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This study aimed to phenotypically and genotypically characterize biofilm-forming\n                      <jats:italic>E. coli<\/jats:italic>\n                      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.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The overall prevalence of\n                      <jats:italic>E. coli<\/jats:italic>\n                      was 26.7% (68\/254), with the highest isolation rates in Giza (37.1%) and Cairo-Alexandria (30.2%) regions. Molecular screening of diarrheagenic\n                      <jats:italic>E. coli<\/jats:italic>\n                      (DEC) virulence genes revealed the presence of\n                      <jats:italic>st<\/jats:italic>\n                      (10.3%),\n                      <jats:italic>stx<\/jats:italic>\n                      <jats:sub>\n                        <jats:italic>2<\/jats:italic>\n                      <\/jats:sub>\n                      (5.9%), and\n                      <jats:italic>eae<\/jats:italic>\n                      (4.4%), whereas all isolates were negative for\n                      <jats:italic>stx<\/jats:italic>\n                      <jats:sub>\n                        <jats:italic>1<\/jats:italic>\n                      <\/jats:sub>\n                      . All isolates harbored the biofilm-associated\n                      <jats:italic>luxS<\/jats:italic>\n                      gene, and 80.9% of isolates also carried\n                      <jats:italic>fimH<\/jats:italic>\n                      . 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\u2013STEC (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 (\n                      <jats:italic>p<\/jats:italic>\n                      \u2009&lt;\u20090.0001). 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.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>\n                      These findings highlight the important contribution of environmental and commensal\n                      <jats:italic>E. coli<\/jats:italic>\n                      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\n                      <jats:italic>E. coli<\/jats:italic>\n                      in dairy herds.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1186\/s12917-026-05598-2","type":"journal-article","created":{"date-parts":[[2026,6,3]],"date-time":"2026-06-03T13:34:16Z","timestamp":1780493656000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Phenotypic and genotypic characterization of biofilm-forming Escherichia coli from subclinical bovine mastitis and association with antimicrobial resistance"],"prefix":"10.1186","volume":"22","author":[{"given":"Hanan S.","family":"Khalefa","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Asmaa M.","family":"Ali","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heba S.","family":"Farag","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohamed S.","family":"Kamel","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amani Abd El\u2013latif","family":"Mosleh","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noha M","family":"Bakry","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,6,3]]},"reference":[{"issue":"1","key":"5598_CR1","doi-asserted-by":"publisher","first-page":"140","DOI":"10.1111\/jam.15192","volume":"132","author":"N Zaatout","year":"2022","unstructured":"Zaatout N, Hezil D. 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