猪链球菌亚种及1、2、9型四重PCR检测方法的建立

Establishment of quadruple PCR for detecting subspecies and type 1,2 and 9 of swine streptococcus suis

根据序列分析设计了4对引物,分别扩增猪链球菌gdh、cps1、cps2J和cps9G基因的725、212、387bp和556bp大小的片段,利用合成的4对引物并通过对反应条件与反应体系的优化建立了四重PCR,结果显示,猪链球菌阳性出现1条725bp目的带,猪链球菌1型阳性出现725bp和212bp两条目的带,猪链球菌2型阳性出现725bp和387bp两条目的带,猪链球菌9型阳性出现725bp和556bp两条目的带。应用该多重PCR检测了分离到的链球菌1095株,检出猪链球菌阳性667株、猪链球菌1型8株、2型33株、9型16株。研究结果表明,该方法特异性高、敏感性强,可广泛应用于猪链球菌病的快速诊断及流行病学研究。

According to the analysis of sequences, four pair primers were designed to amplified 725bp, 212bp, 387bp and 556bp fragments of four swine streptococcus suis genes including gdh, cps1I, cps2J and cps9G, respectively. At the same time, a quadruple PCR was established by adjusting reaction condition and system of four pairs of primers. The results showed that the aiming band of positive swine streptococcus suis was 725bp, and that of positive streptococcus suis type 1, 2 and 9 were 725bp and 212bp, 725bp and 387bp, 725bp and 556bp, respectively. 1059 isolated strains of streptococcus suis was detected by quadruple PCR, and it included 667 strains of positive streptococcus suis of swine, 8 strains of streptococcus suis type 1, 33 strains of type 2 and 16 strains of type 9, respectively. These results indicated that quadruple PCR detection was highly specific and sensitive, and could be applied on rapid diagnosis of disease of swine streptococcus suis and epidemiology research of streptococcus suis.