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A largely understudied microbially mediated mercury (Hg) bioremediative pathway includes the volatilization of Hg 2+ to Hg⁰. Therefore, studies on Hg resistant bacteria (HgR), isolated from historically long-term contaminated environments, can serve as models to understand mechanisms underpinning Hg cycling. Towards this end, a mercury resistant bacterial strain, identified as Stenotrophomonas sp., strain MA5, was isolated from Mill Branch on the Savannah River Site (SRS); an Hg-impacted ecosystem. Minimum inhibitory concentration (MIC) analysis showed Hg resistance of up to 20 µg/mL by MA5 with 95% of cells retaining viability. Microcosm studies showed that the strain depleted more than 90% of spiked Hg 2+ within the first 24 h of growth and the detection of volatilized mercury indicated that the strain was able to reduce Hg 2+ to Hg⁰. To understand molecular mechanisms of Hg volatilization, a draft whole genome sequence was obtained, annotated and analyzed, which revealed the presence of a transposon-derived mer operon ( merRTPADE ) in MA5, known to transport and reduce Hg 2+ into Hg⁰. Based on the whole genome sequence of strain MA5, qRT-PCR assays were designed on merRTPADE , we found a ~40-fold higher transcription of mer  T, P, A, D and E when cells were exposed to 5 µg/mL Hg 2+ . Interestingly, strain MA5 increased cellular size as a function of increasing Hg concentrations, which is likely an evolutionary response mechanism to cope with Hg stress. Moreover, metal contaminated environments are shown to co-select for antibiotic resistance. When MA5 was screened for antibiotic resistance, broad resistance against penicillin, streptomycin, tetracycline, ampicillin, rifampicin, and erythromycin was found; this correlated with the presence of multiple gene determinants for antibiotic resistance within the whole genome sequence of MA5. Overall, this study provides an in-depth understanding of the underpinnings of Stenotrophomonas -mercury interactions that facilitate cellular survival in a contaminated soil habitat.

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Multiple Lines of Evidences Reveal Mechanisms Underpinning Mercury Resistance and Volatilization by Stenotrophomonas sp. MA5 Isolated from the Savannah River Site (SRS), USA