J Infect Dis, 2001 Feb 15, 183(4), 662 - 5 Epub 2001 Jan 11. Altered expression of CD88 and associated impairment of complement 5a-induced neutrophil responses in human immunodeficiency virus type 1-infected patients with and without pulmonary tuberculosis; Meddows-Taylor S et al.; The effect of infection with human immunodeficiency virus type 1 (HIV patient group), infection with Mycobacterium tuberculosis (TB patient group), and coinfection with both of these organisms (HIV/TB patient group) on the expression of CD88 on polymorphonuclear leukocytes (PMNL) was determined by using a receptor-specific monoclonal antibody and flow cytometry . A significant reduction in the fluorescence intensity of CD88 on PMNL was observed in the HIV and HIV/TB groups, compared with both the healthy donor (HD) and TB groups . Furthermore, when degranulation of PMNL was induced by ligation of CD88 by complement 5a (C5a), a large proportion of patients in the HIV and the HIV/TB groups was found to have reciprocal degranulation responses . Patients in the 2 HIV groups also were found to have significantly reduced C5a-induced chemotactic responses and significantly elevated peripheral levels of C5a des Arg, compared with the HD and TB groups . These differences may contribute to the increased susceptibility of HIV-1-infected individuals to secondary microbial infections. Eur J Immunol, 2000 Dec, 30(12), 3591 - 7 Bacterial CpG-DNA activates dendritic cells in vivo: T helper cell-independent cytotoxic T cell responses to soluble proteins; Sparwasser T et al.; Receptors for conserved molecular patterns associated with microbial pathogens induce synthesis of co-stimulatory molecules and cytokines in immature dendritic cells (DC), as do antigen-reactive CD4 T helper cells via CD40 signaling . Once activated, antigen-presenting DC may activate CD8 T cell responses in a T helper cell-independent fashion . Using immunostimulatory CpG-oligonucleotides (ODN) mimicking bacterial CpG-DNA, we tested whether CpG-DNA bypasses the need for T helper cells in CTL responses towards proteins by directly activating antigen-presenting DC to transit into professional APC . We describe that immature DC in situ constitutively process soluble proteins and generate CD8 T cell determinants yet CD8 T cell responses remain abortive . Induction of primary antigen-specific CD8 cytotoxic T lymphocyte (CTL)-mediated responses becomes initiated in wild-type as well as T helper cell-deficient mice, provided soluble protein and CpG-ODN are draining into the same lymph node . Specifically we show that CpG-ODN trigger antigen-presenting immature DC within the draining lymph node to acutely up-regulate co-stimulatory molecules and produce IL-12 . These results provide new insights for generating in vivo efficient CTL responses to soluble proteins which may influence vaccination strategies. J Appl Microbiol, 2001 Feb, 90(2), 294 - 300 Epithelium-associated bacteria in the gastrointestinal tract of Arctic charr (Salvelinus alpinus L.) . An electron microscopical study; Ringo E et al.; AIMS: The primary aim was to use transmission electron microscopy (TEM) and scanning electron microscopy (SEM) to define the location of epithelium-associated bacteria in the digestive tract of the salmonid fish, Arctic charr (Salvelinus alpinus) . METHODS AND RESULTS: TEM and SEM examination of the gastrointestinal tract demonstrated substantial numbers of ovoid and rod-shaped bacterial cells associated with the microvillous brush borders of enterocytes . Bacteria were found at the tips of microvilli as well as between adjacent microvilli . Endocytosis of bacteria by epithelial cells was observed in two regions (pyloric caeca and midgut) . CONCLUSION: Electron microscope examination of the gut is an important tool for evaluating the microbial ecology of the fish digestive tract ecosystem . SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the current study clearly demonstrate that the intestine is involved in bacterial endocytosis. Eur J Biochem, 2001 Jan, 268(2), 374 - 83 Mannosylerythritol lipid induces characteristics of neuronal differentiation in PC12 cells through an ERK-related signal cascade; Wakamatsu Y et al.; Rat pheochromocytoma PC12 cells undergo neuronal differentiation in response to nerve growth factor (NGF) . The differentiation involves protein kinase cascades that include the kinases MEK and ERK, as well as activation of the transcription factors c-Jun and c-Fos . We show here, that exposure of PC12 cells to mannosylerythritol lipid (MEL), a yeast extracellular glycolipid, enhances the activity of acetylcholinesterase and interrupts the cell cycle at the G1 phase, with resulting outgrowth of neurites and partial cellular differentiation . Treatment with MEL stimulates the phosphorylation of ERK to a similar extent as treatment with NGF, although, the appearance of phosphorylated ERK is somewhat delayed . Both the MEL-induced outgrowth of neurites and the increase in the activity of acetylcholinesterase are prevented by PD98059, a specific inhibitor of MEK . Northern blotting analysis of c-jun transcripts and analysis of transcription in PC12 cells of a c-jun/CAT reporter construct demonstrated a significant increase in the rate of transcription of the c-jun gene upon treatment with MEL . The sequence elements required for the MEL-mediated activation of transcription of the c-jun gene are located between nucleotides -126 and -79 in the 5' flanking region . Our results suggest that MEL induces characteristics of neuronal differentiation in PC12 cells, with transactivation of the c-jun gene, via an ERK-related signal cascade that is partially overlapping the pathways activated in response to NGF . These results might provide the groundwork for the use of microbial extracellular glycolipids as novel reagents for the treatment of cancer cells. Clin Oral Implants Res, 2000, 11 Suppl 1, 33 - 58 The scientific basis for and clinical experiences with Straumann implants including the ITI Dental Implant System: a consensus report; Cochran DL; Successful endosseous implant therapy requires integration of the implant with bone, soft connective tissue and epithelium . This report from a consensus conference on Straumann dental implants including the ITI Dental Implant System documents the interaction of these nonsubmerged one-piece implants with the oral tissues and reviews clinical studies supporting the high success achievable with these implants in patients . Light and electron microscopy reveal that epithelial structures similar to teeth are found around the implants . A connective tissue zone exists between the apical extension of the junctional epithelium and the alveolar bone . This connective tissue comprises a dense circular avascular zone of connective tissue fibers surrounded by a loose vascular connective tissue . The histologic dimensions of the epithelium and connective tissue comprising the biologic width are similar to the same tissues around teeth . The nonsubmerged one-piece design of the Straumann implants, which have been used for over 20 years, has set a standard in implant dentistry, with other implants now being manufactured and placed using similar techniques . Straumann implants have an endosseous portion that is either coated with a well-characterized and well-documented titanium plasma-sprayed surface or is sandblasted and acid attacked . Both surfaces have been shown to have advantages for osseous integration compared to machined and other smoother implant surfaces . These advantages include greater amounts of bone-to-implant contact, more rapid integration with bone tissue, and higher removal torque values . The lack of component connection at or below the alveolar crest provides additional benefits . Component connection at the alveolar crest, as seen with submerged implants, results in microbial contamination, crestal bone loss and a more apical epithelial location . Numerous human clinical trials document the successful use of Straumann implants in a variety of indications and areas of the mouth . These include prospective long-term trials using strict criteria of success and life table analyses . Taken together, the clinical studies reveal that Straumann implants can be used predictably in partially edentulous and completely edentulous maxilla and mandibles with high success rates . Furthermore, the animal and microscopic studies reviewed provide a scientific basis for the integration of Straumann implants with bone, connective tissue and epithelium. J Microbiol Methods, 2001 Feb 1, 44(1), 79 - 88 Ecological and physiological analyses of Pseudomonad species within a phenol remediation system; Whiteley AS et al.; A diverse collection of 700 bacteria obtained from an operational phenolic remediating industrial treatment plant was made to select potential strains as microbial biosensors . Pseudomonads were the most abundant group, of which 48 selected from the liquor or suspended solids were assessed for their physiological response to phenolic pollutant loading and niche specialisation . By FAME-MIS identification the Pseudomonads were clustered into six major species groups . Those isolates able to utilise phenol as a sole carbon source predominantly belonged to a non-clonal Pseudomonas pseudoalcaligenes cluster determined by REP-PCR genotyping . Rapid microtitre based respiration assays were developed to contrast activity in response to increasing concentrations of phenol . A considerable range in response for both phenol degrader and non-degrader strains was observed . This natural phenotypic and physiological heterogeneity could facilitate the selection of isolates for the development of a suite of ecologically relevant, custom designed sensors with predictable toxicity susceptibilities to monitor process efficacy. Environ Exp Bot, 2001 Feb, 45(1), 1 - 9 UV-C irradiation reduces microbial populations and deterioration in Cucurbita pepo fruit tissue; Erkan M et al.; Tissue slices of zucchini squash (Cucurbita pepo L., cv . Tigress) fruit were exposed to ultraviolet-C (UV-C) radiation from germicidal lamps for 1, 10 or 20 min; however, only 10 and 20 min UV-C exposure significantly reduced microbial activity and deterioration during subsequent storage at 5 or 10 degrees C . UV-C treated slices had higher respiration rates than controls; however, the ethylene production of the slices was not affected by UV-C treatments . Slight UV-C irradiation damage (reddish brown discoloration) was detected on the surface of 10 and 20 min-treated slices after 12 days of storage at 10 degrees C . Slices stored at 5 degrees C did not show UV-C damage . Chilling injury was not observed until after 20 days of storage at 5 degrees C . The symptoms of chilling injury appeared as dried sunken brown spots on the surface of cortex tissue . UV-C treatments did not affect the degree of chilling injury during storage at 5 degrees C . UV-C treatment also had no consistent effect on sugar or malic acid concentrations . The most pronounced effect of UV-C irradiation was to retard microbial growth thereby providing a basis for the frequently observed delay in senescence and subsequent deterioration in fruit tissues. Mol Immunol, 2000 Jun, 37(9), 545 - 52 Targeting early events in T cell activation to construct improved vaccines; Buchan GS et al.; Live, attenuated vaccines currently offer the best protection against virulent pathogens . Recent advances in Immunology and Molecular Biology provide an opportunity to design vaccines that will be more effective and safer than existing ones . Immunologists are rapidly developing the capacity to identify and construct the minimal immunogenic units from pathogens . The molecular signals required to fully activate antigen presenting cells (APCs) and responder T cells are becoming apparent . Improved vaccine delivery systems are being designed which will mimic the actions of pathogens in vivo . These vaccines will incorporate protective epitopes fused to immunoregulatory cytokines in chimeric proteins . They will be encapsulated in formulations which allow for the slow release of these chimeric proteins thereby inducing the memory T cells required for long-lived immunity . These vaccine formulations will target receptors present on the most active APCs . Here we discuss how these advances will allow us to rationally construct "virtual pathogens" which will provide improved protection against new and old microbial foes. Hum Immunol, 2000 Dec, 61(12), 1270 - 3 Association of IL-10 genotype with sudden infant death syndrome; Summers AM et al.; Sudden infant death syndrome (SIDS) is a major cause of infant death of unknown etiology . We propose that SIDS results from a genetically determined imbalance in the production of inflammatory and anti-inflammatory cytokines in response to the infant's microbial flora . We were especially interested to know the relationship between SIDS and genetically determined higher or lower production of IL-10, an anti-inflammatory cytokine . Biallelic polymorphisms in the promoter region of the IL-10 gene associated with higher or lower production of IL-10 were determined in a SIDS and in a control group using a sequence-specific oligonucleotide approach . One particular allele of the IL-10 gene, the IL-10-592*A allele, was significantly associated with SIDS . Indeed, 70% of the SIDS babies carried the IL-10-592*A allele (p = 0.007 compared with control) . In addition, there was a significant reduction in the frequency of homozygosity for the allele IL-10-592*C (p = 0.001 compared with control) . Carrying the A allele (either A/A or A/C) had an odds ratio of 3.3 (95% confidence interval 1.4-8.0) . In the same patients there was no association with other IL-10 gene polymorphisms nor with other cytokine (TNF-alpha, TGF-beta 1) genotypes, emphasizing the particular relationship between SIDS and the IL-10-592*A allele. Am J Ophthalmol, 2001 Jan, 131(1), 131 - 3 Increased incidence of corneal perforation after topical fluoroquinolone treatment for microbial keratitis; Mallari PL et al.; PURPOSE: To compare the incidence of corneal perforation in eyes treated with topical fluoroquinolone or fortified antibiotics for microbial keratitis . METHODS: A retrospective medical record review of patients hospitalized for bacterial keratitis from January 1991 through November 1999 . RESULTS: Two hundred seventy-seven cases of bacterial keratitis were identified . The incidence of corneal perforations was significantly higher in fluoroquinolone-treated eyes (18 out of 142, 12.7%) compared with eyes treated with fortified antibiotics (1 out of 135, 0.7%) (chi-square = 13.6, degrees of freedom (df) = 2, P <.001) . Within the fluoroquinolone-treated group, all corneal perforations occurred in those eyes treated with ofloxacin (18 out of 125, 14.4%) . Corneal perforations that occurred in the ofloxacin-treated eyes were not associated with any particular microbial isolate . CONCLUSION: Our data suggest an increased risk of corneal perforation after fluoroquinolone treatment for bacterial keratitis compared with treatment with fortified antibiotics . Further studies are warranted to verify this association and establish possible mechanisms by which topical fluoroquinolones may alter corneal collagen or keratocyte function. Ecotoxicol Environ Saf, 2001 Jan, 48(1), 85 - 98 Comparative effects of a genetically engineered insect virus and a growth-regulating insecticide on microbial communities in aquatic microcosms; Kreutzweiser D et al.; The effects of a genetically engineered insect baculovirus on indigenous aquatic microbial communities were determined in closed, recirculating aquatic microcosms, and compared with the effects of a natural strain of the virus and of a growth-regulating insecticide, Dimilin . The recombinant virus was a nuclear polyhedrosis virus (NPV) of the spruce budworm (Choristoneura fumiferana (Cf)) with a lacZ marker gene inserted into the egt region of the CfNPV . The natural virus was Ireland strain CfNPV . Microbial measurement endpoints included decomposition activity (mass loss of organic material), respiration on two different substrates (O2 consumption), heterotrophic bacterial abundance (plate counts), and microbial community metabolic profiles (carbon source utilization patterns in Biolog GN microplates) . Viral DNA of both the natural strain and the recombinant viruses, detected by polymerase chain reaction techniques, settled out of the microcosm water and accumulated on bottom substrates within 3 days of the microcosm inoculations . The viral DNA persisted in bottom substrates for the duration of the 21-day experimental period, although there was some evidence that the recombinant virus was less stable than the natural strain in particulate organic matter . No significant changes in microbial decomposition or respiration activity, bacterial abundance, or average metabolic responses were detected by a time trend analysis in microcosms inoculated with either the lacZ recombinant virus or the natural Ireland strain CfNPV . Significant effects on microbial decomposition and respiration activity were detected in microcosms treated with the growth-regulating insecticide at, and above, the expected environmental concentrations . Despite significant effects on microbial community functional attributes in Dimilin-treated microcosms, there were no detectable changes in community structure in terms of metabolic profiles or bacterial abundance. Occup Environ Med, 2001 Feb, 58(2), 113 - 8 Short term exposure to airborne microbial agents during farm work: exposure-response relations with eye and respiratory symptoms; Eduard W et al.; OBJECTIVES: Exposure to high levels of non-infectious microbial agents is recognised as a cause of respiratory disease in working populations, but except for endotoxins, little is known about exposure-response relations . As these effects do not depend on viability, exposure to non-viable microbial agents is important . Various methods not based on microbial cultures were explored to study the complex microbial exposure of farmers and associations with acute symptoms during work . METHODS: Airborne exposure was measured when farmers carried out specific tasks . Fungal spores, bacteria, endotoxins, beta(1-->3)-glucans, fungal antigens specific for Penicillium and Aspergillus species, and mites were measured by methods not based on microbial cultures . Also silica, inorganic and organic dust, ammonia, hydrogen sulphide, and nitrogen dioxide were measured . Respiratory, and nose and eye symptoms experienced during measurements were recorded by a short questionnaire . Both univariate and multivariate statistical analyses were applied to assess the relations between exposure and acute symptoms . RESULTS: 106 Farmers and their spouses participated in this study . Prevalences of work related symptoms were: wheezing 3%; chest tightness 7%; cough 14%; eye symptoms 18%; and nose symptoms 22% . Prevalence ratios for nose and eye symptoms were 4-8 after exposure to 20-500x10(3) fungal spores/m(3) and higher, and a prevalence ratio for cough was 4 after exposure to 500-17,000x10(3) fungal spores/m(3) . Nose symptoms were also associated with exposure to silica with prevalence ratios of 4-6 after exposure to 0.015-0.075 mg /m(3) and higher . CONCLUSIONS: Farmers had a high occurrence of symptoms of the nose and eyes as well as cough during work . These symptoms were associated in a dose dependent manner with exposure to fungal spores . Nose symptoms were also associated with exposure to silica. J Immunol, 2001 Feb 15, 166(4), 2855 - 62 V beta 6+ and V beta 4+ T cells exert cooperative activity in clearance of secondary infection with Histoplasma capsulatum; Gomez FJ et al.; We previously studied the lung Vbeta TCR repertoire of C57BL/6 mice during primary infection with the pathogen Histoplasma capsulatum . We observed a consistent oligoclonal expansion of Vbeta4(+) T cells during the peak of infection and early stages of resolution . The Vbeta4(+) family played a role in protective immunity against the fungus . Depletion of this subpopulation of T cells hindered optimal clearance of infection from tissues . In this report we analyze the flux of the Vbeta TCR repertoire in the lungs of C57BL/6 mice with reinfection histoplasmosis . We observed a significant increase in Vbeta6(+) T cells on days 7, 10, and 14, the peak and early resolution phases of infection . This skewing was preceded by an increased number of memory T cells within Vbeta6(+) cells . The VDJ sequences of Vbeta6 chains were oligoclonal during the early stages of the infection, suggesting that the expansion was driven by a small number of Ags . More than 96% of the expanded Vbeta6(+) cells were CD4(+) . Depletion of Vbeta6(+) T cells but not Vbeta4(+) T cells induced a modest but significant delay in fungal clearance . Simultaneous depletion of Vbeta4(+) and Vbeta6(+) T cells induced a more pronounced impairment of host resistance . These studies illustrate the dynamic interactions between Vbeta families in the response to microbial challenge. J Immunol, 2001 Feb 15, 166(4), 2444 - 50 Microbial lipopeptides stimulate dendritic cell maturation via Toll-like receptor 2; Hertz CJ et al.; The ability of dendritic cells (DC) to initiate immune responses in naive T cells is dependent upon a maturation process that allows the cells to develop their potent Ag-presenting capacity . Although immature DC can be derived in vitro by treatment of peripheral blood monocytes with GM-CSF and IL-4, additional signals such as those provided by TNF-alpha, CD40 ligand, or LPS are required for complete maturation and maximum APC function . Because we recently found that microbial lipoproteins can activate monocytes and DC through Toll-like receptor (TLR) 2, we also investigated whether lipoproteins can drive DC maturation . Immature DC were cultured with or without lipoproteins and were monitored for expression of cell surface markers indicative of maturation . Stimulation with lipopeptides increased expression of CD83, MHC class II, CD80, CD86, CD54, and CD58, and decreased CD32 expression and endocytic activity; these lipopeptide-matured DC also displayed enhanced T cell stimulatory capacity in MLR, as measured by T cell proliferation and IFN-gamma secretion . The lipid moiety of the lipopeptide was found to be essential for induction of maturation . Preincubation of maturing DC with an anti-TLR2 blocking Ab before addition of lipopeptide blocked the phenotypic and functional changes associated with DC maturation . These results demonstrate that lipopeptides can stimulate DC maturation via TLR2, providing a mechanism by which products of bacteria can participate in the initiation of an immune response. J Immunol, 2001 Feb 15, 166(4), 2291 - 5 Distinct CpG DNA and polyinosinic-polycytidylic acid double-stranded RNA, respectively, stimulate CD11c- type 2 dendritic cell precursors and CD11c+ dendritic cells to produce type I IFN; Kadowaki N et al.; Two classes of nucleic acids, bacterial DNA containing unmethylated CpG motifs and dsRNA in viruses, induce the production of type I IFN that contributes to the immunostimulatory effects of these microbial molecules . Thus, it is important to determine which cells produce type I IFN in response to CpG DNA and dsRNA . CD4(+)CD11c(-) type 2 dendritic cell precursors (pre-DC2) were identified as the main producers of type I IFN in human blood in response to viruses . Here we asked whether pre-DC2 also produce type I IFN in response to CpG DNA and dsRNA . Oligodeoxynucleotides containing particular palindromic CpG motifs induced pre-DC2, but not CD11c(+) blood DC or monocytes, to produce IFN-alpha . In contrast, a synthetic dsRNA, polyinosinic polycytidylic-acid, induced CD11c(+) DC, but not pre-DC2 or monocytes, to produce IFN-alphabeta . These data indicate that CpG DNA and polyinosinic-polycytidylic acid stimulate different types of cells to produce type I IFN and that it is important to select oligodeoxynucleotides containing particular CpG motifs to induce pre-DC2 to produce type I IFN, which may play a key role in the strong adjuvant effects of CpG DNA. Microbiology, 2001 Feb, 147(Pt 2), 373 - 82 Characterization of microbial consortia in a terephthalate-degrading anaerobic granular sludge system; Wu JH et al.; The microbial composition and spatial distribution in a terephthalate-degrading anaerobic granular sludge system were characterized using molecular techniques . 16S rDNA clone library and sequence analysis revealed that 78.5% of 106 bacterial clones belonged to the delta subclass of the class Proteobacteria; the remaining clones were assigned to the green non-sulfur bacteria (7.5%), Synergistes (0.9%) and unidentified divisions (13.1%) . Most of the bacterial clones in the delta-Proteobacteria formed a novel group containing no known bacterial isolates . For the domain Archaea, 81.7% and 18.3% of 72 archaeal clones were affiliated with Methanosaeta and Methanospirillum, respectively . Spatial localization of microbial populations inside granules was determined by transmission electron microscopy and fluorescent in situ hybridization with oligonucleotide probes targeting the novel delta-proteobacterial group, the acetoclastic Methanosaeta, and the hydrogenotrophic Methanospirillum and members of Methanobacteriaceae . The novel group included at least two different populations with identical rod-shape morphology, which made up more than 87% of the total bacterial cells, and were closely associated with methanogenic populations to form a nonlayered granular structure . This novel group was presumed to be the primary bacterial population involved in the terephthalate degradation in the methanogenic granular consortium. J Antimicrob Chemother, 2001 Feb, 47(2), 199 - 202 Antimycobacterial action of B4128, a novel tetramethylpiperidyl-substituted phenazine; Matlola NM et al.; The effects of the novel tetramethylpiperidyl (TMP)-substituted phenazine, B4128 (0.6-2.5 mg/L), on growth, phospholipase A2(PLA2) activity, cation (K+, Ca2+) fluxes and energy metabolism (ATP) of Mycobacterium aurum A(+) and Mycobacterium tuberculosis (H37Ra) have been investigated in vitro . Growth, PLA2 and cation fluxes were measured radiometrically, while microbial ATP was assayed by means of a luciferin/luciferase chemiluminescence method . Exposure of the mycobacteria to B4128 resulted in immediate, dose-related enhancement of microbial PLA2 activity and inhibition of K+ influx, which preceded effects on microbial ATP, influx of Ca2+ and growth . These results demonstrate that B4128 possesses membrane-directed antimycobacterial properties that are similar to those of clofazimine. Appl Environ Microbiol, 2001 Feb, 67(2), 702 - 12 Impact of dilution on microbial community structure and functional potential: comparison of numerical simulations and batch culture experiments; Franklin RB et al.; A series of microcosm experiments was performed using serial dilutions of a sewage microbial community to inoculate a set of batch cultures in sterile sewage . After inoculation, the dilution-defined communities were allowed to regrow for several days and a number of community attributes were measured in the regrown assemblages . Based upon a set of numerical simulations, community structure was expected to differ along the dilution gradient; the greatest differences in structure were anticipated between the undiluted-low-dilution communities and the communities regrown from the very dilute (more than 10(-4)) inocula . Furthermore, some differences were expected among the lower-dilution treatments (e.g., between undiluted and 10(-1)) depending upon the evenness of the original community . In general, each of the procedures used to examine the experimental community structures separated the communities into at least two, often three, distinct groups . The groupings were consistent with the simulated dilution of a mixture of organisms with a very uneven distribution . Significant differences in community structure were detected with genetic (amplified fragment length polymorphism and terminal restriction fragment length polymorphism), physiological (community level physiological profiling), and culture-based (colony morphology on R2A agar) measurements . Along with differences in community structure, differences in community size (acridine orange direct counting), composition (ratio of sewage medium counts to R2A counts, monitoring of each colony morphology across the treatments), and metabolic redundancy (i.e., generalist versus specialist) were also observed, suggesting that the differences in structure and diversity of communities maintained in the same environment can be manifested as differences in community organization and function. Appl Environ Microbiol, 2001 Feb, 67(2), 696 - 701 Identification, purification, and characterization of iminodiacetate oxidase from the EDTA-degrading bacterium BNC1; Liu Y et al.; Microbial degradation of synthetic chelating agents, such as EDTA and nitrilotriacetate (NTA), may help immobilizing radionuclides and heavy metals in the environment . The EDTA- and NTA-degrading bacterium BNC1 uses EDTA monooxygenase to oxidize NTA to iminodiacetate (IDA) and EDTA to ethylenediaminediacetate (EDDA) . IDA- and EDDA-degrading enzymes have not been purified and characterized to date . In this report, an IDA oxidase was purified to apparent homogeneity from strain BNC1 by using a combination of eight purification steps . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single protein band of 40 kDa, and by using size exclusion chromatography, we estimated the native enzyme to be a homodimer . Flavin adenine dinucleotide was determined as its prosthetic group . The purified enzyme oxidized IDA to glycine and glyoxylate with the consumption of O2 . The temperature and pH optima for IDA oxidation were 35 degrees C and 8, respectively . The apparent Km for IDA was 4.0 mM with a kcat of 5.3 s(-1) . When the N-terminal amino acid sequence was determined, it matched exactly with that encoded by a previously sequenced hypothetical oxidase gene of BNC1 . The gene was expressed in Escherichia coli, and the gene product as a C-terminal fusion with a His tag was purified by a one-step nickel affinity chromatography . The purified fusion protein had essentially the same enzymatic activity and properties as the native IDA oxidase . IDA oxidase also oxidized EDDA to ethylenediamine and glyoxylate . Thus, IDA oxidase is likely the second enzyme in both NTA and EDTA degradation pathways in strain BNC1. Bioresour Technol, 2001 Jan, 76(2), 125 - 9 Organic matter components and aggregate stability after the application of different amendments to a horticultural soil; Albiach R et al.; The effects of usual or recommended rates of application of five organic amendments (24 t/ha yr of MSW compost, sewage sludge, and ovine manure, 2.4 t/ha yr of commercial vermicompost, and 100 l/ha yr of a commercial humic acids solution) on the soil contents of organic matter, total humified substances, humic acids, carbohydrates and microbial gums, and the structural stability of aggregates were investigated . Four and five years after the beginning of the experiment, significant increments in most of the parameters studied were found after the application of organic residues, whereas the two commercial amendments did not produce any significant change, suggesting that rates recommended by the producers and imposed by their high prices are too low to be useful . MSW compost yielded the highest increases, even if the amount of organic matter applied as ovine manure was very similar . Organic matter and carbohydrates appeared to be the parameters most closely related to soil aggregate stability. PDA J Pharm Sci Technol, 1999 Sep-Oct, 53(5), 235 - 9 Pharmaceutical container/closure integrity . V: An evaluation of the WILCO "LFC" method for leak testing pharmaceutical glass-stoppered vials; Kirsch LE et al.; The sensitivity, reliability, reproducibility and ease of use of the WILCO LFC package integrity test method was evaluated by preparing and testing a series of rubber-stoppered glass vials which were modified by affixing a glass micropipette through the vial side wall . The test units contained water, 50% aqueous ethanol, 20% lithium chloride or 20% aqueous glycerol . Leakage measurement obtained by LFC testing were compared to helium leak rate measurements . The LFC methods detected all leak > 0.0014 standard cubic centimeters per second (sccs), which represents a sensitivity about fourteen-fold greater than standard vacuum decay methods . The minimum detectable leak corresponded to a nominal micropipette internal diameter of between 1 and 2 microns . The effective detection range corresponded to a leak size associated with a 40 to 100% probability of microbial ingress based on a previously reported logistical regression model between helium leak rate and microbial immersion . The sensitivity did not vary with solvent or testing duration in range of 5 to 10 seconds . The coefficient of variation was about 3% . The LFC operation was rapid and without apparent mechanical or electronic problems over the two day testing period used in these studies. Aquat Toxicol, 2000 Mar 1, 48(2-3), 135 - 150 Responses of aquatic communities to 25-6 alcohol ethoxylate in model stream ecosystems; Belanger SE et al.; A model stream ecosystem evaluation of the non-commercial alcohol ethoxylate 25-6 alcohol ethoxylate (AE) was performed in 1994 . Algal, heterotrophic microbial, protozoan, and invertebrate communities were assessed over an 8-week exposure period that followed an 8-week colonization period . Streams were exposed to nominal concentrations of 0, 12, 37, 111, 333 and 1000 microg AE/l . Confirmed concentrations (8-week means) were 7 (at the detection limit), 13, 36, 76, 259, and 760 microg AE/l as measured at the head of streams on a weekly basis . Microbial communities were initially enhanced by AE exposure (first 2 weeks of exposure), but by the conclusion of the study autotrophic and heterotrophic microbial communities were similar across treatments . In contrast, invertebrate populations and communities responded strongly to AE exposure with adverse effects indicated at 259-760 microg/l by 4 weeks and at 36-760 microg/l by 8 weeks . Key affected groups were the sensitive mayfly, stonefly, and caddisfly fauna . Species richness and abundance of selected populations such as Stenonema (mayfly), Chimarra (caddisfly), and Corbicula (Asiatic clam) were affected . These data indicated a model ecosystem no-observed effect concentration of 13 microg/l for 25-6 AE . In contrast to other published AE model ecosystem studies, 25-6 appears more toxic . Structure-activity-relationships at the model ecosystem level still demonstrate good relationships across a range of surfactants with calculated K(ow)s of 3-6 . Importantly, the collective information on fate and effects measured in laboratory and field systems confirms low risk to the environment. J Agric Food Chem, 2000 Dec, 48(12), 6230 - 3 Substrate specificities of microbial transglutaminase for primary amines; Ohtsuka T et al.; Transglutaminase (epsilon-glutaminyl-peptide:amine gamma-glutaminyl-transferase, EC 2.3.2.13) (TGase) is an enzyme that catalyzes acyl transfer reactions between primary amines and Gln residues in proteins and peptides . The substrate specificity of TGase for primary amines was investigated to incorporate various functional groups into proteins and peptides . In this study, microbial transglutaminase and guinea pig liver transglutaminase were used . For the primary amines to be incorporated into benzyloxycarbonyl-L-Gln-Gly (Z-Gln-Gly), they were required to have more than four carbon chains without side chains between the functional groups . These results suggest that with appropriate primary amines as spacers, various functional groups, carboxyl groups, phosphate groups, saccharides, and so on, can be incorporated into proteins by using TGase. Ann Rheum Dis, 2001 Feb, 60(2), 166 - 9 Effect of interleukin 2 on killer cell inhibitory receptors in patients with rheumatoid arthritis; Kogure T et al.; OBJECTIVE: The genes for killer cell inhibitory receptors (KIRs) have been cloned and their functions and responses to other molecules, including cytokines, have been partially clarified . However, the expression of KIRs has not been analysed in patients with autoimmune diseases, such as rheumatoid arthritis (RA), who are highly susceptible to microbial infection . Therefore, KIR expression on lymphocytes in patients with RA, and the regulation of KIR expression by interleukin 2 (IL2) in RA was investigated . METHODS: CD158a/b expression on peripheral blood mononuclear cells (PBMC) obtained from 25 patients with RA and 14 healthy subjects was analysed by flow cytometry . Additionally, PBMC from the two groups of subjects were cultured in RPMI 1640 medium with or without IL2 for 48 hours, and then their CD158a/b expression was analysed . RESULTS: The rate of CD158a expression on the CD8+ cells was lower in patients with RA than in healthy subjects, though there was no significant difference in the CD158a/b expression on the CD16+ cells between the two groups . The upregulation of CD16+CD158a/b+ cells in response to IL2 was significantly reduced in patients with RA compared with healthy subjects . CONCLUSION: The reduced induction of KIR expression in response to IL2 may provide insight into the reason for the high susceptibility of patients with RA to microbial infection. Int J Syst Evol Microbiol, 2000 Nov, 50 Pt 6, 2043 - 55 New genus-specific primers for the PCR identification of members of the genus Saccharomonospora and evaluation of the microbial diversity of wild-type isolates of Saccharomonospora detected from soil DNAs; Salazar O et al.; Members of the genus Saccharomonospora are isolated infrequently, probably due to the low occurrence of these actinomycetes in the environment . Although members of this genus can easily be identified by micromorphological criteria, the extensive chemotaxonomic characterization of each new isolate is a time-consuming task which cannot always be undertaken when handling large numbers of strains as is the case in natural products screening programmes . In this work, the design of one set of genus-specific oligonucleotides which allows rapid detection of members of the genus Saccharomonospora by means of PCR-specific amplification is presented . The genus specificity of these primers was validated on a wide range of collection and wild-type strains, and subsequently applied to evaluate the presence of representatives of this taxon directly from soil DNAs . Partial 16S rDNA sequencing of representative wild-type strains was used to validate their genus assignment . Further analyses of PCR fingerprinting patterns and 16S-23S internal transcribed spacer sequences were used to determine the diversity of wild-type isolates obtained from soils . This study shows the usefulness of the application of these primers for the direct identification of members of this genus and in assessment of its occurrence within natural microbial habitats. Dermatol Clin, 2001 Jan, 19(1), 79 - 86, viii Dapsone and sulfapyridine; Paniker U et al.; Dapsone and sulfapyridine are structurally related compounds with anti-microbial and anti-inflammatory effects . Dapsone remains the most important drug for leprosy and is useful in the prophylaxis of Pneumocystis pneumonia in patients with HIV disease . The medical treatment of choice for dermatitis herpetiformis is dapsone; and sulfapyridine also can be used for those patients who are intolerant of dapsone . Other neutrophilic disorders also may respond to these drugs . Toxic side effects of both dapsone and sulfapyridine are mediated through the hydroxylamine metabolite . These include hemolysis, methemoglobinemia, and agranulocytosis . Careful monitoring for possible adverse reactions includes frequently performing complete blood counts and regular blood chemistry profile determinations. Curr Opin Immunol, 2001 Feb, 13(1), 109 - 13 CD1 and lipid antigens: intracellular pathways for antigen presentation; Jayawardena-Wolf J et al.; Recently, different members of the CD1 family of MHC-like molecules have been shown to sample different intracellular compartments to present lipid and glycolipid antigens to T cells . Emerging models suggest that CD1 may have evolved to monitor the integrity of membrane lipids and/or to present microbial lipid antigens to both alpha beta and gamma delta T cells. Curr Opin Immunol, 2001 Feb, 13(1), 55 - 62 Innate immunity in plants; Cohn J et al.; Studies of receptors and signal-transduction components that play a role in plant disease resistance have revealed remarkable similarities with innate immunity pathways in insects and mammals . In plants, specific receptors encoded by disease-resistance genes interact with products of microbial effector genes to activate defence responses . Resistance proteins have been found to have motifs in common with components of immune response pathways in mammals and invertebrates, and to rely on similar downstream signalling components . In the future, the sharing of ideas among plant and animal biologists is likely to broaden our understanding of defence responses in diverse organisms. Curr Opin Immunol, 2001 Feb, 13(1), 19 - 25 Presentation of self and microbial lipids by CD1 molecules; Matsuda JL et al.; CD1 molecules present both self lipids and microbial lipids . Recent studies have elucidated novel antigenic structures that can be presented by CD1 for T cell stimulation, as well as new pathways for lipid-antigen presentation . Additionally, the development of lipid-CD1 tetramers now permits the tracking of CD1-reactive T cells during immune responses . Despite this, the roles of CD1-reactive T cells in both host defense and immune regulation remain to be unequivocally defined. Clin Exp Pharmacol Physiol, 2001 Jan-Feb, 28(1-2), 1 - 8 Sunlight, immunosuppression and skin cancer: role of histamine and mast cells; Hart PH et al.; 1 . The development into tumours of skin cells transformed by ultraviolet (UV) B radiation of wavelengths 290-320 nm is enhanced by the ability of UVB to suppress an immune response that would otherwise destroy them . Ultraviolet B-induced immunomodulation may be by multiple mechanisms, but generally manifests in an antigen-presenting cell defect and an altered cytokine environment in the draining lymph nodes . 2 . Immune responses to microbial or self-antigens may be dysfunctional by similar mechanisms following UVB exposure . 3 . Earliest-acting intermediates in the initiation of UVB-induced immunosuppression are the UVB absorbers (photoreceptors) of the skin, notably DNA resulting in immunoregulatory cytokine production, and trans-urocanic acid (UCA), which, upon isomerization to its cis isomer, signals downstream immunosuppressive events . 4 . In mice, dermal mast cells are critical to UVB-induced systemic immunomodulation . In mice, there is a functional link as well as a linear relationship between the prevalence of histamine-staining dermal mast cells and the log of the dose of UVB required for 50% immunosuppression . Studies with histamine receptor antagonists support histamine as the main' product of mast cells involved . Histamine acts in large part via a prostanoid-dependent pathway . 5 . Approximately 50% of humans and greater than 90% of patients with non-melanoma skin cancer are UVB susceptible for suppression of a contact hypersensitivity response . Neither cytokine polymorphisms nor UVB-induced levels of cis-UCA in irradiated skin have been linked to UVB susceptibility . Patients with basal cell carcinomas (BCC) have an increased dermal mast cell prevalence in non-sun-exposed buttock skin . We propose that mast cells function in humans, as in mice, by initiating immunosuppression and, thereby, allowing a permissive environment for BCC development. Bratisl Lek Listy, 2000, 101(8), 445 - 9 {Proteinases and antiproteinases: biomedical correlations}; Kotyza J; Both degradative and limited proteolysis are involved in physiological processes . Once activated, proteinases are controlled by endogenous inhibitors (antiproteinases) . A variety of genetic disorders and exogenous microbial proteinases disturb the balance between proteinases and cognate inhibitors . alpha 1-Antitrypsin deficiency is a model disorder resulting in an insufficient control of leukocyte elastase with a subsequent alveolar tissue damage . It appears that the manifold molar excess of inhibitor required to compensate the deficiency, is the consequence of both the local relative inaccessibility of the serpin, and the mode of proteinase-serpin interaction, in accord with the "branched pathway" mechanism . Slow-binding kinetics and the leak of native proteinase from the complex is illustrated by the peptidolytic action of porcine elastase in the presence of human alpha 1-antitrypsin and by gelatin zymography, respectively . (Tab . 2, Fig . 5, Ref . 44.) Ann Agric Environ Med, 2000, 7(2), 111 - 8 Effects of exposure to flax dust in Polish farmers: work-related symptoms and immunologic response to microbial antigens associated with dust; Skorska C et al.; Medical examinations were performed in a group of 51 Polish farmers heavily exposed to flax dust during harvesting and scutching (threshing) and in a group of 50 healthy urban dwellers not exposed to organic dusts (controls) . The examinations included: interview concerning the occurrence of respiratory disorders and work-related symptoms, physical examination, X-ray examination of chest, lung function tests, oxymetric examinations, determination of the concentration of cytokines (IL-1alpha IL-6, TNFalpha) in blood serum and allergological tests with microbial antigens associated with organic dust, comprising: skin prick test with 4 antigens, agar-gel precipitation test with 12 antigens and test for specific inhibition of leukocyte migration with 4 antigens . As many as 32 farmers (62.7%) reported the occurrence of work-related symptoms during harvesting, transporting and scutching of flax . The most common complaint was general weakness reported by 15 farmers (29.4%), followed by headache reported by 14 (27.5%), blocking of the nose - by 11 (21.6%), dry cough, shivering, and eyes itching - each by 10 (19.8%), chest tightness and hoarseness - each by 9 (17.6%) . No control subjects reported these work-related symptoms . The mean spirometric values in the examined group of farmers were within a normal range and did not show a significant post-shift decline . In contrast, a significant post-shift decline of oxymetric values was found among flax farmers . The farmers showed a frequency of the positive early skin reactions to environmental allergens in the range of 0-19.6%, a frequency of positive precipitin reactions in a range of 0-56.9%, and frequency of positive reactions of leukocyte migration inhibition in a range of 7.8-21.6% . The members of the control group responded to the majority of allergens with a significantly lower frequency of positive results compared to the farmers . Elevated concentrations of IL-1alpha and IL-6, but not TNFalpha, were found in blood sera of flax farmers . In conclusion, farmers engaged in harvesting and scutching of flax represent a group of elevated professional risk because of high incidence of work-related symptoms and high frequency of allergic reactions to bacteria and fungi associated with organic dust. Inorg Chem, 2000 Dec 11, 39(25), 5591 - 602 Kinetics and mechanism of iron(III) dissociation from the dihydroxamate siderophores alcaligin and rhodotorulic acid; Boukhalfa H et al.; The kinetics and mechanism of siderophore ligand dissociation from their fully chelated Fe(III) complexes is described for the highly preorganized cyclic tetradentate alcaligin and random linear tetradentate rhodotorulic acid in aqueous solution at 25 degrees C (Fe2L3 + 6H+ reversible 2 Fe3+ aq + 3 H2L) . At siderophore:Fe(III) ratios where Fe(III) is hexacoordinated, kinetic data for the H(+)-driven ligand dissociation from the Fe2L3 species is consistent with a singly ligand bridged structure for both the alcaligin and rhodotorulic acid complexes . Proton-driven ligand dissociation is found to proceed via parallel reaction paths for rhodotorulic acid, in contrast with the single path previously observed for the linear trihydroxamate siderophore ferrioxamine B . Parallel paths are also available for ligand dissociation from Fe2(alcaligin)3, although the efficiency of one path is greatly diminished and dissociation of the bis coordinated complex Fe(alcaligin)(OH2)2+ is extremely slow (k = 10(-5) M-1 s-1) due to the high degree of preorganization in the alcaligin siderophore . Mechanistic interpretations were further confirmed by investigating the kinetics of ligand dissociation from the ternary complexes Fe(alcaligin)(L) in aqueous acid where L = N-methylacetohydroxamic acid and glycine hydroxamic acid . The existence of multiple ligand dissociation paths is discussed in the context of siderophore mediated microbial iron transport. J Allergy Clin Immunol, 2001 Jan, 107(1), 160 - 4 Antibody responses to bacteriophage phi X-174 in human subjects exposed to the antarctic winter-over model of spaceflight; Shearer WT et al.; BACKGROUND: It has been proposed that exposure to long-term spaceflight conditions (stress, isolation, sleep disruption, containment, microbial contamination, and solar radiation) or to ground-based models of spaceflight will alter human immune responses, but specific antibody responses have not been fully evaluated . OBJECTIVE: We sought to determine whether exposure to the 8-month Antarctic winter-over model of spaceflight would alter human antibody responses . METHODS: During the 1999 Australian National Antarctic Research Expeditions, 11 adult study subjects at Casey, Antarctica, and 7 control subjects at Macquarie Island, sub-Antarctica, received primary and secondary immunizations with the T cell-dependent neoantigen bacteriophage phi X-174 . Periodic plasma samples were analyzed for specific antibody function . RESULTS: All of the subjects from Casey, Antarctica, cleared bacteriophage phi X-174 normally by 1 week after primary immunization, and all had normal primary and secondary antibody responses, including immunologic memory amplification and switch from IgM to IgG antibody production . One subject showed a high normal pattern, and one subject had a low normal pattern . The control subjects from Macquarie Island also had normal immune responses to bacteriophage phi X-174 . CONCLUSIONS: These data do not support the hypothesis that de novo specific antibody responses of subjects become defective during the conditions of the Antarctic winter-over . Because the Antarctic winter-over model of spaceflight lacks the important factors of microgravity and solar radiation, caution must be used in interpreting these data to anticipate normal antibody responses in long-term spaceflight. J Cell Biol, 2001 Jan 8, 152(1), 165 - 80 The phagosome proteome: insight into phagosome functions; Garin J et al.; Phagosomes are key organelles for the innate ability of macrophages to participate in tissue remodeling, clear apoptotic cells, and restrict the spread of intracellular pathogens . To understand the functions of phagosomes, we initiated the systematic identification of their proteins . Using a proteomic approach, we identified >140 proteins associated with latex bead-containing phagosomes . Among these were hydrolases, proton pump ATPase subunits, and proteins of the fusion machinery, validating our approach . A series of unexpected proteins not previously described along the endocytic/phagocytic pathways were also identified, including the apoptotic proteins galectin3, Alix, and TRAIL, the anti-apoptotic protein 14-3-3, the lipid raft-enriched flotillin-1, the anti-microbial molecule lactadherin, and the small GTPase rab14 . In addition, 24 spots from which the peptide masses could not be matched to entries in any database potentially represent new phagosomal proteins . The elaboration of a two-dimensional gel database of >160 identified spots allowed us to analyze how phagosome composition is modulated during phagolysosome biogenesis . Remarkably, during this process, hydrolases are not delivered in bulk to phagosomes, but are instead acquired sequentially . The systematic characterization of phagosome proteins provided new insights into phagosome functions and the protein or groups of proteins involved in and regulating these functions. Diagn Microbiol Infect Dis, 2000 Dec, 38(4), 213 - 21 Discriminative power of fatty acid methyl ester (FAME) analysis using the microbial identification system (MIS) for Candida (Torulopsis) glabrata and Saccharomyces cerevisiae; Peltroche-Llacsahuanga H et al.; Candida (Torulopsis) glabrata is frequently isolated in cases of fungal infection and commonly shows acquired or innate fluconazole resistance . Saccharomyces cerevisiae, an emerging opportunistic yeast pathogen, causes serious systemic infections in immunocompromised, and vaginitis and superficial infections in immunocompetent patients . For both species reliable identification in the routine laboratory is mandatory, but species identification of strains, e.g . trehalose-negative C . glabrata, may be difficult . Therefore, gas-liquid chromatography (GLC) of whole cell fatty acid methyl ester (FAME) profiles, that is independent of assimilation profiles of strains and suitable for reliable and rapid identification of clinically important yeasts, was applied . However, frequent misidentification of C . glabrata as S . cerevisiae has been reported when using the Yeast Clinical Database of MIS . Accuracy of MIS identification may be strongly influenced by the amounts of cell mass analyzed . Therefore, the present study compared the MIS results of these two yeasts achieved with different cell masses . Primarily we optimized, especially with respect to cost-effectiveness, the recommended streaking technique yielding a maximal recovery of 90-130 mg of cell mass from one plate, enabling testing of poor growing strains of C . glabrata . For all C . glabrata strains tested (n = 10) the highest identification scores (SI {Similarity Index} range 0.525-0.963, median 0.832) were achieved with 30 to 45 mg of cell mass . Only 5 of 10 S . cerevisiae strains revealed good library comparisons (SI > or = 0.5) when using 30 mg of cell mass, whereas with 45 mg all strains but two revealed this SI-level . For S . cerevisiae a higher amount of cell mass processed (up to 90 mg) was correlated with better identification scores (SI range using 90 mg: 0.464-0.870, median, 0.737) . Several passages prior to FAME analysis of C . glabrata strains on recommended media revealed narrowing of SI ranges, but differences in SI values were not statistically significant. Crit Rev Immunol, 2000, 20(5), 393 - 405 Critical roles of Toll-like receptors in host defense; Kaisho T et al.; Drosophila Toll is involved not only in dorsoventral patterning of embryos but also in immune responses to microbial infection . Several Toll-like receptors (TLRs) have also been identified in mammals . They are expressed on macrophages or dendritic cells (DCs), which are essential sentinels for innate immunity . These cells utilize TLRs as a recognition and signal transducing receptor for microbial molecular components . The most characterized mammalian TLR, TLR4, is a receptor for lipopolysaccharides (LPS) . TLR2 recognizes other components, such as peptideglycans (PGN) . This recognition, called pattern recognition, is essential for the establishment of innate immunity, which is the basis for host defense . In this article, we review recent findings about this expanding receptor family. Mil Med, 2000 Nov, 165(11 Suppl), 1 - 21 Aeromedical evacuation of biological warfare casualties: a treatise on infectious diseases on aircraft; Withers MR et al.; A basic understanding of the transmission and isolation of infections would be essential to the safe and effective aeromedical evacuation (AE) of biological warfare (BW) casualties . First, the airframe as microbial environment is considered, and relevant preventive and disinfecting measures are discussed . A survey of past infectious disease transmission on civilian aircraft (including tuberculosis, influenza, measles, smallpox, and viral hemorrhagic fevers) is presented, and the communicability and stability of likely BW agents is described . A brief history of U.S . military aeromedical evacuation (as it relates to contagious diseases and U.S . Air Force BW doctrine) is also outlined . Special containment procedures (especially as used by the U.S . Army Aeromedical Isolation Team) are described . Finally, international legal and regulatory aspects of the AE of BW casualties are considered, and some unanswered questions and suggestions for future research are offered . It is concluded that, given adequate foresight, expertise, and resources, the AE of even contagious BW casualties could be safely and effectively accomplished. Biochemistry, 2001 Jan 9, 40(1), 84 - 92 Characterization of BphF, a Rieske-type ferredoxin with a low reduction potential; Couture MM et al.; BphF is a small, soluble, Rieske-type ferredoxin involved in the microbial degradation of biphenyl . The rapid, anaerobic purification of a heterologously expressed, his-tagged BphF yielded 15 mg of highly homogeneous recombinant protein, rcBphF, per liter of cell culture . The reduction potential of rcBphF, determined using a highly oriented pyrolytic graphite (HOPG) electrode, was -157+/- 2 mV vs the standard hydrogen electrode (SHE) (20 mM MOPS, 80 mM KCl, and 1 mM dithiothreitol, pH 7.0, 22 degrees C) . The electron paramagnetic resonance spectrum of the reduced rcBphF is typical of a Rieske cluster while the close similarity of the circular dichroic (CD) spectra of rcBphF and BedB, a homologous protein from the benzene dioxygenase system, indicates that the environment of the cluster is highly conserved in these two proteins . The reduction potential and CD spectra of rcBphF were relatively independent of pH between 5 and 10, indicating that the pK(a)s of the cluster's histidinyl ligands are not within this range . Gel filtration studies demonstrated that rcBphF readily oligomerizes in solution . Crystals of rcBphF were obtained using sodium formate or poly(ethylene glycol) (PEG) as the major precipitant . Analysis of the intermolecular contacts in the crystal revealed a head-to-tail interaction that occludes the cluster, but is very unlikely to be found in solution . Oligomerization of rcBphF in solution was reversed by the addition of dithiothreitol and is unrelated to the noncovalent crystallographic interactions . Moreover, the oligomerization state of rcBphF did not influence the latter's reduction potential . These results indicate that the 450 mV spread in reduction potential of Rieske clusters of dioxygenase-associated ferredoxins and mitochondrial bc(1) complexes is not due to significant differences in their solvent exposure. Clin Diagn Lab Immunol, 2001 Jan, 8(1), 21 - 30 Defective neutrophil degranulation induced by interleukin-8 and complement 5a and down-regulation of associated receptors in children vertically infected with human immunodeficiency virus type 1; Meddows-Taylor S et al.; The polymorphonuclear neutrophils (PMNs) of patients infected with human immunodeficiency virus type 1 (HIV-1) show impaired microbicidal responses . The present study assessed the functional integrity of PMN degranulation responses and the expression of specific receptors that mediate these responses in a group of children vertically infected with HIV-1 . PMN degranulation in response to interleukin-8 (IL-8) and complement 5a (C5a) was measured in a group of HIV-1-infected children with mild and severe clinical disease and in an uninfected control group . In addition, the expression of CXCR1, CXCR2, and CD88 on whole-blood PMNs was quantified by flow cytometry . Although CXCR1 expression was found to be largely unaltered in the HIV-1-infected children relative to that in the control children, the intensity of CXCR2 expression was significantly reduced in those with severe disease . Furthermore, there was a significant reduction in the percentage of cells expressing CD88 and in the intensity of CD88 fluorescence in the HIV-1-infected children compared to that in control children, with CD88 fluorescence intensity more significantly reduced in the presence of severe disease . PMNs from a large proportion of the HIV-1-infected children either showed reciprocal degranulation responses or were unresponsive to IL-8 and C5a, whereas the PMNs from the uninfected children showed positive responses . Inefficient agonist-induced degranulation may contribute to the increased susceptibility of HIV-1-infected children to secondary microbial infections . Furthermore, reduced expression of CXCR2 and CD88 may be suggestive of defects in other functions of PMNs from HIV-1-infected children. Extremophiles, 2000 Dec, 4(6), 343 - 9 Diverse genes of cellulase homologues of glycosyl hydrolase family 45 from the symbiotic protists in the hindgut of the termite Reticulitermes speratus; Ohtoko K et al.; Diverse genes encoding cellulase homologues belonging to glycosyl hydrolase family 45 were identified from the symbiotic protists in the hindgut of the termite Reticulitermes speratus through the use of consensus PCR and the screening of a cDNA library . Fifteen full-length cDNA clones were isolated and sequenced, which encoded polypeptides consisting of 218-221 amino acid residues showing up to 63% identity to known family 45 cellulases . The cellulase sequences of the termite symbiotic protists were phylogenetically monophyletic, showing more than 75% amino acid identity with each other . These enzymes consist of a single catalytic domain, lacking the ancillary domains found in most microbial cellulases . By whole-cell in situ hybridization using oligonucleotide probes specific for regions conserved in some of the sequences, the origin of the genes was identified as symbiotic hypermastigote protists . The presence of diverse cellulase homologues suggests that symbiotic protists of termites may be rich reservoirs of novel cellulase sequences. Int J Food Microbiol, 2000 Dec 5, 62(1-2), 37 - 45 Delimination of brewing yeast strains using different molecular techniques; Tornai-Lehoczki J et al.; In general, the genetic characteristics, the phenotype and the microbial purity of the production brewing yeast strains are among the most important factors in maintaining a consistently good quality of products . Analysis of restriction fragment length polymorphism (RFLP) patterns of 18S rRNA-coding DNA was investigated to group ale and lager strains . All production brewing yeast strains showed the same RFLP pattern as the type strain and synonym type strains of S . cerevisiae, and were quite different from the type and synonym type strains of S . pastorianus . Based on these data, all production brewing yeast strains investigated in this study appeared to belong to S . cerevisiae . Electrophoretic karyotyping and random amplified polymorphic DNA (RAPD) analysis appeared to be suitable methods for distinguishing not only the type and synonym type strain of S . cerevisiae and S . pastorianus, but also the ale and the lager strains. Hum Immunol, 2000 Nov, 61(11), 1150 - 6 HLA-G and HLA-E polymorphisms in an indigenous African population . The ZVITAMBO Study Group; Matte C et al.; Studies of HLA-G and HLA-E polymorphisms in different populations from many industrialized countries have uniformly reported little sequence variation within these genes . To date, the polymorphism of these genes has not been characterized in populations from developing countries where more sequence variation would be anticipated due to greater exposure to microbial pathogens . In order to address this issue, we have recently investigated the polymorphism of these MHC genes in an indigenous African population (Shona ethnic group) . HLA-G and HLA-E alleles were determined by PCR-restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system (ARMS) analysis in 108 unrelated women recruited from maternity clinics and hospitals in Harare (Zimbabwe) . The genotyping analysis identified six different HLA-G alleles and three HLA-E alleles in this population . Of interest, the null-allele HLA-G*0105N was found with 11.1% of frequency, which is higher than in other populations tested so far . We observed the presence of 15 distinct HLA-G genotypes and 6 HLA-E genotypes . These data indicate that this African population contains low levels of allelic polymorphism similar to ethnic groups from industrialized countries . This is the first report describing HLA-G and HLA-E polymorphisms in an indigenous African population. J Exp Biol, 2001 Jan, 204(Pt 2), 349 - 58 Quantifying nutrient production by the microbial symbionts in an aphid; Douglas AE et al.; The symbiotic bacteria Buchnera sp . provide aphids with essential amino acids, nutrients in short supply in the aphid diet of plant phloem sap . The contribution of Buchnera-derived amino acids to net protein growth of the aphid Aphis fabae was quantified from the protein growth of aphids reared on chemically defined diets lacking individual amino acids . The amino acid production rates varied among the nine essential amino acids over the range 8-156 pmol microg(-1)protein day(-1) (for tryptophan and leucine, respectively), equivalent to 0.02-0.33 fmol Buchnera(-1)day(-1) . In a complementary metabolic analysis, the aphids incorporated radioactivity from dietary {(14)C}glutamic acid into the essential amino acids isoleucine, lysine and threonine . Incorporation into isoleucine was significantly elevated by the omission of dietary isoleucine, indicating that dietary supply may affect the biosynthetic rates of certain amino acids by Buchnera . Aphids experimentally deprived of Buchnera did not synthesize essential amino acids from dietary glutamic acid . The mortality of aposymbionts was high over 7 days on the phenylalanine-free diet, and their assimilation of dietary leucine was depressed on the complete diet, suggesting that both the absence of bacteria-derived amino acids and the low rates of assimilation of certain dietary amino acids may contribute to the poor growth of these insects. FEMS Microbiol Ecol, 2001 Jan, 34(3), 255 - 266 Microbial community dynamics in Mediterranean nutrient-enriched seawater mesocosms: changes in abundances, activity and composition; Lebaron P et al.; Quantitative and qualitative changes in bacterial communities from the Mediterranean Sea were compared in duplicate batch mesocosms with or without addition of inorganic nutrients . Methods including traditional microbial ecology techniques, molecular biology and flow cytometry were combined to determine abundances, production, cell size, activity, culturability and taxonomic diversity of bacterial cells . Addition of nutrients and confinement resulted in an increase of bacterial densities which were rapidly controlled by protozoan grazing . Changes in bacterial activity and morphology were observed during the growth phase of bacteria and under grazing pressure . The proportion of medium-size and culturable cells increased during the growth phase . These cells were preferentially consumed by grazers resulting in a strong limitation of bacterial production . As a consequence of the grazing pressure, large cells were produced and contributed to the remaining bacterial productivity after grazing . Grazing had an effect on the taxonomic composition of bacterial communities by preferentially eliminating gamma-Proteobacteria, alpha-Proteobacteria were preserved . It seems that some species from the genera Ruegeria and Cytophaga may have developed defence strategies to escape predation. Plant J, 2000 Dec, 24(6), 837 - 47 Identification of genes encoding receptor-like protein kinases as possible targets of pathogen- and salicylic acid-induced WRKY DNA-binding proteins in Arabidopsis; Du L et al.; To understand how plant host genes are regulated during the activation of plant defence responses, we are studying a group of pathogen- and salicylic acid (SA)-induced DNA-binding proteins containing the novel WRKY domain . To identify downstream target genes of these WRKY proteins, we have searched the Arabidopsis genome and identified four closely linked genes on chromosome IV that contain an unusually large number of the W-box sequences {(T)TGAC(C/T)} recognized by WRKY proteins within a few hundred base pairs upstream of their coding regions . All four genes encode proteins characteristic of receptor-like protein kinases (RLK), each consisting of an N-terminal signal sequence, an extracellular receptor domain, a single transmembrane domain and a C-terminal cytoplasmic serine/threonine protein kinase domain . All four RLK genes were induced by treatment with SA or infection by a bacterial pathogen . Studies with one of the RLK genes (RLK4) indicated that a cluster of W-box elements in its promoter region were recognized by both purified WRKY proteins and SA-induced W-box binding activities from SA-treated Arabidopsis plants . Further analysis using the RLK4 gene promoter fused to a reporter gene in transgenic Arabidopsis indicated that the consensus WRKY protein-binding sites in the RLK4 gene promoter were important for the inducible expression of the reporter gene . These results indicate that pathogen- and SA-induced W-box binding proteins regulate not only genes encoding defence proteins with direct or indirect anti-microbial activities, but also genes encoding proteins with regulatory functions. J Clin Invest, 2001 Jan, 107(1), 121 - 30 EBNA1-specific CD4+ T cells in healthy carriers of Epstein-Barr virus are primarily Th1 in function; Bickham K et al.; The Epstein-Barr virus (EBV) nuclear antigen-1 (EBNA1) maintains the viral episome in all host cells infected with EBV . Recently, EBNA1 was found to be the main EBV latency antigen for CD4+ T cells and could be recognized in cultures from all donors tested . We now identify a polarized Th1 phenotype and obtain evidence for its presence in vivo . When T cells were stimulated with dendritic cells infected with vaccinia vectors expressing EBNA1, 18 of 19 donors secreted IFN-gamma, whereas only two of 19 secreted IL-4 . Magnetic selection was then used to isolate cells from fresh blood based on EBNA1-induced cytokine production . Specific IFN-gamma CD4+ cell lines were established from six of six donors and IL-4 lines from three of six . Only the Th1 lines specifically lysed targets expressing three different sources of EBNA1 protein . When the IgG isotype of EBNA1 plasma Ab's was tested, most specific Ab's were IgG1 and of a high titer, confirming a Th1 response to EBNA1 in vivo . Ab's to other microbial antigens generally were not skewed toward IgG1 . Given emerging evidence that Th1 CD4+ T cells have several critical roles in host defense to viral infection and tumors, we propose that EBNA1-specific CD4+ Th1 cells contribute to resistance to EBV and EBV-associated malignancies. Appl Environ Microbiol, 2001 Jan, 67(1), 100 - 9 Microbial activity in aquatic environments measured by dimethyl sulfoxide reduction and intercomparison with commonly used methods; Griebler C et al.; A new method to determine microbial (bacterial and fungal) activity in various freshwater habitats is described . Based on microbial reduction of dimethyl sulfoxide (DMSO) to dimethyl sulfide (DMS), our DMSO reduction method allows measurement of the respiratory activity in interstitial water, as well as in the water column . DMSO is added to water samples at a concentration (0.75% {vol/vol} or 106 mM) high enough to compete with other naturally occurring electron acceptors, as determined with oxygen and nitrate, without stimulating or inhibiting microbial activity . Addition of NaN(3), KCN, and formaldehyde, as well as autoclaving, inhibited the production of DMS, which proves that the reduction of DMSO is a biotic process . DMSO reduction is readily detectable via the formation of DMS even at low microbial activities . All water samples showed significant DMSO reduction over several hours . Microbially reduced DMSO is recovered in the form of DMS from water samples by a purge and trap system and is quantified by gas chromatography and detection with a flame photometric detector . The DMSO reduction method was compared with other methods commonly used for assessment of microbial activity . DMSO reduction activity correlated well with bacterial production in predator-free batch cultures . Cell-production-specific DMSO reduction rates did not differ significantly in batch cultures with different nutrient regimes but were different in different growth phases . Overall, a cell-production-specific DMSO reduction rate of 1.26 x 10(-17) +/- 0 . 12 x 10(-17) mol of DMS per produced cell (mean +/- standard error; R(2) = 0.78) was calculated . We suggest that the relationship of DMSO reduction rates to thymidine and leucine incorporation is linear (the R(2) values ranged from 0.783 to 0.944), whereas there is an exponential relationship between DMSO reduction rates and glucose uptake, as well as incorporation (the R(2) values ranged from 0.821 to 0.931) . Based on our results, we conclude that the DMSO reduction method is a nonradioactive alternative to other methods commonly used to assess microbial activity. J Antibiot (Tokyo), 2000 Oct, 53(10), 1117 - 22 Fistupyrone, a novel inhibitor of the infection of Chinese cabbage by Alternaria brassicicola, from Streptomyces sp . TP-A0569; Igarashi Y et al.; A new microbial metabolite, designated fistupyrone, was isolated from the culture broth of a plant-associated Streptomyces sp . TP-A0569 . Fistupyrone inhibited the in vivo infection of the seedlings of Chinese cabbage by Alternaria brassicicola TP-F0423, the cause of Alternaria leaf spot, without any in vitro fungicidal activity. Crit Rev Oral Biol Med, 2000, 11(4), 383 - 408 Epithelial structural proteins of the skin and oral cavity: function in health and disease; Presland RB et al.; Epithelial tissues function to protect the organism from physical, chemical, and microbial damage and are essential for survival . To perform this role, epithelial keratinocytes undergo a well-defined differentiation program that results in the expression of structural proteins which maintain the integrity of epithelial tissues and function as a protective barrier . This review focuses on structural proteins of the epidermis and oral mucosa . Keratin proteins comprise the predominant cytoskeletal component of these epithelia . Keratin filaments are attached to the plasma membrane via desmosomes, and together these structural components form a three-dimensional array within the cytoplasm of epithelial cells and tissues . Desmosomes contain two types of transmembrane proteins, the desmogleins and desmocollins, that are members of the cadherin family . The desmosomal cadherins are linked to the keratin cytoskeleton via several cytoplasmic plaque proteins, including desmoplakin and plakoglobin (gamma-catenin) . Epidermal and oral keratinocytes express additional differentiation markers, including filaggrin and trichohyalin, that associate with the keratin cytoskeleton during terminal differentiation, and proteins such as loricrin, small proline-rich proteins, and involucrin, that are cross-linked into the cornified envelope by transglutaminase enzymes . The importance of these cellular structures is highlighted by the large numbers of genetic and acquired (autoimmune) human disorders that involve mutations in, or autoantibodies to, keratins and desmosomal and cornified envelope proteins . While much progress has been made in the identification of the structural proteins and enzymes involved in epithelial differentiation, regulation of this process is less clear . Both calcium and retinoids influence epithelial differentiation by altering the transcription of target genes and by regulating activity of enzymes critical in epithelial differentiation, such as transglutaminases, proteinases, and protein kinases . These studies have furthered our understanding of how epithelial tissue and cell integrity is maintained and provide a basis for the future treatment of skin and oral disorders by gene therapy and other novel therapeutics. J Hepatol, 2000 Dec, 33(6), 926 - 32 Covalent adducts of proteins with acetaldehyde in the liver as a result of acetaldehyde administration in drinking water; Jokelainen K et al.; BACKGROUND/AIMS: Acetaldehyde, the first metabolic product of ethanol, has been suggested to be responsible for several adverse effects of ethanol through its ability to form covalent adducts with proteins and cellular constituents . It has recently been suggested that acetaldehyde derived from microbial ethanol oxidation in the gut could also contribute to the effects of ethanol in the liver . The present work aimed to examine whether modification of proteins by acetaldehyde occurs in rat liver as a result of acetaldehyde administration in drinking water . METHODS: Rats were fed with either 0.7% acetaldehyde (n=10) or water (n=10) for 11 weeks . At the end of the feeding period, liver specimens were processed for immunohistochemistry for protein adducts with acetaldehyde and for hepatic cell type-specific protein markers . RESULTS: Mild fatty change was found in the liver of the acetaldehyde-treated animals but not in the control animals . Immunohistochemical stainings for acetaldehyde adducts revealed intensive positive staining for acetaldehyde adducts in eight (80%) of the animals fed with acetaldehyde . The adducts were predominantly perivenular, although positive staining also occurred along the sinusoids and in the periportal area . Double immunofluorescence staining experiments revealed that hepatocytes were the primary targets of acetaldehyde adduct deposition, although stellate cells and Kupffer cells also showed weak positive reactions . CONCLUSIONS: The present data indicate that acetaldehyde-protein adducts are formed in the liver of animals following acetaldehyde administration in drinking water, which may contribute to the hepatotoxicity of extrahepatic acetaldehyde . These findings should be implicated in studies on the extrahepatic pathways of ethanol oxidation. Pediatr Clin North Am, 2000 Dec, 47(6), 1339 - 54 Complement deficiencies; Frank MM; The complement proteins play an important role in innate immunity, promoting inflammation and microbial killing . They play a role in the adaptive immune response, as well . Inherited total deficiencies of complement proteins are extremely rare . Table 1 lists more than 40 proteins that comprise the elements of the complement system . Deficiency of the proteins that promote lysis and opsonization is so rare that two papers are able to list all the observed cases to 1991 . The exception is mannan-binding lectin, in which deficiency may be commoner . Diseases of regulatory proteins, such as occurs in hereditary angioedema or paroxysmal nocturnal hemoglobinuria, are commoner but still are quite rare . As we learn more about complement proteins and their mechanism of action, we will understand more clearly how these proteins function . Polymorphisms of the proteins exists . Learning how these polymorphisms contribute to the development of disease will be the focus of complement studies in the next decade. Phytochemistry, 2000 Nov, 55(6), 481 - 504 Advances in flavonoid research since 1992; Harborne JB et al.; Some of the recent advances in flavonoid research are reviewed . The role of anthocyanins and flavones in providing stable blue flower colours in the angiosperms is outlined . The contribution of leaf flavonoids to UV-B protection in plants is critically discussed . Advances in understanding the part played by flavonoids in warding off microbial infection and protecting plants from herbivory are described . The biological properties of flavonoids are considered in an evaluation of the medicinal and nutritional values of these compounds. Int Rev Immunol, 2000, 19(6), 557 - 72 Pathogenic thyroglobulin peptides as model antigens: insights on the induction and maintenance of autoimmune thyroiditis; Carayanniotis G et al.; In recent years, the discovery of pathogenic thyroglobulin (Tg) peptides has given a new impetus to study, at the basic level, mechanisms of induction and immunoregulation of autoimmune thyroiditis . The genetic control of the immune response against defined Tg epitopes and the diversity of the T-cell receptor repertoire recruited for their recognition were among the first issues examined . Some of these epitopes contained hormonogenic sites, i.e . thyroxine residues, and thus offered an excellent opportunity to study how post-translational modifications such as iodination, can influence induction of thyroiditogenic cells . The delineation of pathogenic Tg determinants also enabled the search for "molecular mimics" i.e . peptides of microbial origin that may be involved in the pathogenesis of the disease . In addition, factors promoting the generation of pathogenic epitopes during Tg processing in antigen presenting cells could now be systematically investigated . This review summarizes recent findings in these areas. Pharmazie, 2000 Nov, 55(11), 791 - 7 Vermiculine: a diolide with immunoregulatory activity; Proksa B et al.; Vermiculine (1), is a macrodiolide which displays a selective effect on certain reactions of cellular immunity . This paper summarizes data on its biological activity, isolation, microbial production, chemical properties and methods for its synthesis. Curr Top Microbiol Immunol, 2000, 252, 189 - 200 Neo-self antigens and the expansion of B-1 cells: lessons from atherosclerosis-prone mice; Silverman GJ et al.; The pathogenesis of atherosclerosis involves an inflammatory process that is modulated by the immune system, and within these complex responses we have discerned a possible role for an archetypic B-1 clone . We speculate that due to their immunogenicity and in vivo distribution the "neo"-self determinants created in oxidatively modified LDL are highly stimulatory for certain B-1 cell clones . These neo-self determinants, which can be created chemically, by somatic processes, may in fact represent the molecular analogues of somatic maturation, or even aging . These changes, including those on non-protein antigens induced by oxidative metabolism, amongst others, create neo-determinants against which the host no doubt can not develop rigorous B-cell tolerance . The onset of expression of these oxidative neo-determinants relatively late in development may well serve a useful function for the highly evolved mammalian immune system, as targeting by evolutionarily selected B-1 clones may facilitate the amplification of other useful antibody-mediated physiologic functions . As in the case of the T15 clone, these antibodies may aid in protection against common microbial pathogens . Hence we postulate that during the evolution of the adaptive immune system the neo-self antigenic milieu may have been exploited for the natural selection of primordial clonal specificities . The T15 B-1 clone may then illustrate a common paradigm in which there has been natural selection based on utility for the defense of the individual from environmental threats, as well as for possible "housekeeping" role(s) and the maintenance of cellular homeostasis. Plant J, 2000 Dec, 24(5), 569 - 82 Oxidative burst and cognate redox signalling reported by luciferase imaging: identification of a signal network that functions independently of ethylene, SA and Me-JA but is dependent on MAPKK activity; Grant JJ et al.; Recognition of avirulent microbial pathogens activates an oxidative burst leading to the accumulation of reactive oxygen intermediates (ROIs), which are thought to integrate a diverse set of defence mechanisms resulting in the establishment of plant disease resistance . A novel transgenic Arabidopsis line containing a gst1:luc transgene was developed and employed to report the temporal and spatial dynamics of ROI accumulation and cognate redox signalling in response to attempted infection by avirulent strains of Pseudomonas syringae pv . tomato (Pst) . Strong engagement of the oxidative burst was dependent on the presence of functional Pst hrpS and hrpA gene products . Experiments employing pharmacological agents suggested that at least two distinct sources, including an NADPH oxidase and a peroxidase-type enzyme, contributed to the generation of redox cues . The analysis of gst1 and pal1 gene expression in nahG, coi1 and etr1 plants suggested that engagement of the oxidative burst and cognate redox signalling functioned independently of salicylic acid, methyl jasmonate and ethylene . In contrast, studies using a panel of protein kinase and phosphatase inhibitors and in-gel kinase assays in these mutant backgrounds suggested that a 48 kDa mitogen-activated protein kinase (MAPK) activity was required for the activation of gst1 and pal1 in response to redox cues . Thus the engagement of a bifurcating redox signalling pathway possessing a MAPK module may contribute both to the establishment of plant disease resistance, and to the development of cellular protectant mechanisms. J Immunol, 2001 Jan 1, 166(1), 633 - 41 Production of IL-12 by human monocyte-derived dendritic cells is optimal when the stimulus is given at the onset of maturation, and is further enhanced by IL-4; Ebner S et al.; Dendritic cells produce IL-12 both in response to microbial stimuli and to T cells, and can thus skew T cell reactivity toward a Th1 pattern . We investigated the capacity of dendritic cells to elaborate IL-12 with special regard to their state of maturation, different maturation stimuli, and its regulation by Th1/Th2-influencing cytokines . Monocyte-derived dendritic cells were generated with GM-CSF and IL-4 for 7 days, followed by another 3 days +/- monocyte-conditioned media, yielding mature (CD83(+)/dendritic cell-lysosome-associated membrane glycoprotein(+)) and immature (CD83(-)/dendritic cell-lysosome-associated membrane glycoprotein(-)) dendritic cells . These dendritic cells were stimulated for another 48 h, and IL-12 p70 was measured by ELISA . We found the following: 1) Immature dendritic cells stimulated with CD154/CD40 ligand or bacteria (both of which concurrently also induced maturation) secreted always more IL-12 than already mature dendritic cells . Mature CD154-stimulated dendritic cells still made significant levels (up to 4 ng/ml) . 2) Terminally mature skin-derived dendritic cells did not make any IL-12 in response to these stimuli . 3) Appropriate maturation stimuli are required for IL-12 production: CD40 ligation and bacteria are sufficient; monocyte-conditioned media are not . 4) Unexpectedly, IL-4 markedly increased the amount of IL-12 produced by both immature and mature dendritic cells, when present during stimulation . 5) IL-10 inhibited the production of IL-12 . Our results, employing a cell culture system that is now being widely used in immunotherapy, extend prior data that IL-12 is produced most abundantly by dendritic cells that are beginning to respond to maturation stimuli . Surprisingly, IL-12 is only elicited by select maturation stimuli, but can be markedly enhanced by the addition of the Th2 cytokine, IL-4. Clin Exp Immunol, 2000 Dec, 122(3), 477 - 83 Parasite-mediated down-regulation of collagen-induced arthritis (CIA) in DA rats; Mattsson L et al.; Microbial infection can impact on the course of autoimmune disease, both in disease-inducing and disease-protecting capacities . Here we investigated if infection with Trypanosoma brucei brucei (Tbb), the protozoan causative agent of African Sleeping Sickness, could ameliorate the course of CIA in the Dark Agouti rat, an experimental model which shares many features with human rheumatoid arthritis . Infection of animals with living, but not inoculation with dead Tbb resulted in complete or significant reduction of clinical arthritic symptoms . Infection prior to collagen immunization was more effective than a later treatment, and this effect was related to the level of parasitaemia . Using reverse transcriptase-polymerase chain reaction we detected an increase in interferon-gamma mRNA in the draining lymph nodes of Tbb-treated animals relative to controls at day 28 after disease induction . Transforming growth factor-beta could be detected in the lymph nodes in four out of six animals that had received Tbb . In the joints, immunohistochemistry revealed reduced production of tumour necrosis factor-alpha in Tbb-treated animals relative to controls . The most striking difference between Tbb-infected and control groups, as measured by ELISA, was the down-regulation of anti-collagen II IgG antibody responses in parasite-infected animals . We conclude that live parasites can exert an immunomodulatory and protective effect in CIA in which several mechanisms may work in parallel, although the almost complete down-regulation of the anti-collagen antibody response may alone explain the protective effect in CIA . The described model may be useful in further attempts to use the mechanisms involved in parasite immune defence to prevent and treat certain autoimmune conditions. Curr Opin Microbiol, 2000 Dec, 3(6), 625 - 30 Signalling molecules involved in cellular differentiation during Dictyostelium morphogenesis; Weeks G; GSK-3, Dd-STATa, PKA, rZIP and Ras all play important roles in cell type determination of Dictyostelium discoideum . The fact that homologs of these proteins also function in metazoan development emphasizes the importance of Dictyostelium as a model microbial organism for studying the molecular mechanisms that regulate development . The recent elaboration of the central role for GSK-3 in cell type determination has been of particular importance . The stimulatory effect of extracellular cAMP on GSK-3 activity has been shown to act through the cell surface receptor cAR3 and a tyrosine protein kinase ZAK1, which directly activates and phosphorylates GSK-3 . Several proteins, including Dd-STATa, have been identified as substrates for GSK-3, and are therefore potential transducers of the signals involved in cell type determination. Antimicrob Agents Chemother, 2001 Jan, 45(1), 187 - 95 Efficacies of lipophilic inhibitors of dihydrofolate reductase against parasitic protozoa; Lau H et al.; Competitive inhibitors of dihydrofolate reductase (DHFR) are used in chemotherapy or prophylaxis of many microbial pathogens, including the eukaryotic parasites Plasmodium falciparum and Toxoplasma gondii . Unfortunately, point mutations in the DHFR gene can confer resistance to inhibitors specific to these pathogens . We have developed a rapid system for testing inhibitors of DHFRs from a variety of parasites . We replaced the DHFR gene from the budding yeast Saccharomyces cerevisiae with the DHFR-coding region from humans, P . falciparum, T . gondii, Pneumocystis carinii, and bovine or human-derived Cryptosporidium parvum . We studied 84 dicyclic and tricyclic 2,4-diaminopyrimidine derivatives in this heterologous system and identified those most effective against the DHFR enzymes from each of the pathogens . Among these compounds, six tetrahydroquinazolines were effective inhibitors of every strain tested, but they also inhibited the human DHFR and were not selective for the parasites . However, two quinazolines and four tetrahydroquinazolines were both potent and selective inhibitors of the P . falciparum DHFR . These compounds show promise for development as antimalarial drugs. Infect Immun, 2001 Jan, 69(1), 456 - 62 Borrelia spirochetes upregulate release and activation of matrix metalloproteinase gelatinase B (MMP-9) and collagenase 1 (MMP-1) in human cells; Gebbia JA et al.; Borrelia burgdorferi, the spirochetal agent of Lyme disease, stimulated human peripheral blood monocytes to release pro-matrix metalloproteinase-9 (gelatinase B; pro-MMP-9) and active matrix metalloproteinase-1 (collagenase-1; MMP-1) . Human neutrophils also released pro-MMP-9 and a 130-kDa protein with gelatinolytic activity in response to live B . burgdorferi . In addition, U937 cells and human keratinocyte cells were also stimulated to release pro-MMP-9 under the same conditions . However, human umbilical vein endothelial cells (HUVECs) released pro-MMP-9 and pro-MMP-2 in a constitutive manner and were not influenced by live spirochetes . MMPs produced by human monocytes also enhanced the penetration of B . burgdorferi through extracellular matrix component barriers in vitro . Plasmin stabilized on the surface of the Lyme disease spirochete was shown to activate pro-MMP-9 to its active form . This active form was also observed in the plasma of mice infected with a relapsing fever borrelia . These results suggest that borreliae can upregulate MMPs and possibly mediate an activation cascade initiated by plasmin bound to the microbial surface . MMPs may play a role in dissemination of the Lyme disease spirochete and in the pathogenesis of Borrelia infection. Scand J Immunol, 2000 Dec, 52(6), 609 - 17 Expression of CD1d in the duodenum of patients with cow's milk hypersensitivity; Ulanova M et al.; CD1 cell surface glycoproteins represent a family of non-major histocompatibility complex (MHC) encoded antigen-presenting molecules . All members of the CD1 family appear to mediate the recognition of microbial or endogenous lipid and glycolipid antigens . The recognition of CD1d by a unique subset of natural killer (NK) T cells that leads to rapid production of large amounts of both type 1 and type 2 cytokines can be augmented by some synthetic glycolipids . Because of the proposed role of such CD1d-restricted T cells in immunoregulation, we hypothesized that CD1d molecules participate in mucosal immune responses in patients with gastrointestinal symptoms owing to food hypersensitivity . Patients of that category represent a heterogeneous group in which poorly defined immunological mechanisms are believed to contribute to disease pathogenesis . The expression of CD1 in duodenal biopsy samples from six patients with verified intolerance to cow's milk and six healthy controls was studied by immunoperoxidase staining of cryostat sections using a panel of mouse monoclonal antibodies (MoAbs) specific for CD1a, b, c, and d . Large numbers of CD1d positive cells were found in the lamina propria of all the patients, both during the symptomatic and the asymptomatic periods, whereas healthy controls were virtually devoid of CD1d expression in the duodenum . The localization of CD1d positive cells corresponded to areas where B cells, plasma cells and dendritic cells (DC) were present . A positive correlation was found between the numbers of CD1d(+) and CD19(+) cells in the lamina propria . In contrast to previous reports, no CD1d expression was found on the epithelial cells . Although less numerous than CD1d(+) the CD1c(+) cells were also present in all the patients and in five out of six controls . No staining for CD1a or CD1b was detected in the duodenal biopsy samples from any of the subjects . The exclusive presence of CD1d in the duodenal lamina propria of the patients with cow's milk hypersensitivity might suggest the participation of these molecules in the pathogenesis of allergic reactions to food. Virology, 2000 Dec 20, 278(2), 514 - 9 In vitro induction of HIV-1 replication in resting CD4(+) T cells derived from individuals with undetectable plasma viremia upon stimulation with human T-cell leukemia virus type I; Moriuchi H et al.; Microbial coinfections have been associated with transient bursts of human immunodeficiency virus (HIV) viremia in patients . In this study we investigated whether human T-cell leukemia virus type I (HTLV-I), another human retrovirus that is prevalent among certain HIV-infected populations, can induce HIV-1 replication in patients who had been successfully treated with highly active antiretroviral therapy . We demonstrate that supernatants from HTLV-I-producing MT-2 cells can induce in vitro replication of HIV-1 from highly purified, resting CD4(+) T cells obtained from individuals with undetectable plasma viremia . Depletion of proinflammatory cytokines from the supernatants reduced, but did not abrogate, the ability to induce HIV-1 replication, indicating that other factors such as HTLV-I Tax or Env also have a role . The HTLV-I-mediated effect does not require productive infection: exposure to heat-inactivated HTLV-I virions, purified Tax protein, or HTLV-I Env glycoprotein also induced expression of HIV-1 . Furthermore, we demonstrate that coculture of resting CD4(+) T cells with autologous CD8(+) T cells markedly inhibits the HTLV-I-induced virus replication . Our results suggest that coinfection with HTLV-I may induce viral replication in the latent viral reservoirs; however, CD8(+) T cells may play an important role in controlling the spread of virus upon microbial stimulation . Nature, 2000 Nov 30, 408(6812), 580 - 3 Bacterial dehalorespiration with chlorinated benzenes; Adrian L et al.; Chlorobenzenes are toxic, highly persistent and ubiquitously distributed environmental contaminants that accumulate in the food chain . The only known microbial transformation of 1,2,3,5-tetrachlorobenzene (TeCB) and higher chlorinated benzenes is the reductive dechlorination to lower chlorinated benzenes under anaerobic conditions observed with mixed bacterial cultures . The lower chlorinated benzenes can subsequently be mineralized by aerobic bacteria . Here we describe the isolation of the oxygen-sensitive strain CBDB1, a pure culture capable of reductive dechlorination of chlorobenzenes . Strain CBDB1 is a highly specialized bacterium that stoichiometrically dechlorinates 1,2,3-trichlorobenzene (TCB), 1,2,4-TCB, 1,2,3,4-TeCB, 1,2,3,5-TeCB and 1,2,4,5-TeCB to dichlorobenzenes or 1,3,5-TCB . The presence of chlorobenzene as an electron acceptor and hydrogen as an electron donor is essential for growth, and indicates that strain CBDB1 meets its energy needs by a dehalorespiratory process . According to their 16S rRNA gene sequences, strain CBDB1, Dehalococcoides ethenogenes and several uncultivated bacteria form a new bacterial cluster, of which strain CBDB1 is the first, so far, to thrive on a purely synthetic medium. Curr Microbiol, 2001 Jan, 42(1), 32 - 8 Evaluation of nitrogenous substrates such as peptones from fish:a new method based on Gompertz modeling of microbial growth; Dufosse L et al.; Fish peptones from tuna, cod, salmon, and unspecified fish were compared with a casein one by using a new method based on Gompertz modeling of microbial growth . Cumulative results obtained from six species of bacteria, yeasts, and fungi showed that, in most cases, these fish peptones are very effective . Nevertheless, this study raised some questions about the standardization of fish raw material, the enzymatic hydrolysis of fish proteins, and the composition of the culture medium used for testing the peptones. Vaccine, 2000 Nov 22, 19(7-8), 840 - 3 Protection of cattle against rinderpest by intranasal immunisation with a dry powder tissue culture vaccine; Anderson J et al.; Dry powder tissue culture rinderpest vaccine containing 10(2.5) TCID(50) of virus per dose administered intranasally to cattle induced high titre circulating antibody responses and protection against challenge with a virulent strain of rinderpest virus . A reduction in the dose of virus to 10(1.1) TCID(50) resulted in a failure to elicit detectable antibody responses and a lack of protection . Intranasal powder vaccine offers several advantages over conventional needle-administered aqueous rinderpest vaccine, including greater stability in the absence of a cold chain, reduced risk of 'needle transfer' of other microbial agents present in the vaccinated herd and lower cost. Clin Infect Dis, 2000 Oct, 31 Suppl 5, S190 - 201 Endogenous antipyretics; Tatro JB; Fever is the hallmark of the stereotyped host response to microbial infection, although it is just one of a number of high-risk strategies employed by the infected host to clear itself of invading pathogens . The febrile response is accompanied by activation of multiple endogenous antipyretic systems that serve to suppress its magnitude or duration . These include neuroactive substances of neural and humoral origin, some of which (e.g., glucocorticoids, melanocortins, and IL-10) have broad-ranging anti-inflammatory actions . Glucocorticoids, vasopressin, and melanocortins appear to exert their antipyretic effects by acting on receptors within the brain, but beyond this the mechanisms involved are unknown . It is hypothesized, but not proven, that endogenous antipyretic systems protect the host against the destructive consequences of unchecked fever . Importantly, pharmacological blockade of the actions of endogenous antipyretic systems increases fevers of even low to moderate intensity . Therefore, in addition to protecting against catastrophic consequences of high fever, endogenous antipyretic systems seem to play a fundamental physiological role in determining the normal course of fever . Elucidating the neural and biochemical mechanisms involved in suppression of fever by physiological antipyretic systems will yield a rich benefit, both by advancing the basic understanding of host defense strategies, and by permitting the design of novel antipyretic and anti-inflammatory strategies for therapeutic intervention in human disease. Int J Med Microbiol, 2000 Oct, 290(4-5), 307 - 11 Toxin genes on pathogenicity islands: impact for microbial evolution; Dobrindt U et al.; Toxin-specific genes are often located on mobile genetic elements such as phages, plasmids and pathogenicity islands (PAIs) . The uropathogenic E . coli strain 536 carries two alpha-hemolysin gene clusters, which are part of the pathogenicity islands I536 and II536, respectively . Using different genetic techniques, two additional PAIs were identified in the genome of the E . coli strain 536, and it is likely that further PAIs are located on the genome of this strain . Pathogenicity islands are often associated with tRNA genes . In the case of the E . coli strain 536, the PAI-associated tRNA gene leuX, which encodes a minor leucyl-tRNA, affects the expression of various virulence traits including alpha-hemolysin production . The exact mode of action of the tRNA5Leu-dependent gene expression has to be identified in the future. Clin Pediatr (Phila), 2000 Nov, 39(11), 627 - 33 Radiologic differentiation between bacterial and viral lower respiratory infection in children: a systematic literature review; Swingler GH; A systematic literature review was performed to quantify the accuracy of chest radiography in differentiating bacterial from viral lower respiratory infection in children . Relevant studies were identified in a systematic literature search and were included in the review according to predetermined criteria . Five of 13 relevant identified studies met the inclusion criteria . No clinically useful degree of accuracy was demonstrated, but great caution is needed in interpreting the findings because of the suboptimal nature of the reference standards, even in included studies . It is recommended that future surveys of the microbial etiology of pneumonia that employ a credible reference standard (such as lung aspiration) be used as opportunities to perform studies of diagnostic accuracy. Proc Inst Mech Eng {H}, 2000, 214(5), 513 - 8 Influence of gelatin and bovine serum lubricants on ultra-high molecular weight polyethylene wear debris generated in in vitro simulations; Bell J et al.; Ultra-high molecular weight polyethylene (UHMWPE) wear debris induced osteolysis has a major role in the late aseptic loosening and ultimate failure of total hip replacements (THR) . Clinically relevant in vitro simulations of wear are essential to predict the osteolytic potential of bearing surfaces in artificial hip joints . Newborn calf or bovine serum has been accepted as a boundary lubricant for such in vitro tests, but its biological stability has been questioned . This study compared the wear factors, number of wear particles and levels of microbial contamination produced in bovine serum and a gelatin-based lubricant . The wear factors produced by the two lubricants were not significantly different, however the wear debris morphology produced was substantially different . The bovine serum became contaminated with micro-organisms within 28 h, whereas the protein-based lubricant remained uncontaminated . The results showed that bovine serum was not a stable boundary lubricant . They also showed that although the wear factors for the two solutions were not significantly different, the protein-based lubricant was not a suitable alternative to bovine serum because the wear debris produced was not clinically relevant. Int J Hyg Environ Health, 2000 Oct, 203(2), 97 - 104 Exposure to airborne fungi, MVOC and mycotoxins in biowaste-handling facilities; Fischer G et al.; Health impacts due to fungi in indoor air can only be estimated reliably, if both fungal propagules and fungal secondary metabolites are qualified and quantified . In the present study, the fungal species composition in a compost facility is compared to the spectrum of microbial metabolites in the air with regard to the physiological properties of different fungal species . A number of relevant fungi was tested for the production of both volatile and non-volatile metabolites on different substrata . The profiles of mycotoxins and microbial volatile organic compounds (MVOC) turned out to be specific for certain species in pure culture . Consequently, the fungi may have different toxicological health impacts, though information on the relevance of microbial volatiles is still limited. Diagn Microbiol Infect Dis, 2000 Nov, 38(3), 185 - 7 A new diagnostic problem: isolation of Escherichia coli O157:H7 strains with aberrant biochemical properties; Ware JM et al.; Over a five-year period (1995-1999) the Microbial Diseases Laboratory received 34 strains of E . coli O157:H7 each with a single aberrant biochemical property . In addition, 27 O157 strains with negative or delayed motility were noted during the same time period . These observations suggest that there may be an increased likelihood to misdiagnose O157:H7 infections using commercial systems in the future due to increasing phenotypic variability. Int Microbiol, 2000 Sep, 3(3), 139 - 46 DNA methylation in ciliates: implications in differentiation processes; Gutierrez JC et al.; Much experimental evidence on the role of DNA methylation in gene expression has been reported . Here we review reports on DNA methylation in ciliated protozoa, emphasizing its implications in cell differentiation processes . Both types of methylated bases (adenine and cytosine) can be found in macronuclear DNA . The division cycle and conjugation have been studied with regard to adenine methylation, and several different functions have been assigned to the methylation changes detected in these processes . Cytosine methylation changes were analyzed during stomatogenesis of Paramecium and encystment of Colpoda inflata . A comparative analysis with other similar microbial eukaryotic differentiation processes is carried out. Int Microbiol, 2000 Jun, 3(2), 97 - 101 Mineralogical composition and biomass studies of the microbial mats sediments from the Ebro Delta, Spain; Urmeneta J et al.; The mineral composition of the microbial mats at La Banya spit was studied . The spit is formed by a narrow sand bar and a peninsula and is located south of the main body of the Ebro Delta (Tarragona, Spain) . Although quartz was the predominant mineral component in all sampling sites, clay, feldspars, calcite, aragonite, halite, dolomite and gypsum were also found . An increase in both the fine material (clay) and the halite content was observed in the sites influenced by nearby salterns . The amount of each mineral did not differ significantly along a 55 cm deep profile, except for halite and aragonite, which reached a maximum in the surface and decreased with depth . Dolomite, which ranged from 0.5 to 5% (w/w), is a possible indicator of sulfate-reducing bacteria activity in the past . Organic carbon and total nitrogen were quantified for biomass assessment . Total nitrogen ranged from 0.1 to 0.56% in the uppermost layer, where the microbial mat is active, but was undetectable at deeper layers . Organic carbon ranged from 1 to 5.5% in the active microbial mat layers and decreased to 0.3% at deeper layers . During the summer, both organic carbon and total nitrogen contents (biomass) of the microbial mat samples from some sites increase, whereas other sites show constant concentrations throughout the year, and others have a fluctuant biomass content. Int Microbiol, 1998 Mar, 1(1), 27 - 34 Cosmopolitan distribution of the large composite microbial mat spirochete, Spirosymplokos deltaeiberi; Margulis L et al.; Inocula from organic-rich black muds immediately underlying intertidal laminated microbial mats dominated by Microcoleus chthonoplastes yielded large, variable diameter spirochetes . These unusual spirochetes, previously reported only from the Alfacs Peninsula at the delta of the Ebro river in northeast Spain, contain striking arrays of cytoplasmic granules packed into their protoplasmic cylinders . On several occasions, both in summer and winter, the huge spirochetes were recognized in samples from mats growing in the Sippewissett salt marsh at Woods Hole Massachusetts . They were also seen in similar samples from microbial mats at North Pond, Laguna Figueroa, Baja California Norte, Mexico . The identity of these spirochetes was confirmed by electron microscopy: number and disposition of flagella, composite structure, measurements of their distinctive cytoplasmic granules . The granules, larger, more conspicuous and present in addition to ribosomes, are hypothesized to contain ATPases . As culture conditions worsen, these spirochetes retract into membrane-bounded round bodies in which they form refractile inclusions . From morphology and behavior we conclude the North American spirochetes from both Atlantic and Pacific intertidal microbial mats are indistinguishable from those at the delta of the Ebro river . We conclude a cosmopolitan distribution for Spirosymplokos deltaeiberi. Braz J Infect Dis, 1997 Mar, 1(1), 17 - 26 Emerging Pathogens Associated with Tick-Borne Infections; Roberts R et al.; In recent years, several microbial agents have been identified