Identification of Reductive Dehalogenases That Mediate Complete Debromination of Penta- and Tetrabrominated Diphenyl Ethers in Dehalococcoides spp.

Abstract

Polybrominated diphenyl ethers (PBDEs) are persistent, highly toxic, and widely distributed environmental pollutants. The microbial populations and functional reductive dehalogenases (RDases) responsible for PBDEs debromination in anoxic systems remain poorly understood, which confounds bioremediation of PBDE-contaminated sites. Here we report a PBDE-debrominating enrichment culture dominated by a previously undescribed Dehalococcoides mccartyi population. A D. mccartyi strain, designated TZ50, whose genome contains 25 putative RDase encoding genes, was isolated from the debrominating enrichment culture. Strain TZ50 dehalogenated a mixture of penta- and tetra-BDE congeners (total BDEs 1.48 μM) to diphenyl ether within two weeks (0.58 μM Br - /d) via o rtho - and meta - bromine elimination; strain TZ50 also dechlorinated tetrachloroethene (PCE) to vinyl chloride and ethene (260.2 μM Cl - /d). Results of native-PAGE, proteomic profiling, and in vitro enzymatic activity assays implicated involvement of three RDases in PBDEs and PCE dehalogenation. TZ50_0172 (PteA TZ50 ) and TZ50_1083 (TceA TZ50 ), were responsible for debromination of penta- and tetra-BDEs to di-BDE. TZ50_0172 and TZ50_1083 were also implicated in dechlorination of PCE to TCE and of TCE to vinyl chloride/ethene, respectively. The other expressed RDase, TZ50_0090 (designated BdeA), was associated with debromination of di-BDE to diphenyl ether, but its role in PCE dechlorination was unclear. Comparatively few RDases are known to be involved in PBDE debromination and the identification of PteA TZ50 , TceA TZ50 , and BdeA provides additional information for evaluating debromination potential at contaminated sites. Moreover, the ability of PteA TZ50 and TceA TZ50 to dehalogenate both PBDEs and PCE makes strain TZ50 a suitable candidate for remediation of co-contaminated sites. Importance The ubiquity, toxicity, and persistence of polybrominated diphenyl ethers (PBDEs) in the environment have drawn significant public and scientific interest to the need for remediation of PBDEs-contaminated ecosystems. However, the low bioavailability of PBDEs in environmental compartments typically limits bioremediation of PBDEs and has long impeded the study of anaerobic microbial PBDEs removal. In the current study, a novel Dehalococcoides mccartyi , dubbed strain TZ50, that expresses RDases that mediate organohalide respiration of both PBDEs and chloroethenes was isolated and characterized. Strain TZ50 could potentially be used to remediate multiple co-occurring organohalides in contaminated systems.

Microbial Debromination of Polybrominated Diphenyl Ethers by Dehalococcoides-Containing Enrichment Culture

“…Compared with lower BDE congener groups, higher BDEs are more persistent and recalcitrant for biodegradation in the environment. Thus far, a complete debromination of lower BDE congeners (such as penta-BDE 99, 100, and tetra-BDE 47) has been found in the bacterial populations of Dehalococcoides ( Ding et al, 2017 ; Zhao et al, 2021 ), yet difficulties still lie in the discovery of proper bacterial populations to completely detoxify higher BDEs; only slow and partial biodegradation, which often leads to the accumulation of even more toxic congener groups, has been reported ( Zhao et al, 2018 ). In the PB enrichment culture, multiple RDases are suggested to catalyze higher and lower BDEs (i.e., BDE-183 and BDE-47), which is consistent with previous studies that multiple RDases were identified in the debromination of penta-BDEs ( Ding et al, 2017 ; Zhao et al, 2021 ).…”

“…Thus far, a complete debromination of lower BDE congeners (such as penta-BDE 99, 100, and tetra-BDE 47) has been found in the bacterial populations of Dehalococcoides ( Ding et al, 2017 ; Zhao et al, 2021 ), yet difficulties still lie in the discovery of proper bacterial populations to completely detoxify higher BDEs; only slow and partial biodegradation, which often leads to the accumulation of even more toxic congener groups, has been reported ( Zhao et al, 2018 ). In the PB enrichment culture, multiple RDases are suggested to catalyze higher and lower BDEs (i.e., BDE-183 and BDE-47), which is consistent with previous studies that multiple RDases were identified in the debromination of penta-BDEs ( Ding et al, 2017 ; Zhao et al, 2021 ). Taken together, to improve the removal effectiveness and efficiency of higher BDEs in practical application, we may consider the strategy of bio-augmenting cultures with a broader dehalogenating range of PBDE congeners, which could be achieved by integrating multiple bacterial populations with more versatile dehalogenating capabilities on different PBDE congener orthologs.…”

“…Currently, both microcosms established from a versatile environment and isolated microorganisms have shown slower and less extensive debromination on deca- through hexa-BDEs rather than penta- and tetra-BDEs ( Lee and He, 2010 ). So far, only partial and incomplete debromination of deca- through hexa-BDEs ( Robrock et al, 2008 ) and a limited number of microorganisms capable of complete debromination of penta- and tetra-BDEs have been reported ( Ding et al, 2017 ; Zhao et al, 2021 ). Therefore, cultures that can more efficiently debrominate PBDEs and further be applied in in situ warrant further exploration.…”

Offshore Marine Sediment Microbiota Respire Structurally Distinct Organohalide Pollutants

“…Primers that target the 16S rRNA genes of Dehalococcoides, Dehalogenimonas, and Dehalobacter and previously reported RDase genes were used to quantify OHRB and RDase genes, respectively (Table S2). ,,, Standard curve quantification of target sequences was performed using plasmids carrying a target amplicon constructed using the pGEM-T vector system (Promega) and extracted by the QIAprep Spin Miniprep Kit (QIAGEN) as reported . Nuclease-free water was used as a negative control.…”

“…Recent studies found that several D. mccartyi strains (i.e., CG1, CG4, and CG5) and numerous uncultivated Dehalococcoidia can utilize a single RDase to dehalogenate structurally dissimilar (i.e., aliphatic and aromatic) organohalide compounds. − For example, RDases PcbA1, PcbA4, and PcbA5 can dehalogenate both tetrachloroethene (PCE) and PCBs, which likely contributes to the role that Dehalococcoidia play in attenuating organohalides in terrestrial environments. , However, the prevalence and activity of bifunctional or multifunctional RDases among marine OHRB have not been reported.…”