鉴定7种新型猪链球菌荚膜多糖基因型多重PCR方法的建立(英文)

Development of a multiplex PCR assay to identify seven new capsular gene loci of Streptococcus suis

目的建立针对发现的7种新型别荚膜多糖基因簇(cps)的多重PCR(multiplex PCR)鉴定方法。方法选择wzy作为靶基因,先以单重PCR筛选每对引物的扩增效率与特异性,再将筛选出的各对引物组建多重PCR,验证其特异性及敏感性后,用其检测91株血清学不可分型的猪链球菌分离株。结果所建立的多重PCR体系的敏感性为每个反应最低可检测到100pg DNA。在检测的91株菌中,71株可用本研究建立的多重PCR法鉴定,属于新的cps型别。结论成功建立了特异的可鉴定7种新发现cps型别的多重PCR方法,并且成功应用此法鉴定了大部分血清学不可分型的猪链球菌分离菌株。

Streptococcus suis is an important zoonotic agent causing severe diseases in both pigs and humans. To date, 33 serotypes of S. suis have been identified based on antigenic differences in the capsular polysaccharide. The capsular polysac- charide synthesis （cps） locus encodes proteins/enzymes that are responsible for capsular production and structure variation, which is the basis of S. suis serotyping. The cps clusters of 33 reference serotypes were published. Based on serotype-specific gene wzy in the cps gene cluster, we developed multiplex PCR （mPCR） assays for the known 33 serotypes. Seven new cps loci types were discovered, an mPCR assay for identifying them was developed in this study. The detection limit of the mPCR assay for the genomic DNA of representative strains of each new cps loci was 100 pg. Moreover, this mPCR strategy was applied in 91 untypable S. suis strains, and it turned out that 71 of them belong to these new cps types. The multiplex PCR assays deve- loped in this study provide a rapid and specific method for identification of 7 new cps types of S. suis.