体外诱导禽巴氏杆菌链霉素耐药株及耐药机制的初步分析

摘要

为了研究多杀性巴氏杆菌(Pasteurella multocida,Pm)对链霉素(Streptomycin,Str)药物敏感性变化及其耐药机制,采用体外诱导法对禽源Pm亲本敏感株(C48-1)进行诱导后成功获得Str耐药株,并对Str耐药株的最小抑菌浓度(MIC)、生化特性、生长曲线、耐药稳定性、耐药基因点突变以及耐药逆转性进行了研究.结果表明,与亲本株相比,Str耐药株生化特性与生长曲线无显著差异,MIC值由8 μg/mL上升至1024 μg/mL;发现耐药基因StrA、StrB,且核糖体基因rpsL编码的氨基酸发生了Lys43→Thr突变;耐药逆转性试验中羰基氰氯苯腙(CCCP)将Str耐药株MIC值降为16 μg/mL.逐步递增药物浓度可以诱导禽源Pm对氨基糖苷类药物的耐药性,其耐药机理为磷酸转移酶类对氨基糖苷类抗生素结构的修饰作用,核糖体靶位点突变和外排泵的过量表达.%In order to study the drug susceptibility to sreptomycin (Str) and its drug resisitance mechanism of avian Pasteurella multocida (Pm),a Str-resistant avian Pm was obtained via in vitro induction with gradually increasing drug concentrations to the standard strain named C48-1. The minimal inhibitory concentration (MIC),biochemical characteristics, growth curve,stability of drug resistance,drug resistance gene site mutation and drug resistance reversal of the Str-resistant strain was analyzed. The results showed that compared with the parent strain, the biochemical characteristics and growth curve had no significant difference,the MIC of the Str-resistant strain was increased from 8 μg/mL to 1 024 μg/mL. What more,the drug resistance gene StrA and StrB were found and the amino acid sequence encoded by the rpsL gene had an amino acid exchange of Lys43→Thr. In the drug resistance reversal test, carbonyl cyanide-m-chlorophenylhydrazone (CCCP) reduced the MIC of the Str-resistant strain to 16 μg/mL. In total,it shows that gradually increasing drug concentration can induce Pm resistance to aminoglycosides in vitro. Its mechanism is the modification of aminoglycoside antibiotics by phospho-transferase, the mutation of ribosomal protein and the over expression of efflux pumps.