急性死亡奶牛血液中松鼠葡萄球菌的分离与鉴定

Isolation and identification of a strain of Staphylococcus sciuri in blood of acute dead cows

为查找引起河南省某奶牛场产后奶牛急性死亡的病原(前期检测牛流行热病毒、牛病毒性腹泻病毒和牛传染性鼻气管炎病毒核酸均为阴性,肠道内容物中的梭菌、肺脏中的巴氏杆菌核酸为阴性),试验采集猝死奶牛的脾脏、肝脏等组织及血液样本进行细菌的分离培养、革兰氏染色和镜检、生化鉴定、16S rRNA基因的PCR扩增与测序、药物敏感试验和致病性试验。结果表明:从奶牛血液样本中分离到1株细菌,该菌株革兰氏染色镜检呈蓝紫色并排列成葡萄状,且与葡萄球菌的生化特性相符,其16S rRNA基因序列与GenBank数据库中的松鼠葡萄球菌高度同源,相似性在99.7%以上,证实分离菌株为松鼠葡萄球菌,命名为HNSX-B1;HNSX-B1株对头孢类药物(头孢唑林、头孢拉定和头孢曲松)、喹诺酮类药物(诺氟沙星、环丙沙星和氧氟沙星)、四环素类药物(四环素、多西环素和米诺环素)和庆大霉素、红霉素、哌拉西林、苯唑西林敏感,对氨苄西林、头孢他啶和氨基糖苷类药物(卡那霉素、克林霉素)中度敏感,对青霉素和复方新诺明耐药;小鼠在感染HNSX-B1株(1×109cfu/mL,200μL)36 h后全部死亡。说明试验成功分离到1株对小鼠具有明显致病性的松鼠葡萄球菌,该菌可能是引起该奶牛场产后奶牛急性死亡的病原。

In order to find the pathogen of acute death of dairy cows after giving birth in a dairy farm in Henan Province(nucleic acid tests of early bovine epidemic fever virus,bovine viral diarrhea virus and bovine infectious rhinotracheitis virus were negative,and Clostridium in intestinal contents and Pasteurella in lung were negative),in this experiment,the spleen,liver and other tissues and blood samples of sudden death cows were collected for bacterial isolation and culture,Gram staining microscopy,biochemical identification,PCR amplification and sequencing of 16S rRNA gene,and drug susceptibility test and pathogenicity test.The results showed that one strain of bacteria was isolated from cow blood samples,and the Gram staining microscopic examination of this strain was blue-purple and arranged in a grape-like pattern,which was consistent with the biochemical characteristics of Staphylococcus sciuri;its 16S rRNA gene sequence was highly homologous to Staphylococcus sciuri in GenBank database,with a similarity of more than 99.7%,which confirmed that the isolated strain was Staphylococcus sciuri,named HNSX-B1.The HNSX-B1 strain was sensitive to cephalosporins(cefazolin,cefradine and ceftriaxone),quinolones(norfloxacin,ciprofloxacin,and ofloxacin),tetracyclines(tetracycline,doxycycline,and minocycline),and gentamicin,erythromycin,piperacillin,oxacillin,moderately sensitive to ampicillin,ceftazidime,neomycin,and aminoglycosides(kanamycin,clindamycin),and resistant to penicillin and cotrimoxazole.Mice all died after 36 h of infection with HNSX-B1 strain(1x10^(9)cfu/mL,200μL).The results suggested that one strain of Mammalicoccus sciuri with obvious pathogenicity to mice was successfully isolated,which might be the pathogen causing acute death of postpartum cows in the dairy farm.