Genomic, transcriptomic, and phenotypic differences among archetype Shigella flexneri strains of serotypes 2a, 3a, and 6

Abstract

Shigella is one of the leading causes of diarrheal disease in low-to-middle income countries, where it accounts for significant disease burden in children under the age of five. Shigella flexneri , one of four Shigella species, is composed of 15 serotypes and caused 65.9% of all shigellosis cases within the Global Enteric Multicenter Study. S. flexneri serotypes 2a, 3a, and 6 are the three leading serotypes implicated in clinical disease, yet there are limited studies examining the variation of pathogenesis and virulence between serotypes. Our data revealed through in silico and in vitro analysis that several serotype-pecific differences exist between archetype strains 2457T ( S. flexneri serotype 2a [ Sf 2a]), J17B ( S. flexneri serotype 3a [ Sf 3a), and CCH060 ( S. flexneri serotype 6 [ Sf6 ]). Comparative genomics of these archetype strains demonstrated that CCH060 contains the greatest amount of unique genomic content compared to the Sf 3a and Sf 2a archetype strains, as well as lacks several previously identified Shigella virulence genes. The pINV virulence plasmid contains a highly conserved core genome irrespective of the archetype reference strain with the greatest strain-specific genomic features. The transcriptional responses of each archetype strain to bile salt stimulus were unique in a strain-dependent manner. Phenotypic analysis revealed that while archetype strains adhere at similar levels to cultured HT-29 intestinal cells, there are differences in the production and secretion of IpaB, IpaC, and IpaD, as well as reduced invasion by CCH060 compared to 2457T and J17B. Our results identify strain-specific features that support further large-scale analysis to identify unique serotype-specific host-pathogen interactions. Given the genomic diversity between S. flexneri serotypes and the paucity of data to support serotype-specific phenotypic differences, we applied in silico and in vitro functional analyses of archetype strains of 2457T ( Sf 2a), J17B ( Sf 3a), and CH060 ( Sf 6). These archetype strains represent the three leading S. flexneri serotypes recommended for inclusion in multivalent vaccines. Characterizing the genomic and phenotypic variation among these clinically prevalent serotypes is an important step toward understanding serotype-specific host-pathogen interactions to optimize the efficacy of multivalent vaccines and therapeutics. This study underpins the importance for further large-scale serotype-targeted analyses.