油菜菌核病菌抗腐霉利菌株的生理生化特性及抗药机制初步分析

通过室内紫外线和药剂诱导,获得多株高抗腐霉利油菜菌核病菌（Sclerotiniasclerotiorum）突变体。与野生亲本菌株相比,抗性突变株（EC50〉800μg/mL）在高糖（〉40g/L蔗糖）和高盐（〉5g/LNaCl）培养基上生长缓慢,表明对高渗透压比较敏感;菌丝相对电导率较高,显示细胞膜透性增大,胞内电解质渗出较多。试验还发现,在含药培养液中,亲本菌株菌丝内甘油大量合成和积累,增加462.45%,而抗药突变株甘油含量仅稍有增加或有所下降,说明抗腐霉利突变株对外界渗透压的调节能力较低。根据已报道组氨酸激酶（HK）基因序列设计特异引物,经PCR扩增和测序,获得油菜菌核病菌HK基因保守区域序列1605bp。抗、感腐霉利菌株HK基因保守区域在第45位、第81位和第625位碱基不同,但编码的氨基酸一致。因此,油菜菌核病菌对腐霉利的抗性及抗药菌株的高渗敏感性与HK基因保守区域碱基突变无关。

Physiological and biochemical features of rocymidoneresistant mutants to Sclerotinia sclerotiorum and preliminary study of resistance mechanism

Several procymidoneresistant mutants of Sclerotinia sclerotiorum causing rape stem rot were obtained by UV or procymidone induction.They grew slower （EC50 800μg/mL） than their parent on media containing sucrose （〉40g/L） or NaCl （〉5g/L）,indicating that the mutants were more sensitive to high osmotic stress.The relative conductivity of mutant mycelia was higher than that of the parent,which led to cell electrolyte leakage.The glycerol in parent mycelia increased quickly and accumulated by 462.45% in procymidone media,while that of the mutant was hardly changed,which implied weaker regulatory ability of mutant to osmotic stress.Moreover,sequence differences were found in histidine kinase （HK） gene of both wild parent and mutants.A 1 605bp product was amplified from conserved domain of HK gene.Three bases at 45^th,81^th and 625^th sites were different among mutants and parent,although the coded amino acids were the same.It suggested that the procymidone resistance and high osmotic stress sensitivity of S.sclerotiorum mutants had no correlation to the base mutation of HK gene conserved domain.