Combined Use of Diagnostic Fumarate Addition Metabolites and Genes Provides Evidence for Anaerobic Hydrocarbon Biodegradation in Contaminated Groundwater

Abstract

The widespread use of hydrocarbon-based fuels has led to the contamination of many natural environments due to accidental spills or leaks. While anaerobic microorganisms indigenous to many fuel-contaminated groundwater sites can play a role in site remediation (e.g., monitored natural attenuation, MNA) via hydrocarbon biodegradation, multiple lines of evidence in support of such bioremediation are required. In this study, we investigated two fuel-contaminated groundwater sites for their potential to be managed by MNA. Microbial community composition, biogeochemical indicators, fumarate addition metabolites, and genes diagnostic of both alkane and alkyl-monoaromatic hydrocarbon activation were assessed. Fumarate addition metabolites and catabolic genes were detected for both classes of hydrocarbon biodegradation at both sites, providing strong evidence for in situ anaerobic hydrocarbon biodegradation. However, relevant metabolites and genes did not consistently co-occur within all groundwater samples. Using newly designed mixtures of quantitative polymerase chain reaction (qPCR) primers to target diverse assA and bssA genes, we measured assA gene abundances ranging from 105–108 copies/L, and bssA gene abundances ranging from 105–1010 copies/L at the sites. Overall, this study demonstrates the value of investigating fuel-contaminated sites using both metabolites and genes diagnostic of anaerobic hydrocarbon biodegradation for different classes of hydrocarbons to help assess field sites for management by MNA.

Field investigations and reactive transport modelling of biodegrading coal tar compounds at a complex former manufactured gas plant

“…Although the results show some general trends in redox conditions at the site, the results also demonstrate a large level of variability and therefore unpredictability of subsurface conditions at the Griftpark. Similar variations at a single site were documented by Kharey et al at two fuel-contaminated groundwater sites [155]. They found that nitrate, sulphate and iron reduction occurred with varying levels of potential at different locations of the sites, making it challenging to discern clear trends between hydrocarbon concentrations with the electron accepting processes.…”

The Effect of an Adsorbent Matrix on Recovery of Microorganisms from Hydrocarbon-Contaminated Groundwater

“…No nitrate, nitrite, organic acids (such as acetate) or aromatic hydrocarbons were detected in these samples. However, previous groundwater analyses (in 2016) indicated that benzene and toluene levels in this site reached 8.39 mg/L and 0.377 mg/L [ 33 ], both exceeding Canadian maximum allowable concentrations (0.005 and 0.024 mg/L, respectively [ 34 ]). The lack of hydrocarbons detected in the present groundwater samples was likely due to volatilization that occurred during transportation and storage.…”

“…In contrast, microbial communities incubated anoxically failed to oxidize any benzene over 80 days of monitoring ( Figure S3 ). Given the levels of benzene detected at the Stony Plain site in 2016 (8.39 mg/L [ 33 ]), we hypothesized that anaerobic benzene degradation would occur due to its historical exposure to the indigenous microbial population. However, this was not realized within the 80 day incubation period across the anaerobic conditions tested.…”

Increasing in situ bioremediation effectiveness through field-scale application of molecular biological tools

“…MI performed qPCR utilizing commercially available assays for bssA, assA, mnssA, bcrA, ANC, and APS. It is important to note that some of the commercially available assays involve the use of multiple qPCR assays (five assays for bssA, six for assA, two for mnssA, three for bcrA, and seven for APS) for which quantities are added as previously described (Beller et al, 2002;Rossmassler et al, 2019;Kharey et al, 2020).…”