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目的建立可以快速检测不同致病性弧菌的CODEHOP PCR方法。方法以致病性弧菌共有的DNA旋转酶B亚单位氨基酸为目标,采用CODEHOP程序,设计一对引物,建立弧菌CODEHOP PCR检测方法,以8种不同致病性弧菌菌株及其他菌株为模板,评价方法的灵敏度、特异性,并对64份入境船舶压舱水进行致病弧菌检测。结果该方法特异性好,8种致病性弧菌标准菌株均获得特异性扩增条带,检测灵敏度可以达到3×103CFU/ml,从64份入境压舱水中检测到14份阳性样品,PCR产物经TA克隆后,测序分析确定为4种不同致病性弧菌。结论本研究所建立的方法可用于致病性弧菌的快速检测与种类鉴定。
Objective To establish a CODEHOP PCR method that can rapidly detect different pathogenic Vibrio species. Methods Amino acid sequence of DNA gyrase B subunit shared by pathogenic Vibrio was designed. A CODEHOP program was designed to design a pair of primers to detect Vibrio cholerae CODEHOP PCR. Eight different pathogenic Vibrio strains and other strains were identified as The sensitivity and specificity of the method were evaluated. Sixty-four inbound marine ballast water was tested for pathogenic vibrio. Results The specificity of this method was good. The standard strains of 8 strains of pathogenic Vibrio were obtained with specific amplified bands, the detection sensitivity could reach 3 × 103CFU / ml, 14 positive samples were detected from 64 inbound ballast water, PCR The product was cloned by TA, sequencing analysis identified as four different pathogenic Vibrio. Conclusion The method established in this study can be used for the rapid detection and identification of pathogenic Vibrio.