Development of multiplex cross displacement amplification combined with lateral flow biosensor assay for detection of virulent

Shigella sonnei is the most common Shigella spp. in developed areas and the second most common in undeveloped regions. In this study, a multiple cross displacement amplification (MCDA) assay was used in combination with a lateral flow biosensor (LFB) assay to detect virulent S. sonnei strains containing the ipaH and wbgX genes. The multiplex MCDA-LFB assay detected wbgX at ≥1 pg/μL and ipaH at ≥10 fg/μL within 30 min in pure cultures maintained at 63°C. This assay was sensitive for ~37 CFU of virulent S. sonnei and ~3.7 CFU of Shigella spp. and enteroinvasive E. coli in stimulated fecal samples and had 100% specificity among 59 reference strains. The MCDA-LFB assay was also able to differentiate between virulent S. sonnei and other Shigella spp. and enteroinvasive E. coli among 99 clinical isolates. In summary, a multiplex MCDA-LFB assay was developed for rapid, convenient, point-of-care, and accurate identification of virulent S. sonnei within 30 min and at a constant temperature without the need for expensive lab equipment.