Development of loop-mediated isothermal amplification for detection of Escherichia coli producing Shiga toxin II variant

The objective of this study was to establish a loop-mediated isothermal amplification(LAMP) method for the detection of Stx2e-producing Escherichia coli.According to 12 Stx2e sequences and 9 Stx2 sequences available in GenBank,a set of four primers were designed based on the Stx2e gene sequence.Optimal conditions,specificity and sensitivity tests were performed,respectively.In results,ladder-like products were produced with those Stx2e positive samples by LAMP,while no product was generated with other controls.The LAMP assay had a detection limit equivalent to 38 CFU/tube,which was more sensitive than PCR method(3.8×103 CFU/tube).The agreement rate between LAMP and PCR was 100% in detecting simulation samples.In conclusion,the developed LAMP assay provides a novel method for the rapid detection of E.coli producing Stx2e and diagnosis of porcine edema disease.