AUTHOR=Liao Chuan , Pan Lele , Tan Meiying , Zhou Zihan , Long Shaoping , Yi Xueli , Li Xuebin , Wei Guijiang , Liang Lina 

TITLE=A dual RPA-LFD assay for the simultaneous detection of Salmonella typhimurium and Salmonella enteritidis

JOURNAL=Frontiers in Bioengineering and Biotechnology

VOLUME=12

YEAR=2024

URL=https://www.frontiersin.org/journals/bioengineering-and-biotechnology/articles/10.3389/fbioe.2024.1379939

DOI=10.3389/fbioe.2024.1379939

ISSN=2296-4185

ABSTRACT=<p><bold>Introduction:</bold><italic>Salmonella</italic> was one of the most common bacteria that caused foodborne illness, with <italic>S. typhimurium</italic> (<italic>Salmonella typhimurium</italic>) and <italic>S. enteritidis</italic> (<italic>Salmonella enteritidis</italic>) infections accounting for more than 75% of human salmonella infections.</p><p><bold>Methods:</bold> In this study, we developed a method of dual recombinase polymerase amplification (RPA) combined with a lateral flow dipstick for the rapid detection of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in clinical specimens (stool).</p><p><bold>Results:</bold> The entire reaction process, including amplification and result reading, could be completed within 65 min. The detection limits of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in pure culture samples were 5.23 × 10<sup>1</sup> CFU/mL and 3.59 × 10<sup>1</sup> CFU/mL, respectively. The detection limits of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in artificially contaminated samples were 8.30 × 10<sup>1</sup> CFU/mL and 2.70 × 10<sup>2</sup> CFU/mL, respectively. In addition, the method had no cross-reaction with other pathogenic microorganisms. The results in clinical samples were fully consistent with those obtained using Bacterial Analysis Manual, with sensitivity and specificity were 100% (8/8) and 100% (17/17) for <italic>S. typhimurium</italic> and 100% (4/4) and 100% (21/21) for <italic>S. enteritidis</italic>, respectively.</p><p><bold>Discussion:</bold> The detection limits of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in artificially contaminated samples were higher than those in pure culture samples, which might be attributed to the inherent complex composition of artificially contaminated samples. In addition, the detection limits of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in the same sample were also different, which might be attributed to different amplification efficiency of two target genes in the same reaction system.</p><p><bold>Conclusion:</bold> This assay had potential application outdoors, as it could be performed within 1 h at 38°C without a complex instrument, and the results could be observed with the naked eye. In conclusion, the dual RPA-LFD assay established in this study had practical significance for the rapid detection of <italic>S. typhimurium</italic> and <italic>S. enteritidis</italic> in the future.</p>