Orenia metallireducens sp. nov. Strain Z6, a Novel Metal-Reducing Member of the Phylum Firmicutes from the Deep Subsurface

Abstract

A novel halophilic and metal-reducing bacterium, Orenia metallireducens strain Z6, was isolated from briny groundwater extracted from a 2.02 km-deep borehole in the Illinois Basin, IL. This organism shared 96% 16S rRNA gene similarity with Orenia marismortui but demonstrated physiological properties previously unknown for this genus. In addition to exhibiting a fermentative metabolism typical of the genus Orenia, strain Z6 reduces various metal oxides [Fe(III), Mn(IV), Co(III), and Cr(VI)], using H 2 as the electron donor. Strain Z6 actively reduced ferrihydrite over broad ranges of pH (6 to 9.6), salinity (0.4 to 3.5 M NaCl), and temperature (20 to 60°C). At pH 6.5, strain Z6 also reduced more crystalline iron oxides, such as lepidocrocite (␥- IMPORTANCEA novel iron-reducing species, Orenia metallireducens sp. nov., strain Z6, was isolated from groundwater collected from a geological formation located 2.02 km below land surface in the Illinois Basin, USA. Phylogenetic, physiologic, and genomic analyses of strain Z6 found it to have unique properties for iron reducers, including (i) active microbial iron-reducing capacity under broad ranges of temperatures (20 to 60°C), pHs (6 to 9.6), and salinities (0.4 to 3.5 M NaCl), (ii) lack of c-type cytochromes typically affiliated with iron reduction in Geobacter and Shewanella species, and (iii) being the only member of the Halanaerobiales capable of reducing crystalline goethite and hematite. This study expands the scope of phylogenetic affiliations, metabolic capacities, and catalytic mechanisms for iron-reducing microbes.T he reduction of ferric [Fe(III)-bearing] minerals by microorganisms is widespread in both terrestrial and marine environments (1, 2) and is potentially one of the earliest forms of metabolism (3, 4). Due to the abundance of ferric minerals in the Earth's crust and the ubiquity of dissimilatory metal-reducing bacteria (DMRB), Fe(III) reduction is of global environmental significance, particularly in the subsurface. A phylogenetic variety of organisms has been reported for the capacity to reduce iron in a dissimilatory manner (1, 5). Dissimilatory iron reduction can be classified into two major groups. Many of the iron-reducing organisms (e.g., fermentative iron reducers) use Fe(III) as a minor side reaction in their metabolism but do not appear to conserve energy to support growth from this electron transfer. In comparison, the respiratory iron reducers conserve energy to support growth from Fe(III) via an electron transfer chain (1,5). DMRB strongly influence the biogeochemical cycling of metals and mineralization of organic matter in aquatic and sedimentary environments (1,(6)(7)(8). In addition, DMRB play a key role in the bioremediation of hazardous contaminants, such as heavy metals, radionuclides, and hydrocarbons (9). DMRB capable of reducing ferric minerals have previously been isolated from gold mines, petroleum reservoirs, and other

Electron Donor Utilization and Secondary Mineral Formation during the Bioreduction of Lepidocrocite by Shewanella putrefaciens CN32

“…Green rusts were observed as products of lepidocrocite bioreduction by S. putrefaciens CN32 when H 2 , formate, lactate, or NAG were provided as electron donors ( ) is a geologically significant iron carbonate mineral that is of commercial interest due to its use as a minor iron ore. Siderite is commonly reported as a secondary mineral during the bioreduction of Fe(III) oxides in systems with substantial carbonate concentrations (i.e., >20 mM) [9,33,36,[121][122][123]. In our experimental systems, siderite was observed as the sole secondary mineral in pyruvate-and serine-amended systems (note, as discussed above, serine is likely converted to pyruvate by S. putrefaciens CN32).…”

“…As such, the presence of Fe(II) in suboxic to anoxic near surface environments is typically the result of the activity of dissimilatory iron-reducing (DIR) bacteria and archaea. These phylogenetically diverse microorganisms can couple the oxidation of organic compounds or hydrogen (H 2 ) to the reduction of Fe(III) to Fe(II) [6][7][8][9][10][11][12][13][14][15][16][17][18][19]. As a group, dissimilatory iron-reducing bacteria (DIRB) are able to use soluble Fe(III) complexes (e.g., ferric citrate), Fe(III) oxides, and Fe(III)-bearing clay minerals as terminal electron acceptors for anaerobic respiration [20][21][22][23][24][25][26][27][28].…”

Labilibaculum manganireducens gen. nov., sp. nov. and Labilibaculum filiforme sp. nov., Novel Bacteroidetes Isolated from Subsurface Sediments of the Baltic Sea

“…Thus, the strains represent the “fermentative metal reducer type,” which transfers only a small part of the electron equivalents to the metal (typically <5%) while most of the electron equivalents are recovered in fermentation products (Lovley, 2013 ). However, even if only a small part of electron equivalents was transferred to iron oxides, it has been shown to improve the fermentative balance (less formation of acetate, butyrate, or hydrogen) resulting in thermodynamically more favorable conditions (Lehours et al, 2010 ; Dong et al, 2016 ). Thus, the available substrates were used more efficiently by the fermenter, which might be an important competitive advantage in carbon-limited environments like the deep biosphere.…”

“…The capability of iron and manganese reduction has rarely been tested for Bacteroidetes species, and only one species, Bacteroides hypermegas , appears in the list of iron-reducing organisms (Jones et al, 1984 ; Lovley, 2013 ). Bacteria of the deep biosphere have been shown previously to reduce metals including members of the Firmicutes, Proteobacteria, Deferribacteres and Deinococcus-Thermus (Roden and Lovley, 1993 ; Greene et al, 1997 ; Kieft et al, 1999 ; Dong et al, 2016 ; Vandieken et al, 2017 ), supporting the suggestion that iron and manganese oxides in Baltic Sea sediments (Hardisty et al, 2016 ) might be reduced by fermenters in order to more efficiently use the scarce organic material for survival in the deep biosphere.…”

Denitrification by Anaeromyxobacter dehalogenans, a Common Soil Bacterium Lacking the Nitrite Reductase Genes nirS and nirK

“…4. Additionally, not all Fe(III)reducing bacteria require c-type cytochromes (39), suggesting that the prediction of potential iron-reducing bacteria by searching for c-type cytochromes is a conservative approach and likely an underestimation of potential chemodenitrifiers. Physiological studies and measurements of the NO 3 Ϫ reduction products of potential chemodenitrifiers will reveal if these organisms indeed utilize the chemodenitrifier ecological strategy demonstrated for A. dehalogenans.…”