Clinical Distribution and Virulence Gene Profiles of CNF1- and CNF2-Positive Escherichia coli in Calves

Cytotoxic necrotizing factor (CNF)-associated Escherichia coli (E. coli) occur in enteric and extraintestinal disease in calves, but the distribution of CNF1 and CNF2 and their virulence profiles remain incompletely defined. We analysed E. coli from 544 calves: 131 healthy, 210 diarrhoeic, 173 with septicaemia, and 30 with central nervous system (CNS) infection. Isolates were classified by PCR detection of cnf1 or cnf2, and only CNF-positive isolates were screened for eight accessory virulence genes. In age-adjusted models restricted to calves aged 2–19 days, overall CNF positivity (aOR 2.83, 95% CI 1.25–6.44; Holm-adjusted p = 0.039) and CNF2 positivity (aOR 3.14, 95% CI 1.18–8.31; adjusted p = 0.043) were associated with diarrhoea, whereas CNF1 was not. CNF2-positive isolates were enriched for f17A, afa8E, and cdtIII, whereas CNF1-positive isolates more often carried papC, sfa/focDE, hlyA, and kpsMT K1. In invasive cases, CNF1 was more frequent in CNS infection than in septicaemia (33.3% vs. 12.7%; adjusted p = 0.034). Among CNF1-positive invasive isolates, kpsMT K1 occurred in 90.0% of CNS versus 9.1% of septicaemia isolates. These findings support distinct CNF2-associated enteric and CNF1-associated invasive virulence profiles; PCR detection does not establish expression, phenotype, causality, or genomic linkage.