Rapid culture-based identification of Shiga toxin-producing Escherichia coli and Shigella spp./Enteroinvasive E. coli using the eazyplex® EHEC complete assay

Abstract

Shiga toxin-producing Escherichia coli (STEC) and Shigella spp./enteroinvasive E. coli (EIEC) are common diarrheagenic bacteria that cause sporadic diseases and outbreaks. Clinical manifestations vary from mild symptoms to severe complications. For microbiological diagnosis, culture confirmation of a positive stool screening PCR test is challenging because of time-consuming methods for isolation of strains, wide variety of STEC pathotypes, and increased emergence of non-classical strains with unusual serotypes. Therefore, molecular assays for the rapid identification of suspect colonies growing on selective media are very useful. In this study, the performance of the newly introduced eazyplex® EHEC assay based on loop-mediated isothermal amplification (LAMP) was evaluated using 18 representative STEC and Shigella strains and 31 isolates or positive-enrichment broths that were collected from clinical stool samples following screening by BD MAX™ EBP PCR. Results were compared to real-time PCR as a reference standard. Overall, sensitivities and specificities of the eazyplex® EHEC were as follows: 94.7% and 100% for Shiga toxin 1 (stx1), 100% and 100% for stx2, 93.3% and 97.1% for intimin (eae), 100% and 100% for enterohemolysin A (ehlyA), and 100% and 100% for invasion-associated plasmid antigen H (ipaH) as Shigella spp./EIEC target, respectively. Sample preparation for LAMP took only some minutes, and the time to result of the assay ranged from 8.5 to 13 min. This study shows that eazyplex® EHEC is a very fast and easy to perform molecular assay that provides reliable results as a culture confirmation assay for the diagnosis of STEC and Shigella spp./EIEC infections.

Keywords: E. coli; LAMP; Shiga toxin; Shigella; Stool culture.