兔源多杀性巴氏杆菌的分离鉴定及耐药性分析

Isolation,Identification and Drug Resistance Analysis of Pasteurella multocida from Rabbits

为了对疑似感染巴氏杆菌的病死兔进行病原的分离、鉴定并分析其耐药情况,本试验对送检的病死兔进行剖检,结合传统分离培养及16S rRNA和荚膜抗原A(capA)基因的PCR扩增等鉴定病原菌,PCR方法扩增其23个毒力基因,最后进行药敏试验。结果显示,分离菌在血琼脂培养基的菌落特征为光滑、湿润、半透明、灰白色、圆形、露珠状,染色镜检可观察到革兰阴性、两端染色较深的红色短杆菌。PCR扩增及测序结果显示,该分离菌株的16S rRNA和capA基因与多杀性巴氏杆菌Q菌株(GenBank登录号:CP033597)同源性分别为99.86%和100%。23个毒力基因中仅pfhA、oma 87、fur、pmHAS基因检测出目的条带,其余19个毒力基因均未检出。该分离菌株对环丙沙星、氧氟沙星、头孢拉定、氨苄西林、卡那霉素、阿米卡星、庆大霉素、红霉素和多黏菌素B共9种药物敏感,对四环素、复方新诺明和克林霉素等耐药。结果表明本试验从病死兔中分离鉴定出1株荚膜血清A型的多杀性巴氏杆菌,可选用环丙沙星和红霉素等进行治疗。该结果可为临床巴氏杆菌病的治疗提供一定的理论依据。

This study aimed to isolate and identify pathogenic Pasteurella multocida from dead rabbits through dissection of dead rabbits,traditional isolation and culture,PCR amplification of 16S rRNA,capA and 23 virulence genes,and drug sensitivity test.The results showed that the colony characteristics of the isolated bacteria in blood agar medium were smooth,moist,translucent,gray-white,round and dew-like.Gram-negative,darkly stained red brevibacterium could be observed by Gram staining and microscope.PCR amplification and sequencing results showed that the 16S rRNA and capA genes of the isolate shared 99.86%and 100%homology with Pasteurella multocida Q strain(GenBank accession number:CP033597),respectively.Of the 23 virulence genes,only pfhA,oma 87,fur and pmHAS genes were detected,while the other 19 virulence genes were absent.The isolate was sensitive to ciprofloxacin,ofloxacin,cefradine,ampicillin,kanamycins,amikacin,gentamicin,erythromycin and polymyxin B,but resistant to tetracycline,trimethoprim/sulfamethoxazole and clindamycin.Thus,a strain of Pasteurella multocida with capsular serotype A was isolated and identified from dead rabbits,and could be treated with ciprofloxacin and erythromycin.The results can provide a theoretical basis for the treatment of clinical pasteurellosis.