Polymer biodegradation by Halanaerobium promotes reservoir souring during hydraulic fracturing

Hydraulically fractured shale reservoirs have facilitated studies of unexplored niches in the continental deep biosphere. In high-salinity North American shale systems, members of the genus Halanaerobium seem to be ubiquitous. Polymers like guar gum used as gelling agents in hydraulic fracturing fluids are known to be fermentable substrates, but metabolic pathways encoding these processes have not been characterized. To explore this, produced water samples from the Permian Basin were incubated both at 30°C to simulate above-ground storage conditions and at 60°C to simulate subsurface reservoir conditions. Guar metabolism coincided with Halanaerobium growing only at 30°C, revealing genes for polymer biodegradation through the mixed-acid fermentation pathway in different metagenome-assembled genomes (MAGs). Whereas thiosulfate reduction to sulfide is often invoked to explain the dominance of Halanaerobium in these settings, genes for thiosulfate metabolism were lacking in Halanaerobium genomes with high estimated completeness. Sulfide production was observed in 60°C incubations, with corresponding enrichment of Desulfohalobium and Desulfovibrionaceae that possess complete pathways for coupling mannose and acetate oxidation to sulfate reduction. These findings outline how production of fermentation intermediates (mannose and acetate) by Halanaerobium in topside settings can result in reservoir souring when these metabolites are introduced into the subsurface through produced water reuse.

Hydraulically fractured shale oil reservoirs are ideal for studying extremophiles such as thermohalophiles. During hydraulic fracturing, reservoir production water is stored in surface ponds prior to reuse. Microorganisms in these systems therefore need to withstand various environmental changes such as the swing between warm downhole oil reservoir temperatures and cooler surface conditions. While most studies on hydraulically fractured oil reservoirs mimic the environmental conditions found in oil wells, this study follows this water cycle during fracking and the associated microbial metabolic potential during topside-produced water storage and subsurface oil reservoir conditions. Of particular interest are members of the genus Halanaerobium that have been reported to reduce thiosulfate contributing to souring of oil reservoirs. Here, we show that some Halanaerobium strains were unable to grow at hotter temperatures reflective of oil reservoir conditions and lack genes for thiosulfate reduction, despite the proposed importance of this metabolism in other studies. Rather, it is likely that these organisms metabolize complex organics in fracking fluids at lower temperatures, thereby generating substrates that support reservoir souring by thermophilic sulfate-reducing bacteria at higher temperatures. In this way, Halanaerobium promotes souring indirectly by feeding sulfate-reducing microorganisms fermentation products (e.g., acetate and hydrogen) rather than via direct sulfidogenesis via thiosulfate reduction. Therefore, the novelty of this research is not within the detection of known oil reservoir colonizing bacteria but rather in the relationship between bacteria and the indirect involvement of Halanaerobium, promoting souring throughout the produced water reuse cycle.