Metagenomics Microbial Characterization of Production and Process Fluids in the Powder River Basin: Identification and Sources of Problematic Microorganisms Associated with SWD Facilities
Authors: Enzien Michael , Starustka Sadie , Gurecki Michael , Fincher-Miller Trinity , Kuhn Bryce , Sowecke Carly , Jones Kody , O'Sullivan Kevin , Norris Kyle , Stidham Jason 
Published: November 2021

Abstract
Inconsistent bacterial control and monitoring led to variability in Salt Water Disposal (SWD) well performance and injectivity creating excess costs in biocide applications and remedial work. A metagenomics study using Whole Genome Sequencing (WGS) was conducted to determine the source(s) of problematic microorganisms throughout the process life cycle: Freshwater&gt; Drilling&gt; Completion&gt; Flowback&gt; Produced water&gt; SWD.
A total of 30 metagenomes were collected from the 6 process stages and identification and quantification of the major microbial taxa from each of these stages were identified. "Taxonomy to Function" associations were identified for all the major taxa found in the SWD fluids. WGS was performed on positive Sulfate Reducing Bacteria (SRB) and Acid Producing Bacteria (APB) media bottles inoculated in the field for a Flowback sample.
Four of the six major taxa found in SWD samples are considered groups of microorganisms known to cause microbiologically influenced corrosion (MIC): Clostridia, methanogens, SRB and Iron Reducing bacteria. Thermovirga and Thermotagae, were the two most abundant taxa found in SWD samples, both thermophilic halophilic fermenting bacteria. The Fe reducing bacteria Shewanella was only detected in Drilling and SWD fluids suggesting its source was Drilling fluids. Completion fluid metagenome profiles from two separate sites followed similar patterns. During middle of completions Proteobacteria phyla were dominant taxa represented mostly by Pseudomonas. Other abundant phyla were all characteristic of polymer degrading bacteria. None of these taxa were dominant populations identified in SWD waters. Fresh water only shared similar taxa with Drilling and Completion fluids. A few minor taxa from Drilling and Completion stages show up as significant taxa in SWD fluids. The majority of taxa found in SWD samples appear to originate from Flowback and Produced waters, although at lower abundances than found in SWD samples. It cannot be determined if the microorganisms found in Flowback and Produced waters were endemic to the formation or come from contaminated source waters, i.e. process equipment used to store and transport water sources.
Petrotoga mobilis was the dominant population of bacteria that grew in both media bottles, 96% and 77% for SRB and APB, respectively, while Petrotoga was detected at 14% in the field sample. The most abundant bacteria detected in field sample were Clostridia (38%) while only 2.7% were detected in APB media. SRB media bottle had 0.18% SRB detected by WGS; APB media had 9% SRB population abundance. No SRB were detected in corresponding field sample or below detectable limits (BDL) for WGS methods (&lt;0.01%).
WGS was forensically used to successfully identify type and source of problematic microorganism in SWD facilities. Results from media bottle and field sample comparisons stress the importance of developing improved field monitoring techniques that more accurately detect the dominant microorganisms.