关键词: 沙门氏菌, 鸡肉, 实时荧光定量环介导等温扩增, 检测

Abstract: In this study, a real-time loop-mediated isothermal amplification (Rti-LAMP) assay system was developed for the rapid detection of Salmonella enterica in chicken. It showed that the lowest level of 6 CFU/reaction for pure Salmonella culture could be detected in 45 minutes by the Rti-LAMP targeting the invA gene using Midori Green as nucleic acid dye. Furthermore, 25 g of chicken was artificially with various numbers (CFU) of Salmonella, followed by enrichment culture at 37 ℃ for 4 h and then filtration through Whirl-Pak bag and the bacterial cells were pelleted by centrifugation at 13 000 g. The resulting pellets were suspended in saline and processed for cell lysis and DNA purification. Finally, 2 μL of purified DNA sample was incorporated into 25 μL of Rti-LAMP reactions at 65 ℃ for 60 min. The lowest level of 450 CFU/g of Salmonella was consistently detected by the Rti-LAMP assay. The entire assay could be completed in 7 h. A total of 21 chicken samples purchased from local market were detected by the Rti-LAMP assay using Salmonella culture as positive control. The same one sample was detected positive to Salmonella by the Rti-LAMP assay and the method described in the Chinese national standard. It suggested that the sensitivity and accuracy of the Rti-LAMP assay were comparable to those of the national standard method, and the former took only 7 h, which was highly time-saving compared with the latter.

Key words: Salmonella enterica ser. Enteritidis, chicken, real-time loop-mediated amplification, detection