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目的建立一种快速有效地检测出口冷冻淡水水产品中单核细胞增生性李斯特菌的方法。方法以李斯特菌属特异性iap基因和种特异性hly基因为靶序列,建立双重PCR方法;比较和优化3种提取模板的方法;通过干扰实验、模拟污染实验等验证了所建方法的稳定性、特异性、敏感性;并与SN0184-93方法和mini-VIDAS方法进行比较。结果建立的方法稳定性强、特异性和敏感性好,检测限度为3cfu/25g样品,检测结果与SN0184-93方法和mini-VIDAS方法的一致性为100%;该方法可直接使用单核细胞增生性李斯特菌菌液作为模板进行扩增。结论建立的双重PCR方法可快速、敏感,特异地检测出口冷冻淡水水产品中单核细胞增生性李斯特菌。
Objective To establish a rapid and effective method for the detection of Listeria monocytogenes in frozen freshwater aquatic products for export. Methods The double-PCR method was established by using the Listeria-specific iap gene and the species-specific hly gene as target sequences. Three methods of extracting the template were compared and optimized. The stability of the constructed method was verified by interference experiments and simulated contamination experiments Sex, specificity, sensitivity; and compared with SN0184-93 method and mini-VIDAS method. Results The established method was stable, specific and sensitive. The limit of detection was 3 cfu / 25 g. The concordance was 100% with the SN0184-93 method and the mini-VIDAS method. The method could be used directly by monocytes Listeria monocytogenes bacteria as a template for amplification. Conclusion The established dual PCR method can rapidly, sensitively and specifically detect Listeria monocytogenes in frozen freshwater aquatic products.