噬菌体特异性侵染耦合荧光RPA的沙门菌活菌快速检测
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1江西省疾病预防控制中心重大疫情防控江西省重点实验室,江西 南昌 330029;2南昌大学公共卫生学院,江西 南昌 330019;3营养膳食与健康江西省卫生健康重点实验室,江西 南昌 330019;4江西师范大学生命科学学院,江西 南昌 330022;5福建林业职业技术学院,福建 南平 353000

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钟子清(1988—),女,福建林业职业技术学院讲师,博士。E-mail:crystal1439@163.com

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江西省重点研发计划(编号:20243BBH81007);江西省自然科学基金重点项目—南昌(编号:20253BAC280055);江西省自然科学基金青年基金项目(编号:20232BAB21605,20232BAB216106)


Rapid detection of viable Salmonella based on specific phage infection combined with fluorescent RPA
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1Jiangxi Provincial Key Laboratory of Major Epidemics Prevention and Control, Jiangxi Provincial Center for Disease Control and Prevention, Nanchang, Jiangxi 330029, China;2School of Public Health, Nanchang University, Nanchang, Jiangxi 330019, China;3Jiangxi Key Laboratory of Preventive Medicine, Nanchang, Jiangxi 330019, China;4College of Life Sciences, Jiangxi Normal University, Nanchang, Jiangxi 330022, China;5Fujian Forestry Vocational Technical College, Nanping, Fujian 353000, China

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    摘要:

    目的 针对传统分子生物学检测方法难以便捷区分食源性致病菌死活的技术痛点,建立一种基于噬菌体特异侵染与重组酶聚合酶扩增(RPA)的沙门菌活菌快速检测方法。方法 依托O-1噬菌体仅在活菌中增殖的特性,将其接种于增菌后的待检样本中,温育培养后提取总DNA;通过荧光RPA检测噬菌体DNA增殖情况,以判断原样本中是否存在沙门菌活菌。结果 经优化,确定最佳检测条件为:加入100 μL、3×105 PFU/mL的O-1噬菌体,侵染增殖时间3 h。该方法噬菌体核酸检出限为0.07 fg/μL,沙门菌活菌检出限为1.58×102 CFU/mL;对37株不同血清型沙门菌检测的属内包容性为100%,对17株非沙门菌检测的属间特异性为100%。结论 该方法利用噬菌体只在活菌宿主内快速增殖的特性,无需PCR热循环仪,可在4 h内实现沙门菌活菌高灵敏、高特异性检测,适用于食品生产、流通等环节中沙门菌活菌的现场快速筛查与鉴定,为食品中沙门菌活菌即时检测提供了新技术方案。

    Abstract:

    Objective To establish a rapid detection method for viable Salmonella based on specific phage infection and recombinase polymerase amplification (RPA) in response to the key challenge that traditional molecular biological detection methods have difficulty in conveniently distinguishing between viable and non-viable foodborne pathogens.Methods Taking advantage of the characteristic that O-1 phage only proliferates in viable bacteria, the phage is added to the enriched sample. After further incubation, total DNA is extracted. Fluorescent RPA is used to determine whether the phage has significantly proliferated, so as to judge the presence of viable Salmonella in the original sample.Results After optimization, the optimal detection conditions are as follows: 100 μL of O-1 phage with a concentration of 3×105 PFU/mL is added, and the infection and proliferation time is 3 hours. The detection limit of the method for phage DNA is 0.07 fg/μL, and that for viable Salmonella is 1.58×102 CFU/mL. The intra-generic inclusivity of the method is 100% when testing 37 Salmonella strains of different serotypes, and the inter-generic specificity is 100% when testing 17 non-Salmonella strains.Conclusion The method utilizes the characteristic that phages only rapidly proliferate in viable host bacteria, enabling highly sensitive and specific detection of viable Salmonella within 4 hours without a PCR thermal cycler. The method is suitable for on-site rapid screening and identification of viable Salmonella in food production and circulation processes, providing a new approach for the on-site detection of viable Salmonella in food.

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刘道峰,翟玉杰,谭俣宬,等.噬菌体特异性侵染耦合荧光RPA的沙门菌活菌快速检测[J].食品与机械,2026,(7):42-51.
Liu Daofeng, Zhai Yujie, Tan Yucheng, et al. Rapid detection of viable Salmonella based on specific phage infection combined with fluorescent RPA[J]. Food & Machinery,2026,(7):42-51.

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  • 收稿日期:2025-07-03
  • 最后修改日期:2026-05-01
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  • 在线发布日期: 2026-07-25
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