Biodegradation plays a crucial role in the removal of sulfonamides (SAs) from soils. However, a complete pathway for SA biodegradation by Microbial communities remains unclear. Using [phenyl-U-14C]-labeled SAs, we investigated the degradation of sulfadiazine (SDZ), sulfamonomethoxine (SMM), and sulfamethoxazole (SMX) in an enrichment of SDZ-degrading soil microbial community. The extent of phenyl ring mineralization for the three SAs was 60.4–65.4% in the enrichment culture when SAs served as the sole carbon source. Meanwhile, 16, 14, and 10 metabolites of SDZ, SMM, and SMX were identified, respectively. These metabolites belong to twelve primary transformation pathways, including sulfonamide bond cleavage, desulfonylation, para-amino group modification, and heterocyclic moiety modification. Notably, two desulfonylation pathways were revealed for the first time as routes leading to mineralization. Using 13C-Stable isotope probing and activity assays, we identified a novel species strain, Candidatus Leucobacter nanjingensis H4, as a key sulfadiazine degrader for assimilating its phenyl moiety. The sulfonamide-degrading enzyme in strain H4 belongs to a distinct branch of flavin monooxygenase SadA, revealing that the evolutionary aquisition of sulfonamide degradation capability in Microbacteriaceae and Micrococcaceae families has occurred multiple times. Furthermore, 2-aminopyrimidine, the dead-end metabolite produced by sulfadiazine degradation by strain H4, was fully catabolized by another novel species strain, Ca. Nocardioides nanjingensis T5. These results provide critical insights into the mechanisms of bacterial sulfonamide degradation and the natural attenuation process of sulfadiazine in the environment.

Sulfonamide antibiotics,Biotransformation,Soil microorganisms,DNA-SIP