摘要
为分析牛呼吸道多杀性巴氏杆菌的生物学特性及其在规模化牛场的流行情况,对内蒙古呼和浩特规模化牛场中患呼吸道疾病的病死牛的肺脏、肝脏样品进行细菌分离培养、形态学观察,通过生化试验、16S rRNA基因测序、特异性引物PCR鉴定、荚膜血清型分型、致病性试验、毒力基因检测、药敏试验和耐药基因检测方法对分离株进行研究.结果显示,从病死牛的肺脏中分离鉴定出6株荚膜血清A型多杀性巴氏杆菌,细菌16S rRNA序列测序后比对发现6株多杀性巴氏杆菌与重庆分离株CQ2(CP033599.1)的亲缘关系最为接近.小鼠致病性试验及毒力基因检测结果显示,分离株均有致病性,并携带有exbB、nanB、sodC、oma87等至少16个及以上相关毒力基因,但均未检出hsf1和toxA.药敏试验及耐药基因检测结果显示,分离株对环丙沙星、氧氟沙星、头孢噻肟等抗菌药物有不同程度敏感,对链霉素、克林霉素和林可霉素耐药,检出strA、strB和tet(H)耐药基因.综上,结果表明牛A型多杀性巴氏杆菌的致病性与毒力基因,耐药表型与耐药基因具有一定相关性,临床上推荐使用喹诺酮类(环丙沙星等)和头孢类(头孢噻肟等)抗菌药物,可为牛场防治由多杀性巴氏杆菌引发的呼吸道疾病提供科学依据和防控方案,为牛呼吸道多杀性巴氏杆菌病的流行病学监测奠定基础.
Abstract
To analyze the biological characteristics of Pasteurella multocida in bovine respiratory tract and its prevalence in large-scale cattle farms,bacterial isolation,culture,and morphological observation were conducted on the lungs and liver samples of dead cows suffering from respiratory diseases in Hohhot,Inner Mongolia.The isolated strains were studied through biochemical testing,16S rRNA gene sequencing,specific primer PCR identification,capsule serotyping,pathogenicity testing,virulence gene testing,drug sensitivity testing,and drug resistance gene detection methods.The results showed that six strains of Pasteurella multocida serotype A were isolated and identi-fied from the lungs of diseased and dead cows.After sequencing the 16S rRNA sequence of the bac-teria,it was found that the six strains of Pasteurella multocida had the closest genetic relationship with the Chongqing isolate CQ2(CP033599.1).The results of mouse pathogenicity test and viru-lence gene detection showed that all isolates were pathogenic and carried at least 16 or more related virulence genes such as exbB,nanB,sodC,oma 87,etc.,but no hsf1 and toxA were detected.The results of drug sensitivity tests and resistance gene detection showed that the isolated strains were sensitive to different degrees of antibiotics such as ciprofloxacin,ofloxacin,and cefotaxime.They were resistant to streptomycin,clindamycin,and lincomycin,and resistance genes of str A,strB,and tet(H)were detected.The results indicate that there is a certain correlation between the pathoge-nicity and virulence genes,drug resistance phenotype,and drug resistance genes of Pasteurellamultocida type A in cattle.It is recommended to use quinolones(such as ciprofloxacin)and cepha-losporins(such as cefotaxime)antibacterial drugs in clinical practice,which can provide scientific basis and prevention and control plans for the prevention and treatment of respiratory diseases caused by Pasteurella multocida in cattle farms,and lay a foundation for the epidemiological mo-nitoring of bovine respiratory multocida pasteurellosis.
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