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用16S-23S基因间隔区聚合酶链反应和限制性片段长度多态性分析对猪链球菌菌株进行分子特征鉴定

Molecular characterization of Streptococcus suis strains by 16S-23S intergenic spacer polymerase chain reaction and restriction fragment length polymorphism analysis.

作者信息

Marois Corinne, Le Devendec Laëtitia, Gottschalk Marcelo, Kobisch Marylène

机构信息

Agence Française de Sécurité Sanitaire des Aliments, Unité de Mycoplasmologie-Bactériologie, BP53, 22440 Ploufragan, France.

出版信息

Can J Vet Res. 2006 Apr;70(2):94-104.

Abstract

We developed a new molecular method of typing Streptococcus suis based on polymerase chain reaction (PCR) amplification of a large fragment of rRNA genes, including a part of the 16S and 23S genes and the 16S-23S intergenic spacer region (ISR), followed by restriction fragment length polymorphism (RFLP) analysis with RsaI or MboII endonuclease. The 16S-23S ISRs of 5 S. suis isolates were sequenced and compared. Size and sequence polymorphisms were observed between the S735 reference strain and the 4 wild-type strains. The genetic relationships between 138 independent S. suis strains belonging to various serotypes, isolated from swine or human cases, were determined. The discriminatory power of the method was > 0.95, the threshold value for interpreting typing results with confidence (0.954 with RsaI and 0.984 with RsaI plus MboII). The in vitro reproducibility was 100%. The strains isolated from humans were less genetically diverse than the strains isolated from pigs. For the first time, 2 molecular patterns (R6, M9) were significantly associated with S. suis serotype 2 strains. This genetic tool could be valuable in distinguishing individual isolates of S. suis during epidemiologic investigations.

摘要

我们开发了一种基于聚合酶链反应(PCR)扩增rRNA基因大片段的猪链球菌新型分子分型方法,该大片段包括部分16S和23S基因以及16S - 23S基因间隔区(ISR),随后用RsaI或MboII内切酶进行限制性片段长度多态性(RFLP)分析。对5株猪链球菌分离株的16S - 23S ISR进行了测序和比较。在S735参考菌株和4株野生型菌株之间观察到大小和序列多态性。确定了从猪或人类病例中分离出的138株属于不同血清型的独立猪链球菌菌株之间的遗传关系。该方法的鉴别力>0.95,这是自信解释分型结果的阈值(用RsaI时为0.954,用RsaI加MboII时为0.984)。体外重现性为100%。从人类分离出的菌株在遗传上比从猪分离出的菌株多样性更低。首次发现2种分子模式(R6、M9)与猪链球菌2型菌株显著相关。这种遗传工具在流行病学调查中区分猪链球菌的各个分离株时可能很有价值。

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