempted to prepare microspheres from a microbial biodegradable polyester, i.e . polyhydroxybutyrate (PHB) as a potential chemoembolization agent . The drug loaded PHB microspheres were prepared by a solvent evaporation technique, in which methylene chloride, distilled water, and polyvinyl alcohol were utilized as the solvent, dispersion medium, and emulsifier, respectively . Microspheres were obtained within a size range of 5-100 microns by changing the initial polymer/solvent ratio, emulsifier concentration, stirring rate, and initial drug concentration . It was possible to obtain PHB with very narrow size distributions by applying gravity field-flow fractionation technique . Very high drug loadings of up to 407.6 mg rifampicin/g PHB were achieved . Drug release rates were very rapid . Almost 90% of the drug loaded was released in about 24 h . Both the size and drug content of PHB microspheres were found to be effective in controlling the drug release from these microspheres. Am J Kidney Dis, 1997 Dec, 30(6), 859 - 71 National Kidney Foundation report on dialyzer reuse . Task Force on Reuse of Dialyzers, Council on Dialysis, National Kidney Foundation; Mechanisms of action of anti-GM1 and anti-GQ1b ganglioside antibodies in Guillain-Barré syndrome; University Department of Neurology, Institute of Neurological Sciences, Southern General Hospital, Glasgow, United KingdomAnti-GM1 and anti-GQ1b ganglioside antibodies are found in association with acute and chronic peripheral neuropathies, including Guillain-Barre syndrome . They are believed to arise as a result of molecular mimicry with immunogenic microbial polysaccharides . Although anti-ganglioside antibodies are suspected to play a causal role in neuropathy pathogenesis, the details of this have yet to be proven . The approach in this laboratory to solving this issue has been to generate anti-GM1 and anti-GQ1b monoclonal antibodies from peripheral blood lymphocytes of affected patients and to study their immunolocalization in peripheral nerve and their electrophysiologic effects in animal models in which peripheral nerve sites are exposed to anti-ganglioside antibodies . These data show that anti-ganglioside antibody-reactive epitopes are widely distributed in peripheral nerve and can cause electrophysiologic abnormalities in a variety of model systems; thus, these data support the view that anti-ganglioside antibody-reactive epitopes may directly contribute to neuropathy pathogenesis. JAMA, 1997 Dec 10, 278(22), 1946 - 55 Immunopathogenesis of gastrointestinal and hepatobiliary diseases; Galperin C et al.; The largest lymphoid organ in the body is the gut and the gut-associated lymphoid tissue . The mucosal immune system faces many challenges in protecting the body from microbial invasion . Its chief function is to maintain a diverse population of mature lymphocytes capable of responding to foreign antigens . This task is accomplished with a variety of unique features that distinguish the mucosal from the systemic immune system . In addition, the mucosal immune system plays a role in inflammatory bowel disease, Whipple disease, autoimmune gastritis, Helicobacter pylori infection, immunoproliferative small intestinal disease, hepatitis A, B, C, D, E, F, and G, autoimmune hepatitis, primary biliary cirrhosis, progressive sclerosing cholangitis, and vanishing bile duct syndrome. Ann Acad Med Singapore, 1997 Jul, 26(4), 395 - 400 Penetrating keratoplasty in the Singapore National Eye Centre and donor cornea acquisition in the Singapore Eye Bank; Chan CM et al.; We analyzed all penetrating keratoplasties performed in the Singapore National Eye Centre from 1 January 1991 to 31 December 1995, using records of the Singapore Eye Bank Registry, evaluating the indications, complications, causes of graft failure, visual outcome and graft survival rate . We also looked into donor cornea acquisition in the Singapore Eye Bank and its influence on the development of corneal transplantation in the Singapore National Eye Centre . A total of 327 penetrating keratoplasties were performed during the 5-year period . Bullous keratopathy was an indication in 26.3% of cases . Of these, aphakic bullous keratopathy accounted for 11.6% of all cases, while pseudophakic bullous keratopathy accounted for 11.3% . Other indications were regrafts (11.9%), corneal dystrophies (10.4%), traumatic corneal scarring (10.1%) and keratoconus (9.8%) . Graft rejection was a complication in 20% of all cases . Of these, 40.9% led to graft failure . Other major complications were raised intraocular pressure (18%), epithelium-related problems (7.3%), wound dehiscence (4.3%), cataract (3.3%) and microbial keratitis (3.1%) . The main causes of graft failure were graft rejection (8.2%), endothelial failure (2.4%), infection (2.4%) and glaucoma (2.1%) . Of the 327 grafts, 40.3% achieved best corrected visual acuity of 6/12 or better; 70.8% achieved vision of 6/24 or better . The overall graft survival rate was 82.3% after a mean follow-up period of 2 years . Donor corneas for the penetrating keratoplasties were obtained from foreign eye banks as well as locally, with the local donation rate steadily increasing from 1991 to 1996, with the establishment of proper eye banking facilities and the Singapore Eye Bank . These results show that the indications and outcome of penetrating keratoplasty in the Singapore National Eye Centre are similar and comparable to that of other centres with established corneal grafting programmes . The establishment of the Singapore Eye Bank has ensured the proper co-ordination of acquisition of donor material which has been vital to the development of corneal transplantation in the Singapore National Eye Centre. Am J Gastroenterol, 1997 Dec, 92(12 Suppl), 5S - 11S Pathogenesis and immune mechanisms of chronic inflammatory bowel diseases; Sartor RB; The inflammatory bowel diseases (IBDs) are characterized by intestinal inflammation of unknown etiology . Two distinct disorders, Crohn's disease and ulcerative colitis, have been identified . Three theories of IBD etiology are currently under consideration: 1) reaction to a persistent intestinal infection, 2) existence of a defective mucosal barrier to luminal antigens, and 3) a dysregulated host immune response to ubiquitous antigens . In each of these theories, either pathogenic or resident luminal bacteria constantly stimulate the mucosal and systemic immune systems to perpetuate the inflammatory cascade . Chronicity of inflammation results from an interaction of the persistent stimulus of microbial antigens with genetically determined host susceptibility factors that determine the individual's immune response or mucosal barrier function . The pathogenesis of IBD involves a series of steps, beginning with the breach of the intestinal mucosal barrier by infectious agents or toxins . The defective barrier exposes lamina propria immune cells to the continual presence of resident luminal bacteria, bacterial products, or dietary antigens, which perpetuates the inflammatory cascade . Many immunoregulatory abnormalities are noted in IBD, including the ratio of proinflammatory to immunosuppressive cytokines, selective activation of T(H) lymphocyte subsets, and abnormalities in epithelial antigen presentation . When activated during the initial inflammatory process, macrophages and T lymphocytes secrete a host of cytokines, which recruit other inflammatory cell types, thereby continuing the process . Tissue injury is the net result of the soluble products of the activated inflammatory cells . Knowledge of the pathogenesis in IBD suggests that the ultimate goals of therapy should be to block the proinflammatory mediators toward the proximal, rather than the distal, end of the cascade, to decrease the constant antigenic drive of luminal bacteria, and to correct the dysregulated immune response. Eur J Immunol, 1997 Nov, 27(11), 2959 - 64 Hypermutation, diversity and dissemination of human intestinal lamina propria plasma cells; Dunn-Walters DK et al.; In this work we have microdissected lamina propria plasma cells and used polymerase chain reaction and sequencing to investigate immunoglobulin (Ig) gene rearrangements and mutations in human intestine . In addition, specific primers were designed for individual Ig gene rearrangements to analyze the distribution of related B cell and plasma cell clones at different sites along the bowel . Confirming our earlier work, intestinal IgVH genes were highly mutated in plasma cells from older individuals (> 30 years) . IgVH genes were significantly less mutated in samples taken from patients aged 11-30 years, and there were fewer mutations again in samples from young children (< 11 years) . In age-matched specimens the number of mutations was equivalent in the duodenum and colon . Using complementarity-determining region 3 primers to amplify specific Ig gene rearrangements, evidence was also found for the existence of related lamina propria plasma cells along the small bowel and colon, although these were quite scarce . In addition, analysis of the numbers of related clones in a random sampling from discrete areas of lamina propria indicates that the local population is diverse . These results suggest that the highly mutated IgVH genes in adult intestinal plasma cells are a consequence of chronic antigen exposure with age . Duodenal plasma cells are as highly mutated as colonic plasma cells, despite the fact that the upper bowel has no indigenous microbial flora (the stimulus for intestinal plasma cells) . They also show that the plasma cell population is diverse and can be widely disseminated along the bowel. Eur J Immunol, 1997 Nov, 27(11), 2812 - 21 Control of self-reactive cytotoxic T lymphocytes expressing gamma delta T cell receptors by natural killer inhibitory receptors; Halary F et al.; The majority of peripheral blood gamma delta T cells in human adults expresses T cell receptors (TCR) with identical V regions (V gamma 9 and V delta 2) . These V gamma 9 V delta 2 T cells recognize the major histocompatibility complex (MHC) class I-deficient B cell line Daudi and broadly distributed nonpeptidic antigens present in bacteria and parasites . Here we show that unlike alpha beta or V gamma 9- gamma delta T cells, the majority of V gamma 9V delta 2 T cells harbor natural killer inhibitory receptors (KIR) (mainly CD94/NKG2A heterodimers), which are known to deliver inhibitory signals upon interaction with MHC class I molecules . Within V gamma 9V delta 2 T cells, KIR were mainly expressed by clones exhibiting a strong lytic activity against Daudi cells . In stark contrast, almost all V gamma 9V delta 2 T cell clones devoid of killing activity were KIR-, thus suggesting a coordinate acquisition of KIR and cytotoxic activity within V gamma 9V delta 2 T cells . In functional terms, KIR inhibited lysis of MHC class I-positive tumor B cell lines by V gamma 9V delta 2 cytotoxic T lymphocytes (CTL) and raised their threshold of activation by microbial antigens presented by MHC class I-positive cells . Furthermore, masking KIR or MHC class I molecules revealed a TCR-dependent recognition by V gamma 9V delta 2 CTL of ligands expressed by activated T lymphocytes, including the effector cells themselves . Taken together, these results suggest a general implication of V gamma 9V delta 2 T cells in immune response regulation and a central role of KIR in the control of self-reactive gamma delta CTL. J Oral Maxillofac Surg, 1997 Dec, 55(12 Suppl 5), 25 - 30 The influence of 0.12% chlorhexidine digluconate rinses on the incidence of infectious complications and implant success; Lambert PM et al.; The effect of perioperative chlorhexidine on the frequency of infectious complications through stage II was examined . Chlorhexidine was used perioperatively in 54.6% of patients (52.5% of implants) in a Dental Implant Clinical Research Group study with a database of 2,641 implants (595 patients) . With chlorhexidine, there was a significant reduction in the number of infectious complications (4.1% vs 8.7%) . Two percent of implants failed in the absence of an infectious complication, whereas 12% with infectious complications failed . This sixfold difference is highly significant . Chlorhexidine may reduce microbial complications when used in the immediate perioperative period. J Dent Res, 1997 Dec, 76(12), 1854 - 61 Prevalence and depth of artificial caries-like lesions adjacent to cavities prepared in roots and restored with a glass ionomer or a dentin-bonded composite material; Gilmour AS et al.; One potential advantage of glass-ionomer materials for the treatment of root caries is their ability to release fluoride and so resist cariogenic attack . A commercially available composite material has also been reported to release fluoride which reduced caries lesions in the tooth tissue adjacent to it . The aim of this study was to assess the effectiveness of a conventional glass-ionomer restoration compared with a dentin-bonded, fluoride-releasing, composite restoration when exposed to a microbial artificial caries system . Artificial caries-like lesions produced in relation to the restorations were examined and classified either as outer (surface) lesions or as wall lesions . A split-unit experimental design allowed for within-tooth comparisons of the 2 experimental restorations at different sites on the root surface . These were either totally within the root surface or positioned at the amelo-cemental junction . Outer lesion depths were significantly (p < 0.001) shallower at all sites adjacent to the glass ionomer when compared with the composite restorations . Wall lesions were significantly (p < 0.01) more prevalent adjacent to the composite material . In addition, the cavity margin position significantly (p < 0.05) affected the incidence of wall lesions, particularly in the composite group . In conclusion, glass ionomer was successful in reducing the caries-like lesion production in the adjacent root surface . This resulted from improved marginal integrity and fluoride release from this material when compared with the composite bonding system used. J Dent Res, 1997 Dec, 76(12), 1833 - 9 Profile of cytokine mRNA expression in chronic adult periodontitis; Roberts FA et al.; Chronic inflammation induced by bacteria often leads to host-mediated destruction of tissues adjacent to the sites of microbial insult . The chronic inflammatory process of adult periodontitis results in the destruction of supporting osseous and connective tissues of the teeth . We hypothesized that virulence factors of periodontal pathogens such as lipopolysaccharide stimulate inflammatory cytokine expression by mononuclear cells of the host which contribute to disease development . In this study, to elucidate the role of these cytokines in chronic adult periodontitis, we tested whether the prevalence of mRNA for inflammatory cytokines generally associated with mononuclear phagocytes was higher in diseased than in healthy gingival tissue . Gingival mononuclear cells or whole gingival biopsies from 32 adult periodontitis patients and five healthy individuals used as controls were evaluated for inflammatory cytokine mRNA expression by reverse-transcription polymerase chain-reaction (RT-PCR) procedures . The cytokines assessed included IL-1 alpha, IL-1 beta, IL-1ra, IL-6, IL-8, IL-12, IL-13, TNF-alpha, TGF-beta, and IFN-gamma . The monocyte/macrophage lipopolysaccharide (LPS) receptor CD14 was also assessed . Results showed that TNF-alpha mRNA was present significantly more frequently in diseased than in healthy biopsies, whereas IL-1 alpha, IL-1 beta, and IL-1ra mRNA were found in most (from 80 to 100%) healthy tissues . Message for CD14 was present in both healthy and diseased tissue samples (100%) . This study provides evidence for a major role of TNF-alpha in chronic adult periodontitis . Moreover, our results suggest that the mononuclear cells derived from periodontal tissues have the capacity to respond to components of periodontal pathogens and express both pro- and anti-inflammatory cytokines in these tissues. Plant Physiol, 1997 Nov, 115(3), 949 - 57 Starches from A to C . Chlamydomonas reinhardtii as a model microbial system to investigate the biosynthesis of the plant amylopectin crystal; Buleon A et al.; Wide-angle powder x-ray diffraction analysis was carried out on starch extracted from wild-type and mutant Chlamydomonas reinhardtii cells . Strains containing no defective starch synthases as well as mutants carrying a disrupted granule-bound starch synthase structural gene displayed the A type of diffraction pattern with a high degree of crystallinity . Mutants carrying a defect for the major soluble starch synthase (SSS), SSS II, were characterized by a switch to the B type of diffraction pattern with very low crystallinity . Mutant strains carrying SSS I as the only glucan elongation enzyme regained some of their crystallinity but switched to the C type of diffraction pattern . Differential scanning calorimetry analysis correlated tightly with the x-ray diffraction results . Together with the electron microscopy analyses, these results establish C . reinhardtii as a microbial model system displaying all aspects of cereal starch synthesis and structure . We further show that SSS II is the major enzyme involved in the synthesis of crystalline structures in starch and demonstrate that SSS I alone builds a new type of amylopectin structure. J Anim Sci, 1997 Nov, 75(11), 2986 - 93 Utilization of phytate and nonphytate phosphorus in chicks as affected by source and amount of vitamin D3; Biehl RR et al.; Commercial and laboratory-strain crossbred chicks responded (P < .01) markedly to 1alpha-hydroxycholecalciferol (1alpha-OH D3) during the 2nd and 3rd wk of life . Bone-ash responses exceeded 50% when this compound was added at 20 microg/kg to phosphorus (P)-deficient corn-soybean meal diets containing surfeit levels (25 microg/kg) of cholecalciferol (D3) . Phosphorus excretion was decreased (P < .01) and, thus, retention was increased (P < .01) when 1alpha-OH D3 was supplemented . A P-deficient (.10% P) casein-amino acid purified diet, devoid of D3, was used to determine whether 15 microg/kg of D3 was sufficient to facilitate optimal absorption of the nonphytate P contained in this diet . Bone ash responded to .075% P addition (KH2PO4), and chicks fed diets with .175% nonphytate P exhibited further bone-ash responses to 15 microg/kg of D3 or 10 microg/kg 1alpha-OH D3 . Higher levels of either of these D3 compounds did not produce additional responses . This suggested that 15 to 25 microg/kg of D3 in a P-deficient corn-soybean meal diet (.28% phytate P and .14% nonphytate P) is more than adequate to facilitate optimal absorption of the nonphytate P present in the diet . A P-deficient casein-dextrose diet (.13% nonphytate P and 15 microg/kg D3) was fed in the final chick assay, and chicks fed this diet did not show bone ash responses to 1alpha-OH D3 or to microbial-derived phytase (1,470 units/kg) . Thus, with P-deficient corn-soybean meal diets containing at least 15 microg D3/kg, 1alpha-OH D3 supplementation markedly increased weight gain and bone ash because it increased the utilization of phytate P. Acta Ophthalmol Scand, 1997 Aug, 75(4), 418 - 22 Graft survival and risk factors of penetrating keratoplasty for microbial keratitis; Lomholt JA et al.; PURPOSE: To evaluate graft survival rates and prognostic factors in relation to penetrating keratoplasty for microbial keratitis . METHODS: The records of 95 patients treated with penetrating keratoplasty for microbial keratitis during a twenty-year period were reviewed . Data were analysed by construction of survival curves using the Kaplan-Meier non parametric method . RESULTS: The overall survival of a clear cornea was 72% after one year, 71% after two years and 52% after five years . A significantly lower survival rate (p<0.05) was found in the presence of preoperative local risk factors such as wear of contact lenses and trauma and in inflamed eyes . In contrast, systemic diseases like diabetes, cancer or rheumatoid arthritis did not affect survival and neither did recipient age nor the degree of vascularization of the eye . Male donor buttons showed superior survival compared to female ones (p < 0.05), while females seemed to constitute the best recipients . Recurrence rates of microbial keratitis postoperatively were 11%, 16% and 24% after one, two and five years, respectively . Corresponding graft rejection rates were 9%, 15% and 27% . CONCLUSIONS: Patients suffering from microbial keratitis have a relatively high risk of graft failure . To improve the prognosis care should be taken to minimize local risk factors . Surgery should whenever possible be performed on quiet eyes . The unexpected finding of a better prognosis for male donor buttons might suggest the preferred use of male donors in patients suffering from microbial keratitis, but the observation needs further documentation. Immunotechnology, 1995 Dec, 1(3-4), 211 - 9 Induction of mucosal anti-HIV antibodies by facilitated transfection of airway epithelium with lipospermine/DNA complexes; Mitchell WM et al.; BACKGROUND: Expression of microbial protein sequences in eukaryotic cells transfected by transcriptional/translational permissive cDNA constructs can induce systemic humoral and cellular responses in vivo . Two methods of in vivo transfection have been described to date . One method uses large quantities of naked DNA injected into skeletal muscle . The second method uses relatively small quantities of DNA complexed to gold particles for bollistic penetration of the plasma membrane of keratinocytes . The major disadvantage of the bolistic method is that instrumentation is required which is not generally available . OBJECTIVES: The objectives of this study were to determine whether the use of DNA complexed with a cationic lipopolyamine could reduce the quantity of DNA required to induce systemic humoral responses following muscle transfection and whether similar DNA/lipopolyamine complexes could induce mucosal humoral responses following airway exposure . STUDY DESIGN: Balb/c mice were exposed by nasal aerosol or intramuscular inoculation to a mammalian transcriptional/translational permissive DNA construct containing the entire sequence for the HIV-1 envelope polyprotein . Experimental animals were further segregated by the number of exposures at 3-week intervals and whether the DNA was complexed to dioctadecylamidoglycylspermine (DOGS) at a 5:1 molar charge ratio of DOGS/DNA . RESULTS: DOGS facilitated in vivo transfection of mouse muscle reduced the quantity of DNA required for a systemic humoral response to surface expressed HIV-envelope proteins by one order of magnitude . Exposure of airway mucosa to both 10 micrograms and 1 microgram quantities of DNA complexed to DOGS produced systemic humoral responses to HIV-envelope as well as mucosal antibodies in pulmonary and colonic epithelia . Molecular modeling of DOGS/DNA complexes at the 5:1 charge ratio used in this study indicates that the DNA component is not exposed to aqueous solvents and may be relatively resistant to degradation under common biological environments . CONCLUSION: Facilitated transfer of DNA by DOGS to transcriptional/translational competent cells offers several distinct advantages to the use of DNA alone . Since significantly smaller amounts of DNA are required, the potential for the induction of antibodies against DNA itself lessens the likelihood for the development of a lupus-like syndrome . More importantly, however, is the apparent ability to transfect mucosal cells which results in the development of specific mucosal immune responses . This procedure may allow the development of general methods for the induction of mucosal immunity at the level of entry for mucosal pathogens without the disadvantages inherent in live attenuated vectors. Curr Opin Hematol, 1994 Nov, 1(6), 443 - 51 Leukocyte depletion of cellular blood components; Lane TA; Clinical studies have indicated that the use of leukocyte-reduced cellular blood components produced in the laboratory may prevent febrile reactions and delay or prevent alloimmunization to HLA antigens and refractoriness to platelet transfusion . Additional investigations regarding the effects of the use of leukocyte-reduced blood components were reported during the past year . A recent study in patients with hematologic malignancy that employed the commonly used bedside leukocyte-reduction filters failed to confirm a decrease in the rate of alloimmunization, except in a subgroup of patients with acute myelogenous leukemia . Another major multicenter trial confirmed the effectiveness of leukocyte-reduced blood components in the prevention of cytomegalovirus infection . The effect of allogeneic leukocytes in transfused blood on immune function in patients undergoing colorectal surgery continues to receive attention . Whereas one study failed to demonstrate an adverse effect of standard blood components on disease recurrence or survival, a second study demonstrated a marginally significant decrease in infectious complications in patients who received only leukocyte-reduced blood . Increasingly efficient leukocyte-reduction filters have been developed for cellular blood components, many of which are best suited for laboratory filtration of unstored blood . Laboratory studies indicate that prestorage leukocyte-reduction of cellular blood components does not impair erythrocyte or platelet function and will not increase the incidence of microbial contamination of blood . New methods that employ flow cytometry should enable improved quality control of blood components rendered leukocyte-reduced by the newer, more efficient filters . Finally, a cost-benefit analysis suggests that the appropriate use of leukocyte-reduction filters for acute leukemia patients may reduce the cost of health care to these patients. FEBS Lett, 1997 Oct 20, 416(2), 139 - 42 An estrogen inducible 104 kDa chaperone glycoprotein binds ferric iron containing proteins: a possible role in intracellular iron trafficking; Poola I; We have previously described an estrogen inducible, intracellular, homodimeric membrane glycoprotein (subunit Mr 104 kDa) which is structurally related to 'chaperone' proteins (Poola, I . and Kiang J.G., J . Biol . Chem . 269 (1994) 21762-21769) . In this report we describe a novel finding that the 104 kDa chaperone protein exhibits affinity for iron containing proteins such as transferrins from several species, human lactoferrin and microbial ferric binding protein (FBP) . A single ferric ion in the above proteins appears to be sufficient for binding . It also binds to immobilized ferritin . However, it does not exhibit any affinity for apotransferrins, apolactoferrin, apoferritin and apoFBP . This is the first report of a chaperone protein that exhibits affinity for iron containing proteins. Emerg Infect Dis, 1997 Oct-Dec, 3(4), 551 - 4 Strategies for rapid response to emerging foodborne microbial hazards; Majkowski J; The foodborne outbreak paradigm has shifted . In the past, an outbreak affected a small local population, had a high attack rate, and involved locally prepared food products with limited distribution . Now outbreaks involve larger populations and may be multistate and even international; in many the pathogenic organism has a low infective dose and sometimes is never isolated from the food product . Delay in identifying the causative agent can allow the outbreak to spread, increasing the number of cases . Emergency intervention should be aimed at controlling the outbreak, stopping exposure, and perhaps more importantly, preventing future outbreaks . Using epidemiologic data and investigative techniques may be the answer . Even with clear statistical associations to a contaminated food, one must ensure that the implicated organism could logically and biologically have been responsible for the outbreak. Eur J Immunol, 1997 Oct, 27(10), 2666 - 72 Secreted proteophosphoglycan of Leishmania mexicana amastigotes activates complement by triggering the mannan binding lectin pathway; Peters C et al.; Cutaneous lesions induced by infection of mice with the protozoan parasite, Leishmania mexicana, contain abundant amounts of a high molecular mass proteophosphoglycan (PPG), which is secreted by the amastigote stage residing in phagolysosomes of macrophages and can then be released into the tissue upon rupture of the infected cells . Amastigote PPG forms sausage-shaped but soluble particles and belongs to a novel class of serine-rich proteins that are extensively O-glycosylated by phosphooligosaccharides capped by mannooligosaccharides . The purified molecule is shown here to efficiently activate complement (C) and deplete hemolytic activity of normal serum and may prevent the opsonization of L . mexicana amastigotes . Complement activation is Ca2+ dependent but does not depend on antibodies or the complement component C1 . PPG binds to serum mannan binding protein (MBP), thus activating the MBP-associated serine protease, P100 . Subsequently, the C cascade is triggered through C4 leading to covalent modification probably of carbohydrate hydroxyls of PPG by C3 fragments . Thus, PPG is able to activate C via the mannan binding lectin pathway which is unusual for secreted, soluble products of microbial origin . The proteophosphoglycan-induced complement activation is postulated to contribute to the lesion development and pathology caused by the parasite. Emerg Infect Dis, 1997 Oct-Dec, 3(4), 417 - 23 Infectious disease as an evolutionary paradigm; Lederberg J; The basic principles of genetics and evolution apply equally to human hosts and to emerging infections, in which foodborne outbreaks play an important and growing role . However, we are dealing with a very complicated coevolutionary process in which infectious agent outcomes range from mutual annihilation to mutual integration and resynthesis of a new species . In our race against microbial evolution, new molecular biology tools will help us study the past; education and a global public health perspective will help us deal better with the future. Chirality, 1997, 9(7), 667 - 71 Chirality of the gamma-lactones produced by Sporidiobolus salmonicolor grown in two different media; Dufosse L et al.; Sporidiobolus salmonicolor is an aroma-producing yeast which gives a peach-like smell to the culture media . The enantiomeric ratios of the five gamma-lactones produced by this yeast cultivated in two different media were determined by multidimensional gas chromatography (MDGC) on a fused silica capillary column coupled to a modified beta-cyclodextrin column . These ratios remain constant during growth and are not affected by the composition of the medium . The (R)-enantiomer is highly predominant (99%) for gamma-decalactone and predominant (68-88%) for gamma-octalactone, gamma-nonalactone, and (Z6)-gamma-dodecenolactone . A ratio close to racemic was found for gamma-dodecalactone . A discussion on the metabolic origin of these lactones is based on the analysis of the enantiomeric ratios obtained . With respect to consumers' preference for products considered as "natural," microbial lactone production may represent a valuable alternative to fruit flavors . The enantiomeric lactone ratios produced by Sporidiobolus salmonicolor are compared with those reported from some fruits. AIDS, 1997 Nov, 11(13), 1575 - 81 HIV-1 detection in cervicovaginal secretions during pregnancy; Loussert-Ajaka I et al.; OBJECTIVE: To assess the reproducibility of and factors associated with HIV detection in cervicovaginal secretions (CVS) . DESIGN: Longitudinal study of 43 HIV-1-infected pregnant women in Paris . METHODS: HIV DNA was detected in peripheral blood mononuclear cells (PBMC) by Amplicor and gag nested polymerase chain reaction (PCR) assays . The HIV genotype was determined by heteroduplex mobility assay . Amplicor and gag nested PCR assays were performed on serial CVS samples for HIV DNA detection, and the HIV Monitor test was used for HIV RNA detection in plasma and CVS . RESULTS: A total of 144 CVS samples were collected from the women included in the study . HIV-1 DNA was detected in 36 (25%) of the 144 samples, from 16 (37.2%) of the 43 women . Results of HIV-1 DNA detection were concordant in the first two samples in 27 (84.4%) of the 32 women with at least two CVS samples . The last CVS sample collected in each woman was HIV-1 DNA-positive in 13 (30.2%) of the 43 women . Three factors were found to be independently associated with HIV-1 DNA detection in CVS: HIV-1 subtype B, absence of zidovudine therapy, and microbial cervicovaginal infection . HIV RNA was detected in CVS from 10 (23.3%) out of 43 women and correlated with DNA detection in the same sample and HIV RNA detection in plasma . CONCLUSIONS: DNA and RNA PCR can be used to detect HIV in cells and supernatants of CVS . These techniques may be useful in cohort studies to investigate HIV transmission and to evaluate the efficacy of antiretroviral drugs to reduce HIV excretion. J Biol Chem, 1997 Nov 14, 272(46), 28933 - 8 Functional complementation of the yeast divalent cation transporter family SMF by NRAMP2, a member of the mammalian natural resistance-associated macrophage protein family; Pinner E et al.; The mammalian NRAMP gene family has two members, NRAMP1 and NRAMP2 that encode integral membrane proteins . Nramp1 is expressed exclusively in macrophages where it is found in the phagosomal membrane, and NRAMP1 mutations cause susceptibility to infection by abrogating the capacity of macrophages to control intracellular microbial replication . Nramp2 is highly similar to Nramp1, but is expressed in several tissues and cell types . The Nramp protein family is remarkably conserved throughout evolution, and recent data suggest that the mammalian Nramp2 and the yeast homologues Smf1 and Smf2 transport divalent cations . We tested whether structural similarity between the mammalian Nramp and the yeast Smf proteins results in functional complementation in yeast . Wild-type and mutant variants of the Nramp1 and Nramp2 proteins were expressed in a yeast mutant bearing null alleles at the SMF1 and SMF2 loci, and complementation of the phenotypes of this yeast mutant was investigated . Nramp2, but not Nramp1, was found to complement hypersensitivity to EGTA of the smf1/smf2 mutant under oxidative stress conditions (methyl viologen) . We also observed that the smf1/smf2 double mutant is hypersensitive to growth at alkaline pH (pH 7.9) and that Nramp2 could complement this phenotype as well . Complementation by Nramp2 was specific and required a functional protein as independent mutations in residues highly conserved in all members of the Nramp family abrogated Nramp2 complementation . Since Mn2+ was the only divalent cation capable of completely suppressing both the EGTA and pH phenotypes, our results suggest that Nramp2 can transport Mn2+ in yeast. Life Sci, 1997, 61(18), 1843 - 50 Nicotinamide inhibits inducible nitric oxide synthase enzyme activity in macrophages by allowing nitric oxide to inhibit its own formation; Andrade J et al.; Nitric oxide (NO) production by macrophages is mainly regulated by induction of nitric oxide synthase (iNOS) by cytokines and microbial products . Nicotinamide (NIC) inhibits NO production by activated macrophages in a dose dependent manner . NIC also inhibits NOS enzyme activity in extracts from activated macrophages . The inhibition was noncompetitive with L-arginine (Ki 13.37 +/- 4.40 mM, n=3), uncompetitive versus NADPH (Ki 3.06 +/- 0.17 mM, n=3) and tetrahydrobiopterin . Finally, the inhibition by nicotinamide was fully reversed by scavenging NO with hemoglobin . We suggest that NIC acts by allowing NO to inhibit its own formation. J Dairy Sci, 1997 Oct, 80(10), 2449 - 53 Nutritional risk factors in the etiology of left displaced abomasum in dairy cows: a review; Shaver RD; The transition period occurring 2 wk prepartum through 2 to 4 wk postpartum is the major risk period in the etiology of left displaced abomasum . The prepartum depression of intake and the slow postpartum increase in intake are risk factors causing lower ruminal fill, reduced forage to concentrate ratio, and increased incidence of other postpartum disorders . Uncomplicated ketosis, retained placenta, metritis, and hypocalcemia at parturition are risk factors for left displaced abomasum . Excessive amounts of concentrate during the prepartum period increase the risk of left displaced abomasum, which may occur from the lower ruminal fill caused by greater prepartum intake depression and reduced forage to concentrate ratio, decreased ruminal motility from lower ruminal fill and higher volatile fatty acid concentration, and decreased abomasal motility and emptying from higher concentrations of volatile fatty acids . Effects of volatile fatty acids on motility may be exacerbated by low ruminal absorption of volatile fatty acids during the transition period . Minimal intake of concentrate during the prepartum period may increase the risk of left displaced abomasum through failure to increase the absorptive capacity of the ruminal papillae and failure of the microbial population of the rumen to adapt prior to the intake of high energy postpartum diets . Increased risk of left displaced abomasum in cows that are hypocalcemic at parturition may be due to decreased ruminal and abomasal motility. Nat Med, 1997 Nov, 3(11), 1266 - 70 A novel protein that participates in nonself discrimination of malignant cells by homologous complement; Matsumoto M et al.; The human complement (C) system protects an individual against substances of nonself origin, including xenografts and microbial pathogens . Human cells express C-regulatory proteins, CD46 and CD55, thereby circumventing attack by C3, a major effector of C . Nevertheless, certain malignant cells, particularly those undergoing apoptotic stress, can activate homologous C, overcoming the regulatory actions of CD46 and/or CD55 . The molecular mechanisms whereby malignant cells are tagged by homologous C3 remain largely unknown . We identified a novel gene product that converts human cells into targets for homologous complement . Only malignant cells and cell lines exposed to Fas or X-irradiation stimuli produced this protein, designated M161Ag, which was an unglycosylated 43-kDa protein . Analysis of cloned cDNAs indicated that this molecule was a secretory protein containing five amino acids encoded by TGA codons . Its functions were unique in that once secreted from the tumor cells, it bound back to the surface of these cells and activated homologous complement (C3) via the alternative pathway, allowing for C3 deposition on the membrane . This molecule may offer new insight into innate immunity; surveillance of tumor cells by complement is a common feature in the human immune system. J Nat Prod, 1997 Oct, 60(10), 959 - 66 Isolation, structure elucidation, and biological activity of the steroid oligoglycosides and polyhydroxysteroids from the Antarctic starfish Acodontaster conspicuus; De Marino S et al.; A total of 19 steroids, of which 13 steroidal oligoglycosides (nine new and four known) and six polyhydroxylated steroids (four new and two known), has been isolated from the Antarctic starfish Acodontaster conspicuus . The mixture is dominated by glycosides composed of steroidal aglycons having the hydroxyl groups typically disposed on one side of the tetracyclic nucleus, i.e., 3 beta,4 beta,6 alpha,8,15 beta-, with some having a sulfate at C-6, and differing in the side chains and/or in the disaccharide moieties that are usually attached at C-26, with some at C-28 and C-29 . Those compounds are accompanied by minute amounts of glycosides with a delta 8(14)-double bond in the steroid, which is a structural feature not previously found among polyhydroxysteroids derived from starfish . Small amounts of six related unglycosidated polyhydroxysteroids and three higher-molecular-weight asterosaponins complete the composition of the mixture . The structures of the new compounds were determined by interpretation of their spectral data and by comparison with spectral data of known compounds . Eighteen of these compounds were evaluated for their ability to inhibit growth in Antarctic marine bacteria isolated from either the water column or the surfaces of benthic marine invertebrates . Of these compounds, 50% were active against at least one Antarctic marine bacterium . This suggests that these compounds may play an important role in deterring microbial fouling. PDA J Pharm Sci Technol, 1997 Sep-Oct, 51(5), 203 - 7 Pharmaceutical container/closure integrity . III: Validation of the helium leak rate method for rigid pharmaceutical containers; Kirsch LE et al.; Validation of a helium leak rate method for pharmaceutical container/closure integrity quality assurance required the demonstration that this physical testing method was as good or better than microbial immersion challenge testing in detecting potential integrity failures . One lot of rubber-stoppered, broth-filled glass vials also containing defective vials with known leaks were subjected to both helium leak rate and microbial challenge testing . The defective vials were prepared by affixing glass micropipettes (0.1 to 10 microns) into the vial side walls . The validation lot included a 10% seeded defect rate of which about 50% contained leaks with a predicted probability of failing a microbial challenge (> 10%) . Helium tracer was placed in the test units by charging them for 4 hours under a 40 psi helium pressure . The critical leak rate after charging was determined to be 10(-7) standard cc/second, and test units with measured leak rates greater than this value were considered helium leak rate failures . Microbial immersion challenge was conducted by exposing the test units in a bath inoculated with 10(9-10) viable E . coli and B . diminuta organisms for 24 hours followed by a 13 day (35 degrees C) incubation . Microbial failures were determined visually . The helium and microbial leak test methods were compared statistically using mean failure rates . The mean helium failure rate was 6.9%, whereas the mean microbial failure rate was 2.8% . The difference between helium and microbial failure rates was significantly greater than zero . Thus, helium leak rate testing was demonstrated to be a suitable pharmaceutical container/closure integrity method for microbial quality assurance of rigid containers. PDA J Pharm Sci Technol, 1997 Sep-Oct, 51(5), 195 - 202 Pharmaceutical container/closure integrity . II: The relationship between microbial ingress and helium leak rates in rubber-stoppered glass vials; Kirsch LE et al.; Helium leak rate measurements were quantitatively correlated to the probability of microbial ingress for rubber-stoppered glass vials subjected to immersion challenge . Standard 10-mL tubing glass vials were modified by inserting micropipettes of various sizes (0.1 to 10 microns nominal diameter) into a side wall hole and securing them with epoxy . Butyl rubber closures and aluminum crimps were used to seal the vials . The test units were sealed in a helium-filled glove bag, then the absolute helium leak rates were determined . The test units were disassembled, filled with media, resealed, and autoclaved . The test units were thermally treated to eliminate airlocks within the micropipette lumen and establish a liquid path between microbial challenge media and the test units' contents . Microbial challenge was performed by immersing the test units in a 35 degrees C bath containing magnesium ion and 8 to 10 logs of viable P . diminuta and E . coli for 24 hours . The test units were then incubated at 35 degrees C for an additional 13 days . Microbial ingress was detected by turbidity and plating on blood agar . The elimination of airlocks was confirmed by the presence of magnesium ions in the vial contents by atomic absorption spectrometry . A total of 288 vials were subjected to microbial challenge testing . Those test units whose contents failed to show detectable magnesium ions were eliminated from further analysis . At large leak rates, the probability of microbial ingress approached 100% and at very low leak rates microbial ingress rates were 0% . A dramatic increase in microbial failure occurred in the leak rate region 10(-4.5) to 10(-3) std cc/sec, which roughly corresponded to leak diameters ranging from 0.4 to 2 microns . Below a leak rate of 10(-4.5) std cc/sec the microbial failure rate was < 10% . The critical leak rate in our studies, i.e . the value below which microbial ingress cannot occur because the leak is too small, was observed to be between 10(-5) and 10(-5.8) std cc/sec, which corresponds to an approximate leak diameter of 0.2-0.3 micron. PDA J Pharm Sci Technol, 1997 Sep-Oct, 51(5), 187 - 94 Pharmaceutical container/closure integrity . I: Mass spectrometry-based helium leak rate detection for rubber-stoppered glass vials; Kirsch LE et al.; The development of mass spectrometry-based leak detection for pharmaceutical container integrity was undertaken to provide an alternative to microbial challenge testing . Standard 10-mL vials were modified to contain pinholes (0.5 to 10 microns) by affixing micropipettes with epoxy into 2-mm vial side wall holes . The absolute leak rate was determined using vials that were sealed in a tracer (helium) environment with butyl rubber stoppers and crimps . Alternatively leak rates were determined using