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Detection of Listeria monocytogenes in Food Using the Proofman-LMTIA Assay
Chunmei Song1, Borui Wang2, Yongzhen Wang1
1Key Laboratory of Biomarker Based Rapid-Detection Technology for Food Safety of Henan Province, Xuchang University, Xuchang 461000, China.
Abstract:
Microbial factors, including bacteria, viruses, and other pathogens, are significant contributors to foodborne illnesses, posing serious food safety risks due to their potential for rapid growth and contamination. Listeria monocytogenes is one of the most common types of foodborne bacteria that can cause serious foodborne diseases or even fatalities. In this study, a novel nucleic acid amplification method called Proofman-LMTIA was employed to detect Listeria monocytogenes contamination in food. This method combines proofreading enzyme-mediated probe cleavage with ladder-shape melting temperature isothermal amplification. A positive recombinant plasmid was used as a control to ensure the accuracy of the detection results, and primers and Proofman probes were specifically designed for the LMTIA. Genomic DNA was extracted, the reaction temperature was optimized, and the primers' specificity was verified using foodborne pathogens like Staphylococcus aureus, Escherichia coli O157:H7, and Salmonella. The sensitivity was assessed by testing serial dilutions of genomic DNA, and the method's applicability was confirmed by detecting artificially contaminated fresh pork. The established LMTIA method exhibited both high specificity and sensitivity. At the optimal reaction temperature of 63 °C, the primers specifically identified Listeria monocytogenes contamination in pork at a concentration of 8.0 ± 0.7 colony-forming units (CFUs) per 25 g. Furthermore, the Proofman-LMTIA method was applied to test Listeria monocytogenes DNA in 30 food samples purchased from a Chinese retail market, and reassuringly, all results indicated no contamination. Proofman-LMTIA can serve as a reliable and rapid method for detecting Listeria monocytogenes in food, contributing to public health by safeguarding consumers from foodborne illnesses, and strengthening food safety regulations.
Insights
A new method, Proofman-LMTIA, accurately detects Listeria monocytogenes, a dangerous foodborne bacterium. This rapid nucleic acid amplification test enhances food safety by identifying contamination in food products.
Area of Science:
- Food Safety and Microbiology
- Molecular Diagnostics
- Nucleic Acid Amplification Techniques
Background:
- Foodborne illnesses pose significant public health risks, often caused by microbial pathogens like *Listeria monocytogenes*.
- *Listeria monocytogenes* is a prevalent foodborne bacterium capable of causing severe diseases and fatalities.
- Accurate and rapid detection methods are crucial for preventing *Listeria monocytogenes* contamination in food.
Purpose of the Study:
- To develop and validate a novel nucleic acid amplification method for the sensitive and specific detection of *Listeria monocytogenes* in food.
- To assess the performance characteristics of the new method, including specificity, sensitivity, and applicability to real food samples.
- To provide a reliable tool for food safety monitoring and regulatory compliance.
Main Methods:
- Development of Proofman-LMTIA, a novel isothermal amplification method combining proofreading enzyme-mediated probe cleavage and ladder-shape melting temperature analysis.
- Specific primer and probe design for *Listeria monocytogenes* detection.
- Optimization of reaction conditions, specificity testing against related foodborne pathogens, sensitivity assessment using serial dilutions, and validation on artificially contaminated pork and retail food samples.
Main Results:
- The Proofman-LMTIA method demonstrated high specificity and sensitivity for detecting *Listeria monocytogenes*.
- The method accurately identified *Listeria monocytogenes* contamination in pork at a low concentration (8.0 ± 0.7 CFU/25 g) at an optimal temperature of 63 °C.
- Analysis of 30 retail food samples from a Chinese market yielded no detectable *Listeria monocytogenes* contamination using the developed method.
Conclusions:
- Proofman-LMTIA is a reliable and rapid method for detecting *Listeria monocytogenes* in food matrices.
- This technology can significantly contribute to public health by enhancing food safety and strengthening regulatory measures against foodborne pathogens.
- The method offers a promising alternative for routine food safety testing and surveillance programs.
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