Novel dichloromethane-fermenting bacteria in the Peptococcaceae family

Dichloromethane (DCM; CH2Cl2) is a toxic groundwater pollutant that also has a detrimental effect on atmospheric ozone levels. As a dense non-aqueous phase liquid, DCM migrates vertically through groundwater to low redox zones, yet information on anaerobic microbial DCM transformation remains scarce due to a lack of cultured organisms. We report here the characterisation of DCMF, the dominant organism in an anaerobic enrichment culture (DFE) capable of fermenting DCM to the environmentally benign product acetate. Stable carbon isotope experiments demonstrated that the organism assimilated carbon from DCM and bicarbonate via the Wood–Ljungdahl pathway. DCMF is the first anaerobic DCM-degrading population also shown to metabolise non-chlorinated substrates. It appears to be a methylotroph utilising the Wood–Ljungdahl pathway for metabolism of methyl groups from methanol, choline, and glycine betaine. The flux of these substrates from subsurface environments may either directly (DCM, methanol) or indirectly (choline, glycine betaine) affect the climate. Community profiling and cultivation of cohabiting taxa in culture DFE without DCMF suggest that DCMF is the sole organism in this culture responsible for substrate metabolism, while the cohabitants persist via necromass recycling. Genomic and physiological evidence support placement of DCMF in a novel genus within the Peptococcaceae family, ‘Candidatus Formimonas warabiya’.

A novel, dichloromethane-fermenting bacterium in the Peptococcaceae family, ‘Candidatus Formamonas warabiya’, gen. nov. sp. nov.

Dichloromethane (DCM; CH2Cl2) is a toxic groundwater pollutant that also has a detrimental effect on atmospheric ozone levels. As a dense non-aqueous phase liquid, DCM migrates vertically through groundwater to low redox zones, yet information on anaerobic microbial DCM transformation remains scarce due to a lack of cultured organisms. We report here the characterisation of strain DCMF, the dominant organism in an anaerobic enrichment culture (DFE) that is capable of fermenting DCM to the environmentally benign product acetate. Stable carbon isotope experiments demonstrated that the organism assimilated carbon from DCM and bicarbonate via the Wood-Ljungdahl pathway. Strain DCMF is the first anaerobic DCM-degrading bacterium also shown to metabolise non-chlorinated substrates. It appears to be a methylotroph utilising the Wood-Ljungdahl pathway for metabolism of methyl groups from methanol, choline, and glycine betaine, which has implications for the flux of climate-active compounds from subsurface environments. Community profiling and enrichment of the cohabiting taxa in culture DFE to the exclusion of strain DCMF suggest that it is the sole organism in this culture responsible for substrate metabolism, while the cohabitants persist via necromass recycling. Genomic and physiological evidence support placement of strain DCMF in a novel genus, ‘Candidatus Formamonas warabiya’.

Sustained dechlorination of vinyl chloride to ethene inDehalococcoides-enriched cultures grown without addition of exogenous vitamins and at low pH

ABSTRACTTrichloroethene (TCE) is a ubiquitous groundwater pollutant. Successful TCE bioremediation has been demonstrated at field sites using specialized microbial consortia harboring TCE-respiringDehaloccocoideswhose growth is cobalamin (vitamin B12)-dependent. Bioaugmentation cultures grown ex situ with ample exogenous vitamins in the medium and at neutral pH may become vitamin-limited or inhibited by acidic pH once injected into field sites, resulting in incomplete TCE dechlorination and accumulation of more toxic vinyl chloride (VC). Here, we report growth of theDehalococcoides-containing bioaugmentation culture KB-1 in a TCE-amended mineral medium devoid of vitamins and in a VC-amended mineral medium at low pH (6.0 and 5.5). In cultures grown without exogenous vitamins or cobalamin,Acetobacterium, which can synthesize 5,6-dimethylbenzimidazole (DMB), the lower ligand of cobalamin, andSporomusaare the dominant acetogens. At neutral pH, a growingAcetobacteriumpopulation supports complete TCE dechlorination byDehalococcoidesat millimolar levels with a substantial increase in the amount of measured cobalamin (~20-fold). Sustained dechlorination of VC to ethene was achieved at a pH as low as 5.5, yet at low pHAcetobacteriumis less abundant, potentially affecting the production of DMB and/or cobalamin. However, dechlorination activity at very low pH (< 5.0) was not stimulated by DMB supplementation, but was restored by raising pH to neutral. Assays in cell extracts revealed that vinyl chloride reductase (VcrA) activity declines significantly below pH 6.0 and is undetectable below pH 5.0. This study highlights the roles of and interplay between vitamin-producing populations and pH in microbial dechlorinating communities, and their importance for successful chlorinated ethenes bioremediation at field sites.

Use of C–Cl CSIA to elucidate origin and fate of DCM in complex contaminated field sites

We used C-Cl dual isotope analysis and microcosm studies for elucidating the origin and fate of the common groundwater pollutant dichloromethane (DCM) in two different multi-contaminant field sites in Catalonia, Spain; where DCM contamination could be the result of direct solvent releases and/or chloroform (CF) transformation. Known commercial solvents isotopic compositions as well as characteristic C-Cl dual isotope slopes from our anaerobic enrichment culture containing Dehalobacterium sp., capable of fermenting DCM, and other bacteria from the literature were used for field data interpretation.

Nanoporous networks as effective stabilisation matrices for nanoscale zero-valent iron and groundwater pollutant removal

Nanoporous networks exhibit effective stabilisation properties for nanoscale zero-valent iron (nZVI) and nZVI, with its reductive potentials and wide availability, offers degradative remediation of environmental contaminants.

Functional Genotyping of Sulfurospirillum spp. in Mixed Cultures Allowed the Identification of a New Tetrachloroethene Reductive Dehalogenase

ABSTRACTReductive dehalogenases are the key enzymes involved in the anaerobic respiration of organohalides such as the widespread groundwater pollutant tetrachloroethene. The increasing number of available bacterial genomes and metagenomes gives access to hundreds of new putative reductive dehalogenase genes that display a high level of sequence diversity and for which substrate prediction remains very challenging. In this study, we present the development of a functional genotyping method targeting the diverse reductive dehalogenases present inSulfurospirillumspp., which allowed us to unambiguously identify a new reductive dehalogenase from our tetrachloroethene-dechlorinating SL2 bacterial consortia. The new enzyme, named PceATCE, shows 92% sequence identity with the well-characterized PceA enzyme ofSulfurospirillum multivorans, but in contrast to the latter, it is restricted to tetrachloroethene as a substrate. Its apparent higher dechlorinating activity with tetrachloroethene likely allowed its selection and maintenance in the bacterial consortia among other enzymes showing broader substrate ranges. The sequence-substrate relationships within tetrachloroethene reductive dehalogenases are also discussed.