施氏假单胞菌YC-YH1对甲基对硫磷的降解及其代谢产物检测

利用实验室已获取的菌株YC-YH1对甲基对硫磷进行降解,以高效液相色谱结合质谱分析(HPLC-MS)测定菌株YC-YH1对甲基对硫磷的降解速率并检测其代谢产物,通过基因克隆研究菌株YC-YH1对甲基对硫磷降解的分子机制,同时分析代谢产物对菌株YC-YH1生长的影响.结果表明,菌株YC-YH1为施氏假单胞菌(Pseudomonas stutzeri),能够高效降解甲基对硫磷,其同时含有甲基对硫磷水解酶基因mpd和有机磷水解酶基因ophC2.经质谱分析确定,菌株YC-YH1将甲基对硫磷水解为对硝基苯酚和二甲基硫代硫酸酯,其中对硝基苯酚显著抑制YC-YH1的生长.综上,施氏假单胞菌YC-YH1能够高效地降解甲基对硫磷,但对硝基苯酚作为其代谢产物对YC-YH1的生长存在显著抑制作用.

Biodegradation of Methyl Parathion by Pseudomonas stutzeri YC-YH1 and Detection of Its Metabolites

A newly isolated bacterial strain YC-YH1, which could degrade methyl-parathion efficiently, was identified according to 16S rRNA gene analysis in combination with physiological, biochemical and Biolog tests. The present study was focused on its capability of degrading methyl parathion. The metabolites of methyl parathion were identified by HPLC-MS. Changes of methyl parathion and its metabolite concentrations in the inorganic medium were determined by HPLC. The primers of genes involved in the methyl parathion degradation were designed and the target fragments were obtained by gene cloning. The influence of degradation intermediates on the growth of the strain YC-YH1 was examined using filter paper and plate methods. The colonic and microscopic morphologies of the strain YC-YH1 was determined. The 16S rRNA gene sequence showed that YC-YH1 had 98% similarity to Pseudomonas stutzeri. Pseudomonas stutzeri YC-YH1 showed high degradation efficiency for parathion-methyl. p-nitrophenol and dimethyl phosphorothioate were identified as the intermediates. p-nitrophenol concentrations increased along with decrease of methyl parathion concentrations in the inorganic medium. However, p-nitrophenol significantly inhibited the growth of the strain YC-YH1 in the inorganic medium. Methyl parathion hydrolase gene mpd and organophosphorus pesticides hydrolase gene ophC2 were both cloned from the strain YC-YH1. These results suggest that the strain YC-YH1 could efficiently degrade methyl parathion, but the degradation intermediate p-nitrophenol could significantly inhibit the growth of the strain YC-YH1.