Study on drug resistance, biofilm formation, phylogenetic and virulence gene analysis of Escherichia coli from diarrheic lambs

This study comprehensively characterized E. coli from diarrheal lambs in Aksu, China. We evaluated growth kinetics, phylogenetic groups, virulence and resistance genes, antimicrobial susceptibility, biofilm formation, and pathogenicity in mice to determine their potential risks. Fresh fecal samples from diarrheic lambs were collected for bacterial isolation. E. coli was identified via specific PCR and 16 S rRNA sequencing. A microbial growth analyzer was used to determine growth curves, and motility medium was used to detect active motility. Biofilm formation was assessed by crystal violet staining. The Kirby-Bauer disk diffusion method was used to test the antimicrobial resistance of the strains, while PCR was performed to identify phylogenetic groups, virulence genes, and resistance genes. Pathogenicity was confirmed via a murine infection model. A total of 28 E. coli strains were isolated from 21 diarrheal lambs. Growth kinetic analysis revealed that all the strains entered the logarithmic growth phase after approximately 5 h of cultivation. Among the strains, 53.6% exhibited active motility. Phylogenetic classification revealed a predominance of Group B1 (53.6%), followed by Group D (35.7%) and Group A (10.7%). Thirteen virulence genes and nine resistance genes were detected. The murine infection model demonstrated that 39.2% of the strains tested were pathogenic, with significant pathological lesions observed in the liver, lungs, spleen, kidneys, and small intestine of infected mice. Additionally, 64.3% of the strains were multidrug resistant (MDR), and the detection rate of extended-spectrum β-lactamase (ESBL)-producing strains was 53.6%. blaCTX-M4 was the key determinant mediating E. coli resistance to β-lactam antibiotics. Notably, complete phenotype‒genotype concordance was observed for blaNDM-mediated imipenem resistance and cmlA-mediated chloramphenicol resistance. A statistically significant correlation was found between biofilm formation capacity and resistance patterns: strains with stronger biofilm formation were more likely to be MDR- and ESBL-positive. E. coli from diarrheal lambs poses significant risks to sheep farming and may represent a zoonotic reservoir. These findings highlight the need for effective measures to control E. coli infections on sheep farms.