Project description:The community composition (in terms of abundance, distribution and contribution of diverse clades) of bacteria involved in nitrogen transformations in the oxygen minimum zones may be related to the rates of fixed N loss in these systems. The abundance of both denirifying and anammox bacteria, and the assemblage composition of denitrifying bacteria were investigated in the Eastern Tropical South Pacific and the Arabian Sea using assays based on molecular markers for the two groups of bacteria. The abundance and distribution of bacteria associated with the fixed N removal processes denitrification and anammox were investigated using quantitative PCR for genes encoding nitrite reductase (nirK and nirS) in denitrifying bacteria and hydrazine oxidase(hzo) and 16S rRNA genesin anammox bacteria. All of these genes had depth distributions with maxima associated with the secondary nitrite maximum in low oxygen waters. NirS was mch more abundant than nirK, and much more abundant than the 16S rRNA gene from anammox bacteria. The ratio of hzo:16S rRNA for anammox was low and variable implying greater unexplored diversity in the the hzo gene. Assemblage composition of the abundant nirS-type denitrifiers was evaluated using a funcitonal gene microarray. Of the nirS archetypes represented on the microarray, very few occurred speficically in one region or depth interval, but the assemblages varied significantly. Community composition of denitrifiers based on microarray analysis of the nirS gene was most different between geographical regions. Within each region, the surface layer and OMZ assemblages clustered distinctly. Thus, in addition to spatial and temporal variation in denitrificaiton and anammox rates, both microbial abundance and community composition also vary between OMZ regions and depths.
Project description:Freshwater ecosystems can be largely affected by neighboring agriculture fields where potential fertilizer nitrate run-off may leach into surrounding water bodies. To counteract this eutrophic driver, farmers often utilize denitrifying woodchip bioreactors (WBRs) in which a consortium of microorganisms convert the nitrate into nitrogen-gases in anoxia, fueled by the degradation of lignocellulose. Polysaccharide-degrading strategies have been well-described for various aerobic and anaerobic systems, including the use of carbohydrate-active enzymes, utilization of lytic polysaccharide monooxygenases (LPMOs) and other redox enzymes, as well as the use of cellulosomes and polysaccharide utilization loci. However, for denitrifying microorganisms, the lignocellulose-degrading strategies remain largely unknown. Here, we have applied a combination of enrichment techniques, gas measurements, multi-omics approaches, and amplicon sequencing of fungal ITS and procaryotic 16S rRNA genes to highlight microbial drivers for lignocellulose transformation in woodchip bioreactors with the aim to provide an in-depth characterization of the indigenous microorganisms and their active enzymes. Our findings highlight a microbial community enriched for lignocellulose-degrading denitrifiers with key players from Giesbergeria, Cellulomonas, Azonexus, and UBA5070, including polysaccharide utilization loci from Bacteroidetes. A wide substrate specificity is observed among the many expressed carbohydrate active enzymes (CAZymes), evidencing a swift degradation of lignocellulose, including even enzymes with auxiliary activities whose functionality is still puzzling under strict anaerobic conditions.
Project description:The community composition (in terms of abundance, distribution and contribution of diverse clades) of bacteria involved in nitrogen transformations in the oxygen minimum zones may be related to the rates of fixed N loss in these systems. The abundance of both denirifying and anammox bacteria, and the assemblage composition of denitrifying bacteria were investigated in the Eastern Tropical South Pacific and the Arabian Sea using assays based on molecular markers for the two groups of bacteria. The abundance and distribution of bacteria associated with the fixed N removal processes denitrification and anammox were investigated using quantitative PCR for genes encoding nitrite reductase (nirK and nirS) in denitrifying bacteria and hydrazine oxidase(hzo) and 16S rRNA genesin anammox bacteria. All of these genes had depth distributions with maxima associated with the secondary nitrite maximum in low oxygen waters. NirS was mch more abundant than nirK, and much more abundant than the 16S rRNA gene from anammox bacteria. The ratio of hzo:16S rRNA for anammox was low and variable implying greater unexplored diversity in the the hzo gene. Assemblage composition of the abundant nirS-type denitrifiers was evaluated using a funcitonal gene microarray. Of the nirS archetypes represented on the microarray, very few occurred speficically in one region or depth interval, but the assemblages varied significantly. Community composition of denitrifiers based on microarray analysis of the nirS gene was most different between geographical regions. Within each region, the surface layer and OMZ assemblages clustered distinctly. Thus, in addition to spatial and temporal variation in denitrificaiton and anammox rates, both microbial abundance and community composition also vary between OMZ regions and depths. Two color array (Cy3 and Cy5): the universal standard 20-mer oligo is printed to the slide with a 70-mer oligo (an archetype). Environmental DNA sequences (fluoresced with Cy3) within 15% of the 70-mer conjugated to a 20-mer oligo (fluoresced with Cy5) complementary to the universal standard will bind to the oligo probes on the array. Signal is the ratio of Cy3 to Cy5. Three replicate probes were printed for each archetype. Two replicate arrays were run on duplicate targets.
Project description:Resistance to agricultural fungicides in the field has created a need for discovering fungicides with new modes of action. DNA microarrays, because they provide information on expression of many genes simultaneously, could help to identify the modes of action. To begin an expression pattern database for agricultural fungicides, transcriptional patterns of Saccharomyces cerevisiae strain S288C genes were analysed following 2-h treatments with I50 concentrations of ergosterol biosynthesis inhibitors commonly used against plant pathogenic fungi. Eight fungicides, representing three classes of ergosterol biosynthesis inhibitors, were tested. To compare gene expression in response to a fungicide with a completely different mode of action, a putative methionine biosynthesis inhibitor (MBI) was also tested. Expression patterns of ergosterol biosynthetic genes supported the roles of Class I and Class II inhibitors in affecting ergosterol biosynthesis, confirmed that the putative MBI did not affect ergosterol biosynthesis, and strongly suggested that in yeast, the Class III inhibitor did not affect ergosterol biosynthesis. The MBI affected transcription of three genes involved in methionine metabolism, whereas there were essentially no effects of ergosterol synthesis inhibitors on methionine metabolism genes. There were no consistent patterns in other up- or downregulated genes between fungicides. These results suggest that inspection of gene response patterns within a given pathway may serve as a useful first step in identifying possible modes of action of fungicides. agricultural sterol biosynthesis inhibitor fungicides. Keywords = agriculture Keywords = ergosterol Keywords = methionine Keywords = fungicide Keywords = Saccharomyces cerevisiae S288C Keywords = biosynthesis
Project description:Global gene expression was compared between Nitrosomonas europaea wild-type and a nitrite reductase-deficient mutant using a genomic microarray. Keywords: wild-type to mutant global transcriptome comparison
Project description:Nitrogen metabolism in Aspergillus nidulans is subject to regulation by the GATA transcription factor AreA which is required for the utilization of a wide range of nitrogen sources other than glutamine or ammonium. The level of AreA activity is regulated by intracellular glutamine levels that vary in response to nitrogen supplementation. For nitrate assimilation, which involves two transporters (CrnA, CrnB), nitrate reductase (NiaD) and nitrite reductase (NiiA), the respective genes are subject to regulation at the level of transcription, including nitrogen metabolite repression mediated by AreA and induction mediated by nitrite or nitrate, mediated by a second transcription factor, NirA. Both transcription factors act synergistically to regulate the expression of all four structural genes when nitrogen is limiting or either nitrate or nitrite is available. In this study we dissect the nitrogen limitation effect mediated by AreA form the nitrate/nitrite specific effect mediated by NirA on the transcriptome level. Keywords: Nitrate/nitrogen limitation response