Isolation, Classification And Degradation Characterisics Study Of Nicosulfuron-degrading Strain

Sulfonylurea herbicides have been widely used in agriculture since 1970 s for its high efficiency, broad-spectrum activity and low toxicity to mammals. In 2007, the utilization of commercial herbicides have accounted for approximately 11 % of the global herbicide utilization. However, with the overuse of sulfonylurea herbicides, the residual sulfonylurea herbicides in the soil showed toxic effect to the subsequential crop generation, lead to a decline of yield, and even threaten the aquatic environment through surface runoff and biogeochemical cycle. It is proved that biocatalysis of residual sulfonylurea herbicides existing in the environment is a major way to solve this problem.In this study, strain LAM-WHM-ZC and LAM0415 Twere identified and named as Oceanisphaera psychrotolerans and Bacillus luzhouensis, according to various taxonomy principles such as morphological properties, chemical and molecular analysis, respectively.Strain LAM-WHM-ZC degraded 83.8 % of nicosulfuron in a minimal culture medium M1 with 50 mg/L nicosulfuron as sole carbon source within 8 d. In addition,strain LAM-WHM-ZC had a broad degrading spectrum of herbicide including tribenuron methyl, rimsulfuron, chlorsulfuron and bensulfuron methyl. Then strain LAM-WHM-ZC was used to remediate a simulated nicosulfuron-contaminated soil in lab-scale. The results showed that LAM-WHM-ZC degraded 73.7 % of 20 mg/kg nicosulfuron after 15 d. The metabolic pathway of nicosulfuron by strain LAM-WHM-ZC was investigated and nine intermediate metabolites were identified by HPLC-MS. The mainly degrading process was the cleavage of the sulfonylurea bridge, then forming non-activated intermediate metabolites.Genome sequences of strain LAM-WHM-ZC was obtained from Ion Torrent sequencing platform. The genome of strain LAM-WHM-ZC has 3,733 genes with a total sequence length of 3.348 Mb. 9 esterase genes were predicted based on the swiss-prot datebass in the genome sequences.A new nicosulfuron gene e3 was cloned from LAM-WHM-ZC genome, and it consists of 774 bp, encoding a protein of 257 amino acids. Not only Ser-Glu-His catalytic triad but also the conserved pentapeptide sequence(Gly-X1-Ser-X2-Gly)around the catalytic serine residue of most alpha/beta-hydrolase fold proteins hasbeen found in E3, which was cloned and purified in current study. The effects of temperature, p H, metal ions and surfactants on enzymatic activity were invsetigaed.The results showed that E3 exhibited a high efficiency in degrading nicosulfuron at a broad temperature(25-60 oC) and p H(7.0-8.5). All the results suggested that protein E3 has a good potential for practical use in bioremediation of nicosulfuron-contaminated wastes.