Project description:Nitrification, the oxidation of ammonia via nitrite to nitrate, has always been considered to be a two-step process catalysed by chemolithoautotrophic microorganisms oxidizing either ammonia or nitrite. No known nitrifier carries out both steps, although complete nitrification should be energetically advantageous. This functional separation has puzzled microbiologists for a century. Here we report on the discovery and cultivation of a completely nitrifying bacterium from the genus Nitrospira, a globally distributed group of nitrite oxidizers. The genome of this chemolithoautotrophic organism encodes the pathways both for ammonia and nitrite oxidation, which are concomitantly activated during growth by ammonia oxidation to nitrate. Genes affiliated with the phylogenetically distinct ammonia monooxygenase and hydroxylamine dehydrogenase genes of Nitrospira are present in many environments and were retrieved on Nitrospira contigs in new metagenomes from engineered systems. These findings fundamentally change our picture of nitrification and point to completely nitrifying Nitrospira as key components of nitrogen-cycling microbial communities.
Project description:The ecophysiology of complete ammonia oxidizing Nitrospira (CMX) and their widespread occurrence in groundwater suggests that CMX bacteria have a competitive advantage over ammonia-oxidizing bacteria (AOB) and archaea (AOA) in these environments. However, the relevance of their activity from the ecosystem-level process perspective has remained unclear. We investigated oligotrophic carbonate rock aquifers as a model system to assess the contribution of CMX, AOA and AOB to nitrification and to identify the environmental drivers of their niche differentiation at different levels of ammonium and oxygen. CMX accounted for up to 95% of the ammonia oxidizer communities. Nitrification rates were positively correlated to CMX clade A-associated phylotypes and AOB affiliated with Nitrosomonas ureae. Surprisingly, short-term incubations amended with the nitrification inhibitors allylthiourea and chlorate suggested that AOB contributed more than 90% to overall ammonia oxidation, while metaproteomics analysis confirmed an active role of CMX in both ammonia and nitrite oxidation. Ecophysiological niche differentiation of CMX clades A and B, AOA and AOB was linked to their requirements for ammonium, oxygen tolerance, and metabolic versatility. Our results demonstrate that despite numerical predominance of CMX, the first step of nitrification in oligotrophic groundwater is primarily governed by AOB. Higher growth yields at lower NH4+ turnover rates and energy derived from nitrite oxidation most likely enable CMX to maintain consistently high populations. Activity measurements combined with differential inhibition allowed a refined understanding of ammonia oxidizer coexistence, competition and cooperation beyond the insights from molecular data alone.
Project description:B. cenocepacia is an opportunistic human pathogen that is particularly problematic for patients suffering from cystic fibrosis (CF). In the CF lung, bacteria grow to high densities within the viscous mucus that is limited in oxygen. Pseudomonas aeruginosa, the dominant pathogen in CF patients, is known to grow and survive under oxygen-limited to anaerobic conditions by using micro-oxic respiration, denitrification and fermentative pathways. In contrast, inspection of the genome sequences of available B. cenocepacia strains suggested that B. cenocepacia is an obligate aerobic and non-fermenting bacterium. In accordance with the bioinformatics analysis, we observed that B. cenocepacia H111 is able to grow with as little as 0.1% O2 but not under strictly anoxic conditions. Phenotypic analyses revealed that H111 produced larger amounts of biofilm, pellicle and proteases under micro-oxic conditions (0.5% - 5% O2, i.e. conditions that mimic those encountered in CF lung infection), and was more resistant to several antibiotics. RNA-Seq and shotgun proteomics analyses of cultures of B. cenocepacia H111 grown under micro-oxic and aerobic conditions showed up-regulation of genes involved in the synthesis of the exopolysaccharide (EPS) cepacian as well as several proteases, two isocitrate lyases and other genes potentially important for life in micro-oxia. Oxygen regulation in Burkholderia cenocepacia was investigated using RNA-Seq of cells grown under aerobic or micro-oxic conditions.
Project description:Gambelli et al., 2021, investigate methylomirabilis bacteria, which perform anaerobic methane oxidation coupled to nitrite reduction via an intra-aerobic pathway to produce carbon dioxide and dinitrogen gas. These bacteria possess an unusual polygonal cell shape with sharp ridges that run along the cell body. Previously, a putative surface protein layer (S-layer) was observed as the outermost cell layer of these bacteria which has been further investigated in this study. Corresponding author Laura van Niftrik, Department of Microbiology, Faculty of Science, Radboud University, Nijmegen, 6525 AJ, the Netherlands.
Project description:Chemosynthetic symbioses occur worldwide in marine habitats, but comprehensive physiological studies of chemoautotrophic bacteria thriving on animals are scarce. Stilbonematinae are coated by monocultures of thiotrophic Gammaproteobacteria. As these nematodes migrate through the redox zone, their ectosymbionts experience varying oxygen concentrations. Here, by applying omics, Raman microspectroscopy and stable isotope labeling, we investigated the effect of oxygen on the metabolism of Candidatus Thiosymbion oneisti. Unexpectedly, sulfur oxidation genes were upregulated in anoxic relative to oxic conditions, but carbon fixation genes and incorporation of 13C-labeled bicarbonate were not. Instead, several genes involved in carbon fixation, organic carbon assimilation and polyhydroxyalkanoate (PHA) biosynthesis, as well as nitrogen fixation and urea utilization were upregulated in oxic conditions. Furthermore, in the presence of oxygen, stress-related genes were upregulated together with vitamin biosynthesis genes likely necessary to withstand its deleterious effects, and fewer symbionts were detected to divide. Based on this first global physiological study of an uncultured chemosynthetic ectosymbiont, we propose that, in anoxic sediment, its proliferation is powered by anaerobic sulfur oxidation coupled to denitrification, whereas in upper layers it makes use of aerobic respiration to facilitate assimilation of carbon and nitrogen, and to survive oxidative stress. The ectosymbiont’s versatile metabolism is thus well-adapted to exploiting a highly changeable environment.
Project description:A comparative transcriptome approach was used to assess genes involved in metabolism and pathogenesis that are specifically activated during anaerobic growth of the spore-forming food-borne human pathogen Bacillus cereus ATCC 14579. Growth under anaerobic conditions in Brain Heart Infusion broth revealed a reduced growth rate and a lower yield as compared to that under aerobic conditions. Comparative transcriptome analysis of cells harvested at early- and mid-exponential growth phase, transition phase and stationary phase, subsequently showed hundreds of genes to be induced under anaerobic condition. These included novel genes identified for anaerobic growth of B. cereus, encoding metabolic pathways, such as the arginine deiminase pathway (ArcABDC), a formate dehydrogenase (FdhF) and a pyruvate fomate lyase (Pfl), and alternative respiratory proteins, such as arsenate reductases. Furthermore, the nitrosative stress response was induced in the anaerobic transition phase of growth, conceivably due to the production of nitric oxide as a by-product of nitrite and nitrate respiration. Notably, both hemolytic enzyme and enterotoxin encoding genes were activated in different oxygen limiting conditions, i.e. hemolytic enzyme encoding genes were induced during anaerobic growth, whereas enterotoxin encoding genes were induced in the transition and stationary phase of aerobic cultures reaching a high cell density. These data point to metabolic rearrangements, stress adaptation and activation of the virulent status of B. cereus under anaerobic conditions, such as encountered in the human GI-tract. B. cereus ATCC 14579 was grown in BHI in 50 ml. Aerobic in a Erlenmeyer flask, shaking at 200 rpm. Anaerobic in a closed flask, flushed with Nitrogen-gas for 30 min, also shaking at 200 rpm. Transcriptome analyses Phase compared to mid-exponential phase Anaerobic (OD600) 0.2 compared to 0.4 Early-exponential 1.0 compared to 0.4 Transition 1.1 compared to 0.4 Stationary Aerobic (OD600) 0.2 compared to 0.8 Early-exponential 4.0 compared to 0.8 Transition 8.0 compared to 0.8 Stationary Aerobic to anaerobic (OD600) Anaerobic 0.6 to aerobic 0.6
Project description:B. cenocepacia is an opportunistic human pathogen that is particularly problematic for patients suffering from cystic fibrosis (CF). In the CF lung, bacteria grow to high densities within the viscous mucus that is limited in oxygen. Pseudomonas aeruginosa, the dominant pathogen in CF patients, is known to grow and survive under oxygen-limited to anaerobic conditions by using micro-oxic respiration, denitrification and fermentative pathways. In contrast, inspection of the genome sequences of available B. cenocepacia strains suggested that B. cenocepacia is an obligate aerobic and non-fermenting bacterium. In accordance with the bioinformatics analysis, we observed that B. cenocepacia H111 is able to grow with as little as 0.1% O2 but not under strictly anoxic conditions. Phenotypic analyses revealed that H111 produced larger amounts of biofilm, pellicle and proteases under micro-oxic conditions (0.5% - 5% O2, i.e. conditions that mimic those encountered in CF lung infection), and was more resistant to several antibiotics. RNA-Seq and shotgun proteomics analyses of cultures of B. cenocepacia H111 grown under micro-oxic and aerobic conditions showed up-regulation of genes involved in the synthesis of the exopolysaccharide (EPS) cepacian as well as several proteases, two isocitrate lyases and other genes potentially important for life in micro-oxia.
Project description:Mapping the occupancy of ArcA throughout the genome of Escherchia coli MG1655 K-12 using an affinity purified antibody under anaerobic and aerobic growth conditions. As a control, we also performed ChIP-chip onArcA in a M-bM-^HM-^FarcA mutant strain of Escherchia coli MG1655 K-12. Described in the manuscript The response regulator ArcA uses a diverse binding site architechture to globally regulate carbon oxidation in E. coli Mapping of occupancy of ArcA in the genome of Escherchia coli MG1655 K-12 during anaerobic fermentation and aerobic respiration. Immunoprecipitated DNA compared to INPUT for each sample.