Targeted Acclimation Unlocks Adaptive Evolution of a Methanotrophic Consortium Enabling 3A5MI Elimination and Enhanced Sulfamethoxazole Biodegradation

Targeted pollutant exposure is widely used to acclimate microbial communities for enhanced biodegradation of recalcitrant contaminants, yet the evolutionary mechanisms underlying functional reinforcement remain poorly understood. Here, we acclimated a methanotrophic consortium achieving efficient removal of 3-amino-5-methyl-isoxazole (3A5MI) (>90%, >5 mg/L/d) and elucidated the adaptive evolutionary processes behind it. Analyses of mobile genetic elements (MGEs) and horizontal gene transfer (HGT) revealed that dominant Methylococcaceae members served as genetic exchange hubs in the acclimation bioreactor. Integrated metagenomic and metatranscriptomic analyses showed that prolonged 3A5MI exposure activated their MGEs and promoted extensive HGT of genes related to energy generation, oxidative stress defense, and biosynthesis. This adaptive evolution enabled community-level metabolic rewiring, including optimized carbon metabolism to relieve energy limitation, niche differentiation, and specialized transcription of C–N bond catalytic functions. Furthermore, batch experiments and transformation product analyses confirmed that 3A5MI-induced functional traits (e.g., heterocycle hydroxylation and C–N bond catalysis) facilitated complete sulfamethoxazole (SMX) biodegradation. Overall, this study demonstrates the evolutionary plasticity of methanotrophic consortia under targeted acclimation and highlights MGE-driven genetic exchange and metabolic adaptation as key mechanisms that both underpin functional enhancement and support the development of methanotroph-based strategies for the biodegradation of recalcitrant isoxazole-based pollutants.