Abstract:
Engine oil is one of the most significant sources of energy on the planet. As the usage of petroleum
hydrocarbon products increases, soil contamination with petroleum oil is becoming one of the
major environmental problems. As a result of this, global interest in the microbial biodegradation
of petroleum pollutants has grown recently. The aim of this study was isolation and molecular
characterizations of engine oil degrading bacteria isolates from engine oil-contaminated soil in
Gondar city, Ethiopia. Soil bacteria were isolated by an enrichment culture technique using
minimal salt medium with 1% engine oil as a carbon source, and Screening of potential engine oil
degrading bacterial isolates were performed by using hole plate diffusion techniques. Engine oil
percentage degradation was determined spectrophotometrically as the relative decrease of
absorbance wavelength of the medium before and after treatment at OD420 nm. The potential
bacterial isolates were molecularly identified using 16S rRNA gene sequencing. A total of 41
bacterial cultures were isolated. Out of 41 bacterial isolates seventeen isolates had able to form a
clear zone around the hole on 1% oil medium, but only three Gram-negative bacterial isolates
Ar3A, Az2B, and Ar2B showed a better clear zone on 1% engine oil medium with diameters of
36.33±306 mm, 26.0±2.000 mm, and 22.67±306 mm, respectively. Bacterial isolate Ar3A was
found to be the best oil degrading isolate in this study with 70.033±1.155% degradation after a
21day incubation period while 65.923±0.414% and 64.39±1.507% degradation was observed
using isolate Az2B and Ar2B, respectively. The 16S rRNA gene sequences percent identity of Ar3A
was 94.5 % with Stenotrophomonas acidaminiphila, Az2B was 91.53% with Citrobacter
amalonaticus and Ar2B was 92.9% with Klebsiella pneumoniae. PCR experiments with specific
primers for catabolic genes of AlkB and C23O suggested that only Stenotrophomonas
acidaminiphila strain Ar3A can possess both genes with 660 bp and 500 pb, respectively. In
conclusion, the three bacteria isolates should be considered as good prospects for their
application in the bioremediation of engine oil sludges. However, Further studies could be
conducted on the possibility of cloning AlkB and C23O genes of Stenotrophomonas acidaminiphila
into other bacteria for more efficiency and effectiveness in the bioremediation process.
Keywords: AlkB gene, Bioremediation, C23O gene, and engine oil-degrading bacteria
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1. INTRODUCTION
1.1. Background of the Study
The global environmental problem of engine oil contamination and its derivatives is a
