一株分离自流浪猫的猪链球菌9型的分子生物学鉴定

Isolation and characterization of a Streptococcus suis serotype 9 from a wild cat

摘要:

[目的] 猪链球菌是一种能感染人和猪的人畜共患病病原,并且还可零星感染多种哺乳动物。本试验旨在调查流浪猫携带猪链球菌的情况。[方法] 在流浪猫身上分离猪链球菌,经血清凝集实验和PCR检测,鉴定其血清型;经多序列位点分型分析,鉴定其ST型;将所分离的细菌与GenBank上已公布的猪链球菌构建16S rRNA的系统发育树,分析该菌株与其他猪链球菌的亲缘关系;药敏纸片法分析其耐药性;小鼠攻毒试验分析其毒力。[结果] 本试验在流浪猫身上分离到一株猪链球菌,命名为m70,其血清型为9型。多序列位点分型显示,m70株属于一个新的ST型。与GenBank上已公布的猪链球菌16S rRNA进行系统发育树分析,结果显示m70属于一个单独的分支。m70与临床菌株的耐药情况相似,对四环素耐药,对红霉素中介耐药,对氨苄西林敏感。小鼠攻毒试验显示,感染10⁸ CFU剂量m70的小鼠,死亡率达到60%-80% (3/5-4/5),3次攻毒试验的平均LD₅₀为5.1×10⁷ CFU;而本实验室保存的猪链球菌强毒株HA9801感染小鼠的平均LD₅₀为3.9×10⁷ CFU,两者之间没有显著差异(P < 0.05)。[结论] 从流浪猫身上分离得到的猪链球菌m70属于优势血清型,且毒力较强,提示一些流行血清型的猪链球菌强毒株具有从流浪猫传染人的潜在风险。

Abstract:

[Objective] Streptococcus suis (S. suis) is an emerging zoonotic pathogenic bacterium capable of infecting piglets and human and with sporadic infections in a variety of mammalian species. The aim of this study is to investigate the prevalence of S. suis in wild cats. [Methods] We isolated an S. suis strain from a wild cat. We tested the serotype of the isolated strain by anti-serum agglutination and PCR. We determined the sequence type (ST) of the isolated strain by multilocus sequence typing tests (MLST). We constructed the 16S rRNA phylogenetic tree of the isolation and S. suis strains in NCBI database to demonstrated genetic relationship of different strains. We measured the antibiotic resistance of the isolated strain by triple disk diffusion method. We detected the virulence of the isolated strain by mice infection experiments. [Results] We isolated an S. suis strain m70 from a wild cat, which belongs to serotype 9. MLST showed that m70 fell into a new ST. The 16S rRNA phylogenetic tree of m70 and S. suis strains in NCBI database demonstrated that m70 was in a separate cluster. m70 was resistant to tetracycline, intermediate to erythromycin, and sensitive to ampicillin, corresponding to clinical S. suis isolates in China. The mortality of mice infected with 108 CFU of m70 was achieved 60%-80% (3/5-4/5). The mean LD₅₀ of mice infected with m70 was 5.1×10⁷ CFU, while the mean LD₅₀ of virulent S. suis strain HA9801 was 3.9×10⁷ CFU. There is no significant difference between the LD₅₀ of the two strains(P<0.05). [Conclusion] We isolated an S. suis strain from a wild cat, which belongs to the prevalent serotype and was a virulent strain, indicating the potential of transmission of S. suis from wild cats to humans, especially some prevalent serotype strains.