Development of a real-time loop-mediated isothermal amplification assay for the sensitive and rapid detection of Listeria monocytogenes

L Ye; Y Li; J Zhao; Z Zhang; H Meng; H Yan; S-I Miyoshi; L Shi

doi:10.1111/lam.12429

Development of a real-time loop-mediated isothermal amplification assay for the sensitive and rapid detection of Listeria monocytogenes

Lett Appl Microbiol. 2015 Jul;61(1):85-90. doi: 10.1111/lam.12429. Epub 2015 May 10.

Authors

L Ye¹, Y Li¹, J Zhao², Z Zhang³, H Meng¹, H Yan¹, S-I Miyoshi⁴, L Shi¹

Affiliations

¹ College of Light Industry and Food Sciences, South China University of Technology, Guangzhou, China.
² College of Environmental and Biological Engineering, Guangdong University of Petrochemical Technology, Maoming, China.
³ State Key Laboratory of Food Safety Technology for Meat Products, Xiamen Yinxiang Group Co., Ltd., Xiamen, China.
⁴ Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Tsushima-Naka, Okayama, Japan.

PMID: 25868481
DOI: 10.1111/lam.12429

Abstract

A real-time loop-mediated isothermal amplification (RealAmp) assay for the detection of Listeria was developed. The RealAmp assay, using primers specific for the hemolysin-encoding hlyA gene, was verified using Listeria monocytogenes strains (n = 58) from different regions of the world. Both the sensitivity and specificity of the RealAmp assay were high. The RealAmp assay could detect 10(3) CFU ml(-1) within 30 min. A comparative evaluation of the RealAmp assay, the API Listeria assay, and the real-time PCR assay revealed that the RealAmp assay is simpler, faster, and has a higher specificity than the other two assays.

Significance and impact of the study: Conventional culture and molecular detection methods are always time consuming and require a specific laboratory infrastructure, thereby restricting their use for the rapid detection and diagnosis of pathogens. A real-time loop-mediated isothermal amplification (RealAmp) assay performed by ESEtube scanner to rapidly detect Listeria monocytogenes isolated from food was developed. The results showed that the RealAmp assay using the tube scanner was more efficient and precise than the conventional API Listeria assay and the real-time PCR assay.

Keywords: API Listeria; ESEQuant tube scanner; Listeria monocytogenes; RealAmp; real-time PCR.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

DNA Primers
Food Microbiology / methods*
Hemolysin Proteins / genetics
Humans
Listeria monocytogenes / isolation & purification*
Nucleic Acid Amplification Techniques / methods*
Real-Time Polymerase Chain Reaction

Substances

DNA Primers
Hemolysin Proteins