A Novel Loop-Mediated Isothermal Amplification (LAMP) Assay for Detecting Salmonella Ser. Typhimurium in Egg Products.

Abstract

Background: As a leading cause of foodborne illness worldwide, detection of Salmonella enterica subsp. enterica serovar Typhimurium is essential for food safety and public health.

Objective: This study aimed to develop a loop-mediated isothermal amplification (LAMP) assay for the rapid and sensitive detection of Salmonella ser. Typhimurium in egg products.

Methods and results: The primer set targeting the open reading frame STM3845 of Salmonella ser. Typhimurium was designed using PrimerExplorer V4. The LAMP assay was optimized by adjusting reagent concentrations, reaction temperature, and incubation time, achieving the highest amplification/fluorescence in a 25.0 µL reaction at 65 °C for 30 minutes. Results indicated that the newly designed assay could successfully detect and differentiate Salmonella ser. Typhimurium from other Salmonella serotypes and non-Salmonella bacterial pathogens except for Salmonella serotypes Montevideo, Michigan, and Senftenberg after testing 73 Salmonella ser. Typhimurium, 100 non-Typhimurium Salmonella, and 35 non-Salmonella bacterial pathogens of pure cultures. The LAMP assay was further compared with a commercial real-time PCR and FDA BAM culture method by testing pure culture and 200 inoculated (1-5 CFU/25g) egg and egg product samples, and proved to be comparable to FDA BAM culture method, it also demonstrated 100 times more sensitive than the real-time PCR assay in pure culture testing, with a detection limit of 0.56 log CFU/ml.

Conclusion: The newly developed LAMP assay offers a rapid, specific, and sensitive method for detecting Salmonella ser. Typhimurium in egg products. Its simplicity, speed, and sensitivity position it as a powerful tool for routine monitoring, outbreak investigation, and on-site testing in the food industry.