一株猪源致病性蜡样芽孢杆菌的分离与鉴定

Isolation and characterization of a pathogenic Bacillus cereus from a piglet with diarrhea

无菌采集流行性腹泻耐过仔猪的肝脏样本,经无氧及有氧培养获得一株疑似蜡样芽孢杆菌的纯培养物,命名为HN1203。16SrDNA序列(GenBank登录号为JX294967)分析表明,HN1203为蜡样芽孢杆菌群第7基因型菌株。生化鉴定结果表明,该菌的生化特性符合蜡样芽孢杆菌群成员的特征。药敏试验和伴孢晶体蛋白检测结果表明,该菌对链霉素和四环素敏感,对青霉素不敏感,其芽孢不形成伴孢晶体。多位点序列分型(MLST)检测结果表明,该菌带有蜡样芽孢杆菌看家基因pta新的等位基因pta153,序列基因型为ST611。毒素基因的PCR检测结果表明,该菌携带肠毒素基因(hbl、nhe和entFM)和细胞毒素K基因(cytK)。综合以上结果,分离菌为一株新的蜡样芽孢杆菌,且提示分离培养蜡样芽孢杆菌时,不宜采用常用的有氧条件。进一步动物试验结果表明,该分离菌株能够致死实验小白鼠。提示生产实践中应该重视监控猪源致病性蜡样芽孢杆菌的潜在危害。

Through aerobic and anaerobic cultivation, pure culture of a bacterial strain named HN1203 suspected to be Bacillus cereus （B. cereus） was isolated from liver sample of a piglet recovered from severe post-wean diarrhea syndrome. Analyzing of 16S rRNA sequence （GenBank accession number JX294967） of HN1203 showed this strain belongs to type 7 of Bacillus cereus group. Biochemical identification indicated biochemical characteristics of HN 1203 are accord with that of the strains of B. cereus group. Drug susceptibility testing and parasporal crystal protein test showed that this strain was sensitive to tetracycline and streptomycin, resistant to penicillin, and no parasporal crystal was formed on the spore of HN1203. Multilocus sequence typing （MLST） assays revealed a new allele of house-keeping genepta, which was assigned a new sequence type （ST） of 611. PCR detection showed HN1203 harbors virulence-coding genes hblCDA, nheABC, eytK and entFM. These results showed HNI203 is a now B. cereus strain, and isolation of which indicated that normally used aerobic condition is not suitable for isolation orB. cereus. Further animal test showed HN1203 could be fatal to mice. These findings highlight the need of increased awareness of pathogenic B. cereus in pig and its zoonotic potential.