猪多杀性巴氏杆菌的分离鉴定及oppA基因遗传进化分析  

Isolation and Identification of Pasteurella multocida from Swine and Genetic Evolution Analysis of oppA Gene

作　　者：李红婕 董紫凡 车勇良[2] 周伦江[2] 王隆柏[2] 白丁平[1] LI Hongjie;DONG Zifan;CHE Yongliang;ZHOU Lunjiang;WANG Longbai;BAI Dingping(College of Animal Sciences(College of Bee Science),Fujian Agriculture and Forestry University,Fuzhou 350002,China;Fujian Animal Diseases Control Technology Development Center,Institute of Animal Husbandry and Veterinary Medicine,Fujian Academy of Agricultural Sciences,Fuzhou 350013,China)

机构地区：[1]福建农林大学动物科学学院(蜂学学院),福州350002 [2]福建省农业科学院畜牧兽医研究所,福建省畜禽疫病防治工程技术研究中心,福州350013

出　　处：《中国畜牧兽医》2022年第12期1686-1694,共9页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(31872502);福建省科技重大专项专题项目(2021NZ029023);福建省农业科学院科研项目(CXLM2021004)。

摘　　要：【目的】了解福建省猪场猪多杀性巴氏杆菌(Pasteurella multocida,Pm)的流行及oppA基因遗传进化情况。【方法】本研究采用细菌分离培养、生化试验、16S rRNA PCR扩增测序、PCR荚膜分型、oppA基因克隆及相似性分析、动物回归试验等方法对分离菌株进行鉴定和分析。【结果】本研究共分离到10株菌,分离菌在血平板上形成淡灰白色、湿润光滑、奶油露珠状菌落;分离菌株能酵解蔗糖、果糖、麦芽糖和甘露醇,不能分解葡萄糖、枸橼酸盐、乳糖、硫化氢等,与多杀性巴氏杆菌生化特性基本一致;分离菌株16S rRNA序列与GenBank中登录的多杀性巴氏杆菌相似性达99.9%以上,10株分离菌均为多杀性巴氏杆菌;PCR荚膜分型显示,6株分离菌为荚膜A型,4株为荚膜D型;基于oppA基因的遗传进化树显示,10株分离菌均位于同一分支内;动物回归试验结果显示,在24 h内攻毒小鼠死亡率较高(21/30),分离菌有较强的致病力。【结论】福建省猪场猪多杀性巴氏杆菌流行菌株的荚膜血清型主要是A和D型,且大部分菌株都来源于共同的祖先,本研究结果丰富了猪多杀性巴氏杆菌的流行病学资料,并为该病的防控奠定基础。【Objective】 The purpose of the experiment was to understand and master the epidemic situation of Pasteurella multocida(Pm) and the genetic evolution of oppA gene in pig farms in Fujian province.【Method】 In this study, bacterial isolation and culture, biochemical test, 16 S rRNA PCR amplification and sequencing, PCR capsule typing, oppA gene cloning and similarity analysis, animal regression test and other methods were used to identify and analyze the isolated strains.【Result】 A total of 10 strains were isolated in this study.The isolated bacteria formed pale grayish white, moist and smooth, butter dew beads on the blood plate.The results of biochemical tests showed that the isolated strains could ferment sucrose, fructose, maltose and mannitol, and could not decompose glucose, citrate, lactose and hydrogen sulfide, which were basically consistent with the biochemical characteristics of Pasteurella multocida.The 16 S rRNA sequence of the isolated strains was more than 99.9% similarity to that of Pasteurella multocida in GenBank, and 10 strains were Pasteurella multocida.PCR capsular typing showed that 6 isolates were capsular serotype A and 4 isolates were capsular serotype D.The genetic evolution tree based on oppA gene showed that all 10 isolates were located in the same branch.Animal regression test demonstrated that the mortality of infected mice was higher within 24 h(21/30),and the isolated bacteria strains had strong pathogenicity.【Conclusion】 The results indicated that the capsular serotypes of Pasteurella multocida strains in Fujian pig farms were mainly types A and D,and most of the strains were from common ancestors, which enriched the epidemiology of Pasteurella multocida and laid the foundation for the prevention and control of Pasteurella multocida infection.

关 键 词：猪 多杀性巴氏杆菌 荚膜分型 分离鉴定 oppA基因 遗传进化 

分 类 号：S852.612[农业科学—基础兽医学]