|dc.description.abstract||Human activity in Antarctica has resulted in accidental fuel spills on soils. These are broadly damaging to the Antarctic ecosystem, and expensive to clean up completely by physical removal, therefore, alternative remediation techniques, like bioremediation, are required. Hydrocarbon-degrading bacteria have been isolated from Antarctic soils, but few biotreatability studies have been described. On the basis that limiting factors to bioremediation can be manipulated to enhance biodegradation of contaminated soils, this thesis research project aimed to test these limiting factors to contribute to the development of bioremediation protocols for hydrocarbon-contaminated soils from the Ross Sea Region, Antarctica.
Hydrocarbon-contaminated and control uncontaminated soils collected from Scott Base, Marble Point and Wright Valley were chemically analysed for total hydrocarbons and chemical properties, and analysed for total heterotrophic and hydrocarbon-degrading microbes. Both contaminated and uncontaminated control soils were screened for the alkane catabolic genes Pp alkB, Rh alkBJ, Rh alkB2 and Rh alkB194 by polymerase chain reaction (PCR) and sequenced to determine the soils' alkane-degrading genetic potential. Additionall, hydrocarbondegrading bacteria isolated from Scott Base soils were characterised morphologically and screened for alkB gene homologues. The degradative ability of microbes was assessed by ¹⁴C-hexadecane mineralisation in soil microcosms at 15°C in all untreated soils, water and nitrogen amended Scott Base soils, Scott Base contaminated soil diluted 1 : 1 with uncontaminated soil, and Bull Pass soil bioaugmented with strains Rhodococcus sp. 5/1 and SB0-1, a hydrocarbondegrading microbial isolate from this study. Molecular analysis of 16S rDNA of SB0-1 showed DNA sequence homology of 99% with Rhodococcus sp. 5/14. Changes in microbial populations in the most successful enhancement treatments were detected by denaturant gradient gel electrophoresis (DGGE).
Hydrocarbon-degraders were found only in Marble Point and Scott Base soils (from 10³ to 10⁶ colonies per gram dry weight soil). The most abundant alk gene in the soils was Pp a/kB, followed (in deceasing order of abundance) by Rh a/kB 1, Rh alkB2 and Rh alkB194. This indicated the possible presence of Pseudomonas and Rhodoeoeuss species in these soils. High rates of mineralisation were detected in Marble Point, ea. 60% in 75 days, and low rates in Scott Base soil, ea. 4% in 75 days. Nitrogen overfertilisation and water adjustments decreased mineralisation, but addition of 2500 mg N/kg-H20-soil combined with 10% moisture enhanced mineralisation ea. 4 times in a recent and old spill site in Scott Base soil, indicating that nitrogen was a limiting factor. Dilution of this soil with uncontaminated soil resulted in enhanced mineralisation, 8 times that of unamended soil, reaching ea. 40% mineralisation. DGGE showed that the diluted soil was enriched in a Rhodoeoeeus species. Dilution of the soil, and consequently of the contaminant, was the most successful treatment for Scott Base soil. This indicated that contaminant concentration was a limiting factor in this soil. Bioaugmentation of Bull Pass soil was partially successful; mineralisation was enhanced after addition of inoculum, but reduction in total hydrocarbons was minimal and attributed to abiotic loss.
Overall, limiting factors to bioremediation can be manipulated to enhance biodegradation by nutrient amendments, dilution of contaminant, and inoculation of hydrocarbon-degraders. The implementation of these treatments may aid in remediation of contaminated soils from the Ross Sea Region.||