Complete biodegradation of fungicide carboxin and its metabolite aniline by Delftia sp. HFL-1

Fungicide carboxin was commonly used in the form of seed coating for the prevention of smut, wheat rust and cotton damping-off, leading carboxin and its probable carcinogenic metabolite aniline to directly enter the soil with the seeds, causing residual pollution. In this study, a novel carboxin degrading strain, Delftia sp. HFL-1, was isolated. Strain HFL-1 could use carboxin as the carbon source for growth and completely degrade 50 mg/L carboxin and its metabolite aniline within 24 h. The optimal temperatures and pH for carboxin degrading by strain HFL-1 were 30 to 42 degrees C and 5 to 9, respectively. Furthermore, the complete mineralization pathway of carboxin by strain HFL-1 was revealed by High Resolution Mass Spectrometer (HRMS). Carboxin was firstly hydrolyzed into aniline and further metabolized into catechol through multiple oxidation processes, and finally converted into 4-hydroxy-2-oxopentanoate, a precursor of the tricarboxylic acid cycle. Genome sequencing revealed the corresponding degradation genes and cluster of carboxin. Among them, amidohydrolase and dioxygenase were key enzymes involved in the degradation of carboxin and aniline. The discovery of transposons indicated that the aniline degradation gene cluster in strain HFL-1 was obtained via horizontal transfer. Furthermore, the degradation genes were cloned and overexpressed. The in vitro test showed that the expressed degrading enzyme could efficiently degrade aniline. This study provides an efficient strain resource for the bioremediation of carboxin and aniline in contaminated soil, and further revealing the molecular mechanism of biodegradation of carboxin and aniline.