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. Members of the genus Halanaerobium are ubiquitous in these environments. 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 at 30{degrees}C to simulate above-ground storage conditions, and at 60{degrees}C to simulate subsurface reservoir conditions. Guar metabolism coincided with Halanaerobium enrichment only at 30{degrees}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, Halanaerobium genomes did not uncover genes for this metabolism. Sulfide production was observed in 60{degrees}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, acetate) by Halanaerobium in topside settings can result in reservoir souring when these metabolites are introduced into the subsurface through produced water re-use. ImportanceHydraulically 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 re-use. 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. This study follows this water cycle during fracking and the associated microbial metabolic potential. 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 under oil reservoir temperatures and do not possess genes for thiosulfate reduction. 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.