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PubMed2003

MBGD: microbial genome database for comparative analysis.

MBGD is a workbench system for comparative analysis of completely sequenced microbial genomes. The central function of MBGD is to create an orthologous gene classification table using precomputed all-against-all similarity relationships among genes in multiple genomes. In MBGD, an automated classification algorithm has been implemented so that users can create their own classification table by specifying a set of organisms and parameters. This feature is especially useful when the user's interest is focused on some taxonomically related organisms. The created classification table is stored into the database and can be explored combining with the data of individual genomes as well as similarity relationships among genomes. Using these data, users can carry out comparative analyses from various points of view, such as phylogenetic pattern analysis, gene order comparison and detailed gene structure comparison. MBGD is accessible at http://mbgd.genome.ad.jp/.

Algorithms↗
PubMed2006

Reduced CD1d expression in colonic epithelium in microscopic colitis.

CD1d is a member of a major histocompatibility complex class I-like molecule family. Its function may involve presenting microbial nonpeptide or lipidic antigens to T lymphocytes, therefore to serve as an important factor in normal mucosal immunity of the gastrointestinal tract. In this study, the expression level of CD1d in microscopic colitis (ie, collagenous and lymphocytic colitis) was examined, and compared with that in normal colonic mucosa. Formalin-fixed, paraffin-embedded colon biopsies with diagnosis of lymphocytic colitis (19 cases), collagenous colitis (6 cases), and no pathologic change (20 cases) were studied immunohistochemically using monoclonal antibodies against human CD1d, CD3, CD4, and CD8. CD1d staining in the epithelium and lamina propria was graded along a scale of 0 to 4. Intraepithelial CD3-positive lymphocytes were counted in an area of 300 epithelial cells for each specimen. The results show that CD1d was expressed in normal colonic epithelial cells, primarily on the basolateral membranes with a concentrated intracellular pool in the subnuclear region. The expression level was markedly reduced in both lymphocytic colitis (P<0.001) and collagenous colitis (P<0.001), along with a significant increase in the number of intraepithelial CD3/CD8 lymphocytes (P<0.001). These findings suggest that microscopic colitis is associated with decreased epithelial expression of CD1d, an important immunoregulatory molecule in the gastrointestinal tract.

Adult↗
PubMed2005

Immunoglobulin a modulates inflammatory responses in an in vitro model of pneumonia.

BACKGROUND: Preservation of mucosal immunity has been shown to affect the risk and outcome of pneumonia in severely injured patients. Secretory immunoglobulin A (SIgA) is the principle humoral defense of mucosal surfaces in the body and has several antiinflammatory properties. Polymorphonuclear neutrophils (PMN) function to kill invading microorganisms, but their exaggerated inflammatory responses may cause tissue injury to the host. The purpose of this study was to compare the ability of different immunoglobulin (Ig) isotypes to modulate PMN cytotoxic potential cocultured with respiratory epithelial cells challenged with bacteria. METHODS: Calu-3 cell monolayers were established on membranes (0.1-microm pore) in a two-chamber culture system. Escherichia coli (EC) incubated with either polyclonal SIgA or IgG was inoculated into the apical chamber and PMNs (10/mL) added to the basal chamber. PMN cytotoxic potential was indexed by % CD11b expression, superoxide anion (O2) production, and elastase release. Dextran flux was used to index Calu-3 monolayer permeability. RESULTS: Addition of EC to PMN-Calu-3 cell coculture increased % CD11b expression, O2 production, and elastase release. IgG had no effect on PMN activation after EC challenge. SIgA abrogated PMN activation and the increase in Calu-3 cell monolayer permeability noted with EC or EC + IgG treatment groups. CONCLUSION: PMN cytotoxic potential was decreased by the presence of SIgA but not IgG in an in vitro model to simulate pneumonia in vivo. SIgA may not only function to protect against microbial invasion of mucosal surfaces, but may also protect against tissue injury from an exaggerated inflammatory response.

Apoptosis↗
PubMed2002

Cross-scale ecological dynamics and microbial size spectra in marine ecosystems.

Evaluating the component features of 'scaling' planktonic size spectra, commonly observed in marine ecosystems, is crucial for understanding the ecological and evolutionary processes from which they emerge. Here, we develop a theoretical framework that describes such spectra in terms of the size distributions of individual species, and test it against actual datasets of microbial size spectra from the Atlantic Ocean. We describe characteristics of size probability distributions of component species that are sufficient to support the observational evidence and infer that, when a power law describes the community size spectrum (thus suggesting critical self-organization of microbial ecosystem structure and function), a related power law links the total number of individuals of a given species to its mean size.

Animals↗
PubMed2005

Pulmonary infections in swine induce altered porcine surfactant protein D expression and localization to dendritic cells in bronchial-associated lymphoid tissue.

Surfactant protein D (SP-D) is a pattern-recognition molecule of the innate immune system that recognizes various microbial surface-specific carbohydrate and lipid patterns. In vitro data has suggested that this binding may lead to increased microbial association with macrophages and dendritic cells. The aim of the present in vivo study was to study the expression of porcine SP-D (pSP-D) in the lung during different pulmonary bacterial infections, and the effect of the routes of infection on this expression was elucidated. Furthermore, the aim was to study the in vivo spatial relationship among pSP-D, pathogens, phagocytic cells and dendritic cells. Lung tissue was collected from experimental and natural bronchopneumonias caused by Actinobacillus pleuropneumoniae or Staphylococcus aureus, and from embolic and diffuse interstitial pneumonia, caused by Staph. aureus or Arcanobacterium pyogenes and Streptococcus suis serotype 2, respectively. By comparing normal and diseased lung tissue from the same lungs, increased diffuse pSP-D immunoreactivity was seen in the surfactant in both acute and chronic bronchopneumonias, while such increased expression of pSP-D was generally not present in the interstitial pneumonias. Co-localization of pSP-D, alveolar macrophages and bacteria was demonstrated, and pSP-D showed a patchy distribution on the membranes of alveolar macrophages. SP-D immunoreactivity was intracellular in dendritic cells. The dendritic cells were identified by their morphology, the absence of macrophage marker immunoreactivity and the presence of dendritic cell marker immunoreactivity. Increased expression of pSP-D in the surfactant coincided with presence of pSP-D-positive dendritic cells in bronchus-associated lymphoid tissue (BALT), indicating a possible transport of pSP-D through the specialized M cells overlying (BALT). In conclusion, we have shown that pSP-D expression in the lung surfactant is induced by bacterial infection by an aerogenous route rather than by a haematogenous route, and that the protein interacts specifically with alveolar macrophages and with dendritic cells in microbial-induced BALT. The function of the interaction between pSP-D and dendritic cells in BALT remain unclear, but pSP-D could represent a link between the innate and adaptive immune system, facilitating the bacterial antigen presentation by dendritic cells in BALT.

Actinobacillus Infections↗
PubMed2005

Novel hydrolase diversity retrieved from a metagenome library of bovine rumen microflora.

A metagenome expression library of bulk DNA extracted from the rumen content of a dairy cow was established in a phage lambda vector and activity-based screening employed to explore the functional diversity of the microbial flora. Twenty-two clones specifying distinct hydrolytic activities (12 esterases, nine endo-beta-1,4-glucanases and one cyclodextrinase) were identified in the library and characterized. Sequence analysis of the retrieved enzymes revealed that eight (36%) were entirely new and formed deep-branched phylogenetic lineages with no close relatives among known ester- and glycosyl-hydrolases. Bioinformatic analyses of the hydrolase gene sequences, and the sequences and contexts of neighbouring genes, suggested tentative phylogenetic assignments of the rumen organisms producing the retrieved enzymes. The phylogenetic novelty of the hydrolases suggests that some of them may have potential for new applications in biocatalysis.

Animals↗
PubMed2006

Interactions between arbuscular mycorrhizal fungi and bacteria and their potential for stimulating plant growth.

Arbuscular mycorrhizal (AM) fungi and bacteria can interact synergistically to stimulate plant growth through a range of mechanisms that include improved nutrient acquisition and inhibition of fungal plant pathogens. These interactions may be of crucial importance within sustainable, low-input agricultural cropping systems that rely on biological processes rather than agrochemicals to maintain soil fertility and plant health. Although there are many studies concerning interactions between AM fungi and bacteria, the underlying mechanisms behind these associations are in general not very well understood, and their functional properties still require further experimental confirmation. Future mycorrhizal research should therefore strive towards an improved understanding of the functional mechanisms behind such microbial interactions, so that optimized combinations of microorganisms can be applied as effective inoculants within sustainable crop production systems. In this context, the present article seeks to review and discuss the current knowledge concerning interactions between AM fungi and plant growth-promoting rhizobacteria, the physical interactions between AM fungi and bacteria, enhancement of phosphorus and nitrogen bioavailability through such interactions, and finally the associations between AM fungi and their bacterial endosymbionts. Overall, this review summarizes what is known to date within the present field, and attempts to identify promising lines of future research.

Bacterial Physiological Phenomena↗
PubMed2005

Host-bacterial mutualism in the human intestine.

The distal human intestine represents an anaerobic bioreactor programmed with an enormous population of bacteria, dominated by relatively few divisions that are highly diverse at the strain/subspecies level. This microbiota and its collective genomes (microbiome) provide us with genetic and metabolic attributes we have not been required to evolve on our own, including the ability to harvest otherwise inaccessible nutrients. New studies are revealing how the gut microbiota has coevolved with us and how it manipulates and complements our biology in ways that are mutually beneficial. We are also starting to understand how certain keystone members of the microbiota operate to maintain the stability and functional adaptability of this microbial organ.

Anaerobiosis↗
PubMed1996

Competition and coexistence of sulfate-reducing and methanogenic populations in anaerobic biofilms.

The microbial population structure and function of natural anaerobic communities maintained in laboratory fixed-bed biofilm reactors were tracked before and after a major perturbation, which involved the addition of sulfate to the influent of a reactor that had previously been fed only glucose (methanogenic), while sulfate was withheld from a reactor that had been fed both glucose and sulfate (sulfidogenic). The population structure, determined by using phylogenetically based oligonucleotide probes for methanogens and sulfate-reducing bacteria, was linked to the functional performance of the biofilm reactors. Before the perturbation, the methanogenic reactor contained up to 25% methanogens as well as 15% sulfate-reducing bacteria, even though sulfate was not present in the influent of this reactor. Methanobacteriales and Desulfovibrio spp. were the most abundant methanogens and sulfate-reducing bacteria, respectively. The presence of sulfate-reducing bacteria (primarily Desulfovibrio spp. and Desulfobacterium spp.) in the absence of sulfate may be explained by their ability to function as proton-reducing acetogens and/or fermenters. Sulfate reduction began immediately following the addition of sulfate consistent with the presence of significant levels of sulfate-reducing bacteria in the methanogenic reactor, and levels of sulfate-reducing bacteria increased to a new steady-state level of 30 to 40%; coincidentally, effluent acetate concentrations decreased. Notably, some sulfate-reducing bacteria (Desulfococcus/Desulfosarcina/Desulfobotulus group) were more competitive without sulfate. Methane production decreased immediately following the addition of sulfate; this was later followed by a decrease in the relative concentration of methanogens, which reached a new steady-state level of approximately 8%. The changeover to sulfate-free medium in the sulfidogenic reactor did not cause a rapid shift to methanogenesis. Methane production and a substantial increase in the levels of methanogens were observed only after approximately 50 days following the perturbation.

Journal Article↗
PubMed2001

Cross-epithelial hydrogen transfer from the midgut compartment drives methanogenesis in the hindgut of cockroaches.

In the intestinal tracts of animals, methanogenesis from CO(2) and other C(1) compounds strictly depends on the supply of electron donors by fermenting bacteria, but sources and sinks of reducing equivalents may be spatially separated. Microsensor measurements in the intestinal tract of the omnivorous cockroach Blaberus sp. showed that molecular hydrogen strongly accumulated in the midgut (H(2) partial pressures of 3 to 26 kPa), whereas it was not detectable (<0.1 kPa) in the posterior hindgut. Moreover, living cockroaches emitted large quantities of CH(4) [105 +/- 49 nmol (g of cockroach)(-1) h(-1)] but only traces of H(2). In vitro incubation of isolated gut compartments, however, revealed that the midguts produced considerable amounts of H(2), whereas hindguts emitted only CH(4) [106 +/- 58 and 71 +/- 50 nmol (g of cockroach)(-1) h(-1), respectively]. When ligated midgut and hindgut segments were incubated in the same vials, methane emission increased by 28% over that of isolated hindguts, whereas only traces of H(2) accumulated in the headspace. Radial hydrogen profiles obtained under air enriched with H(2) (20 kPa) identified the hindgut as an efficient sink for externally supplied H(2). A cross-epithelial transfer of hydrogen from the midgut to the hindgut compartment was clearly evidenced by the steep H(2) concentration gradients which developed when ligated fragments of midgut and hindgut were placed on top of each other-a configuration that simulates the situation in vivo. These findings emphasize that it is essential to analyze the compartmentalization of the gut and the spatial organization of its microbiota in order to understand the functional interactions among different microbial populations during digestion.

Animals↗
PubMed2002

Sequencing and transcriptional analysis of the chlorite dismutase gene of Dechloromonas agitata and its use as a metabolic probe.

The dismutation of chlorite into chloride and O(2) represents a central step in the reductive pathway of perchlorate that is common to all dissimilatory perchlorate-reducing bacteria and is mediated by a single enzyme, chlorite dismutase. The chlorite dismutase gene cld was isolated and sequenced from the perchlorate-reducing bacterium Dechloromonas agitata strain CKB. Sequence analysis identified an open reading frame of 834 bp that would encode a mature protein with an N-terminal sequence identical to that of the previously purified D. agitata chlorite dismutase enzyme. The predicted translation product of the D. agitata cld gene is a protein of 277 amino acids (aa), including a leader peptide of 26 aa. Primer extension analysis identified a single transcription start site directly downstream of an AT-rich region that could represent the -10 promoter region of the D. agitata cld gene. Northern blot analysis indicated that the cld gene was transcriptionally up-regulated when D. agitata cells were grown in perchlorate-reducing versus aerobic conditions. Slot blot hybridizations with a D. agitata cld probe demonstrated the conservation of the cld gene among perchlorate-reducing bacteria. This study represents the first description of a functional gene associated with microbial perchlorate reduction.

Amino Acid Sequence↗
PubMed2006

Unexpected diversity of bacteria capable of carbon monoxide oxidation in a coastal marine environment, and contribution of the Roseobacter-associated clade to total CO oxidation.

The species diversity, phylogenetic affiliations, and physiological activity rates of carbon monoxide-oxidizing microorganisms were investigated, using new isolates from surface waters collected from the coast of New England and type strains from established collections. A direct isolation method allowed the simultaneous recovery of organisms with different growth rates and nutritional requirements and the identification of marine microorganisms that oxidize CO at an environmentally relevant concentration (42 nM CO). Isolates that oxidized CO at environmentally relevant rates (>4.5x10(-11) nmol CO oxidized cell-1 h-1) were taxonomically diverse, with representatives in the alpha and gamma subclasses of the Proteobacteria and the phylum Bacteroidetes, and represent a hitherto unreported metabolic function for several diverse microbial types. Isolates and type strains having the greatest specific rates of CO metabolism (1.1x10(-10) to 2.3x10(-10) nmol CO oxidized cell-1 h-1) belonged to the Roseobacter-associated clade (RAC) of the alpha subclass of the Proteobacteria. By using triple-labeled slide preparations, differential counts of active CO-oxidizing RAC cells, total RAC cells, and total bacterial cell counts in environmental samples were obtained. RAC organisms were a major component of total cell numbers (36%). Based on the density of active CO-oxidizing RAC cells in natural samples and RAC-specific metabolic activities determined for pure cultures, active CO-oxidizing RAC cells may contribute up to 15% of the total CO oxidation occurring in coastal waters.

Carbon Monoxide↗
PubMed1985

Mathematical model for determining the effects of intracytoplasmic inclusions on volume and density of microorganisms.

Procaryotic microorganisms accumulate several polymers in the form of intracellular inclusions as a strategy to increase survival in a changing environment. Such inclusions avoid osmotic pressure increases by tightly packaging certain macromolecules into the inclusion. In the present paper, a model describing changes in volume and density of the microbial cell as a function of the weight of the macromolecule forming the inclusion is derived from simple theoretical principles. The model is then tested by linear regression with experimental data from glycogen accumulation in Escherichia coli, poly-beta-hydroxybutyrate accumulation in Alcaligenes eutrophus, and sulfur accumulation in Chromatium spp. The model predicts a certain degree of hydration of the polymer in the inclusion and explains both the linear relationship between volume of the cell and weight of the polymer and the hyperbolic relationship between density of the cell and weight of the polymer. Other implications of the model are also discussed.

Alcaligenes↗
PubMed1989

Cleavage of small peptides in vitro by human rhinovirus 14 3C protease expressed in Escherichia coli.

The 3C region of human rhinovirus 14 was expressed in Escherichia coli. The microbially synthesized protease was functional, since the expressed precursor underwent autoproteolytic processing to generate mature molecules of the expected molecular weight and antigenicity. Mutation of the putative active-site Cys-146 residue to an alanine resulted in the synthesis of unprocessed precursor molecules. Large quantities of the 20-kilodalton protease were purified by a simple purification protocol, and the resulting molecule was shown to be biologically active in vitro against synthetic peptides corresponding to the 2C-3A cleavage site. This site was cleaved with high efficiency and fidelity and was used to generate kinetic data on the 3C protease. The protease exhibited sensitivity to Zn2+, was capable of cleaving five of seven rhinovirus cleavage site peptides tested with variable efficiency, and could distinguish authentic substrate peptides from control peptides containing the dipeptide cleavage sequence pair Gln-Gly.

3C Viral Proteases↗
PubMed2003

Granulocyte function in grancalcin-deficient mice.

Grancalcin, one of the penta-EF-hand Ca(2+) binding proteins, is expressed at high levels in polymorphonuclear granulocytes (neutrophils). EF-hand proteins are implicated in the regulation of diverse processes including cell migration, apoptosis, and mobilization of neutrophil effector functions. To determine the role of grancalcin in vivo, we inactivated the gene encoding grancalcin (Gca) in embryonic stem cells and generated grancalcin-deficient mice. Homozygous Gca mutants appeared healthy and reproduced normally. Leukocyte recruitment into the peritoneal cavity upon induction of inflammation was not significantly affected by the absence of grancalcin. The mutants also resisted systemic fungal infection similarly to wild-type mice, and in vitro killing of Staphylococcus aureus by inflammatory cells was not significantly impaired. While marginally increased survival rates of mutants faced with endotoxic shock may indicate a contribution of grancalcin to immunopathogenesis, it is not essential for vital leukocyte effector functions required to control microbial infections.

Animals↗
PubMed2004

Selective decontamination of the digestive tract attenuated the myocardial inflammation and dysfunction that occur with burn injury.

This study examined the effects of oral antibiotics to selectively decontaminate the digestive tract (SDD) on postburn myocardial signaling, inflammation, and function. We hypothesized that antibiotic therapy to eliminate pathogens from the gastrointestinal (GI) tract would reduce myocardial inflammatory responses and improve postburn myocardial performance. Sprague-Dawley rats received polymyxin E (15 mg), tobramycin (6 mg), and 5-flucytosin (100 mg) by oral gavage twice daily for 3 days preburn and 24 h postburn. Experimental groups included 1) sham burn given vehicle (3 ml water), 2) sham plus SDD, 3) burn over 40% total body surface area (TBSA) plus SDD, and 4) burn over 40% TBSA given vehicle. All burns received lactated Ringer solution (4 mg.kg(-1).%burn(-1)); myocardial signaling (PKCepsilon/p38 MAPK/NF-kappaB) was studied 2, 4, and 24 h postburn; and cytokine secretion (systemic and myocyte secreted cytokines, ELISA) and cardiac function were examined 24 h postburn. Vehicle-treated burn injury increased myocardial PKCepsilon/p38 MAPK expression, promoted NF-kappaB nuclear translocation, promoted TNF-alpha, IL-1beta, IL-6, and IL-10 secretion, and impaired myocardial function. SDD attenuated burn-related proinflammatory myocardial signaling, cytokine secretion, and myocardial contractile defects. Our data suggest that burn-related loss of GI barrier function and translocation of microbial products serve as upstream mediators of postburn myocardial inflammatory signaling and dysfunction.

Animals↗
PubMed1997

Of mites and men: trypsin-like proteases in the lungs.

It is no surprise to anyone who has tried to memorize clotting and complement cascades that there are a great many trypsin-like proteases-too many, it may seem. This fecund enzyme family encompasses such a range of biological roles that other important members and functions are sure to await discovery. Of known trypsin-like proteases, few, if any, are lung-specific. Nonetheless, many contribute in ways critical to lung function, such as fighting microbial invaders, regulating the formation and removal of polymerized fibrin, and rejecting tumors or transplanted tissues. Some of these enzymes, such as plasminogen activators and mast cell tryptases, are native to the lung and live where they work; identical enzymes live and work in other tissues. Other enzymes, such as most complement and hemostatic proteases, are migrants, typically born elsewhere (e.g., the liver). They pass through the lung, looking for something to do; if nothing is found, they pass on, circulating, perhaps to return later. Still others are unwanted, uninvited intruders, such as dust mite proteases. This minireview provides a selective glimpse of the lives of some of the trypsin-like enzymes at work in the lung and airways.

Animals↗
PubMed2006

Waterborne microbial risk assessment: a population-based dose-response function for Giardia spp. (E.MI.R.A study).

BACKGROUND: Dose-response parameters based on clinical challenges are frequently used to assess the health impact of protozoa in drinking water. We compare the risk estimates associated with Giardia in drinking water derived from the dose-response parameter published in the literature and the incidence of acute digestive conditions (ADC) measured in the framework of an epidemiological study in a general population. METHODS: The study combined a daily follow-up of digestive morbidity among a panel of 544 volunteers and a microbiological surveillance of tap water. The relationship between incidence of ADC and concentrations of Giardia cysts was modeled with Generalized Estimating Equations, adjusting on community, age, tap water intake, presence of bacterial indicators, and genetic markers of viruses. The quantitative estimate of Giardia dose was the product of the declared amount of drinking water intake (in L) by the logarithm of cysts concentrations. RESULTS: The Odds Ratio for one unit of dose [OR = 1.76 (95% CI: 1.21, 2.55)] showed a very good consistency with the risk assessment estimate computed after the literature dose-response, provided application of a 20 % abatement factor to the cysts counts that were measured in the epidemiological study. Doing so, a daily water intake of 2 L and a Giardia concentration of 10 cysts/100 L, would yield an estimated relative excess risk of 12 % according to the Rendtorff model, against 11 % when multiplying the baseline rate of ADC by the corresponding OR. This abatement parameter encompasses uncertainties associated with germ viability, infectivity and virulence in natural settings. CONCLUSION: The dose-response function for waterborne Giardia risk derived from clinical experiments is consistent with epidemiological data. However, much remains to be learned about key characteristics that may heavily influence quantitative risk assessment results.

Adolescent↗