Rapid detection of Salmonella in soil by recombinase aided amplification method

Abstract:

[Background] Salmonella is a pathogenic bacterium that can cause human and animal diseases, and is also one of the most important foodborne bacteria. Salmonella in soil causes food poisoning in humans through plants such as vegetables. However, due to the nature of the soil and interference from other microorganisms, how to quickly identify whether the soil is contaminated with salmonella remains a challenge. [Objective] To develop a real-time recombinase aided amplification (RT-RAA) method for rapid and sensitive detection of Salmonella in soil. [Methods] A pair of specific primers and a probe were designed targeting the sequence of invA gene in Salmonella, and a recombinant plasmid containing invA gene fragment was constructed to evaluate the sensitivity of RT-RAA method. The specificity of RT-RAA method was performed by amplification of template DNA from standard strains including Salmonella enteritidis, Escherichia coli, Shigella flexneri and Staphylococcus aureus. RT-RAA method was used to detect Salmonella in soil of tomato and ginger, and plate culture method were used to verify the results. [Results] RT-RAA method can be used to detect invA gene fragment in the recombinant plasmid at a minimum copy number of 10 per reaction within 20 min at 39 °C, without cross-reaction with Escherichia coli, Shigella flexneri and Staphylococcus aureus. RT-RAA detection of the DNA in the tested soil samples showed that the tomato soil was contaminated with Salmonella, while the ginger soil was not, which was consistent with the results of plate culture. [Conclusion] RT-RAA method has the characteristics of high sensitivity and specificity, and can be used for rapid detection of soil salmonella contamination.

Key words:recombinase aided amplification, Salmonella, molecular detection, soil microorganism