猪胸膜肺炎放线杆菌血清8型菌株的分离鉴定及耐药性研究

Isolation,Identification and Drug Sensitive Tests of Actinbacillus pleuropneumoniae Serotype 8

为了确定引起2018年5月江苏泰州某规模化养猪场育肥猪发病死亡的病原,本试验无菌采取病死猪肺脏组织,通过病原分离培养、染色观察、生化试验、PCR扩增等方法进行鉴定,随后对分离菌株进行致病性和耐药性试验。结果显示,该分离菌株在病原分离培养过程中,需在含有NAD的培养基中才能生长;该分离菌通过革兰氏染色,显微镜观察细菌呈红色,可判定新分离的菌株为革兰氏阴性菌;根据生化试验和PCR结果可判定该分离菌株为猪传染性胸膜肺炎放线杆菌;PCR血清型分型结果显示,分离菌株为猪胸膜肺炎放线杆菌血清8型菌株;小鼠毒力试验结果显示,当分离株菌液浓度达到3×10^8 CFU/mL时便可引起BALB/c小鼠死亡,浓度为2×10^9 CFU/mL时对BALB/c小鼠的致死率达到100%,表明该分离菌株对BALB/c小鼠有较强的致病力及致死性;药敏试验结果表明,该分离菌株对头孢噻吩、头孢噻肟、氨曲南、头孢吡肟、头孢曲松、氧氟沙星等14种抗菌药物高度敏感,对氨苄西林中度敏感,对链霉素耐药。综合以上试验结果表明该猪场存在猪胸膜肺炎放线杆菌血清8型菌株感染情况。

In order to ascertain the pathogen causing the death of fatten pigs in a large-scale pig farm in Taizhou,Jiangsu in May 2018,the lung tissues of the dead pigs were sterilized by the pathogen isolation,culture,stain observation,biochemical test and PCR amplification,and then the pathogenicity and drug resistance of the isolates were tested.The results showed that the isolated strain could only grow in the medium containing NAD.The bacteria could be found red through the Gram staining and microscope.The isolates were Gram-negative bacteria.According to the results of biochemical tests and PCR,the isolates were identified as Actinobacillus pleuropneumoniae.Serotype PCR showed that the isolates were Actinobacillus pleuropneumoniae serotype 8.The results of virulence test showed that BALB/c mice died when the concentration of the isolated strain reached 3×10^8 CFU/mL,and the mortality rate of BALB/c mice was 100%when the concentration was 2×10^9 CFU/mL.The results of susceptibility test showed that the strain was highly sensitive to 14 antimicrobial agents,such as cefotaxime,cefotaxime,aztreonam,cefepime,ceftriaxone and ofloxacin,and was moderately sensitive to ampicillin and resistant to streptomycin.The above results showed that there was infection of Actinobacillus pleuropneumoniae serotype 8 in the pig farm.