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Updated: Oct 5, 2025
Loop-Mediated Isothermal Amplification for Screening Salmonella in Animal Food and Confirming Salmonella from Culture Isolation
Published on: May 20, 2020
Isothermal RNA Amplification for the Detection of Viable Pathogenic Bacteria to Estimate the Salmonella Virulence for
Ting Xue1, Ying Lu1, Hao Yang2
1Targeted Tracer Research and Development Laboratory, Institute of Respiratory Health, State Key Laboratory of Biotherapy, National Clinical Research Center for Geriatrics, West China Hospital, Sichuan University, Chengdu 610041, China.
Abstract:
Viable foodborne pathogens can cause intestinal infection and food poisoning. Herein, we reported an RNA assay allowing for sensitive (close to 1 CFU and 1% viable bacteria detectable) and rapid (within 2.5 h) detection of viable pathogenic bacteria by coupling isothermal RNA amplification (nucleic acid sequence-based amplification, NASBA) with a CRISPR/Cas13a system. NASBA allowed direct amplification of 16S rRNA extracted from viable S. enterica (RNAs degrade rapidly in dead bacteria), and the specificity of amplification was ensured using Cas13a/crRNA to recognize the amplicons. We used the CRISPR/Cas13-based NASBA assay (termed cNASBA assay) to investigate the in vivo colonization and intestinal infection of S. enterica in mice. We found that S. enterica was mainly colonized at the cecum, colon, and rectum, and the severity of enteritis caused by S. enterica was determined by the number of viable S. enterica rather than the total count of S. enterica. The cNASBA assay can quantify viable S. enterica and thus can improve the accuracy of virulence estimation compared to qPCR. It shows promise as a reliable tool for monitoring pathogen contamination and biosafety control.
