食品中志贺氏菌快速检测免疫磁分离样品前处理技术研究

doi:10.3969/j.issn.1000-1158.2023.03.02

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Abstract Anti-Shigella multi-target immunomagnetic beads were prepared by using Protein A magnetic beads and anti-Shigella mixed serum, and a 25mL immunomagnetic separation system for Shigella was established. Combined with determining the growth curve of Shigella bacteria in representative foods (milk, fruit, bread), a feasible strategy for the pretreatment of food samples was designed, which was integrated with the LAMP technology to conduct rapid detection of Shigella in food. The results showed that the multi-target immunomagnetic beads could simultaneously capture four Shigella species, including flexneri, sonnei, Bowman, and dysentery, with a capture efficiency of 35%~94% and a non-specificity of less than 1% (except for Staphylococcus aureus 10%). Above 10²CFU/mL of target Shigella could be obtained when 1~2×10¹CFU Shigella in 25mL milk/in 25g bear/bread +25mL Shigella enrichent broth base by culturing for 6h following the operation of immunomagnetic separation, the time of samples pretreatment for Shigella detection is shortened from 48h to within 8h, which can meet the requirements of the detection limit of technologies subsequently used to realize the rapid detection of Shigella in food.

Key words： metrology food detection Shigella immunomagnetic separation capture efficiency protein A magnetic beads multivalent serum

Received: 01 August 2022 Published: 08 March 2023

PACS:

TB99

Fund:National Natural Science Foundation of China (general project);High-Level Innovation Team Program;BJAST Budding Talent Program

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	Articles by authors
	LI Jing-wen
	CHEN Er-ning
	KANG Fu-ying
	HONG Tian
	FU Ying-hui
	DU Mei-hong

Cite this article:

LI Jing-wen,CHEN Er-ning,KANG Fu-ying, et al. Study on Samples Pretreatment Technology Based on Immunomagnetic Separation for Rapid Detection of Shigella in Food[J]. Acta Metrologica Sinica, 2023, 44(3): 326-333.

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http://jlxb.china-csm.org:81/Jwk_jlxb/EN/10.3969/j.issn.1000-1158.2023.03.02 OR http://jlxb.china-csm.org:81/Jwk_jlxb/EN/Y2023/V44/I3/326

［4］	Meng Q, Bai X N, Zhao A, et al. Characterization of Shiga toxin-producing Escherichia coli isolated from healthy pigs in China［J］. Bmc Microbiology, 2014,14:5-19.
［6］	GB 4789.5—2012 食品微生物检验-志贺氏菌检验［S］.
［1］	Farshad S, Sheikhi R, Japoni A, et al. Characterization of Shigella strains in Iran by plasmid profile analysis and PCR amplification of ipa genes［J］. Journal of Clinical Microbiology, 2006,44(8):2879-2883.
［10］	Du M H, Li J W, Liu Q J, et al. Rapid detection of trace Salmonella in milk using an effective pretreatment combined with droplet digital polymerase chain reaction［J］. Microbiol Res, 2021,251:126838.
［13］	Islam D, Lindberg A A. Detection of Shigella-dysenteriae type-1 and Shigella-flexneri in feces by immunomagnetic isolation and polymerase chain-reaction［J］. Journal of Clinical Microbiology, 1992,30(11):2801-2806.
［15］	Achi-Berglund R, Lindberg A A. Rapid and sensitive detection of Shigella sonnei in feces by the use of an O-antigen-specific monoclonal antibody in a combined immunomagnetic separation-polymerase chain reaction assay［J］. Clin Microbiol Infect, 1996,2(1): 55-58.
	Wu H J, Sun Y P, Fan T L, et al. Detection of Live Listeria monocytogenes by EMA-qPCR［J］. Journal of Food Science and Biotechnology,2020,39(11):65-70.
	Li D H, Lu J F, Zeng J L, et al. Evaluation of Uncertainty in Determination of Total Residues of Ribavirin and its Metabolites in Eggs by LC-MS/MS［J］. Acta Metrologica Sinica, 2021, 42 (9): 1250-1256.
［19］	张明明,肖剑,林秀敏,等. 多重微滴数字PCR同时定量检测三种食源性致病菌DNA拷贝数［J］.农业生物技术学报,2022,30(3):606-618.
［7］	GB 4789.31—2003 食品微生物学检验-沙门氏菌、志贺氏菌和致泻大肠埃希氏菌的肠杆菌科噬菌体诊断检验［S］.
［16］	Achi R, Mata L, Siles X, et al. Immunomagnetic separation and PCR detection show Shigellae to be common faecal agents in children from urban marginal communities of Costa Rica［J］. Journal of Infection, 1996,32(3): 211-218.
［22］	王斌, 隋志伟, 刘思渊,等. 基于流式分析技术的奶粉中金黄色葡萄球菌活菌快速定量检测方法研究［J］. 计量学报, 2021, 42 (2): 250-258.
［2］	Lee J Y, Kang D H. Development of an improved selective medium for the detection of Shigella spp［J］. Lwt-Food Science and Technology, 2016,65:311-317.
［3］	Iseki H, Alhassan A, Ohta N, et al. Development of a multiplex loop-mediated isothermal amplification (mLAMP) method for the simultaneous detection of bovine Babesia parasites［J］. Journal of Microbiological Methods, 2007,71(3):281-287.
［5］	Lauri A, Castiglioni B, Mariani P. Comprehensive analysis of Salmonella sequence polymorphisms and development of a LDR-UA assay for the detection and characterization of selected serotypes［J］. Applied Microbiology and Biotechnology, 2011,91(1):189-210.
［9］	Kaluzewski S, Sienicka J. Quantitative evaluation of growth-promoting properties of selected culture media used for isolation of Salmonella and Shigella strains［J］. Med Dosw Mikrobiol, 1990,42(1-2):35-43.
［11］	Li J W, Liu Q J, Wan Y P, et al. Rapid detection of trace Salmonella in milk and chicken by immunomagnetic separation in combination with a chemiluminescence microparticle immunoassay［J］. Anal Bioanal Chem, 2019,411:6067-6080.
［23］	李德恒,陆晶芳,曾金林,等.液相色谱串联质谱法测定鸡蛋中利巴韦林及其代谢物总残留的不确定度评定［J］. 计量学报, 2021,42 (9): 1250-1256.
［8］	Mokhtari W, Nsaibia S, Majouriet D, et al. Detection and characterization of Shigella species isolated from food and human stool samples in Nabeul, Tunisia, by molecular methods and culture techniques［J］. Journal of Applied Microbiology, 2012,113(1):209-222.
［12］	Du M H, Li J W, Zhao R X, et al. Effective pre-treatment technique based on immune-magnetic separation for rapid detection of trace levels of Salmonella in milk［J］. Food Control, 2018, 91:92-99.
［14］	Islam D, Tzipori S, Islam M, et al. Rapid Detection of Shigella-dysenteriae and Shigella-Flexneri infeces by an immunomagnetic assay with monoclonal-antibodies［J］. European Journal of Clinical Microbiology & Infectious Diseases, 1993,12(1):25-32.
［17］	Ma K, Deng Y, Bai Y, et al. Rapid and simultaneous detection of Salmonella, Shigella, and Staphylococcus aureus in fresh pork using a multiplex real-time PCR assay based on immunomagnetic separation［J］. Food Control,2014,42:87-93.
［18］	Xia S Q, Yu Z B, Liu D F, et al. Developing a novel immunochromatographic test strip with gold magnetic bifunctional nanobeads (GMBN) for effificient detection of Salmonella choleraesuis in milk［J］. Food Control, 2016,59,507-512.
［21］	Dolka B, Cisek A A, Szeleszczuk P. The application of the loop-mediated isothermal amplification (LAMP) method for diagnosing Enterococcus hirae-associated endocarditis outbreaks in chickens［J］.BMC Microbiology,2019,19:48.
	Zhang M M, Xiao J, Lin X M, et al. Simultaneous Quantification Method of DNA Copy Number for Three Food-borne Pathogens by Multiplex Droplet Digital PCR［J］. Journal of Agricultural Biotechnology, 2022,30(3):606-618.
［20］	吴海江,孙玉萍,范田丽,等. EMA结合实时荧光PCR方法检测单核细胞增生李斯特氏菌［J］.食品与生物技术学报,2020,39(11):65-70.
	Wang B, Sui Z W, Liu S Y, et al. Rapid and Quantitative Detection of Viable Staphylococcus aureus in Milk Powder Based on Flow Cytometry Technology［J］. Acta Metrologica Sinica, 2021, 42 (2): 250-258.