Time Course-Dependent Methanogenic Crude Oil Biodegradation: Dynamics of Fumarate Addition Metabolites, Biodegradative Genes, and Microbial Community Composition.

Courtney R A Toth,Lisa M Gieg

doi:10.3389/fmicb.2017.02610

Abstract

Biodegradation of crude oil in subsurface petroleum reservoirs has adversely impacted most of the world's oil, converting this resource to heavier forms that are of lower quality and more challenging to recover. Oil degradation in deep reservoir environments has been attributed to methanogenesis over geological time, yet our understanding of the processes and organisms mediating oil transformation in the absence of electron acceptors remains incomplete. Here, we sought to identify hydrocarbon activation mechanisms and reservoir-associated microorganisms that may have helped shape the formation of biodegraded oil by incubating oilfield produced water in the presence of light (°API = 32) or heavy crude oil (°API = 16). Over the course of 17 months, we conducted routine analytical (GC, GC-MS) and molecular (PCR/qPCR of assA and bssA genes, 16S rRNA gene sequencing) surveys to assess microbial community composition and activity changes over time. Over the incubation period, we detected the formation of transient hydrocarbon metabolites indicative of alkane and alkylbenzene addition to fumarate, corresponding with increases in methane production and fumarate addition gene abundance. Chemical and gene-based evidence of hydrocarbon biodegradation under methanogenic conditions was supported by the enrichment of hydrocarbon fermenters known to catalyze fumarate addition reactions (e.g., Desulfotomaculum, Smithella), along with syntrophic bacteria (Syntrophus), methanogenic archaea, and several candidate phyla (e.g., “Atribacteria”, “Cloacimonetes”). Our results reveal that fumarate addition is a possible mechanism for catalyzing the methanogenic biodegradation of susceptible saturates and aromatic hydrocarbons in crude oil, and we propose the roles of community members and candidate phyla in our cultures that may be involved in hydrocarbon transformation to methane in crude oil systems.

Highlights

Methanogenic hydrocarbon degradation is the leading model to explain the widespread occurrence of biodegraded oils and gas formation in oxidant-free reservoirs, whereby lighter oil components are transformed to heavier forms that are of lower quality and more challenging to recover (Head et al, 2003, 2014; Larter et al, 2008)
We identified a total of four peaks in the light oil-amended culture with MS fragment ions corresponding to propane or butane fumarate addition products, suggesting that hydrocarbon activation was occurring at both the primary and secondary carbon atom (Figure 3; Kniemeyer et al, 2007)
At T8 and T12, the concentration of most alkylsuccinates was below detectable limits (≤10 nM), and cumulative aromatic/other hydrocarbon metabolites decreased to 2.6–7.6 μM; up to 52 times less than concentrations observed at T2 and T4 (Figure 2)

Summary

Introduction

Methanogenic hydrocarbon degradation is the leading model to explain the widespread occurrence of biodegraded oils and gas formation in oxidant-free reservoirs, whereby lighter oil components (e.g., saturates and aromatic hydrocarbons) are transformed to heavier forms that are of lower quality and more challenging to recover (Head et al, 2003, 2014; Larter et al, 2008). Research in the past two decades has demonstrated fumarate addition as a possible anaerobic activation mechanism for nalkanes and alkyl-substituted aromatics (Beller and Spormann, 1997; Annweiler et al, 2000; Beller and Edwards, 2000; Kropp et al, 2000; Rabus et al, 2001; Kniemeyer et al, 2003; RiosHernandez et al, 2003; Wilkes et al, 2003; Cravo-Laureau et al, 2005; Davidova et al, 2005).

Methods

Results

Conclusion