Syntrophic pathways for microbial mercury methylation

Abstract

Exposure to dietary sources of methylmercury (MeHg) is the focus of public health concerns with environmental mercury (Hg) contamination. MeHg is formed in anoxic environments by anaerobic microorganisms. This process has been studied mostly with single-species culture incubations, although the relevance of such studies to Hg(II)-methylation in situ is limited because microbial activities in the environment are critically modulated by interactions among microbial functional groups. Here we describe experiments in which Hg(II)-methylation was examined within the context of various microbial syntrophies. We show enhanced Hg(II)-methylation under conditions that established syntrophy by interspecies hydrogen and acetate transfer. Relative to activity of monocultures, interactions of Hg(II) methylating sulfate-reducing bacteria with a methanogen stimulated potential Hg(II)-methylation rates 2-fold to 9-fold, and with Syntrophobacter sp. 1.7-fold to 1.8-fold; those of a Hg(II) methylating Syntrophobacter sp. with a methanogen increased Hg(II)-methylation 2-fold. Under sulfate-depleted conditions, higher Hg(II)-methylation rates in the syntrophic incubations corresponded to higher free energy yields (ΔG°') than in the monocultures. Based on energetic considerations, we therefore propose that syntrophic microbial interactions are likely a major source of MeHg in sulfate- and iron-limited anoxic environments while in sulfate-replete environments, MeHg formation via sulfate reduction dominates.

Fig. 1

The effects on Hg(II)-methylation of interspecies hydrogen and acetate transfer between a methanogen M. hungatei JF-1 and SRB D. desulfuricans ND132 (a), and D. africanus subsp africanus (b), in a lactate-bicarbonate medium. Samples were withdrawn for determination of MeHg and protein concentrations (c, d) every day for 5 days. Heat-killed (1 h at 80 °C) and medium-only (blank) controls were included. Averages and SD of three replicate cultures are shown

Fig. 2

Hg(II)-methylation by Syntrophobacter spp. MeHg synthesis (pmol of mg initial protein) (a) and cell protein contents (b) were analyzed in monocultures of S. wolinii DB, S. sulfatireducens TB1806, S. fumaroxidans MPOB, and D. africanus. Cultures were grown in sulfate-reducing media as recommended for each by the DSMZ (Na₂S as a reducing agent). Averages and standard deviations of three replicate cultures are shown

Fig. 3

The effects on Hg(II)-methylation of interspecies hydrogen and acetate transfer between the syntrophs S. wolinii DSM 2805 (a), S. sulfatireducens DSM 16706 (b), and S. fumaroxidans DSM 10017 (c) and the methanogen M. hungatei JF-1. Samples were withdrawn for determination of MeHg concentrations and protein content (d–f) in a sulfate-free propionate Syntrophobacter medium every day for 5 days. Heat-killed (1 h at 80 °C) controls were included. Averages and standard deviations of three replicate cultures are shown

Fig. 4

The effects on Hg(II)-methylation (a, b) and growth (c, d) of syntrophy between the methylating SRB strains D. desulfuricans ND132 or D. africanus subsp. africanus and the syntroph S. sulfatireducens TB8061 when grown in a propionate-sulfate SRB medium (at an initial sulfate concentration of 3.94 mM). Samples were withdrawn for determination of MeHg concentrations and cell counts by flow cytometry-cell sorting (Figure S1) every day for 5 days. Heat-killed (1 h at 80 °C) and medium-only (blank) controls were included. Averages and standard deviations (positive only for clarity) of three replicate cultures are shown

Fig. 5

Free energy yields of reaction for individual Hg(II) methylating anaerobes and syntrophies likely to be important in freshwater (low sulfate and iron) and marine systems (low iron). A hypothetical scheme predicting the relative contribution of different anaerobes and their interactions to Hg(II) methylation (a). Entries in bold were tested in this paper. Microbes or syntrophies with higher (more negative) free energy yields (ΔG°') should outcompete those with lower ΔG°' at the specified environmental concentrations of substrates and products (Table S4). Relationship between Hg(II)-methylation rates in low iron and sulfate freshwater systems (Table 1) and the expected free energy yields of their metabolisms (Table S4) (b). Open circle-methanogens; open triangle-syntrophs grown by fermentation; filled triangle-syntrophs grown with methanogens; open squares-SRB grown by fermentation; filled squares-SRB grown with methanogens. The two pairs of symbols depicting SRB potential methylation rates are those observed with strains D. desulfuricans and D. africanus. The relationship between methylation rate and free energy yield is log (methylation rate) = −0.079ΔG°' - 0.4 (R² = 0.66, p < 0.05). Such a log-linear relationship is in accord with theoretical linear free energy relationships between reaction kinetics and thermodynamics [46]