ABSTRACT Chloroflexi small-subunit (SSU) rRNA gene sequences are frequently recovered from subseafloor environments, but the metabolic potential of the phylum is poorly understood. The phylum Chloroflexi is represented by isolates with diverse metabolic strategies, including anoxic phototrophy, fermentation, and reductive dehalogenation; therefore, function cannot be attributed to these organisms based solely on phylogeny. Single-cell genomics can provide metabolic insights into uncultured organisms, like the deep-subsurface Chloroflexi . Nine SSU rRNA gene sequences were identified from single-cell sorts of whole-round core material collected from the Okinawa Trough at Iheya North hydrothermal field as part of Integrated Ocean Drilling Program (IODP) expedition 331 (Deep Hot Biosphere). Previous studies of subsurface Chloroflexi single amplified genomes (SAGs) suggested heterotrophic or lithotrophic metabolisms and provided no evidence for growth by reductive dehalogenation. Our nine Chloroflexi SAGs (seven of which are from the order Anaerolineales ) indicate that, in addition to genes for the Wood-Ljungdahl pathway, exogenous carbon sources can be actively transported into cells. At least one subunit for pyruvate ferredoxin oxidoreductase was found in four of the Chloroflexi SAGs. This protein can provide a link between the Wood-Ljungdahl pathway and other carbon anabolic pathways. Finally, one of the seven Anaerolineales SAGs contains a distinct reductive dehalogenase homologous ( rdhA ) gene. IMPORTANCE Through the use of single amplified genomes (SAGs), we have extended the metabolic potential of an understudied group of subsurface microbes, the Chloroflexi . These microbes are frequently detected in the subsurface biosphere, though their metabolic capabilities have remained elusive. In contrast to previously examined Chloroflexi SAGs, our genomes (several are from the order Anaerolineales ) were recovered from a hydrothermally driven system and therefore provide a unique window into the metabolic potential of this type of habitat. In addition, a reductive dehalogenase gene ( rdhA ) has been directly linked to marine subsurface Chloroflexi , suggesting that reductive dehalogenation is not limited to the class Dehalococcoidia . This discovery expands the nutrient-cycling and metabolic potential present within the deep subsurface and provides functional gene information relating to this enigmatic group.
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Anaerobic Microbial Metabolism of Dichloroacetate
ABSTRACT Dichloroacetate (DCA) commonly occurs in the environment due to natural production and anthropogenic releases, but its fate under anoxic conditions is uncertain. Mixed culture RM comprising “ Candidatus Dichloromethanomonas elyunquensis” strain RM utilizes DCA as an energy source, and the transient formation of formate, H 2 , and carbon monoxide (CO) was observed during growth. Only about half of the DCA was recovered as acetate, suggesting a fermentative catabolic route rather than a reductive dechlorination pathway. Sequencing of 16S rRNA gene amplicons and 16S rRNA gene-targeted quantitative real-time PCR (qPCR) implicated “ Candidatus Dichloromethanomonas elyunquensis” strain RM in DCA degradation. An ( S )-2-haloacid dehalogenase (HAD) encoded on the genome of strain RM was heterologously expressed, and the purified HAD demonstrated the cofactor-independent stoichiometric conversion of DCA to glyoxylate at a rate of 90 ± 4.6 nkat mg −1 protein. Differential protein expression analysis identified enzymes catalyzing the conversion of DCA to acetyl coenzyme A (acetyl-CoA) via glyoxylate as well as enzymes of the Wood-Ljungdahl pathway. Glyoxylate carboligase, which catalyzes the condensation of two molecules of glyoxylate to form tartronate semialdehyde, was highly abundant in DCA-grown cells. The physiological, biochemical, and proteogenomic data demonstrate the involvement of an HAD and the Wood-Ljungdahl pathway in the anaerobic fermentation of DCA, which has implications for DCA turnover in natural and engineered environments, as well as the metabolism of the cancer drug DCA by gut microbiota. IMPORTANCE Dichloroacetate (DCA) is ubiquitous in the environment due to natural formation via biological and abiotic chlorination processes and the turnover of chlorinated organic materials (e.g., humic substances). Additional sources include DCA usage as a chemical feedstock and cancer drug and its unintentional formation during drinking water disinfection by chlorination. Despite the ubiquitous presence of DCA, its fate under anoxic conditions has remained obscure. We discovered an anaerobic bacterium capable of metabolizing DCA, identified the enzyme responsible for DCA dehalogenation, and elucidated a novel DCA fermentation pathway. The findings have implications for the turnover of DCA and the carbon and electron flow in electron acceptor-depleted environments and the human gastrointestinal tract.
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- Award ID(s):
- 1831599
- NSF-PAR ID:
- 10352594
- Editor(s):
- Lovley, Derek R.
- Date Published:
- Journal Name:
- mBio
- Volume:
- 12
- Issue:
- 2
- ISSN:
- 2150-7511
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1128/mBio.00537-21
