山东省某规模化羊场肺炎克雷伯菌和多杀性巴氏杆菌的分离鉴定及耐药性检测

Isolation and Identification of Klebsiella pneumoniae and Pasteurella multocida and Detection of Drug Resistance in an Intensive Sheep Farm in Shandong Province

为查明引起山东省阳谷县某规模化羊场羊群咳嗽、呼吸急促、腹泻的病原体,采集病死羊的鼻腔咽拭子以及肺脏、脾脏、肝脏和气管组织样品,利用形态学观察、生化鉴定、16Sr RNAPCR检测和遗传进化分析等方法进行细菌分离鉴定,并对分离菌株进行耐药性检测和致病性试验。结果显示:分离得到2种致病菌,分别命名为LCh/2023-1和XT/2023-2,其培养特性和生化特性分别符合多杀性巴氏杆菌和肺炎克雷伯菌;系统进化树发现,LCh/2023-1与多杀性巴氏杆菌J-4103株同源性最高,XT/2023-2与肺炎克雷伯菌HE1株同源性最高;耐药性检测发现,LCh/2023-1对复方新诺明、氨苄西林、氟苯尼考、卡那霉素、新霉素、四环素、恩诺沙星、红霉素、阿奇霉素耐药,XT/2023-2对复方新诺明、羧苄西林、氟苯尼考、卡那霉素、新霉素、四环素、多西环素、恩诺沙星、阿奇霉素耐药;小鼠致病性试验可见,攻毒后小鼠全部死亡。结果说明:该羊场发病由肺炎克雷伯菌和多杀性巴氏杆菌混合感染所致,分离菌毒力较强,且耐药性较为严重。

In order to identify the pathogens causing coughpolypnea and diarrhea in sheep in an intensive farm in Yanggu County,Shandong Province,nasal pharyngeal swabs and tissue samples of lungs,spleens,livers and tracheas collected from dead sheep to isolate and identify bacteria using morphology observation,biochemical characterization,16S r RNA PCR and genetic evolutionary analysis,followed by test of drug resistance and pathogenicity.The results showed that two kinds of bacteria were isolated and named as LCh/2023-1 and XT/2023-2,respectively,with bacterial culture and biochemical characteristics in compliance with Pasteurella multocida and Klebsiella pneumoniae,respectively;phylogenetic tree revealed that LCh/2023-1 was the highest identical with Pasteurella multocida strain J-4103,and XT/2023-2 was the highest identical with Klebsiella pneumoniae strain HE1;drug resistance analysis results showed that LCh/2023-1 was resistant to cotrimoxazole,ampicillin,florfenicol,kanamycin,neomycin,tetracycline,enrofloxacin,erythromycin and azithromycin,and XT/2023-2 was resistant to cotrimoxazole,carbenicillin,florfenicol,kanamycin,neomycin,tetracycline,doxycycline,enrofloxacin and azithromycin;mouse pathogenicity test results showed that all mice died after the challenge.In conclusion,the disease was caused by mixed infection of Klebsiella pneumoniae and Pasteurella multocida,and the isolates were more virulent with strong drug resistance.