Development of a filtration-based LAMP–LFA method as sensitive and rapid detection of E. coli O157:H7

  • Jin-Hee Kim
  • Se-Wook OhEmail author
Original Article


Rapid detection of low number of pathogenic bacteria in food is difficult. This study tested the filter-based loop-mediated isothermal amplification-lateral flow immunoassay (LAMP–LFA) method for rapid detection of pathogens in real food. Escherichia coli O157:H7 was inoculated on 25 g of beef and the homogenized sample was filtered with 0.45 μm cellulose nitrate filter, and concentrated E. coli was recovered and DNA was extracted and analyzed by LAMP. LFA reaction was performed by hybridization of digoxygenin-labeled LAMP amplicon and biotinylated probe. The sensitivity of the filtered sample was 100 times more sensitive than that of the unfiltered sample. The total reaction time used for detection from sample preparation to confirmation of E. coli was within 3 h. These results suggest that the LAMP–LFA method can be used in real food systems as point-of-care testing for E. coli O157:H7 in beef.


Loop-mediated isothermal amplification Lateral flow assay Filtration Escherichia coli O157:H7 



This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2017R1D1A1B03030859).


  1. Avery SM, Hudson JA, Penney N (1994) Inhibition of Listeria monocytogenes on normal ultimate pH beef (pH 5.3–5.5) at abusive storage temperatures by saturated carbon dioxide controlled atmosphere packaging. J Food Prot 57:331–333CrossRefGoogle Scholar
  2. Chapman P, Ellin M, Ashton R (2001) A comparison of immunomagnetic separation and culture, RevealTM and VIPTM for the detection of E. coli O157 in enrichment cultures of naturally-contaminated raw beef, lamb and mixed meat products. Lett Appl Microbiol 32:171–175CrossRefGoogle Scholar
  3. Dong H-J, Cho A-R, Hahn T-W, Cho S (2014) Development of a multiplex loop-mediated isothermal amplification assay to detect shiga toxin-producing Escherichia coli in cattle. J Vet Sci 15:317–325CrossRefGoogle Scholar
  4. D’Urso OF, Poltronieri P, Marsigliante S, Storelli C, Hernández M, Rodríguez-Lázaro D (2009) A filtration-based real-time PCR method for the quantitative detection of viable Salmonella enterica and Listeria monocytogenes in food samples. Food Microbiol 26:311–316CrossRefGoogle Scholar
  5. Kaevska M, Slana I (2015) Comparison of filtering methods, filter processing and DNA extraction kits for detection of mycobacteria in water. Ann Agric Environ Med 22:429–432CrossRefGoogle Scholar
  6. Kim SY, Kang DH, Kim JK, Ha YG, Hwang JY, Kim T, Lee SH (2011) Antimicrobial activity of plant extracts against Salmonella Typhimurium, Escherichia coli O157: H7, and Listeria monocytogenes on fresh lettuce. J Food Sci 76:M41–M46CrossRefGoogle Scholar
  7. Ibekwe AM, Watt PM, Grieve CM, Sharma VK, Lyons SR (2002) Multiplex fluorogenic real-time PCR for detection and quantification of Escherichia coli O157:H7 in dairy wastewater wetlands. Appl Environ Microbiol 68:4853–4862CrossRefGoogle Scholar
  8. Lee D, Kim YT, Lee JW, Seo TS (2016) An integrated direct loop-mediated isothermal amplification microdevice incorporated with an immunochromatographic strip for bacteria detection in human whole blood and milk without a sample preparation step. Biosens Bioelectron 79:273–279CrossRefGoogle Scholar
  9. Maheux AF, Bissonnette L, Boissinot M, Bernier J-LT, Huppé V, Picard FJ, Bérubé È, Bergeron MG (2011) Rapid concentration and molecular enrichment approach for sensitive detection of Escherichia coli/Shigella in potable water samples. Appl Environ Microbiol 77:6199–6207CrossRefGoogle Scholar
  10. Plutzer J, Törökné A, Karanis P (2010) Combination of ARAD microfibre filtration and LAMP methodology for simple, rapid and cost-effective detection of human pathogenic Giardia duodenalis and Cryptosporidium spp. in drinking water. Lett Appl Microbiol 50:82–88CrossRefGoogle Scholar
  11. Prompamorn P, Sithigorngul P, Rukpratanporn S, Longyant S, Sridulyakul P, Chaivisuthangkura P (2011) The development of loop-mediated isothermal amplification combined with lateral flow dipstick for detection of Vibrio parahaemolyticus. Lett Appl Microbiol 52:344–351CrossRefGoogle Scholar
  12. Rossen L, Nørskov P, Holmstrøm K, Rasmussen OF (1992) Inhibition of PCR by components of food samples, microbial diagnostic assays and DNA-extraction solutions. Int J Food Microbiol 17:37–45CrossRefGoogle Scholar
  13. Sagong H-G, Lee S-Y, Chang P-S, Heu S, Ryu S, Choi Y-J, Kang D-H (2011) Combined effect of ultrasound and organic acids to reduce Escherichia coli O157: H7, Salmonella Typhimurium, and Listeria monocytogenes on organic fresh lettuce. Int J Food Microbiol 145:287–292CrossRefGoogle Scholar
  14. Shan S, Lai W, Xiong Y, Wei H, Xu H (2015) Novel strategies to enhance lateral flow immunoassay sensitivity for detecting foodborne pathogens. J Agric Food Chem 63:745–753CrossRefGoogle Scholar
  15. Singh J, Sharma S, Nara S (2015) Evaluation of gold nanoparticle based lateral flow assays for diagnosis of enterobacteriaceae members in food and water. Food Chem 170:470–483CrossRefGoogle Scholar
  16. Surasilp T, Longyant S, Rukpratanporn S, Sridulyakul P, Sithigorngul P, Chaivisuthangkura P (2011) Rapid and sensitive detection of Vibrio vulnificus by loop-mediated isothermal amplification combined with lateral flow dipstick targeted to rpoS gene. Mol Cell Probes 25:158–163CrossRefGoogle Scholar
  17. Techathuvanan C, Draughon FA, D’Souza DH (2010) Loop-mediated isothermal amplification (LAMP) for the rapid and sensitive detection of Salmonella Typhimurium from pork. J Food Sci 75:M165–M172CrossRefGoogle Scholar
  18. Wang F, Jiang L, Ge B (2012) Loop-mediated isothermal amplification assays for detecting Shiga toxin-producing Escherichia coli in ground beef and human stools. J Clin Microbiol 50:91–97CrossRefGoogle Scholar
  19. Wang F, Yang Q, Qu Y, Meng J, Ge B (2014) Evaluation of a loop-mediated isothermal amplification suite for the rapid, reliable, and robust detection of Shiga toxin-producing Escherichia coli in produce. Appl Environ Microbiol 80:2516–2525CrossRefGoogle Scholar
  20. Wolffs PF, Glencross K, Thibaudeau R, Griffiths MW (2006) Direct quantitation and detection of salmonellae in biological samples without enrichment, using two-step filtration and real-time PCR. Appl Environ Microbiol 72:3896–3900CrossRefGoogle Scholar
  21. Yamazaki W, Kumeda Y, Uemura R, Misawa N (2011) Evaluation of a loop-mediated isothermal amplification assay for rapid and simple detection of Vibrio parahaemolyticus in naturally contaminated seafood samples. Food Microbiol 28:1238–1241CrossRefGoogle Scholar
  22. Yongkiettrakul S, Jaroenram W, Arunrut N, Chareanchim W, Pannengpetch S, Suebsing R, Kiatpathomchai W, Pornthanakasem W, Yuthavong Y, Kongkasuriyachai D (2014) Application of loop-mediated isothermal amplification assay combined with lateral flow dipstick for detection of Plasmodium falciparum and Plasmodium vivax. Parasitol Int 63:777–784CrossRefGoogle Scholar
  23. Zhao X, Li Y, Wang L, You L, Xu Z, Li L, He X, Liu Y, Wang J, Yang L (2010) Development and application of a loop-mediated isothermal amplification method on rapid detection Escherichia coli O157 strains from food samples. Mol Biol Rep 37:2183–2188CrossRefGoogle Scholar

Copyright information

© Association of Food Scientists & Technologists (India) 2019

Authors and Affiliations

  1. 1.Department of Foods and NutritionKookmin UniversitySeoulKorea

Personalised recommendations