Prevalence of noncanonical virulence factors in the adherence and invasion by enterotoxigenic Escherichia coli in the small and large intestines.

INTRODUCTION: Enterotoxigenic Escherichia coli (ETEC) expresses a diverse set of noncanonical genes that increase its virulence and are involved in mucin degradation, epithelial invasion, and biofilm formation. Collectively, these factors enable effective intestinal colonization, leading to watery diarrhea. AIM: This study investigated the prevalence of the noncanonical genes tia, tibA, and leoA and their effects on the ability of ETEC to adhere, invade, and form biofilms. MATERIALS AND METHODS: A total of 244 ETEC strains were analyzed by PCR to confirm the presence of noncanonical virulence genes (tia, tibA, and leoA) and toxins, including heat-labile toxins (LTs) and/or heat-stable toxins (STs). The ability of the strains to adhere to and invade HT-29 colorectal adenocarcinoma cells (large intestine) and HuTu80 duodenal adenocarcinoma cells (small intestine) was evaluated, along with their ability to form biofilms. The susceptibility of these strains to 18 antibiotics was evaluated in accordance with the Clinical & Laboratory Standards Institute (CLSI) 2025 guidelines. RESULTS: Our data revealed that the tia, tibA and leoA genes were amplified in 11.34% (11/97), 15.46% (15/97), and 7.21% (7/97) of the ETEC strains, respectively. ETEC tia+ strains showed adherence values ranging from 3.6 × 105 to 34.1 × 105 CFUs/mL in HT-29 cells and from 4.1 × 105 to 16.6 × 105 CFUs/mL in HuTu80 cells. ETEC tibA + strains exhibited adherence values ranging from 11.16 × 105 to 65.1 × 105 CFUs/mL in HT-29 cells and from 5.6 × 105 to 27.1 × 105 CFUs/mL in HuTu80 cells. Among the ETEC tia + strains, 54.54% (6/11) were invasive, with invasion frequencies ranging 0.17% and 1.01% in HT-29 cells and between 0.20% and 0.404% in HuTu80 cells. Between the strains expressing the tibA gene, 47.05% carried both LTs and STs. Among the ETEC tibA+ strains, 20% were strongly biofilm-producing, 13.33% were moderately biofilm-producing, and 66.66% were weakly biofilm-producing. ETEC tia+/tibA+ strains exhibited high rates of multidrug resistance (MDR). CONCLUSIONS: Our data suggest that noncanonical ETEC strains are highly efficient because their adhesion genes (tia, tibA, and leoA), toxins (lt and st), and adherence and biofilm-forming capabilities work together to cause infection, even in the absence of some classical virulence factors.