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Pathogenic Escherichia coli strains are responsible for food-borne disease outbreaks upon consumption of fresh vegetables and fruits. The aim of this study was to establish the transmission route of E. coli strain 0611, as proxy for human pathogenic E. coli, via manure, soil and plant root zones to the above-soil plant compartments. The ecological behavior of the introduced strain was established by making use of a combination of cultivation-based and molecular targeted and untargeted approaches. Strain 0611 CFUs and specific molecular targets were detected in the root zones of lettuce and leek plants, even up to 272 days after planting in the case of leek plants. However, no strain 0611 colonies were detected in leek leaves, and only in one occasion a single colony was found in lettuce leaves. Therefore, it was concluded that transmission of E. coli via manure is not the principal contamination route to the edible parts of both plant species grown under field conditions in this study. Strain 0611 was shown to accumulate in root zones of both species and metagenomic reads of this strain were retrieved from the lettuce rhizosphere soil metagenome library at a level of Log 4.11 CFU per g dry soil.

The genomes of three Golubevia isolates (BC0812, BC0850, and BC0902) that have been shown to reduce conidiation of Blumeria graminis f. sp. tritici were sequenced using a dual-platform approach. The assembled genomes will help to elucidate the molecular mechanisms underlying the biocontrol effect of this understudied group.

The genomes of three Golubevia isolates (BC0812, BC0850, and BC0902) that have been shown to reduce conidiation of Blumeria graminis f. sp. tritici were sequenced using a dual-platform approach. The assembled genomes will help to elucidate the molecular mechanisms underlying the biocontrol effect of this understudied group.

In nature anaerobic ammonium oxidation (anammox) and denitrification processes convert fixed nitrogen to gaseous nitrogen compounds, which are then released to the atmosphere. While anammox bacteria produce N2 from ammonium and nitrite, in the denitrification process nitrate and nitrite are converted to N2 and the greenhouse gas nitrous oxide (N2O). Furthermore, nitrite needed by the anammox bacteria can be supplied by nitrate reduction to nitrite. Consequently, the interplay between nitrogen-transforming microorganisms control the amount of harmless N2 or the greenhouse gas N2O released to the atmosphere. Therefore, it is important to understand the interactions of these microorganisms in the natural environment, where dynamic conditions result in fluctuating substrate concentrations. Here, we studied the interactions between the sulfide-oxidizing denitrifier Sedimenticola selenatireducens and the anammox bacterium Scalindua brodae in a bioreactor mimicking the marine environment by creating sulfide, ammonium and nitrate limitation in distinct operational phases. Through a microbial interaction, Se. selenatireducens reduced nitrate to nitrite, which together with the supplied ammonium was converted to N2 by Sc. Brodae. Using comparative transcriptomics, we determined that Sc. Brodae and Se. selenatireducens had significant responses to ammonium and nitrate limitation, respectively, indicating that the activities of these microorganisms are regulated by different nitrogen compounds.

Biocontrol agents are living organisms with the potential to suppress populations of plant pathogens or pests in a cropping system. The complex interplay between the different players and the changing environment, results in a combination of different modes of action. Here, we applied an mRNA-based systems approach to gain insight into the antagonist-pathogen-host interaction of Golubevia sp. isolates BC0812 and BC0850 with the causal agent of wheat powdery mildew, Blumeria graminis f. sp. tritici, in planta over time. Bioassays were performed on potted wheat plants (water-treated control, antagonist, pathogen, antagonist+pathogen) under controlled conditions. A significantly higher percentage of mildew conidia were parasitized after treatment with Golubevia sp. BC0812 with 26% and BC0850 with 16% compared to the water control with 1%. Differential gene expression analysis of antagonists, pathogen and host 5, 6, 7, and 11 days after inoculation (dai) with the antagonist pointed to a combination of different modes of action: An interplay of modulating plant defense responses, impairing conidiogenesis of the pathogen by scavenging H2O2, facultative hyperparasitism and nitrogen competition. Microscopic observations supported the suggested hyperparasitism as thin mycelium could be observed on Bgt conidia at 6 dai and later. Taken together the results allowed the formulation of new hypothesis regarding modes of action and the interplay between antagonist, pathogen and host. It showed that a solid molecular understanding of the antagonist-pathogen relationship over time is essential for less biased mode of action studies. Understanding this complex interplay is the basis for targeted optimization strategies and allows discovery of new potential targets and markers for future biocontrol development.

Summary Fixed nitrogen is released by anaerobic ammonium oxidation (anammox) and/or denitrification from (marine) ecosystems. Nitrite, the terminal electron acceptor of the anammox process, occurs in nature at very low concentrations and is produced via (micro)aerobic oxidation of ammonium or nitrate reduction. The coupling of sulfide-dependent denitrification to anammox is particularly interesting because besides hydrogen, sulfide is the most important reductant at the chemocline of anoxic marine basins and is abundant within sediments. Although at μM concentrations, sulfide may be toxic and inhibiting anammox activity, a denitrifying microorganism could convert sulfide and nitrate at sufficiently high rates to allow anammox bacteria to stay active despite an influx of sulfide. To test this hypothesis, a laboratory scale model system containing a co-culture of anammox bacteria and the autotrophic denitrifier Sulfurimonas denitrificans DSM1251 was started. Complementary techniques revealed that the gammaproteobacterial Sedimenticola sp. took over the intended role of Su. denitrificans. A stable coculture of anammox bacteria and Sedimenticola sp. consumed sulfide, nitrate, ammonium and CO2. Anammox bacteria contributed 65–75% to the nitrogen loss from the reactor. The cooperation between anammox and sulfide-dependent denitrification may play a significant role in environments where sulfur cycling is active and where actual sulfide concentrations stay below μM range.

Contains fulltext : 117292.pdf (Publisher’s version ) (Open Access) Anaerobic ammonium-oxidizing (anammox) bacteria are responsible for a significant portion of the loss of fixed nitrogen from the oceans, making them important players in the global nitrogen cycle. To date, marine anammox bacteria found in marine water columns and sediments worldwide belong almost exclusively to the 'Candidatus Scalindua' species, but the molecular basis of their metabolism and competitive fitness is presently unknown. We applied community sequencing of a marine anammox enrichment culture dominated by 'Candidatus Scalindua profunda' to construct a genome assembly, which was subsequently used to analyse the most abundant gene transcripts and proteins. In the S. profunda assembly, 4756 genes were annotated, and only about half of them showed the highest identity to the only other anammox bacterium of which a metagenome assembly had been constructed so far, the freshwater 'Candidatus Kuenenia stuttgartiensis'. In total, 2016 genes of S. profunda could not be matched to the K. stuttgartiensis metagenome assembly at all, and a similar number of genes in K.stuttgartiensis could not be found in S. profunda. Most of these genes did not have a known function but 98 expressed genes could be attributed to oligopeptide transport, amino acid metabolism, use of organic acids and electron transport. On the basis of the S. profunda metagenome, and environmental metagenome data, we observed pronounced differences in the gene organization and expression of important anammox enzymes, such as hydrazine synthase (HzsAB), nitrite reductase (NirS) and inorganic nitrogen transport proteins. Adaptations of Scalindua to the substrate limitation of the ocean may include highly expressed ammonium, nitrite and oligopeptide transport systems and pathways for the transport, oxidation, and assimilation of small organic compounds that may allow a more versatile lifestyle contributing to the competitive fitness of Scalindua in the marine realm.

Anaerobic ammonium-oxidizing bacteria were recently shown to use short-chain organic acids as additional energy source. The AMP-forming acetyl-CoA synthetase gene (acs) of Kuenenia stuttgartiensis, encoding an important enzyme involved in the conversion of these organic acids, was identified and heterologously expressed in Escherichia coli to investigate the activation of several substrates, that is, acetate, propionate and butyrate. The heterologously expressed ACS enzyme could complement an E. coli triple mutant deficient in all pathways of acetate activation. Activity was observed toward several short-chain organic acids, but was highest with acetate. These properties are in line with a mixotrophic growth of anammox bacteria. In addition to acs, the genome of K. stuttgartiensis contained the essential genes of an acetyl-CoA synthase/CO dehydrogenase complex and genes putatively encoding two isoenzymes of archaeal-like ADP-forming acetyl-CoA synthetase underlining the importance of acetyl-CoA as intermediate in the carbon assimilation metabolism of anammox bacteria. Electronic supplementary material The online version of this article (doi:10.1007/s00203-012-0829-7) contains supplementary material, which is available to authorized users.

We present the draft genome of anammox bacterium “ Candidatus Scalindua brodae,” which at 282 contigs is a major improvement over the highly fragmented genome assembly of related species “ Ca. Scalindua profunda” (1,580 contigs) which was previously published.

Fixed nitrogen is released by anaerobic ammonium oxidation (anammox) and/or denitrification from (marine) ecosystems. Nitrite, the terminal electron acceptor of the anammox process, occurs in nature at very low concentrations and is produced via (micro)aerobic oxidation of ammonium or nitrate reduction. The coupling of sulfide-dependent denitrification to anammox is particularly interesting because besides hydrogen, sulfide is the most important reductant at the chemocline of anoxic marine basins and is abundant within sediments. Although at μM concentrations, sulfide may be toxic and inhibiting anammox activity, a denitrifying microorganism could convert sulfide and nitrate at sufficiently high rates to allow anammox bacteria to stay active despite an influx of sulfide. To test this hypothesis, a laboratory scale model system containing a co-culture of anammox bacteria and the autotrophic denitrifier Sulfurimonas denitrificans DSM1251 was started. Complementary techniques revealed that the gammaproteobacterial Sedimenticola sp. took over the intended role of Su. denitrificans. A stable coculture of anammox bacteria and Sedimenticola sp. consumed sulfide, nitrate, ammonium and CO2 . Anammox bacteria contributed 65-75% to the nitrogen loss from the reactor. The cooperation between anammox and sulfide-dependent denitrification may play a significant role in environments where sulfur cycling is active and where actual sulfide concentrations stay below μM range.

International audience; Hydrothermally active sediments are highly productive, chemosynthetic areas which are characterized by the rapid turnover of particulate organic matter under extreme conditions in which ammonia is liberated. These systems might be suitable habitats for anaerobic ammonium oxidizing (anammox) bacteria but this has not been investigated in detail. Here we report the diversity and abundance of anammox bacteria in sediments that seep cold hydrocarbon-rich fluids and hydrothermal vent areas of the Guaymas Basin in the Cortés Sea using the unique functional anammox marker gene, hydrazine synthase (hzsA). All clones retrieved were closely associated to the "Candidatus Scalindua" genus. Phylogenetic analysis revealed two distinct clusters of hzsA sequences (Ca. Scalindua hzsA cluster I and II). Comparison of individual sequences from both clusters showed that several of these sequences had a similarity as low as 76% on nucleotide level. Based on the analysis of this phylomarker, a very high interspecies diversity within the marine anammox group is apparent. Absolute numbers of anammox bacteria in the sediments samples were determined by amplification of a 257 bp fragment of the hszA gene in a qPCR assay. The results indicate that numbers of anammox bacteria are generally higher in cold hydrocarbon-rich sediments compared to the vent areas and the reference zone. Ladderanes, lipids unique to anammox bacteria were also detected in several of the sediment samples corroborating the hzsA analysis. Due to the high concentrations of reduced sulfur compounds and its potential impact on the cycling of nitrogen we aimed to get an indication about the key players in the oxidation of sulfide in the Guaymas Basin sediments using the alpha subunit of the adenosine-5'-phosphosulfate (APS) reductase (aprA). Amplification of the aprA gene revealed a high number of gammaproteobacterial aprA genes covering the two sulfur-oxidizing bacteria aprA lineages as well as sulfate-reducers.

International audience; The complete and assembled genome sequences were determined for six strains of the alphaproteobacterial genus Methylobacterium, chosen for their key adaptations to different plant-associated niches and environmental constraints.

Many extremophilic organisms require unusual enzymes to help them survive in harsh environments. For example, acid-loving hyperthermophilic Archaea found in the bubbling mud of volcanic solfataras are able to oxidize reduced sulphur compounds. The X-ray crystal structure of a carbon disulphide (CS2) hydrolase from an Acidianus strain isolated from the Solfatara volcano near Naples, Italy, has now been determined. The enzyme, which converts CS2 into hydrogen sulphide and carbon dioxide, has a typical carbonic anhydrase fold and active site, although CO2 is not a substrate for the enzyme. This suggests that CS2 hydrolase is an example of divergent evolution, where a new enzyme has emerged through the evolution of a new quaternary structure rather than through mutations of the active site. Extremophilic organisms require specialized enzymes for their exotic metabolisms. Acid-loving thermophilic Archaea that live in the mudpots of volcanic solfataras obtain their energy from reduced sulphur compounds such as hydrogen sulphide (H2S) and carbon disulphide (CS2)1,2. The oxidation of these compounds into sulphuric acid creates the extremely acidic environment that characterizes solfataras. The hyperthermophilic Acidianus strain A1-3, which was isolated from the fumarolic, ancient sauna building at the Solfatara volcano (Naples, Italy), was shown to rapidly convert CS2 into H2S and carbon dioxide (CO2), but nothing has been known about the modes of action and the evolution of the enzyme(s) involved. Here we describe the structure, the proposed mechanism and evolution of a CS2 hydrolase from Acidianus A1-3. The enzyme monomer displays a typical β-carbonic anhydrase fold and active site, yet CO2 is not one of its substrates. Owing to large carboxy- and amino-terminal arms, an unusual hexadecameric catenane oligomer has evolved. This structure results in the blocking of the entrance to the active site that is found in canonical β-carbonic anhydrases and the formation of a single 15-A-long, highly hydrophobic tunnel that functions as a specificity filter. The tunnel determines the enzyme’s substrate specificity for CS2, which is hydrophobic. The transposon sequences that surround the gene encoding this CS2 hydrolase point to horizontal gene transfer as a mechanism for its acquisition during evolution. Our results show how the ancient β-carbonic anhydrase, which is central to global carbon metabolism, was transformed by divergent evolution into a crucial enzyme in CS2 metabolism.