Revisiting the dihaloelimination potential of Dehalococcoides revealed by genomic and proteomic analyses.

Abstract

Dehalococcoides (Dhc) has recently gained significant attention in microbial reductive dehalogenation due to its specialized ability to metabolize various halocarbons. While previous reports have documented Dhc-mediated dihaloelimination of 1,2-dichloroethane (1,2-DCA), 1,2-dichloropropane (1,2-DCP), and 1,2-dibromoethane, this study reveals an expansion of Dhc's dihaloelimination substrate range. We successfully established PJ_DCA and PJ_TCA enrichment cultures from petroleum-contaminated soil, exhibiting dihaloelimination activity toward 1,1,2-trichloroethane (1,1,2-TCA), 1,2,3-trichloropropane (1,2,3-TCP), and 1,1,2,2-tetrachloroethane (1,1,2,2-TeCA)-substrates previously only known to be transformed by other organohalide-respiring bacteria (OHRB). Amplicon sequencing revealed the predominance of Dhc as the primary OHRB within the PJ_DCA and PJ_TCA cultures, leading to the identification of two novel Dhc populations, designated as strains PJ_DCA and PJ_TCA, respectively. These strains showed robust growth yields of 4.1 ± 0.4 × 10⁷ and 7.6 ± 0.4 × 10⁷ cells per μmol Cl^- released when using 1,2-DCA and 1,1,2-TCA as electron acceptors, respectively. Genomic analysis revealed a reductive dehalogenase (RDase) homologous to the characterized DcpA, an enzyme known for dihaloelimination activity. Proteomic studies confirmed the expression of this DcpA-like RDase during dihaloelimination of 1,2-DCA, 1,1,2-TCA, and 1,2-DCP. This work identifies Dhc strains that retain dihaloelimination activity toward classical substrates while expanding this capability to 1,1,2-TCA, 1,2,3-TCP, and 1,1,2,2-TeCA. The discovery of these versatile Dhc strains enhances our understanding of microbial dehalogenation potential across diverse geological settings and improves prospects for bioremediation of halogenated alkane contaminants.

Importance: This study identifies Dehalococcoides strains capable of dihaloeliminating diverse chlorinated alkanes (1,1,2-TCA, 1,2,3-TCP, 1,1,2,2-TeCA), expanding their bioremediation potential. These compounds are persistent groundwater contaminants with high toxicity. The discovery of Dhc populations (PJ_DCA/PJ_TCA) with robust growth yields (10⁷ cells/μmol Cl^-) and broad substrate range offers new solutions for detoxifying complex halogenated pollutant mixtures. The identification and proteomic confirmation of DcpA-like RDases (DheA) provide the genetic and functional basis for this expanded dihaloelimination capacity. These findings advance strategies for in situ remediation of industrial sites contaminated with C₂-C₃ halocarbons, reducing ecological and human health risks.