Sulfonamide Per- and Polyfluoroalkyl Substances Can Impact Microorganisms Used in Aromatic Hydrocarbon and Trichloroethene Bioremediation.

Per- and polyfluoroalkyl substances (PFASs) from aqueous film forming foams (AFFFs) can hinder bioremediation of co-contaminants such as trichloroethene (TCE) and benzene, toluene, ethylbenzene, and xylene (BTEX). Anaerobic dechlorination can require bioaugmentation of Dehalococcoides, and for BTEX, oxygen is often sparged to stimulate in situ aerobic biodegradation. We tested PFAS inhibition to TCE and BTEX bioremediation by exposing an anaerobic TCE-dechlorinating coculture, an aerobic BTEX-degrading enrichment culture, and an anaerobic toluene-degrading enrichment culture to n-dimethyl perfluorohexane sulfonamido amine (AmPr-FHxSA), perfluorohexane sulfonamide (FHxSA), perfluorohexanesulfonic acid (PFHxS), or nonfluorinated surfactant sodium dodecyl sulfate (SDS). The anaerobic TCE-dechlorinating coculture was resistant to individual PFAS exposures but was inhibited by >1000× diluted AFFF. FHxSA and AmPr-FHxSA inhibited the aerobic BTEX-degrading enrichment. The anaerobic toluene-degrading enrichment was not inhibited by AFFF or individual PFASs. Increases in amino acids in the anaerobic TCE-dechlorinating coculture compared to the control indicated stress response, whereas the BTEX culture exhibited lower concentrations of all amino acids upon exposure to most surfactants (both fluorinated and nonfluorinated) compared to the control. These data suggest the main mechanisms of microbial toxicity are related to interactions with cell membrane synthesis as well as protein stress signaling.

Syntrophic Interactions Ameliorate Arsenic Inhibition of Solvent-Dechlorinating Dehalococcoides mccartyi.

Interactions and nutrient exchanges among members of microbial communities are important for understanding functional relationships in environmental microbiology. We can begin to elucidate the nature of these complex systems by taking a bottom-up approach utilizing simplified, but representative, community members. Here, we assess the effects of a toxic stress event, the addition of arsenite (As(III)), on a syntrophic co-culture containing lactate-fermenting Desulfovibrio vulgaris Hildenborough and solvent-dechlorinating Dehalococcoides mccartyi strain 195. Arsenic and trichloroethene (TCE) are two highly prevalent groundwater contaminants in the United States, and the presence of bioavailable arsenic is of particular concern at remediation sites in which reductive dechlorination has been employed. While we previously showed that low concentrations of arsenite (As(III)) inhibit the keystone TCE-reducing microorganism, D. mccartyi, this study reports the utilization of physiological analysis, transcriptomics, and metabolomics to assess the effects of arsenic on the metabolisms, gene expression, and nutrient exchanges in the described co-culture. It was found that the presence of D. vulgaris ameliorated arsenic stress on D. mccartyi, improving TCE dechlorination under arsenic-contaminated conditions. Nutrient and amino acid export by D. vulgaris may be a stress-ameliorating exchange in this syntrophic co-culture under arsenic stress, based on upregulation of transporters and increased extracellular nutrients like sarcosine and ornithine. These results broaden our knowledge of microbial community interactions and will support the further development and implementation of robust bioremediation strategies at multi-contaminant sites.

Fate of Dissolved Nitrogen in a Horizontal Levee: Seasonal Fluctuations in Nitrate Removal Processes.

Horizontal levees are a nature-based approach for removing nitrogen from municipal wastewater effluent while simultaneously providing additional benefits, such as flood control. To assess nitrogen removal mechanisms and the efficacy of a horizontal levee, we monitored an experimental system receiving nitrified municipal wastewater effluent for 2 years. Based on mass balances and microbial gene abundance data, we determined that much of the applied nitrogen was most likely removed by heterotrophic denitrifiers that consumed labile organic carbon from decaying plants and added wood chips. Fe(III) and sulfate reduction driven by decay of labile organic carbon also produced Fe(II) sulfide minerals. During winter months, when heterotrophic activity was lower, strong correlations between sulfate release and nitrogen removal suggested that autotrophic denitrifiers oxidized Fe(II) sulfides using nitrate as an electron acceptor. These trends were seasonal, with Fe(II) sulfide minerals formed during summer fueling denitrification during the subsequent winter. Overall, around 30% of gaseous nitrogen losses in the winter were attributable to autotrophic denitrifiers. To predict long-term nitrogen removal, we developed an electron-transfer model that accounted for the production and consumption of electron donors. The model indicated that the labile organic carbon released from wood chips may be capable of supporting nitrogen removal from wastewater effluent for several decades with sulfide minerals, decaying vegetation, and root exudates likely sustaining nitrogen removal over a longer timescale.

Biotransformation of 6:2 Fluorotelomer Thioether Amido Sulfonate in Aqueous Film-Forming Foams under Nitrate-Reducing Conditions.

Despite the prevalence of nitrate reduction in groundwater, the biotransformation of per- and polyfluoroalkyl substances (PFAS) under nitrate-reducing conditions remains mostly unknown compared with aerobic or strong reducing conditions. We constructed microcosms under nitrate-reducing conditions to simulate the biotransformation occurring at groundwater sites impacted by aqueous film-forming foams (AFFFs). We investigated the biotransformation of 6:2 fluorotelomer thioether amido sulfonate (6:2 FtTAoS), a principal PFAS constituent of several AFFF formulations using both quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) and qualitative high-resolution mass spectrometry analyses. Our results reveal that the biotransformation rates of 6:2 FtTAoS under nitrate-reducing conditions were about 10 times slower than under aerobic conditions, but about 2.7 times faster than under sulfate-reducing conditions. Although minimal production of 6:2 fluorotelomer sulfonate and the terminal perfluoroalkyl carboxylate, perfluorohexanoate was observed, fluorotelomer thioether and sulfinyl compounds were identified in the aqueous samples. Evidence for the formation of volatile PFAS was obtained by mass balance analysis using the total oxidizable precursor assay and detection of 6:2 fluorotelomer thiol by gas chromatography-mass spectrometry. Our results underscore the complexity of PFAS biotransformation and the interactions between redox conditions and microbial biotransformation activities, contributing to the better elucidation of PFAS environmental fate and impact.

Biological and Chemical Transformation of the Six-Carbon Polyfluoroalkyl Substance N-Dimethyl Ammonio Propyl Perfluorohexane Sulfonamide (AmPr-FHxSA).

Sites impacted by aqueous film-forming foam (AFFF) contain co-contaminants that can stimulate biotransformation of polyfluoroalkyl substances. Here, we compare how microbial enrichments from AFFF-impacted soil amended with diethyl glycol monobutyl ether (found in AFFF), aromatic hydrocarbons (present in co-released fuels), acetate, and methane (substrates used or formed during bioremediation) impact the aerobic biotransformation of an AFFF-derived six-carbon electrochemical fluorination (ECF) precursor N-dimethyl ammonio propyl perfluorohexane sulfonamide (AmPr-FHxSA). We found that methane- and acetate-oxidizing cultures resulted in the highest yields of identifiable products (38 and 30%, respectively), including perfluorohexane sulfonamide (FHxSA) and perfluorohexane sulfonic acid (PFHxS). Using these data, we propose and detail a transformation pathway. Additionally, we examined chemical oxidation products of AmPr-FHxSA and FHxSA to provide insights on remediation strategies for AmPr-FHxSA. We demonstrate mineralization of these compounds using the sulfate radical and test their transformation during the total oxidizable precursor (TOP) assay. While perfluorohexanoic acid accounted for over 95% of the products formed, we demonstrate here for the first time two ECF-based precursors, AmPr-FHxSA and FHxSA, that produce PFHxS during the TOP assay. These findings have implications for monitoring poly- and perfluoroalkyl substances during site remediation and application of the TOP assay at sites impacted by ECF-based precursors.

Effects of Arsenic on Trichloroethene-Dechlorination Activities of Dehalococcoides mccartyi 195.

Arsenic and trichloroethene (TCE) are among the most prevalent groundwater contaminants in the United States. Co-contamination of these two compounds has been detected at 63% of current TCE-contaminated National Priorities List sites. When in situ TCE reductive dechlorination is stimulated by the addition of fermentable substrates to generate a reducing environment, the presence of arsenic can be problematic because of the potential for increased mobilization and toxicity caused by the reduction of arsenate [As(V)] to arsenite [As(III)]. This study assesses the effects of arsenic exposure on the TCE-dechlorinating activities of Dehalococcoides mccartyi strain 195. Our results indicate that 9.1 µM As(III) caused a 50% decrease in D. mccartyi cell growth. While As(V) concentrations up to 200 µM did not initially impact TCE dechlorination, inhibition was observed in cultures amended with 200 µM As(V) and 100 µM As(V) in 12 and 17 days, respectively, corresponding with the accumulation of As(III). Transcriptomic and metabolomic analyses were performed to evaluate cellular responses to both As(V) and As(III) stress. Amendment of amino acids enhanced arsenic tolerance of D. mccartyi. Results from this study improve our understanding of potential inhibitions of D. mccartyi metabolism caused by arsenic and can inform the design of bioremediation strategies at co-contaminated sites.

A novel Chromatiales bacterium is a potential sulfide oxidizer in multiple orders of marine sponges.

Sponges are benthic filter feeders that play pivotal roles in coupling benthic-pelagic processes in the oceans that involve transformation of dissolved and particulate organic carbon and nitrogen into biomass. While the contribution of sponge holobionts to the nitrogen cycle has been recognized in past years, their importance in the sulfur cycle, both oceanic and physiological, has only recently gained attention. Sponges in general, and Theonella swinhoei in particular, harbour a multitude of associated microorganisms that could affect sulfur cycling within the holobiont. We reconstructed the genome of a Chromatiales (class Gammaproteobacteria) bacterium from a metagenomic sequence dataset of a T. swinhoei-associated microbial community. This relatively abundant bacterium has the metabolic capability to oxidize sulfide yet displays reduced metabolic potential suggestive of its lifestyle as an obligatory symbiont. This bacterium was detected in multiple sponge orders, according to similarities in key genes such as 16S rRNA and polyketide synthase genes. Due to its sulfide oxidation metabolism and occurrence in many members of the Porifera phylum, we suggest naming the newly described taxon Candidatus Porisulfidus.

Growth of Dehalococcoides mccartyi species in an autotrophic consortium producing limited acetate.

The dechlorinating Dehalococcoides mccartyi species requires acetate as carbon source, but little is known on its growth under acetate limiting conditions. In this study, we observed growth and dechlorination of a D. mccartyi-containing mixed consortium in a fixed-carbon-free medium with trichloroethene in the aqueous phase and H2/CO2 in the headspace. Around 4 mM formate was produced by day 40, while acetate was constantly below 0.05 mM. Microbial community analysis of the consortium revealed dominance by D. mccartyi and Desulfovibrio sp. (57 and 22% 16S rRNA gene copies, respectively). From this consortium, Desulfovibrio sp. strain F1 was isolated and found to produce formate and acetate (1.2 mM and 48 µM, respectively, by day 24) when cultivated alone in the above mentioned medium without trichloroethene. An established co-culture of strain F1 and D. mccartyi strain 195 demonstrated that strain 195 could grow and dechlorinate using acetate produced by strain F1; and that acetate was constantly below 25 µM in the co-culture. To verify that such low level of acetate is utilizable by D. mccartyi, we cultivated strain 195 alone under acetate-limiting conditions and found that strain 195 consumed acetate to below detection (5 µM). Based on the acetate consumption and cell yield of D. mccartyi, we estimated that on average 1.2 × 108 acetate molecules are needed to supply carbon for one D. mccartyi cell. Our study suggests that Desulfovibrio may supply a steady but low amount of fixed carbon to dechlorinating bacteria, exhibiting important implications for natural bio-attenuation when fixed carbon is limited.

Effects of Sulfate Reduction on Trichloroethene Dechlorination by Dehalococcoides-Containing Microbial Communities.

In order to elucidate interactions between sulfate reduction and dechlorination, we systematically evaluated the effects of different concentrations of sulfate and sulfide on reductive dechlorination by isolates, constructed consortia, and enrichments containing Dehalococcoides sp. Sulfate (up to 5 mM) did not inhibit the growth or metabolism of pure cultures of the dechlorinator Dehalococcoides mccartyi 195, the sulfate reducer Desulfovibrio vulgaris Hildenborough, or the syntroph Syntrophomonas wolfei In contrast, sulfide at 5 mM exhibited inhibitory effects on growth of the sulfate reducer and the syntroph, as well as on both dechlorination and growth rates of D. mccartyi Transcriptomic analysis of D. mccartyi 195 revealed that genes encoding ATP synthase, biosynthesis, and Hym hydrogenase were downregulated during sulfide inhibition, whereas genes encoding metal-containing enzymes involved in energy metabolism were upregulated even though the activity of those enzymes (hydrogenases) was inhibited. When the electron acceptor (trichloroethene) was limiting and an electron donor (lactate) was provided in excess to cocultures and enrichments, high sulfate concentrations (5 mM) inhibited reductive dechlorination due to the toxicity of generated sulfide. The initial cell ratio of sulfate reducers to D. mccartyi (1:3, 1:1, or 3:1) did not affect the dechlorination performance in the presence of sulfate (2 and 5 mM). In contrast, under electron donor limitation, dechlorination was not affected by sulfate amendments due to low sulfide production, demonstrating that D. mccartyi can function effectively in anaerobic microbial communities containing moderate sulfate concentrations (5 mM), likely due to its ability to outcompete other hydrogen-consuming bacteria and archaea.IMPORTANCE Sulfate is common in subsurface environments and has been reported as a cocontaminant with chlorinated solvents at various concentrations. Inconsistent results for the effects of sulfate inhibition on the performance of dechlorination enrichment cultures have been reported in the literature. These inconsistent findings make it difficult to understand potential mechanisms of sulfate inhibition and complicate the interpretation of bioremediation field data. In order to elucidate interactions between sulfate reduction and reductive dechlorination, this study systematically evaluated the effects of different concentrations of sulfate and sulfide on reductive dechlorination by isolates, constructed consortia, and enrichments containing Dehalococcoides sp. This study provides a more fundamental understanding of the competition mechanisms between reductive dechlorination by Dehalococcoides mccartyi and sulfate reduction during the bioremediation process. It also provides insights on the significance of sulfate concentrations on reductive dechlorination under electron donor/acceptor-limiting conditions during in situ bioremediation applications. For example, at a trichloroethene-contaminated site with a high sulfate concentration, proper slow-releasing electron donors can be selected to generate an electron donor-limiting environment that favors reductive dechlorination and minimizes the sulfide inhibition effect.

Metagenomic and Metatranscriptomic Analyses Reveal the Structure and Dynamics of a Dechlorinating Community Containing Dehalococcoides mccartyi and Corrinoid-Providing Microorganisms under Cobalamin-Limited Conditions.

The aim of this study is to obtain a systems-level understanding of the interactions between Dehalococcoides and corrinoid-supplying microorganisms by analyzing community structures and functional compositions, activities, and dynamics in trichloroethene (TCE)-dechlorinating enrichments. Metagenomes and metatranscriptomes of the dechlorinating enrichments with and without exogenous cobalamin were compared. Seven putative draft genomes were binned from the metagenomes. At an early stage (2 days), more transcripts of genes in the Veillonellaceae bin-genome were detected in the metatranscriptome of the enrichment without exogenous cobalamin than in the one with the addition of cobalamin. Among these genes, sporulation-related genes exhibited the highest differential expression when cobalamin was not added, suggesting a possible release route of corrinoids from corrinoid producers. Other differentially expressed genes include those involved in energy conservation and nutrient transport (including cobalt transport). The most highly expressed corrinoid de novo biosynthesis pathway was also assigned to the Veillonellaceae bin-genome. Targeted quantitative PCR (qPCR) analyses confirmed higher transcript abundances of those corrinoid biosynthesis genes in the enrichment without exogenous cobalamin than in the enrichment with cobalamin. Furthermore, the corrinoid salvaging and modification pathway of Dehalococcoides was upregulated in response to the cobalamin stress. This study provides important insights into the microbial interactions and roles played by members of dechlorinating communities under cobalamin-limited conditions.IMPORTANCE The key chloroethene-dechlorinating bacterium Dehalococcoides mccartyi is a cobalamin auxotroph, thus acquiring corrinoids from other community members. Therefore, it is important to investigate the microbe-microbe interactions between Dehalococcoides and the corrinoid-providing microorganisms in a community. This study provides systems-level information, i.e., taxonomic and functional compositions and dynamics of the supportive microorganisms in dechlorinating communities under different cobalamin conditions. The findings shed light on the important roles of Veillonellaceae species in the communities compared to other coexisting community members in producing and providing corrinoids for Dehalococcoides species under cobalamin-limited conditions.

Concomitant Leaching and Electrochemical Extraction of Rare Earth Elements from Monazite.

Rare earth elements (REEs) have become increasingly important in modern day technologies. Unfortunately, their recycling is currently limited, and the conventional technologies for their extraction and purification are exceedingly energy and chemical intensive. New sustainable technologies for REE extraction from both primary and secondary resources would be extremely beneficial. This research investigated a two-stage recovery strategy focused on the recovery of neodymium (Nd) and lanthanum (La) from monazite ore that combines microbially based leaching (using citric acid and spent fungal supernatant) with electrochemical extraction. Pretreating the phosphate-based monazite rock (via roasting) dramatically increased the microbial REE leaching efficiency. Batch experiments demonstrated the effective and continued leaching of REEs by recycled citric acid, with up to 392 mg of Nd L-1 and 281 mg of La L-1 leached during seven consecutive 24 h cycles. Neodymium was further extracted in the catholyte of a three-compartment electrochemical system, with up to 880 mg of Nd L-1 achieved within 4 days (at 40 A m-2). Meanwhile, the radioactive element thorium and counterions phosphate and citrate were separated effectively from the REEs in the anolyte, favoring REE extraction and allowing sustainable reuse of the leaching agent. This study shows a promising technology that is suitable for primary ores and can further be optimized for secondary resources.

Acetylene Fuels TCE Reductive Dechlorination by Defined Dehalococcoides/Pelobacter Consortia.

Acetylene (C2H2) can be generated in contaminated groundwater sites as a consequence of chemical degradation of trichloroethene (TCE) by in situ minerals, and C2H2 is known to inhibit bacterial dechlorination. In this study, we show that while high C2H2 (1.3 mM) concentrations reversibly inhibit reductive dechlorination of TCE by Dehalococcoides mccartyi isolates as well as enrichment cultures containing D. mccartyi sp., low C2H2 (0.4 mM) concentrations do not inhibit growth or metabolism of D. mccartyi. Cocultures of Pelobacter SFB93, a C2H2-fermenting bacterium, with D. mccartyi strain 195 or with D. mccartyi strain BAV1 were actively sustained by providing acetylene as the electron donor and carbon source while TCE or cis-DCE served as the electron acceptor. Inhibition by acetylene of reductive dechlorination and methanogenesis in the enrichment culture ANAS was observed, and the inhibition was removed by adding Pelobacter SFB93 into the consortium. Transcriptomic analysis of D. mccartyi strain 195 showed genes encoding for reductive dehalogenases (e.g., tceA) were not affected during the C2H2-inhibition, while genes encoding for ATP synthase, biosynthesis, and Hym hydrogenase were down-regulated during C2H2 inhibition, consistent with the physiological observation of lower cell yields and reduced dechlorination rates in strain 195. These results will help facilitate the optimization of TCE-bioremediation at contaminated sites containing both TCE and C2H2.

Incomplete Wood-Ljungdahl pathway facilitates one-carbon metabolism in organohalide-respiring Dehalococcoides mccartyi.

The acetyl-CoA "Wood-Ljungdahl" pathway couples the folate-mediated one-carbon (C1) metabolism to either CO2 reduction or acetate oxidation via acetyl-CoA. This pathway is distributed in diverse anaerobes and is used for both energy conservation and assimilation of C1 compounds. Genome annotations for all sequenced strains of Dehalococcoides mccartyi, an important bacterium involved in the bioremediation of chlorinated solvents, reveal homologous genes encoding an incomplete Wood-Ljungdahl pathway. Because this pathway lacks key enzymes for both C1 metabolism and CO2 reduction, its cellular functions remain elusive. Here we used D. mccartyi strain 195 as a model organism to investigate the metabolic function of this pathway and its impacts on the growth of strain 195. Surprisingly, this pathway cleaves acetyl-CoA to donate a methyl group for production of methyl-tetrahydrofolate (CH3-THF) for methionine biosynthesis, representing an unconventional strategy for generating CH3-THF in organisms without methylene-tetrahydrofolate reductase. Carbon monoxide (CO) was found to accumulate as an obligate by-product from the acetyl-CoA cleavage because of the lack of a CO dehydrogenase in strain 195. CO accumulation inhibits the sustainable growth and dechlorination of strain 195 maintained in pure cultures, but can be prevented by CO-metabolizing anaerobes that coexist with D. mccartyi, resulting in an unusual syntrophic association. We also found that this pathway incorporates exogenous formate to support serine biosynthesis. This study of the incomplete Wood-Ljungdahl pathway in D. mccartyi indicates a unique bacterial C1 metabolism that is critical for D. mccartyi growth and interactions in dechlorinating communities and may play a role in other anaerobic communities.

Bioleaching of rare earth elements from monazite sand.

Three fungal strains were found to be capable of bioleaching rare earth elements from monazite, a rare earth phosphate mineral, utilizing the monazite as a phosphate source and releasing rare earth cations into solution. These organisms include one known phosphate solubilizing fungus, Aspergillus niger ATCC 1015, as well as two newly isolated fungi: an Aspergillus terreus strain ML3-1 and a Paecilomyces spp. strain WE3-F. Although monazite also contains the radioactive element Thorium, bioleaching by these fungi preferentially solubilized rare earth elements over Thorium, leaving the Thorium in the solid residual. Adjustments in growth media composition improved bioleaching performance measured as rare earth release. Cell-free spent medium generated during growth of A. terreus strain ML3-1 and Paecilomyces spp. strain WE3-F in the presence of monazite leached rare earths to concentrations 1.7-3.8 times those of HCl solutions of comparable pH, indicating that compounds exogenously released by these organisms contribute substantially to leaching. Organic acids released by the organisms included acetic, citric, gluconic, itaconic, oxalic, and succinic acids. Abiotic leaching with laboratory prepared solutions of these acids was not as effective as bioleaching or leaching with cell-free spent medium at releasing rare earths from monazite, indicating that compounds other than the identified organic acids contribute to leaching performance.

Perfluoroalkyl Acids Inhibit Reductive Dechlorination of Trichloroethene by Repressing Dehalococcoides.

The subsurface recalcitrance of perfluoroalkyl acids (PFAAs) derived from aqueous film-forming foams could have adverse impacts on the microbiological processes used for the bioremediation of co-mingled chlorinated solvents such as trichloroethene (TCE). Here, we show that reductive dechlorination by a methanogenic, mixed culture was significantly inhibited when exposed to concentrations representative of PFAA source zones (>66 mg/L total of 11 PFAA analytes, 6 mg/L each). TCE dechlorination, cis-dichloroethene and vinyl chloride production and dechlorination, and ethene generation were all inhibited at these PFAA concentrations. Phylogenetic analysis revealed that the abundances of 65% of the operational taxonomic units (OTUs) changed significantly when grown in the presence of PFAAs, although repression or enhancement resulting from PFAA exposure did not correlate with putative function or phylogeny. Notably, there was significant repression of Dehalococcoides (8-fold decrease in abundance) coupled with a corresponding enhancement of methane-generating Archaea (a 9-fold increase). Growth and dechlorination by axenic cultures of Dehalococcoides mccartyi strain 195 were similarly repressed under these conditions, confirming an inhibitory response of this pivotal genus to PFAA presence. These results suggest that chlorinated solvent bioattenuation rates could be impeded in subsurface environments near PFAA source zones.

Effects of Aqueous Film-Forming Foams (AFFFs) on Trichloroethene (TCE) Dechlorination by a Dehalococcoides mccartyi-Containing Microbial Community.

The application of aqueous film-forming foams (AFFFs) to extinguish chlorinated solvent-fueled fires has led to the co-contamination of poly- and perfluoroalkyl substances (PFASs) and trichloroethene (TCE) in groundwater and soil. Although reductive dechlorination of TCE by Dehalococcoides mccartyi is a frequently used remediation strategy, the effects of AFFF and PFASs on TCE dechlorination are not well-understood. Various AFFF formulations, PFASs, and ethylene glycols were amended to the growth medium of a D. mccartyi-containing enrichment culture to determine the impact on dechlorination, fermentation, and methanogenesis. The community was capable of fermenting organics (e.g., diethylene glycol butyl ether) in all AFFF formulations to hydrogen and acetate, but the product concentrations varied significantly according to formulation. TCE was dechlorinated in the presence of an AFFF formulation manufactured by 3M but was not dechlorinated in the presence of formulations from two other manufacturers. Experiments amended with AFFF-derived PFASs and perfluoroalkyl acids (PFAAs) indicated that dechlorination could be inhibited by PFASs but that the inhibition depends on surfactant concentration and structure. This study revealed that the fermentable components of AFFF can stimulate TCE dechlorination, while some of the fluorinated compounds in certain AFFF formulations can inhibit dechlorination.

Identification of specific corrinoids reveals corrinoid modification in dechlorinating microbial communities.

Cobalamin and other corrinoids are essential cofactors for many organisms. The majority of microbes with corrinoid-dependent enzymes do not produce corrinoids de novo, and instead must acquire corrinoids produced by other organisms in their environment. However, the profile of corrinoids produced in corrinoid-dependent microbial communities, as well as the exchange and modification of corrinoids among community members have not been well studied. In this study, we applied a newly developed liquid chromatography tandem mass spectrometry-based corrinoid detection method to examine relationships among corrinoids, their lower ligand bases and specific microbial groups in microbial communities containing Dehalococcoides mccartyi that has an obligate requirement for benzimidazole-containing corrinoids for trichloroethene respiration. We found that p-cresolylcobamide ([p-Cre]Cba) and cobalamin were the most abundant corrinoids in the communities. It suggests that members of the family Veillonellaceae are associated with the production of [p-Cre]Cba. The decrease of supernatant-associated [p-Cre]Cba and the increase of biomass-associated cobalamin were correlated with the growth of D. mccartyi by dechlorination. This supports the hypothesis that D. mccartyi is capable of fulfilling its corrinoid requirements in a community through corrinoid remodelling, in this case, by importing extracellular [p-Cre]Cba and 5,6-dimethylbenzimidazole (DMB) (the lower ligand of cobalamin), to produce cobalamin as a cofactor for dechlorination. This study also highlights the role of DMB, the lower ligand produced in all of the studied communities, in corrinoid remodelling. These findings provide novel insights on roles played by different phylogenetic groups in corrinoid production and corrinoid exchange within microbial communities. This study may also have implications for optimizing chlorinated solvent bioremediation.

Efficient metabolic exchange and electron transfer within a syntrophic trichloroethene-degrading coculture of Dehalococcoides mccartyi 195 and Syntrophomonas wolfei.

Dehalococcoides 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 µmol day(-1)) and cell yield [(1.1 ± 0.3) × 10(8) cells µmol(-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 H2 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 µmol per bottle, which was lower than that observed for strain 195 in isolation. The minimum H2 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 H2 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.

Development of a fluorescence-activated cell sorting method coupled with whole genome amplification to analyze minority and trace Dehalococcoides genomes in microbial communities.

Dehalococcoides mccartyi are functionally important bacteria that catalyze the reductive dechlorination of chlorinated ethenes. However, these anaerobic bacteria are fastidious to isolate, making downstream genomic characterization challenging. In order to facilitate genomic analysis, a fluorescence-activated cell sorting (FACS) method was developed in this study to separate D. mccartyi cells from a microbial community, and the DNA of the isolated cells was processed by whole genome amplification (WGA) and hybridized onto a D. mccartyi microarray for comparative genomics against four sequenced strains. First, FACS was successfully applied to a D. mccartyi isolate as positive control, and then microarray results verified that WGA from 10(6) cells or ∼1 ng of genomic DNA yielded high-quality coverage detecting nearly all genes across the genome. As expected, some inter- and intrasample variability in WGA was observed, but these biases were minimized by performing multiple parallel amplifications. Subsequent application of the FACS and WGA protocols to two enrichment cultures containing ∼10% and ∼1% D. mccartyi cells successfully enabled genomic analysis. As proof of concept, this study demonstrates that coupling FACS with WGA and microarrays is a promising tool to expedite genomic characterization of target strains in environmental communities where the relative concentrations are low.

Aerobic Biotransformation of Fluorotelomer Thioether Amido Sulfonate (Lodyne) in AFFF-Amended Microcosms.

The aerobic biotransformation pathways of 4:2, 6:2, and 8:2 fluorotelomer thioether amido sulfonate (FtTAoS) were characterized by determining the fate of the compounds in soil and medium microcosms amended with an aqueous film-forming foam (AFFF) solution. The biotransformation of FtTAoS occurred in live microcosms over approximately 40 days and produced 4:2, 6:2, and 8:2 fluorotelomer sulfonate (FtS), 6:2 fluorotelomer unsaturated carboxylic acid (FtUCA), 5:3 fluorotelomer carboxylic acid (FtCA), and C4 to C8 perfluorinated carboxylic acids (PFCAs). Two biotransformation products corresponding to singly and doubly oxygenated forms of 6:2 FtTAoS were also identified through high resolution mass spectrometry (MS) analysis and liquid chromatography tandem-MS. An oxidative assay was used to indirectly quantify the total concentration of polyfluorinated compounds and check the mass balance. The assay produced near complete mass recovery of FtTAoS after biotransformation, with 10% (mol/mol) of the amended FtTAoS accounted for in FtS, FtCA, and PFCA products. The transformation rates of identified products appear to be slow relative to FtTAoS, indicating that some intermediates may persist in the environment. This study confirms some of the sources of FtS and PFCAs in groundwater and soil at AFFF-impacted sites and suggests that fluorinated intermediates that are not routinely measured during the biotransformation of PFASs may accumulate.