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bioRxiv · 10.64898/2026.01.23.701439

Genomic diversity, antibiotic resistance, and maturation-dependent adhesion of F18 enterotoxigenic Escherichia coli strains in porcine intestinal cells

Liu, D.·Gallina, N.·Li, C.·Irizarry-Tardi, N.·Sayedahmed, M.·Karunathilaka, J. C.·Horn, N.·Wang, W.·Khalek, A. A.·Bhunia, A. K.
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Abstract

Enterotoxigenic Escherichia coli (ETEC) strains expressing F4 and F18 fimbriae are major causes of neonatal and post-weaning diarrhea in swine. Although epithelial maturation influences susceptibility in vivo, its impact on ETEC-host interactions remains poorly defined. This study characterizes emerging F18 ETEC isolates using a differentiated porcine intestinal cell model. Three F18 strains (3EC1, 27EC1, 3247EC), a porcine F4 strain, and human ETEC H10407 were analyzed by comparative genomics for virulence factors, toxin genes, and antimicrobial resistance determinants. Adhesion assays were performed using IPEC-1, IPEC-J2, and Caco-2 cells conditioned to Early (6 days post-confluence, DPC), Mid (9 DPC), and Late (16 DPC) maturation states. Transcription of F18-binding receptors (FUT1, FUT2) was quantified by RT-qPCR. IPEC-1 cells exhibited significantly higher FUT1 and FUT2 expression than IPEC-J2, corresponding to approximately two-fold stronger adhesion by most F18 isolates. Strain 3EC1 showed a distinct adhesion peak at 9 DPC, approaching F4 levels, while F4 and H10407 consistently displayed the highest adhesion across all models. Genomic analyses revealed substantial heterogeneity among F18 strains in fimbrial loci, flagellin, lipopolysaccharide biosynthesis, and antimicrobial resistance. Strain 3EC1 uniquely carried stx2e, and non-classical EAST1 variants were detected in 3EC1 and 3247EC. All F18 isolates encoded hlyE and were {beta}-hemolytic; 3247EC harbored 28 antimicrobial resistance genes. The IPEC-1/IPEC-J2 maturation stages recapitulate age-dependent susceptibility to ETEC, likely driven by FUT1/2 expression levels. The combination of strong adhesion, stx2e, and extensive antimicrobial resistance in F18 strains underscores their evolving virulence and supports this model as a refined platform for studying porcine ETEC pathogenesis. ImportanceETEC remains a leading cause of neonatal and post-weaning diarrhea in swine, yet the biological basis for age-dependent susceptibility is not fully understood. This study demonstrates that maturation of porcine intestinal epithelial cells strongly influences F18 ETEC adhesion, driven in part by developmental regulation of the F18-binding receptors FUT1 and FUT2. By integrating comparative genomics with a physiologically relevant in vitro maturation model, we reveal substantial diversity in virulence and resistance among F18 strains, including strong adhesion capacity, stx2e, and extensive antimicrobial resistance in strain 3EC1. These findings highlight the evolution of ETEC toward increased persistence and pathogenic potential in swine populations. The interaction between the maturation-dependent IPEC-1 and IPEC-J2 cell lines and ETEC offers a valuable tool for evaluating intervention strategies to reduce weaning piglet susceptibility to ETEC infection.

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Liu, D., Gallina, N., Li, C., Irizarry-Tardi, N., Sayedahmed, M., Karunathilaka, J. C., Horn, N., Wang, W., Khalek, A. A., Bhunia, A. K.. 2026-01-25. Genomic diversity, antibiotic resistance, and maturation-dependent adhesion of F18 enterotoxigenic Escherichia coli strains in porcine intestinal cells. https://doi.org/10.64898/2026.01.23.701439

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