基于nuc基因的环介导等温核酸扩增可视化技术检测食源性金黄色葡萄球菌  

Development of Loop Mediated Isothermal Amplifi cation(Lamp)Visualization Technique Based on Nuc Gene for Detection of Foodborne Staphylococcus aureus

作　　者：邹作成 王西 刘雪兰[3] 胡仲皓 田祥强 何晓花 杜晓莉 王佳玲 李柯 刘浩洋 嵇辛勤[1] 胡青海[2] ZOU Zuocheng;WANG Xi;LIU Xuelan;HU Zhonghao;TIAN Xiangqiang;HE Xiaohua;DU Xiaoli;WANG Jialing;LI Ke;LIU Haoyang;JI Xinqin;HU Qinghai(College of Animal Science and Technology,Guizhou University,Guiyang 230036,China;Shanghai Veterinary Research Institute,CAAS,Shanghai 200241,China;College of Animal Science and Technology,Anhui Agricultural University,Hefei 230036,China)

机构地区：[1]贵州大学动物科学学院,贵阳550025 [2]中国农业科学院上海兽医研究所,上海200241 [3]安徽农业大学动物科技学院,合肥230036

出　　处：《中国动物传染病学报》2023年第5期108-113,共6页Chinese Journal of Animal Infectious Diseases

基　　金：上海市科技创新项目(农业领域)(21N31900900);国家重点研发项目(2018YFD0500500)。

摘　　要：金黄色葡萄球菌食物中毒主要是由食用被金黄色葡萄球菌及其所产生的肠毒素污染的奶、肉、蛋、糕点、剩饭等蛋白或淀粉含量丰富的食品引起,快速精准的金黄色葡萄球菌检测技术对于保障人们的健康和食品安全具有重要意义。本研究以nuc基因作为检测靶基因,建立了一种检测金黄色葡萄球菌的环介导等温核酸扩增(LAMP)技术。该方法检测金黄色葡萄球菌基因组DNA样品为阳性,而对单细胞增生李斯特菌、鼠伤寒沙门菌、肠出血性大肠杆菌、空肠弯曲菌、小肠结肠炎耶尔森菌和产气荚膜梭菌等食源性致病菌和猪链球菌、胸膜肺炎放线杆菌和多杀性巴氏杆菌等猪源细菌基因组DNA检测结果均为阴性,表明建立的LAMP方15法具有较好的特异性。灵敏度的检测结果表明,LAMP的检测限可达5.6×10^(1) CFU/mL,而常规PCR的检测限为5.6×10^(5) CFU/mL,即LAMP的检测灵敏度比常规PCR高出10000倍。临床样品的检测结果表明,50份猪肉样品中有4份检出金黄色葡萄球菌。本研究为食源性金黄色葡萄球菌提供了一种快速、可视化、特异性好、灵敏度高、成本低的LAMP检测方法。Staphylococcus aureus food poisoning is mainly caused by the polluted foods with rich proteins or starch contents such as milks,meats,eggs,pastries and leftovers.Therefore,a rapid and accurate detection for S.aureus is particularly important for ensuring people's health and food safety.In this study,we developed a loop mediated isothermal nucleic acid amplifi cation(LAMP)technique for the detection of S.aureus based on the nuc gene.The specifi city of the LAMP detection showed that only the genomic DNA of S.aureus was positive,while those of other foodborne pathogens were negative,such as Listeria monocytogenes,Salmonella typhimurium,Enterohemorrhagic O157:H7,Campylobacter jejuni,Yersinia enterocolitica,Clostridium perfringens and swine origin pathogens such as Streptococcus suis,Actinobacillus pleuropneumoniae,Pasteurella multocida,suggesting that the LAMP developed in this study had good specifi city.The detection limit of LMAP was 5.6×10^(1) CFU/ml while that of the conventional PCR was 5.6×10^(6) CFU/mL,indicating that the sensitivity of LAMP detection was 100000 times higher than that of the conventional PCR.In addition,S.aureus was detected from 4 out of 50 clinical pork samples by both LAMP and conventional PCR.In summary,this study provided a rapid,visual,specifi c,sensitive and low-cost LAMP detection method for foodborne S.aureus.

关 键 词：金黄色葡萄球菌 nuc基因 环介导等温核酸扩增技术 可视化 

分 类 号：TS207.4[轻工技术与工程—食品科学]