Development of loop-mediated isothermal amplification(LAMP) method for detection of Enterobacter sakazakii

Objective To establish a LAMP method for detection of Enterobacter sakazakii.Methods The loop-mediated isothermal amplification(LAMP) amplifying DNA with high specificity and rapidity under an isothermal condition was applied for rapid detection of Enterobacter sakazakii.A set of four primers,two outer and two inner,was designed specifically to recognize the outer membrane protein A gene(OmpA) of Enterobacter sakazakii.The LAMP reaction mix was optimized.Results The most optimal reaction temperature and time of the LAM Passay for the OmpAgene were 58 and 60 minutes, respectively.Genomic DNAs from 26 bacterial strains including 8 Enterobacter sakazakii strains were amplified using LAM P, and no amplicon was observed in other bacterial strains.The detection lmiit of the LAM Passay was a round 101 colonies forming units for pure cultures.In addition, this method was applied to detect food samples.Conclusion The results suggest that detection of Enterobac ter saka zakii by LAMP is an effective and lowcost procedure with high specificity and sensitivity that requires no specialized equipment.The assay is expected to become a valuable tool for rapid detection and identification of Enterobacter sakazakii.