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Background <p>Bovine mastitis is one of the most significant and costly diseases in the dairy industry, and <i>Escherichia coli</i> (<i>E. coli</i>) is a common etiological agent of subclinical mastitis. This study aimed to phenotypically and genotypically characterize biofilm-forming <i>E. coli</i> isolated from subclinical bovine mastitis and to investigate their association with antimicrobial resistance. A total of 254 composite milk samples were collected from cows with subclinical mastitis in five dairy farms in Egypt.</p> Results <p>The overall prevalence of <i>E. coli</i> was 26.7% (68/254), with the highest isolation rates in Giza (37.1%) and Cairo-Alexandria (30.2%) regions. Molecular screening of diarrheagenic <i>E. coli</i> (DEC) virulence genes revealed the presence of <i>st</i> (10.3%), <i>stx</i><sub><i>2</i></sub> (5.9%), and <i>eae</i> (4.4%), whereas all isolates were negative for <i>stx</i><sub><i>1</i></sub>. All isolates harbored the biofilm-associated <i>luxS</i> gene, and 80.9% of isolates also carried <i>fimH</i>. Based on virulence gene profiles, 7.4%, 4.4%, 2.9%, and 2.9% of isolates were classified as ETEC, EPEC, STEC, and hybrid ETEC–STEC (ETST), respectively, while 82.4% were DEC-negative. Phylogrouping showed a predominance of group B1 (51.9%), followed by group A (35.3%). Antimicrobial susceptibility testing revealed high resistance rates to amoxicillin/clavulanic acid (60.3%) and cefuroxime (58.8%), whereas all isolates were susceptible to cefepime, and most were susceptible to enrofloxacin (95.6%) and nalidixic acid (94.1%). Multidrug resistance was detected in 52.9% of isolates, mainly involving resistance to amoxicillin/clavulanic acid, ampicillin, tetracycline, and sulpha/trimethoprim. Biofilm formation assessed using the microtiter plate assay showed that 44.1% of isolates were moderate and 38.2% were strong biofilm producers, with significant variation between farms (<i>p</i> &lt; 0.0001). Hierarchical clustering and chi-square analyses indicated that biofilm strength was associated with resistance to selected beta-lactams (amoxicillin/clavulanic acid, ampicillin, and cefuroxime), phylogenetic background (especially groups A and B1), and farm origin, whereas MDR status and most virulence genes showed weak or no association.</p> Conclusion <p>These findings highlight the important contribution of environmental and commensal <i>E. coli</i> lineages to subclinical mastitis in Egypt and underscore the need for improved farm hygiene and prudent antimicrobial use to limit the spread of biofilm-forming and antimicrobial-resistant <i>E. coli</i> in dairy herds.</p>

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Phenotypic and genotypic characterization of biofilm-forming Escherichia coli from subclinical bovine mastitis and association with antimicrobial resistance

  • 期刊论文

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  • 发表日期：

    2026年6月3日

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  • Hanan S. Khalefa,
  • Asmaa M. Ali,
  • Heba S. Farag,
  • Mohamed S. Kamel,
  • Amani Abd El–latif Mosleh,
  • Noha M Bakry

摘要

Background

Bovine mastitis is one of the most significant and costly diseases in the dairy industry, and Escherichia coli (E. coli) is a common etiological agent of subclinical mastitis. This study aimed to phenotypically and genotypically characterize biofilm-forming E. coli isolated from subclinical bovine mastitis and to investigate their association with antimicrobial resistance. A total of 254 composite milk samples were collected from cows with subclinical mastitis in five dairy farms in Egypt.

Results

The overall prevalence of E. coli was 26.7% (68/254), with the highest isolation rates in Giza (37.1%) and Cairo-Alexandria (30.2%) regions. Molecular screening of diarrheagenic E. coli (DEC) virulence genes revealed the presence of st (10.3%), stx2 (5.9%), and eae (4.4%), whereas all isolates were negative for stx1. All isolates harbored the biofilm-associated luxS gene, and 80.9% of isolates also carried fimH. Based on virulence gene profiles, 7.4%, 4.4%, 2.9%, and 2.9% of isolates were classified as ETEC, EPEC, STEC, and hybrid ETEC–STEC (ETST), respectively, while 82.4% were DEC-negative. Phylogrouping showed a predominance of group B1 (51.9%), followed by group A (35.3%). Antimicrobial susceptibility testing revealed high resistance rates to amoxicillin/clavulanic acid (60.3%) and cefuroxime (58.8%), whereas all isolates were susceptible to cefepime, and most were susceptible to enrofloxacin (95.6%) and nalidixic acid (94.1%). Multidrug resistance was detected in 52.9% of isolates, mainly involving resistance to amoxicillin/clavulanic acid, ampicillin, tetracycline, and sulpha/trimethoprim. Biofilm formation assessed using the microtiter plate assay showed that 44.1% of isolates were moderate and 38.2% were strong biofilm producers, with significant variation between farms (p < 0.0001). Hierarchical clustering and chi-square analyses indicated that biofilm strength was associated with resistance to selected beta-lactams (amoxicillin/clavulanic acid, ampicillin, and cefuroxime), phylogenetic background (especially groups A and B1), and farm origin, whereas MDR status and most virulence genes showed weak or no association.

Conclusion

These findings highlight the important contribution of environmental and commensal E. coli lineages to subclinical mastitis in Egypt and underscore the need for improved farm hygiene and prudent antimicrobial use to limit the spread of biofilm-forming and antimicrobial-resistant E. coli in dairy herds.