Highly efficient in-situ cleaner degradation of difenoconazole by two novel dominant strains: Microflora diversity, monoclonal isolation, growth factor optimization, intermediates, and pathways

The non-point source pollution of difenoconazole (DIF) has become a serious environmental issue, increasingly causes indelible damages to eco-environment and human health due to its toxicity, persistence, and biomagnification. An eco-friendly, cost-effective, and efficient control technology is imperative towards a cleaner and sustainable agricultural production. Herein, a dominant microflora of efficiently degrading DIF was successfully screened, and its microbial diversity was investigated. Two novel degrading strains were isolated and identified as Phyllobacterium sp. (T-1) and Aeromonas sp. (T-2). The results of growth factor optimization indicated that the degradation rates of DIF (C₀ = 20 mg/L) by strain T-1 and T-2 were up to 96.32% and 97.86% within 14 d, respectively, under the optimal conditions. Moreover, there no obvious synergy between strain T-1 and strain T-2. From catalytic kinetics of enzymes, the intracellular enzyme of strain T-1 dominated the degradation of DIF (C₀ = 20 mg/L) entirely with the degradation rate of 82.4% (48 h), the extracellular enzyme showed little catalytic activity. However, the degrade rates of DIF (C₀ = 20 mg/L) by both intracellular and extracellular enzymes of strain T-2 were 77.99% and 26.73% within 48 h, respectively. Moreover, these enzymes remained an undiminished catalytic activity within 48 h. DIF was degraded by strain T-1 to three main transformation products (DIF-TPs 406, DIF-TPs 216, and DIF-TPs 198) undergoing hydroxyl substitution, hydrolysis, cleavage of ether bond between benzene rings, and rearrangement, while two additional products (DIF-TPs 281 and DIF-TPs 237) were generated with the biodegradation of strain T-2, excepting for DIF-TPs 406 and DIF-TPs 216, involving hydrolysis, hydroxylation, and ether bond cleavage between benzene rings. Moreover, QSAR simulation showed that the by-products were almost much lower toxicity or even non-toxic to three typical aquatic organisms (fish, daphnia, and green algae) than DIF. This study not only provides an in depth understanding of DIF bioelimination, but also be instrumental in cleaner management of DIF-contaminated soil. This study can promote the sustainable development of agriculture.