Establishment of Rapid Detection of Salmonella with Loop-Mediated Isothermal Amplification

To develop a loop-mediated isothermal amplification(LAMP)assay for the rapid and sensitive detection of Salmonella.4 primers which recognized 6 distinct regions on the invA gene of Salmonella were designed and used for LAMP assay.Salmonella DNA was amplified under isothermal conditions(65°C)for 60 min,then,LAMP results were judged by electrophoretic analysis and restriction digestion.To evaluate the specificity of the LAMP assay,4 strains of Salmonella and 9 strains of non-Salmonella were tested by LAMP and conventional PCR.To compare the sensitivity of the LAMP to that of conventional PCR,S.Enteritidis were 10-fold serially diluted and was amplified by LAMP and PCR.After LAMP reaction,ladder patterns unique to the LAMP assay were observed with 4 strains of Salmonella,amplification was not observed when 9 strains of non-Salmonella were tested.The specificity of LAMP products was confirmed by restriction enzymes.The specificity of LAMP assay was similar to that of a PCR assay,but the sensitivity of LAMP was 10 times higher than that of conventional PCR assay.The detection limit of LAMP assay for invA gene was 10 cfu/ml of S.Enteritidis and that of PCR was 100 cfu/ml.LAMP method is superior to conventional PCR for its rapid detection of S.Enteritidis within 60 min.These results indicate that a rapid,specific,and sensitive LAMP method has been developed for detection of Salmonella.