Detection of Salmonella by reverse transcriptase loop-mediated isothermal amplification

Abstract:

[Background] Salmonella is currently one of most common food-borne pathogens worldwide. Every year, food poisoning caused by Salmonella ranks the first among all bacterial poisoning incidents. [Objective] A reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) method was developed to detect Salmonella. [Methods] A series of primers were designed according to the specific conservative invA gene of Salmonella, Real-time fluorescent RT-LAMP method was established to detect Salmonella by optimizing primer selection and reaction conditions. The specificity and sensitivity of the method were studied by using Salmonella and its artificial contaminated skim milk samples, and compared with the sensitivity of reverse transcription polymerase chain (RT-PCR) method. [Results] The RT-LAMP assay successfully detected invA gene of Salmonella within 30 minutes at 65 °C. Five strains of Salmonella were positive and the other 21 tested strains were negative. In the extracting nucleotide from artificial contaminated skim milk samples, the sensitivity was 60 CFU/g, which was 100-fold higher than that of RT-PCR. [Conclusion] The established real-time fluorescent RT-LAMP method can rapidly detect Salmonella on site.

Key words:Reverse transcriptase loop-mediated, isothermal amplification, Salmonella, Detection, Skim milk