Optimization of a loop-mediated isothermal amplification (lamp) assay for the rapid detection of Clostridium perfringens

IF 1.6 3区农林科学 Q2 AGRICULTURE, DAIRY & ANIMAL SCIENCE

Journal of Applied Poultry Research Pub Date : 2024-12-20 DOI:10.1016/j.japr.2024.100513

Deepa Chaudhary , Sabin Poudel , Linan Jia , Anuraj Theradiyil Sukumaran , Xue Zhang , Wen-Hsing Cheng , Aaron S. Kiess , Kenneth S. Macklin , Li Zhang

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Abstract

This study developed and optimized a loop-mediated isothermal amplification (LAMP) assay for the rapid, sensitive, and specific detection of Clostridium perfringens (C. perfringens), a key pathogen in necrotic enteritis in poultry, causing significant global economic losses. We designed two sets of LAMP primers, ID7 and ID22, targeting the cpa gene of C. perfringens. The CP-LAMP assays were optimized for temperature, time, and primer concentration. We conducted inclusivity tests on 55 C. perfringens strains and exclusivity tests on six other Clostridium species strains and 33 non-Clostridium strains (Escherichia coli, Campylobacter, and Salmonella). Sensitivity was evaluated using serially diluted genomic DNA from C. perfringens ATCC 13124 and compared to PCR. The optimal primer concentrations were 8 μM F3/B3 and 48 μM FIP/BIP. Reactions were performed at 63°C for 30 min. Both primer sets achieved 100 % inclusivity and 88.33 % and 100 % exclusivity for other Clostridium and non-Clostridium isolates, respectively. The CP-LAMP assay's sensitivity and specificity were comparable to PCR, enhancing surveillance and management of necrotic enteritis in poultry farms, potentially reducing its economic impact.

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来源期刊

Journal of Applied Poultry Research 农林科学-奶制品与动物科学

CiteScore

4.10

自引率

10.50%

发文量

审稿时长

104 days

期刊介绍： The Journal of Applied Poultry Research (JAPR) publishes original research reports, field reports, and reviews on breeding, hatching, health and disease, layer management, meat bird processing and products, meat bird management, microbiology, food safety, nutrition, environment, sanitation, welfare, and economics. As of January 2020, JAPR will become an Open Access journal with no subscription charges, meaning authors who publish here can make their research immediately, permanently, and freely accessible worldwide while retaining copyright to their work. Papers submitted for publication after October 1, 2019 will be published as Open Access papers. The readers of JAPR are in education, extension, industry, and government, including research, teaching, administration, veterinary medicine, management, production, quality assurance, product development, and technical services. Nutritionists, breeder flock supervisors, production managers, microbiologists, laboratory personnel, food safety and sanitation managers, poultry processing managers, feed manufacturers, and egg producers use JAPR to keep up with current applied poultry research.