On-Site Study of Lipophilic Bacteria in the Bioremediation of Polycyclic Aromatic Hydrocarbons (PAH) from Automotive Grease / Engine Oil Spills

In this study, the isolated strains that are able to grow on grease / oil contaminated spills and used as a
sole carbon source were isolated from Coimbatore regions. The physicochemical analysis of oil
contaminated soil revealed a pH range of 6.8–8.4, indicating a slightly acidic to neutral environment
suitable for microbial growth. Electrical conductivity remained below 2 dS/m, supporting microbial
viability. Enrichment techniques yielded eight distinct bacterial isolates, initially characterized by
morphology, Gram staining, and biochemical tests. Molecular identification via 16S rRNA confirmed and
MALDI-TOF analysis identified two isolates with high confidence, including species like Pseudomonas
stutzeri and Exiguobacterium aurantiacum. FTIR spectra confirmed the presence of polycyclic aromatic hydrocarbons (PAHs), validating the soil for bioremediation studies. It showed reduction in hydrocarbon
peaks and appearance of new functional groups, suggesting PAH degradation. Spray plate assays
revealed clear degradation zones with Pseudomonas stutzeri and also showing the most effective PAH
breakdown than the Exiguobacterium aurantiacum. The Dioxygenase enzyme assays further supported
these findings, showing highest activity in Pseudomonas stutzeri and Exiguobacterium aurantiacum. SDS
PAGE analysis displayed distinct protein bands between 53–78 kDa, with a prominent 37 kDa band in
strong degraders, possibly linked to degradative enzymes.
Keywords: Polycyclic Aromatic Hydrocarbons, Biodegradation, Spray Plate technique, Dioxygenase
enzyme, Phenanthrene