Classifying all reductive dehalogenase genes from organohalide respiring bacteria, including nine newly closed genomes, predicts function and conserved synteny within species.

The microbial transformation potential of 6:2 chlorinated polyfluorooctane ether sulfonate (6:2 Cl-PFESA) was explored in anaerobic microbial systems. Microbial communities from anaerobic wastewater sludge, an anaerobic digester, and anaerobic dechlorinating cultures enriched from aquifer materials reductively dechlorinated 6:2 Cl-PFESA to 6:2 hydrogen-substituted polyfluorooctane ether sulfonate (6:2 H-PFESA), which was identified as the sole metabolite by non-target analysis. Rapid and complete reductive dechlorination of 6:2 Cl-PFESA was achieved by the anaerobic dechlorinating cultures. The microbial community of the anaerobic dechlorinating cultures was impacted by 6:2 Cl-PFESA exposure. Organohalide-respiring bacteria originally present in the anaerobic dechlorinating cultures, including Geobacter, Dehalobacter, and Dehalococcoides, decreased in relative abundance over time. As the relative abundance of organohalide-respiring bacteria decreased, the rates of 6:2 Cl-PFESA dechlorination decreased, suggesting that the most likely mechanism for reductive dechlorination of 6:2 Cl-PFESA was co-metabolism rather than organohalide respiration. Reductive defluorination of 6:2 Cl-PFESA was not observed. Furthermore, 6:2 H-PFESA exhibited 5.5 times lower sorption affinity to the suspended biosolids than 6:2 Cl-PFESA, with the prospect of increased mobility in the environment. These results show the susceptibility of 6:2 Cl-PFESA to microbially mediated reductive dechlorination and the likely persistence of the product, 6:2 H-PFESA, in anaerobic environments.

Few strains of Dehalococcoides mccartyi harbour and express the vinyl chloride reductase (VcrA) that catalyzes the dechlorination of vinyl chloride (VC), a carcinogenic soil and groundwater contaminant. The vcrA operon is found on a Genomic Island (GI) and therefore believed to participate in horizontal gene transfer. To try to induce horizontal gene transfer of the vcrA-GI, we blended two enrichment cultures in medium without ammonium while providing VC. We hypothesized that these conditions would select for a mutant strain of D. mccartyi that could both fix nitrogen and respire VC. However, after more than 4 years of incubation, we found no evidence for horizontal gene transfer of the vcrA-GI. Rather, we observed VC-dechlorinating activity attributed to the trichloroethene reductase TceA. Sequencing and protein modelling revealed a mutation in the predicted active site of TceA which may have influenced substrate specificity. We also identified two nitrogen-fixing D. mccartyi strains in the KB-1 culture. The presence of multiple strains of D. mccartyi with distinct phenotypes is a feature of natural environments and certain enrichment cultures (such as KB-1) and may enhance bioaugmentation success. The fact that multiple distinct strains persist in the culture for decades and that we could not induce horizontal gene transfer of the vcrA-GI suggests that it is not as mobile as predicted, or that mobility is restricted in ways yet to be discovered to specific sub-clades of Dehalococcoides.

Vinyl chloride (VC) is a human carcinogen that accumulates in soil and groundwater due to incomplete dechlorination of chlorinated ethenes. Some strains of obligate organohalide respiring Dehalococcoides mccartyi can synthesize the VC reductase that catalyzes the dechlorination of VC to ethene. The gene encoding the VC reductase, vcrA, is found on a mobile genetic element called the vcrA-Genomic Island (GI) that may participate in horizontal gene transfer. We designed an experiment to try to induce horizontal gene transfer of the vcrA-GI by mixing two enrichment cultures: one containing the donor D. mccartyi strain with the vcrA-GI that could not fix nitrogen and the second containing the recipient strain devoid of the vcrA-GI that could fix nitrogen. Therefore, mixing the two cultures in medium without ammonium while providing VC as the sole electron acceptor was hypothesized to select for a mutant strain of D. mccartyi that could both fix nitrogen and respire VC. However, after over 4 years of incubation, no evidence for horizontal gene transfer of the vcrA-GI was found. Rather, we observed VC-dechlorinating activity attributed to the TCE reductase, TceA, in the recipient strain. We also observed that D. mccartyi can grow by scavenging low concentrations of fixed nitrogen sources. During this experiment we identified two additional D. mccartyi strains in the KB-1 TCE-enriched culture that could fix nitrogen. The presence of multiple strains of D. mccartyi with distinct phenotypes may enhance bioaugmentation success, but here it may have undermined attempts to force horizontal gene transfer of the vcrA-GI.IMPORTANCEDehalococcoides mccartyi are a powerful bioremediation tool for the degradation of chlorinated solvent contamination in soil and groundwater. Only a few D. mccartyi strains have the ability to dechlorinate toxic chlorinated compounds like vinyl chloride. Interestingly, the genetic ability to dechlorinate vinyl chloride is theorized to be shared among D. mccartyi strains. In this study we attempted to promote the genetic transfer of vinyl chloride degrading ability from one D. mccartyi strain to another. Although we did not observe this exchange, our findings suggest there may be restrictions of genetic transfer between specific clades or sub-groups of D. mccartyi strains. Developing our understanding of genetic transfer among D. mccartyi strains could allow for enhanced degradation of chlorinated solvent contamination in situ.