A low resource requirement molecular diagnostic and surveillance tool for Shigella in the era of vaccination

Abstract

Infection with Shigella bacteria is one of the leading causes of diarrhoeal disease globally, with significant burden in low- and middle-income countries and rising incidence in high-income settings. Effective serotyping is essential for surveillance, outbreak investigation, vaccine targeting, and can inform clinical management. Current discriminative approaches, such as seroagglutination and whole genome sequencing, are constrained by cross-reactivity, infrastructure requirements, and cost. Here, we present the development of a nine-target multiplex PCR lateral flow device assay for the rapid, accurate identification of vaccine-prioritised S. flexneri serotypes 1b, 2a, and 3a, and S. sonnei , as well as clinically relevant antimicrobial resistance genes. Validation using 138 DNA samples from clinical Shigella isolates showed 97.8% sensitivity and 100% specificity at the species level, performing comparably to in silico genomic prediction tools. Serotype classifications interpreted from the PCR assay also successfully aligned with the temporal and geographic trends observed in a large multi-country clinical dataset ( n  = 1,074), supporting its utility for temporospatial surveillance. To facilitate field-deployment of the PCR assay, we integrated an oligo-chromatographic dipstick method, enabling a simple, gel-free readout without need for specialised equipment. This molecular dipstick assay provides a practical and scalable solution for Shigella serotyping and antimicrobial resistance profiling in support of vaccine implementation and surveillance efforts.