Reduction of hexavalent chromium by the thermophilic methanogen Methanothermobacter thermautotrophicus

Abstract

Despite the significant progress on iron reduction by thermophilic microorganisms, studies on their ability to reduce toxic metals are still limited, despite their common co-existence in high temperature environments (up to 70°C). In this study, Methanothermobacter thermautotrophicus, an obligate thermophilic methanogen, was used to reduce hexavalent chromium. Experiments were conducted in a growth medium with H2/CO2 as substrate with various Cr6+ concentrations (0.2, 0.4, 1, 3, and 5 mM) in the form of potassium dichromate (K2Cr2O7). Time-course measurements of aqueous Cr6+ concentrations with the 1, 5-diphenylcarbazide colorimetric method showed complete reduction of the 0.2 and 0.4 mM Cr6+ solutions by this methanogen. However, much lower reduction extents of 43.6%, 13.0%, and 3.7% were observed at higher Cr6+ concentrations of 1, 3 and 5 mM, respectively. These lower extents of bioreduction suggest a toxic effect of aqueous Cr6+ to cells at this concentration range. At these higher Cr6+ concentrations, methanogenesis was inhibited and cell growth was impaired as evidenced by decreased total cellular protein production and live/dead cell ratio. Likewise, Cr6+ bioreduction rates decreased with increased initial concentrations of Cr6+ from 13.3 to1.9 µM h−1. X-ray absorption near-edge structure (XANES) spectroscopy revealed a progressive reduction of soluble Cr6+ to insoluble Cr3+ precipitates, which was confirmed as amorphous chromium hydroxide by X-ray diffraction and selected area electron diffraction pattern. However, a small fraction of reduced Cr occurred as aqueous Cr3+. Scanning and transmission electron microscope observations of M. thermautotrophicus cells after Cr6+ exposure suggest both extra- and intracellular chromium reduction mechanisms. Results of this study demonstrate the ability of M. thermautotrophicus cells to reduce toxic Cr6+ to less toxic Cr3+ and its potential application in metal bioremediation, especially at high temperature subsurface radioactive waste disposal sites, where the temperature may reach ∼70°C.

[Cobalt(III)–EDTA]− reduction by thermophilic methanogen Methanothermobacter thermautotrophicus

“…Measurement of time course protein concentration showed initial inhibition of cell growth at all levels of Co(III), indicating the toxic effects of Co(III). However, at lower Co(III) concentrations, there was a small amount of resumption of cell growth after significant Co(III) reduction occurred, similar to previous studies on microbial reduction of Cr(VI) (Cheung and Gu, 2003;Viamajala et al, 2004;Singh et al, 2015). This inhibition-recovery cycle of M. thermautotrophicus cells during [Co(III)-EDTA] − bioreduction (Fig.…”

“…3 inset) could be attributed to the enzymes present in its genome such as superoxide dismutase which is known to possess the ability to repair damaged cellular components. A similar mechanism has been proposed to explain the Cr(VI) toxicity to M. thermautotrophicus (Singh et al, 2015). However, incomplete reduction at higher Co(III) concentrations (7 and 10 mM) was likely caused by a greater toxicity of cobalt to M. thermautotrophicus, as evidenced by inhibition of cell growth at these concentrations (Fig.…”

“…37°C) have been demonstrated to possess the ability to reduce V(V) (Zhang et al, 2014). More recently, the ability of M. thermautotrophicus to reduce toxic Cr(VI) to non-toxic Cr(III) has been demonstrated, highlighting a potential application for the reduction and immobilization of Cr(VI) at high temperature radioactive disposal sites (Singh et al, 2015).…”

“…Methanogens are known to exist at similar DOE waste disposal sites, such as the Waste Isolation Pilot Plant (WIPP) in southern New Mexico (Wang and Francis, 2005), as well as other subsurface sites such as the Äspö Hard Rock Laboratory in Sweden (Pedersen, 1999;Kotelnikova, 2002). While methanogens have been shown capable of reducing structural Fe(III) in clay minerals (Zhang et al, 2013), and heavy metals V(V) (Zhang et al, 2014) and Cr(VI) (Singh et al, 2015), their ability to reduce and detoxify other heavy metals such as Co, especially at elevated temperatures, has not been explored. Indeed, thermophilic methanogens are among the least studied of any microorganisms that have the ability to carry out important redox reactions in nature.…”

Methanogens, sulphate and heavy metals: a complex system

“…Methanothermobacter thermautotrophicus growing in H 2 /CO 2 , was able to reduce 0.2 and 0.4 mM Cr 6? completely (Lira-Silva et al 2013). Singh et al (2015) tested growing M. thermautotrophicus with higher concentrations of hexavalent chromium; amendment of 1, 3 and 5 mM of Cr 6? resulted in 43.6, 13 and 3.7 % reduction of the metal.…”