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It is known for its resistance to multiple antibiotics, particularly through the production of Extended-Spectrum Beta-Lactamases (ESBLs), and its ability to form biofilms that further complicate treatment. This study aimed to isolate and identify <jats:italic>K. pneumoniae<\/jats:italic> from animal and environmental samples and assess commercial disinfectants\u2019 effectiveness against <jats:italic>K. pneumoniae<\/jats:italic> isolates exhibiting ESBL-mediated resistance and biofilm-forming ability in poultry and equine farms in Giza Governorate, Egypt. A total of 320 samples, including nasal swabs from equine (n\u2009=\u200960) and broiler chickens (n\u2009=\u200990), environmental samples (n\u2009=\u2009140), and human hand swabs (n\u2009=\u200930), were collected. <jats:italic>K. pneumoniae<\/jats:italic> was isolated using lactose broth enrichment and MacConkey agar, with molecular confirmation via PCR targeting the <jats:italic>gyrA<\/jats:italic> and <jats:italic>magA<\/jats:italic> genes. PCR also identified ESBL genes (<jats:italic>bla<\/jats:italic><jats:sub>TEM<\/jats:sub><jats:italic>, bla<\/jats:italic><jats:sub>SHV<\/jats:sub><jats:italic>, bla<\/jats:italic><jats:sub>CTX-M<\/jats:sub><jats:italic>, bla<\/jats:italic><jats:sub>OXA-1<\/jats:sub>) and biofilm genes (<jats:italic>luxS, Uge, mrkD<\/jats:italic>). Antimicrobial susceptibility was assessed, and the efficacy of five commercial disinfectants was evaluated by measuring inhibition zones. <jats:italic>Klebsiella pneumoniae<\/jats:italic> was isolated from poultry (13.3%), equine (8.3%), wild birds (15%), water (10%), feed (2%), and human hand swabs (6.6%). ESBL and biofilm genes were detected in the majority of the isolates, with significant phenotypic resistance to multiple antibiotics. The disinfectants containing peracetic acid and hydrogen peroxide were the most effective, producing the largest inhibition zones, while disinfectants based on sodium hypochlorite and isopropanol showed lower efficacy. Statistical analysis revealed significant differences in the effectiveness of disinfectants against <jats:italic>K. pneumoniae<\/jats:italic> isolates across various sample origins (P\u2009&lt;\u20090.05). The presence of <jats:italic>K. pneumoniae<\/jats:italic> in animal and environmental sources, along with the high prevalence of ESBL-mediated resistance and biofilm-associated virulence genes, underscores the zoonotic potential of this pathogen. The study demonstrated that disinfectants containing peracetic acid and hydrogen peroxide are highly effective against ESBL-producing <jats:italic>K. pneumoniae<\/jats:italic>. Implementing appropriate biosecurity measures, including the use of effective disinfectants, is essential for controlling the spread of resistant pathogens in farm environments.<\/jats:p>","DOI":"10.1038\/s41598-024-84149-x","type":"journal-article","created":{"date-parts":[[2025,1,10]],"date-time":"2025-01-10T13:10:35Z","timestamp":1736514635000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":20,"title":["Emerging biofilm formation and disinfectant susceptibility of ESBL-producing Klebsiella pneumoniae"],"prefix":"10.1038","volume":"15","author":[{"given":"Hanan S.","family":"Khalefa","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amany A.","family":"Arafa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dalia","family":"Hamza","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khaled A. 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