Characterization and Evaluation of the Pathogenicity of a Novel Alpha Family Chaperone-Usher Pilus Present on Porcine Enterotoxigenic Escherichia coli.

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of diarrhea in piglets, imposing significant economic burdens on swine production worldwide. Despite its impact, detailed characterization of intestinal colonization and comprehensive genomic profiling of virulent neonatal porcine ETEC strains remain limited. In this study, we isolated a β-hemolytic ETEC strain, PT4357, from a diarrheic neonatal piglet and conducted whole-genome sequencing (WGS) coupled with experimental infection in neonatal piglets to define its pathogenicity. The PT4357 belonged to serotype O8:H7 and sequence type ST2521 and harbored multiple virulence and antimicrobial resistance (AMR) genes, including the plasmid-borne colistin resistance gene mcr-1. Interestingly, a novel chaperone-usher (CU) assembly pathway pilus has a homolog of the α family usher gene that was identified in the chromosome of PT4357. Compared with the CS1 pilus on ETEC, structural superposition of the CU pathway pilus of PT4357, namely, CS33, showed strong conservation in the major subunit and chaperone and was relatively divergent in the N-terminal of usher and minor subunits while retaining essential core folds. Oral inoculation confirmed villi attachment of the ETEC PT4357, causing watery diarrhea in neonatal piglets without mucosal damage. Furthermore, the adherence of the ETEC PT4357 on porcine IPEC cells but not human Caco-2 cells suggests limited human tropism. Our results indicate that Taiwan porcine ETECs carried AMR genes and alternative CU pathway pilus. The role of the CS33 pilus on the pathogenicity or virulence of the pathogen requires further investigation and surveillance.