Molecular characterization of biosurfactant producing marine bacterium isolated from hydrocarbon-contaminated soil using 16S rRNA gene sequencing

Abstract

Objectives: This study was aimed to isolate and identify the biosurfactant producing marine bacteria from petroleum hydrocarbons contaminated sediments by 16S rRNA gene sequencing. Methods: Soil sample was enriched with diesel (2%) as the sole source of carbon in minimal salt medium for 3 weeks at 28 °C ± 2. Enriched sample was plated on nutrient agar and the organisms are selected based on the colony morphology. Biosurfactant producing efficiency of the isolates was assessed using blood haemolysis, oil displacement test, microplate assay (drop collapse test) and emulsification activity. Identification of the efficient biosurfactant producing isolate was done by gram staining and biochemical tests. Molecular characterization was done using 16S rRNA gene sequencing. Results: Totally seven isolates were selected based on colony morphology and among them, the isolate ADY2b was able to reduce surface tension significantly in the oil displacement test and also formed a stable emulsion. The emulsification index (E24) had shown a promising result of 58.33%. The isolate ADY2b was a motile, gram negative rod shaped bacterium and positive for catalase and oxidase. Sequence alignment of the 16S rRNA of the ADY2b strain and database search revealed 98% similarity to Pseudomonas mendocina by BLAST analysis. Conclusion: Seven bacteria were isolated from petroleum hydrocarbon contaminated sediments. Strain ADY2b had shown promising results on the production of biosurfactants and it is capable of increasing the bioavailability of poorly soluble petroleum hydrocarbons. The isolate Pseudomonas mendocina ADY2b obtained in the study had potential to be used in oil degradation purposes.

Keywords

• Biosurfactants
• Petroleum hydrocarbons
• 16S rRNA
• Microplate assay
• Emulsification activity (E24)
• Pseudomonas species