It is now well accepted that the gut microbiota contributes to our health. However, what determines the microbiota composition is still unclear. Whereas it might be expected that the intestinal niche would be dominant in shaping the microbiota, studies in vertebrates have repeatedly demonstrated dominant effects of external factors such as host diet and environmental microbial diversity. Hypothesizing that genetic variation may interfere with discerning contributions of host factors, we turned to Caenorhabditis elegans as a new model, offering the ability to work with genetically homogenous populations. Deep sequencing of 16S rDNA was used to characterize the (previously unknown) worm gut microbiota as assembled from diverse produce-enriched soil environments under laboratory conditions. Comparisons of worm microbiotas with those in their soil environment revealed that worm microbiotas resembled each other even when assembled from different microbial environments, and enabled defining a shared core gut microbiota. Community analyses indicated that species assortment in the worm gut was non-random and that assembly rules differed from those in their soil habitat, pointing at the importance of competitive interactions between gut-residing taxa. The data presented fills a gap in C. elegans biology. Furthermore, our results demonstrate a dominant contribution of the host niche in shaping the gut microbiota.

2,4,6-Trinitrotoluene (TNT) is released in nature from manufacturing or demilitarization facilities but also after munitions firing/detonation or leakage from explosive remnants of war. Due to its toxicity and recalcitrance, life cycle of TNT-containing products and bioremediation are critical issues. As TNT is a strongly electron-deficient aromatic with a positive molecular quadrupole moment and three electrophilic nitro groups, its environmental fate is contingent upon specific sorptive electron donor-acceptor interactions and nucleophilic, reductive (bio)transformations. The microbial degradation of TNT is governed by cometabolism and therefore depends on the growth substrate(s) available in contaminated environments. Long considered an ecotoxicological safety endpoint, the immobilization of TNT metabolites derived from nitro moiety reduction in soil is controversial because they preferentially bind to the dissolved soil organic matter which can be released into surface and groundwaters. The ever-growing biochemical knowledge of TNT degradation has made bioaugmentation and phytoremediation attractive alternatives. While the discovery and engineering of microorganisms with novel/improved degradative abilities are very challenging, the deciphering of the physiological roles of promiscuous enzymes involved in TNT biodegradation, such as type II hydride transferases of the Old Yellow Enzyme family, opens new perspectives for bioremediation. Finally, transgenic plants have enabled effective phytoremediation at the field scale, which is emerging as the preferable in situ option to rehabilitate TNT-contaminated sites.

bDehalococcoides mccartyi 195 (strain 195) and Syntrophomonas wolfei were grown in a sustainable syntrophic coculture using butyrate as an electron donor and carbon source and trichloroethene (TCE) as an electron acceptor. The maximum dechlorination rate (9.9 ؎ 0.1 mol day ؊1 ) and cell yield [(1.1 ؎ 0.3) ؋ 10 8 cells mol ؊1 Cl ؊ ] of strain 195 maintained in coculture were, respectively, 2.6 and 1.6 times higher than those measured in the pure culture. The strain 195 cell concentration was about 16 times higher than that of S. wolfei in the coculture. Aqueous H 2 concentrations ranged from 24 to 180 nM during dechlorination and increased to 350 ؎ 20 nM when TCE was depleted, resulting in cessation of butyrate fermentation by S. wolfei with a theoretical Gibbs free energy of ؊13.7 ؎ 0.2 kJ mol ؊1 . Carbon monoxide in the coculture was around 0.06 mol per bottle, which was lower than that observed for strain 195 in isolation. The minimum H 2 threshold value for TCE dechlorination by strain 195 in the coculture was 0.6 ؎ 0.1 nM. Cell aggregates during syntrophic growth were observed by scanning electron microscopy. The interspecies distances to achieve H 2 fluxes required to support the measured dechlorination rates were predicted using Fick's law and demonstrated the need for aggregation. Filamentous appendages and extracellular polymeric substance (EPS)-like structures were present in the intercellular spaces. The transcriptome of strain 195 during exponential growth in the coculture indicated increased ATP-binding cassette transporter activities compared to the pure culture, while the membrane-bound energy metabolism related genes were expressed at stable levels. Groundwater contamination by trichloroethene (TCE), a potential human carcinogen, poses a serious threat to human health and can lead to the generation of vinyl chloride (VC), which is a known human carcinogen (1). Strains of Dehalococcoides mccartyi are the only known bacteria that can completely degrade TCE to the benign end product ethene. Biostimulation of indigenous Dehalococcoides spp. and bioaugmentation using Dehalococcoides-containing cultures are recognized as the most reliable in situ bioremediation technologies resulting in the complete dechlorination of TCE to ethene (2). However, the mechanisms that regulate the activity of D. mccartyi within natural ecosystems and shape its functional robustness in disturbed environments are poorly understood due to multiscale microbial community complexity and heterogeneity of biogeochemical processes involved in the sequential degradation (3, 4). D. mccartyi exhibits specific restrictive metabolic requirements for a variety of exogenous compounds, such as hydrogen, acetate, corrinoids, biotin, and thiamine, which can be supplied by other microbial genera through a complex metabolic network (1,(5)(6)(7)(8). Therefore, the growth of D. mccartyi is more robust within functionally diverse microbial communities that are deterministically assembled than in pure cultures (5,8,9). Previous studies have shown t...

Increasing pollution of water and soils by xenobiotic compounds has led in the last few decades to an acute need for understanding the impact of toxic compounds on microbial populations, the catabolic degradation pathways of xenobiotics and the set-up and improvement of bioremediation processes. Recent advances in molecular techniques, including high-throughput approaches such as microarrays and metagenomics, have opened up new perspectives and pointed towards new opportunities in pollution abatement and environmental management. Compared with traditional molecular techniques dependent on the isolation of pure cultures in the laboratory, microarrays and metagenomics allow specific environmental questions to be answered by exploring and using the phenomenal resources of uncultivable and uncharacterized micro-organisms. This paper reviews the current potential of microarrays and metagenomics to investigate the genetic diversity of environmentally relevant micro-organisms and identify new functional genes involved in the catabolism of xenobiotics.

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Escherichia coli grew aerobically with 2,4,6-trinitrotoluene (TNT) as sole nitrogen source and caused TNT's partial denitration. This reaction was enhanced in nongrowing cell suspensions with 0.516 mol nitrite released per mol TNT. Cell extracts denitrated TNT in the presence of NAD(P)H. Isomers of amino-dimethyl-tetranitrobiphenyl were detected and confirmed with U-15 N-labeled TNT.2,4,6-Trinitrotoluene (TNT) is recalcitrant to microbial degradation. Denitration (defined as the release of nitrite) is a critical step for further mineralization of TNT (19). A welldescribed TNT denitration pathway involves a nucleophilic addition of hydride ions to the aromatic ring with subsequent nitrite release. Three enzymes performing this addition in the presence of NAD(P)H have been characterized so far: pentaerythritol tetranitrate reductase of Enterobacter cloacae PB2 (9), xenobiotic reductase B (XenB) of Pseudomonas fluorescens I-C (15), and N-ethylmaleimide (NEM) reductase of Escherichia coli (21). In vitro denitration of TNT with purified NEM reductase was described by Williams et al. (21), but the authors did not provide quantitative data with E. coli cells. Also, several reports have described the reduction of TNT by E. coli but not its denitration (6,14,22,23). Only one recent study has mentioned denitration of TNT by E. coli, but no quantitative data were provided and TNT was not the sole nitrogen source (13). The objectives of this study were to determine the kinetics of TNT denitration by E. coli and identify TNT denitrated metabolites.E. coli strains EPI300 (Epicentre Technologies, Madison, WI) and LK111 (24) were routinely cultivated at 37°C in LuriaBertani (LB) broth. Cells were harvested at mid-exponential phase and washed three times with phosphate-buffered saline (containing, per liter, 7 g of Na 2 HPO 4 ·12H 2 O, 3 g of KH 2 PO 4 , 1 g of NaCl). Cells were resuspended in 20 ml of phosphatebuffered saline and used for TNT biodegradation assays.TNT was obtained from Nobel Explosives (Châtelet, Belgium) and was 99.5% pure by high-performance liquid chromatography. Growing cell experiments were carried out in modified mineral salts medium (10) containing 20 mM of glycerol or glucose and TNT as the sole nitrogen source. The medium was inoculated at an optical density at 600 nm (OD 600 ) of 0.025 and incubated at 37°C and 250 rpm. Controls consisted of flasks without TNT, flasks without cells, flasks without a carbon source, and flasks with 200 ppm of Hg 2 Cl 2 and without a carbon source. Nitrite, TNT, and metabolites were quantitatively determined as previously described (7).Bacterial growth of E. coli EPI300 was observed with glycerol and TNT (606 M on the basis of high-performance liquid chromatography analysis) as the sole nitrogen source (Fig. 1A). The growth was relatively fast over the first 26 h, reaching a plateau of 0.120 OD 600 units after 117 h of incubation. With glucose and TNT (588 M), the bacterial growth profile was similar but the OD 600 reached 0.320 after 117 h (Fig. 1B). Without TNT, no si...

To gain insight into the impact of 2,4,6-trinitrotoluene (TNT) on soil microbial communities, we characterized the bacterial community of several TNT-contaminated soils from two sites with different histories of contamination and concentrations of TNT. The amount of extracted DNA, the total cell counts and the number of CFU were lower in the TNT-contaminated soils. Analysis of soil bacterial diversity by DGGE showed a predominance of Pseudomonadaceae and Xanthomonadaceae in the TNT-contaminated soils, as well as the presence of Caulobacteraceae. CFU from TNT-contaminated soils were identified as Pseudomonadaceae, and, to a lesser extent, Caulobacteraceae. Finally, a pristine soil was spiked with different concentrations of TNT and the soil microcosms were incubated for 4 months. The amount of extracted DNA decreased in the microcosms with a high TNT concentration [1.4 and 28.5 g TNT/kg (dry wt) of soil] over the incubation period. After 7 days of incubation of these soil microcosms, there was already a clear shift of their original flora towards a community dominated by Pseudomonadaceae, Xanthomonadaceae, Comamonadaceae and Caulobacteraceae. These results indicate that TNT affects soil bacterial diversity by selecting a narrow range of bacterial species that belong mostly to Pseudomonadaceae and Xanthomonadaceae.

The denitration of 2,4,6-trinitrotoluene (TNT) can produce mono- or dinitro aromatic compounds susceptible to microbial mineralization. In the present study, denitration of TNT and other nitro aromatic compounds was investigated with a solid-phase extract obtained from the culture supernatant of Pseudomonas aeruginosa ESA-5 grown on a chemically defined aerobic medium. When the C18 solid-phase extract containing extracellular catalysts (EC) was incubated with TNT and NAD(P)H, we observed a significant release of nitrite. The concentration of nitrite released in the reaction medium was strongly dependent on the concentration of NAD(P)H and EC. Denitration also occurred with two TNT-related molecules, 2,4,6-trinitrobenzaldehyde, and 2,4,6-trinitrobenzyl alcohol. The release of nitrite was coupled with the formation of two polar metabolites, and mass spectrometry analyses indicated that each of these compounds had lost two nitro groups from the trinitro aromatic parent molecule. During this process, the production of toxic reduced TNT metabolites was minimal. The incubation of EC with TNT, NAD(P)H, and specific scavengers of reactive oxygen species suggested the involvement of superoxide radicals (O2*-) and hydrogen peroxide in the denitration process. Results obtained in this study reveal for the first time that extracellular small-molecular-weight substance(s) of bacterial origin can serve as green catalyst(s) to initiate TNT denitration. In addition, this study gives clear evidence for the production of a TNT metabolite bearing a single nitro groupfollowing a denitration reaction with catalyst(s) of biotic origin.