Determination of the Bacterial Community Structure in a Crude Oil-inundated Tropical Soil Using Next Generation Sequencing Technique

Abstract

Aim: To identify the community composition of a crude oil impacted soil in Gbarain Kingdom of Bayelsa State, Nigeria. Study Design: A crude oil-impacted soil sample (0-10 cm depth) was collected from Etelebuo-Ogboloma, flow station in Yenagoa L.G.A. of Bayelsa State and taken to the laboratory for various microbiological analyses. Place and Duration of Study: The study was carried out at the Environmental Microbiology Laboratory, University of Port Harcourt, Nigeria, for 35 days. Methodology: Cultural morphology of the isolates was studied based on their physical appearances such as colour, shape, size, elevation and margin. While catalase test, oxidase test, indole test, motility test, Methyl-Red Voges-Proskauer’s (MRVP) test and citrate utilization test were used for the biochemical identification of the isolates. Deoxyribonucleic acid (DNA) extraction from crude oil polluted soil sample was performed using ZYMO soil DNA extraction Kit (Model D 6001, Zymo Research, USA) following the manufacturer’s instructions. DNA sequencing was performed by the Next Generation Sequencing Technique to determine the nucleotide sequence of all microorganisms present in the soil sample using sequencing primer -16S: 27F: 5’-GAGTTTGATCCTGGCTCAG-3’ and 518R: 5’- ATTACCGCGGCTGCTGG-3’. Results: Five different bacterial genera were isolated and identified using the cultural techniques, and they include Acetobacter sp., Pseudomonas sp., Arthrobacter sp., Bacillus sp. and Micrococcus sp. However, the molecular characterization revealed that the soil was mainly dominated by the Alphaproteobacteria (54.64%), followed by the Actinobacteria (9.67%), Gammaproteobacteria (6.55%), Betaproteobacteria (2.27%) and Bacilli (0.95%) as well as Clostridia (0.34%); as the most dominant class of bacteria. The unknown group accounted for 25.28%. A total of one hundred and four (104) diverse bacterial species were identified, in the overall metagenomics. Conclusion: This study has shown the bacterial community composition of the crude oil polluted soil obtained from the Gbarain Kingdom. These findings are fundamental to understanding the biological fate of crude oil in these oil rich regions.