Royal Military College of Canada

Environment

Fate and Effects of Hydrocarbon-Degrading Bacteria Used to Inoculate Soil for On-Site Bioremediation in the Arctic

By: Captain Eric Thomassin-Lacroix

Abstract

Numerous Arctic and sub-Arctic sites have been contaminated with a variety of petroleum hydrocarbons such as diesel and jet fuel. The low ambient temperature and short treatment season in the Arctic is hampering effective biodegradation of hydrocarbon-contaminated soil. The main goal of this project was to study the effect of bioaugmentation for on-site bioremediation of hydrocarbon-contaminated soil at Canadian Forces Station (CFS) Alert, Ellesmere Island, Nunavut.

The inoculum used for this project was enriched from soil at CFS Alert for its capability to degrade jet fuel at low temperature. The three most abundant organisms in the enrichment culture were identified through 16S ribosomal DNA (rDNA) analysis. These organisms showed high 16S rDNA similarity to Rhodococcus erythropolis, Sphingomonas sp. UN1F1, and Pseudomonas synxantha. Three specific polymerase chain reaction (PCR) primer sets were designed for these strains, and a PCR-most probable number (PCR-MPN) assay was developed to monitor their fate, growth, and possible spread to nearby locations during a field experiment.

It has been shown in previous studies that Arctic soil contains indigenous microorganisms with hydrocarbon-degrading capabilities. In experiments done at the University of British Columbia under the supervision of Dr. William Mohn, Eric Thomassin-Lacroix characterized and identified a hydrocarbon-degrading consortium from CFS Alert, Nunavut, sequencing the 16S ribosomal DNA (rDNA) gene. He then used molecular biology tools including the polymerase chain reaction (PCR) to develop probes for species-specific detection and quantification of inoculated bacteria in a field experiment at CFS Alert. His experiment was important in order to determine if bioaugmentation is needed in the bioremediation of hydrocarbon-contaminated soil in the Arctic.

Results showed that both inoculated and inoculated treatments presented and average total petroleum hydrocarbon (TPH) removal of 75% over 65 days. This result suggests that the density of the inoculum that was used (approximately 108 cells per g dry soil) was not large enough to stimulate hydrocarbon removal and that the indigenous population is already well adapted to biodegrade these petroleum hydrocarbons. Two of the three phylotypes were present in greater numbers in the inoculated biopiles at 0 days than in the uninoculated ones.

After 65 days of treatment, populations of phylotypes were similar in both treatments, except for one phylotype (Ale-1.14) which was less abundant in the uninoculated densities nad the fate of the three strains and showed similar results than the field experiment. The PCR assays were also used to measure the dynamics of the three strains in the enrichment culture through time. The results of this investigation showed that bioaugmentation did not stimulate hydrocarbon biodegradation at CFS Alert and also that the indigenous hydrocarbon-degrading microflora is abundant and already adapted to fulfill this task at this location. The conclusion of this work will be directly applied to further on-site applications in the remediation of hydrocarbon-contaminated soil in the Arctic.