Succession of toxicity and microbiota in hydraulic fracturing flowback and produced water in the Denver-Julesburg Basin.

Hydraulic fracturing flowback and produced water (FPW) samples were analyzed for toxicity and microbiome characterization over 220 days for a horizontally drilled well in the Denver-Julesberg (DJ) Basin in Colorado. Cytotoxicity, mutagenicity, and estrogenicity of FPW were measured via the BioLuminescence Inhibition Assay (BLIA), Ames II mutagenicity assay (AMES), and Yeast Estrogen Screen (YES). Raw FPW stimulated bacteria in BLIA, but were cytotoxic to yeast in YES. Filtered FPW stimulated cell growth in both BLIA and YES. Concentrating 25× by solid phase extraction (SPE) revealed significant toxicity throughout well production by BLIA, toxicity during the first 55 days of flowback by YES, and mutagenicity by AMES. The selective pressures of fracturing conditions (including toxicity) affected bacterial and archaeal communities, which were characterized by 16S rRNA gene V4V5 region sequencing. Conditions selected for thermophilic, anaerobic, halophilic bacteria and methanogenic archaea from the groundwater used for fracturing fluid, and from the native shale community. Trends in toxicity echoed the microbial community, which indicated distinct stages of early flowback water, a transition stage, and produced water. Biota in another sampled DJ Basin horizontal well resembled similarly aged samples from this well. However, microbial signatures were unique compared to samples from DJ Basin vertical wells, and wells from other basins. These data can inform treatability, reuse, and management decisions specific to the DJ Basin to minimize adverse environmental health and well production outcomes.