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Updated: Apr 26, 2026
Loop-Mediated Isothermal Amplification for Screening Salmonella in Animal Food and Confirming Salmonella from Culture Isolation
Published on: May 20, 2020
Development of double loop-mediated isothermal amplification to detect Listeria monocytogenes in food
Rina Wu1, Xiang Liu, Bangcheng Guo
1Food Biotechnology and Food Safety Laboratory, College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, People's Republic of China.
Abstract:
In this study, a double loop-mediated isothermal amplification (dLAMP) based on two target genes hlyA and iap was developed for the rapid detection of Listeria monocytogenes in food. The results revealed that the detection time and temperature of our dLAMP assay for L. monocytogenes were 15 min and 63 °C respectively, with a sensitivity of 10 fg DNA of L. monocytogenes per tube. While normal LAMP (nLAMP) of hlyA or iap was 100 fg DNA of L. monocytogenes per tube for 45 min and 63 °C. Furthermore, mineral oil and GoldViewII nucleic acid stain were chosen as the basic materials to develop a simple visualized identification of the positive samples. A total of 450 food samples were tested for L. monocytogenes using the dLAMP protocol developed in this study. The results showed that the accuracy of the dLAMP and the "gold standard" culture-biotechnical method were 100 % identical, suggesting that the modified dLAMP assay would provide a potential for detection of L. monocytogenes in food products.
Insights
A new double loop-mediated isothermal amplification (dLAMP) method rapidly detects Listeria monocytogenes in food. This assay offers high sensitivity and 100% accuracy compared to traditional methods.
Area of Science:
- Food safety
- Microbiology
- Molecular diagnostics
Background:
- Listeria monocytogenes is a significant foodborne pathogen requiring rapid detection methods.
- Existing detection methods can be time-consuming and lack sufficient sensitivity for early-stage contamination.
- Isothermal amplification techniques offer potential for rapid and sensitive pathogen detection.
Purpose of the Study:
- To develop and validate a double loop-mediated isothermal amplification (dLAMP) assay for the rapid detection of Listeria monocytogenes in food samples.
- To compare the sensitivity, specificity, and speed of the dLAMP assay against conventional methods.
- To establish a simple, visual identification method for positive samples.
Main Methods:
- Development of a dLAMP assay targeting two specific genes (hlyA and iap) of Listeria monocytogenes.
- Optimization of reaction conditions including temperature (63 °C) and time (15 min).
- Testing of 450 food samples using the developed dLAMP protocol and comparison with the gold standard culture-biotechnical method.
Main Results:
- The dLAMP assay detected as little as 10 fg of Listeria monocytogenes DNA per tube, significantly more sensitive than normal LAMP (100 fg).
- The assay achieved detection in 15 minutes, substantially faster than the 45 minutes required for normal LAMP.
- Visual identification of positive samples was enabled using mineral oil and GoldViewII nucleic acid stain.
- The dLAMP assay demonstrated 100% accuracy when compared to the gold standard culture-biotechnical method in analyzing 450 food samples.
Conclusions:
- The developed dLAMP assay provides a rapid, highly sensitive, and accurate method for detecting Listeria monocytogenes in food.
- This modified dLAMP assay has significant potential for routine application in food safety testing.
- The assay's simplicity and visual detection capability enhance its practical utility in food product analysis.
