A hybrid approach utilising adaptive laboratory evolution and gene expression analysis: How adaptive evolution improves PAHs biodegradation.

Polycyclic aromatic hydrocarbons (PAHs) are persistent environmental contaminants that pose significant ecological and human health risks due to their toxicity, hydrophobicity, and resistance to degradation. Although microbial biodegradation offers a sustainable remediation strategy, its efficiency in natural environments is often constrained by physicochemical stress factors such as salinity, temperature, and pH fluctuations, which limit microbial activity and stability. Therefore, enhancing microbial adaptability and functional performance under such stress conditions remains a critical challenge. In this study, we developed an integrated approach by integrating adaptive laboratory evolution (ALE), transcriptomic analysis, and bioreactor validation to enhance microbial PAHs degradation. Three bacterial strains, Pseudomonas stutzeri MP6, Stutzerimonas stutzeri MP15, and Stutzerimonas frequens MP37, were subjected to ALE under progressively increasing temperature, salinity, and acidic pH conditions to enhance there stress tolerance. Under progressively intensified environmental stress, the evolved microbial consortium demonstrated improved degradation efficiency relative to individual strains, indicating a synergistic interaction. Transcriptomic analysis further revealed increased expression trends in genes associated with aromatic compound metabolism, including oxygenases and ring-cleavage enzymes, suggesting enhanced metabolic activity linked to PAHs transformation. Validation in a soil-slurry system demonstrated that the evolved consortium achieved 85.65% degradation within 90 days under simulated environmental conditions. Overall, these findings suggest that ALE-driven adaptation enhances microbial resilience and functional performance, supporting its potential application in PAHs bioremediation.