抗生素诱导下的农杆菌耐药性及RAPD分析

Studies on Drug-resistance of Agrobacterium tumefacien Induced by Antibiotics

用头孢噻肟、亚胺培南和美罗培南等抗生素对农杆菌EHA101、LBA4404和GV3101进行诱导,检测其对诱导后3种农杆菌的最小抑菌浓度(MIC)、最小杀菌浓度(MBC)和耐药性变化。结果表明,3种农杆菌在含一定浓度抗生素的培养基上培养12～18d后,表现出MIC和MBC值的增加。经耐药性标准鉴定,达到中度敏感或耐药。对筛选到的耐药菌株基因组进行RAPD-PCR随机扩增,探讨其基因组DNA多态性是否存在差异。结果RAPD-PCR扩增产生的条带多态性发生变化,出现特异性条带的增加和缺失。所筛选到的特异性片段的产生和缺失可能与农杆菌耐药性产生相关。

Agrobacterium tumefaciens strains including EHA101,LBA440d and GV3101 were cultured in LB medium adding cefotaxime, imipenem and meropenem. And the effect of these three antibiotics on drug-resistance of three strains of Agrobacterium tumefaciens was determined according to the variation of MIC and MBC. The results showed that the values of MIC and MBC of these three strains increased when cultured for 12-18 days on the media adding antibiotics. Random amplifications with the 6 RAPD polymorphism primers for the 16 Agrobacterium tumefaciens of moderate sensitive or resistant to cefotaxime, meropenem and imipenem, were preformed in order to explore variations of Agrobacterium tumefaciens genomic DNA polymorphism. The results showed that compared with strains without induction, the strains with susceptibilities to antibiotics had several polymorphism changes with the specific fragments＇ increase or loss after induction by the antibiotic. These specific fragments would have relationship with the production of antibiotic resistance of Agrobacterium tumefaciens.