Rapid detection of viable Salmonella based on specific phage infection combined with fluorescent RPA
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.
Publication Date
7-25-2026
First Page
42
Last Page
51
DOI
10.13652/j.spjx.1003.5788.2025.80609
Recommended Citation
Daofeng, Liu; Yujie, Zhai; Yucheng, Tan; Shan, Shan; Jun, Lu; and Ziqing, Zhong
(2026)
"Rapid detection of viable Salmonella based on specific phage infection combined with fluorescent RPA,"
Food and Machinery: Vol. 42:
Iss.
7, Article 5.
DOI: 10.13652/j.spjx.1003.5788.2025.80609
Available at:
https://www.ifoodmm.cn/journal/vol42/iss7/5
References
[1] 范鹏辉,李红秋,褚遵华,等.2023年中国大陆食源性疾病暴发监测结果分析 [J].中国食品卫生杂志,2024,36(10):1 199-1 208.Fan P H,Li H Q,Chu X H,et al.Analysis of foodborne diseases outbreak surveillance in China's Mainland,2023 [J].Chinese Journal of Food Hygiene,2024,36(10):1 199-1 208.
[2] 段星星,曾晓琮,周露.沙门氏菌检验及鉴定方法研究进展[J].食品安全质量检测学报,2020,11(16):5 364-5 370.Duan X X,Zeng X C,Zhou L.Research progress on the detection and identification methods of Salmonella [J].Journal of Food Safety & Quality,2020,11(16):5 364-5 370.
[3] 刘理慧,储锦华,隋雨欣,等.沙门氏菌中主要毒力因子的研究进展 [J].生物技术通报,2022,38(9):72-83.Liu L H,Chu J H,Sui Y X,et al.Research progress of main virulence factors in Salmonella [J].Biotechnology Bulletin,2022,38(9):72-83.
[4] 王紫璇,孙洁芳,邵兵.食物中典型致病菌的快速检测研究进展[J].中国食品卫生杂志,2022,34(2):382-389.Wang Z X,Sun J F,Shao B.Research progress in rapid detection of typical pathogenic bacteria in food stuff [J].Chinese Journal of Food Hygiene,2022,34(2):382-389.
[5] 王芳妹,钟文涛,李政,等.基于量子点纳米粒子对乳制品中致病菌进行定性分析 [J].食品与机械,2025,41(3):39-44.Wang F M,Zhong W T,Li Z,et al.Qualitative analysis of pathogenic bacteria in dairy products based on quantum dot nanoparticles [J].Food & Machinery,2025,41(3):39-44.|[6] Yang Q,Zu J Q,Zhang S H,et al.An overview of rapid detection methods for Salmonella [J].Food Control,2025,167:110771.
[7] 刘紫洋,胡高爽,郝建雄,等.纳米酶标记免疫分析技术在食品安全检测应用中的研究进展 [J].食品与机械,2025,41(1):234-242.Liu Z Y,Hu G S,Hao J X,et al.Research progress in immunoassay technology based on nanoenzyme labeling in detection of food safety[J].Food & Machinery,2025,41(1):234-242.
[8] 林哲.基于噬菌体生物传感器的沙门氏菌检测方法研究:以生鲜肉类中沙门氏菌活菌检测为例 [J].食品安全导刊,2025(14):144-147.Lin Z.Research on Salmonella detection method based on phage biosensors:a case study of live Salmonella detection in fresh meat [J].China Food Safety Magazine,2025 (14):144-147.
[9] 刘婷宇,董庆利,李嘉铭,等.食品中食源性致病菌活菌检测方法研究进展 [J].食品工业科技,2025,46(13):395-403.Liu T Y,Dong Q L,Li J M,et al.Research progress of detection technology of viable foodborne pathogens [J].Science and Technology of Food Industry,2025,46(13):395-403.
[10] 王纯,张若鸿,王晓芳,等.食源性致病菌活菌检测方法的应用现状及展望 [J].食品研究与开发,2021,42(23):183-189.Wang C,Zhang R H,Wang X F,et al.Application status and prospects of detection methods for viable foodborne pathogens[J].Food Research and Development,2021,42(23):183-189.
[11] 周冬根,牟丹,许恒毅.重组酶介导的扩增技术及活菌检测方法的应用研究进展 [J].中国国境卫生检疫杂志,2023,46(3):295-301.Zhou D G,Mu D,Xu H Y.Application and research progress of recombinase-aid amplification technology and live bacteria detection methods [J].Chinese Journal of Frontier Health and Quarantine,2023,46(3):295-301.
[12] 卫博,王杰,陈亦新,等.基于 PMA-CELL-qPCR 的葡萄酒发酵中酿酒酵母活菌计数方法的开发与应用 [J].食品与发酵工业,2022,48(17):79-86.Wei B,Wang J,Chen Y X,et al.Development and application of Saccharomyces cerevisiae viable cell quantitative method in wine fermentation based on PMA-CELL-qPCR [J].Food and Fermentation Industries,2022,48(17):79-86.
[13] 沈静怡,徐佳瑶,陈赵圆,等.基于 RNA检测畜禽病原菌活菌方法研究进展 [J].现代畜牧兽医,2024 (6):81-85.Shen J Y,Xu J Y,Chen Z Y,et al.Research progress of RNA-based detection of viable pathogens in livestock and poultry [J].Modern Journal of Animal Husbandry and Veterinary Medicine,2024 (6):81-85.
[14] 黎财慧,游淑珠,王小玉,等.食品中沙门氏菌活菌的 Taq Man实时荧光 PCR检测方法研究 [J].中国口岸科学技术,2023,5(4):76-83.Li C H,You S Z,Wang X Y,et al.Study on detection of live Salmonella in food by TaqMan Real-time PCR [J].China Port Science and Technology,2023,5(4):76-83.
[15] 王梓晨,牛洪梅,刘阳泰,等.产志贺毒素大肠埃希氏菌活菌检测方法的应用现状及展望 [J].工业微生物,2024,54(2):92-100.Wang Z C,Niu H M,Liu Y T,et al.Application status and prospects of detection methods for living shiga toxin-producing Escherichia coli [J].Industrial Microbiology,2024,54(2):92-100.
[16] Dion M B,Oechslin F,Moineau S.Phage diversity,genomics and phylogeny [J].Nature Reviews Microbiology,2020,18(3):125-138.
[17] Monika B,Iva S.Phage amplification assay for detection of mycobacterial infection:a review [J].Microorganisms,2021,9(2):237.
[18] 郭歌.基于纳米酶的噬菌体生物传感器特异检测金黄色葡萄球菌 [D].扬州:扬州大学,2024:63-96.Guo G.Specific detection of Staphylococcus aureus by bacteriophage biosensor based on nanozymes [D].Yangzhou:Yangzhou University,2024:63-96.
[19] Šuster K,Podgornik A,Cör A.Quick bacteriophage-mediated bioluminescence assay for detecting Staphylococcus spp.in sonicate fluid of orthopaedic artificial joints [J].The New Microbiologica,2017,40(3):190-196.
[20] Ding Y F,Huang C X,Chen M X,et al.Rapid and simultaneous detection of viable S.aureus and its penicillin susceptibility by phage amplification techniques in different food matrices [J].LWT-Food Science and Technology,2023,176:114526.
[21] 张雪花.基于重组酶聚合酶扩增 （RPA）技术快速检测假结核耶尔森氏菌 [D].武汉:华中科技大学,2024:7-11.Zhang X H.Rapid detection of Yersinia pseudotuberculosis based on recombinase polymerase amplification [D].Wuhan:Huazhong University of Science and Technology,2024:7-11.
[22] 万佳欣,朱寅初,徐杏,等.基于实时荧光 RPA 的鸭瘟强毒株感染快速检测方法的建立 [J/OL ].生物工程学报.(2025 -09-05)
[2025 -10-03].https://doi.org/10.13345 /j.cjb.250357.Wan J X,Zhu Y C,Xu X,et al.Development of a rapid detection method for a virulent strain of duck enteritis virus based on real-time fluorescence recombinase polymerase amplification [J/OL ].Chinese Journal of Biotechnology.(2025 -09-05) [2025 -10-03].https://doi.org/10.13345 /j.cjb.250357.
[23] Whichard J M,Weigt L A,Borris D J,et al.Complete genomic sequence of bacteriophage felix O 1[J].Viruses,2010,2(3):710-730.
[24] Welkos S,Schreiber M,Baer H.Identification of Salmonella with the O-l bacteriophage [J].Applied Microbiology,1974,28(4):618-622.
[25] Bárdy P,Pantůček R,Benešík M,et al.Genetically modified bacteriophages in applied microbiology [J].Journal of Applied Microbiology,2016,121(3):618-633.
[26] 周洁,黄夏宁,卢玲敏,等.基于恒温扩增技术的病原体快速检测研究进展 [J].中国口岸科学技术,2024,6(S2):19-27.Zhou J,Huang X N,Lu L M,et al.Advances in isothermal amplification technologies for rapid detection of pathogens [J].China Port Science and Technology,2024,6(S2):19-27.
