甲型副伤寒沙门氏菌主动外排多重耐药基因与表达研究

目的 调查临床分离甲型副伤寒沙门氏菌对常用抗生素的耐药情况与多重耐药主动外排基因acrB的检测、序列分析及其表达水平。方法 用琼脂二倍稀释法测定7种抗生素对甲型副伤寒沙门氏菌的抗菌活性。以基因库序列为参考设计引物PCR扩增acrB、测序并用RT-PCR方法检测其表达。结果 12株甲型副伤寒沙门氏菌对氧氟沙星、环丙沙星、头孢噻肟、哌拉西林、氯霉素、四环素、庆大霉素的耐药率除氯霉素为0外。其余均为8．33％。对三类不同种类抗菌药物多种耐药者l株。所有细菌均检测到多重耐药外排基因acrB．测序结果与参考沙门氏菌序列(No．AL627267)比较，有l处碱基差异。与大肠埃希氏菌(No．ECU00734)比较，碱基同源性为84．4l％(70／449)，提示其碱基序列同源性均极高。分别选取对两类抗菌药物耐药及敏感甲型副伤寒沙门氏菌各一株进行RT-PCR检测，结果 两株细菌均检测到多重耐药外排基因acrB的表达。结论甲型副伤寒沙门氏菌对喹诺酮类、第三代头孢菌素类等药物耐药率低，但有多重耐药。甲型副伤寒沙门氏菌中均存在acrAB主动外排系统。与大肠埃希氏菌同源性高，可检测到其表达。主动外排机制可能是甲型副伤寒沙门氏菌形成多重耐药的主要原因之一。

Multiple- antibiotic- resistant active efflux gene acrB and its expression in Salmonella paratyphi A

Objective To investigate the antibiotic resistance of Salmonella p aratyphi A clinical isolates and to identify the active efflux gene acrB and its expression.Methods The antimicrobial susceptibilities of 12 strains of S.paraty phi A were tested with NCCLS recommend agar dilution method and the acrB gene were identified and sequenced by PCR DNA sequence technique,the express ion of acrB gene was measured by RT-PCR.Results The antibiotic resistance rate s of S.paratyphi A to ofloxacin,ciprofloxacin,cefotaxime,piperacillin,tetracycli ne and gentamicin were 8.33% .No chloramphenicol resistant strain was found.All the 12 clinical strains of S.paratyphi A had acrB gene. Sequence results of acr B of S.paratyphi A 329 showed that there were one nucleotide base difference com pared with the Genebank sequence(No.AL627267) of S.typhi,and 84.41% identity w ith that of Escherichia coli(Genebank No.ECU00734).The nucleic acid base mutatio n did not lead to the alteration in the amino acid of AcrB.The expression of acr B gene were identified by RT-PCR. Conclusions The resistance rates of S.parat yphi A to fluoroquinolones and the third generation of cephalosporins were low i n Chongqing.The acrB gene of S.paratyphi A exists in all of the clinical strains and has high homology with that of Escherichia coli,the expression of the gene could be responsible for the multiple-antibiotic-resistance.