Molecular characterization and co-occurrence of ESBL, carbapenemase and mcr-1 genes in Klebsiella pneumoniae from poultry production systems in Pakistan.

BACKGROUND AND AIMS: Klebsiella pneumoniae is an important opportunistic pathogen with a high capacity to acquire antimicrobial resistance determinants. However, data on the distribution and co-occurrence of extended-spectrum β-lactamase (ESBL), carbapenemase and plasmid-mediated colistin resistance genes in poultry-associated K. pneumoniae in Pakistan remain limited. This study investigated the prevalence, antimicrobial resistance profiles and molecular characteristics of K. pneumoniae recovered from commercial broiler farms, backyard poultry flocks and retail poultry markets. METHODS: A total of 900 samples were collected from commercial broiler farms, backyard poultry flocks and retail poultry markets in Faisalabad Division, Pakistan, between September 2024 and April 2025. K. pneumoniae isolates were identified using conventional biochemical testing, API 20E, MALDI-TOF MS and rpoB gene amplification. Antimicrobial susceptibility testing was performed according to CLSI 2024 recommendations, and PCR was used to detect ESBL, carbapenemase, plasmid-mediated quinolone, aminoglycoside, sulfonamide, tetracycline, chloramphenicol and colistin resistance genes. RESULTS: K. pneumoniae was recovered from 303/900 samples (33.7%). Among the isolates, 199 (65.7%) were multidrug-resistant (MDR) and 48 (15.8%) were extensively drug-resistant (XDR). Commercial farm isolates showed a greater resistance burden than isolates from backyard flocks and retail markets. The most frequently detected ESBL genes were blaCTXM-15 and blaTEM, while blaNDM, blaKPC and blaOXA-48 were detected among carbapenem-resistant isolates. Colistin resistance was detected in 5.9% of isolates, with mcr-1 identified among the resistant isolates. Twenty-one distinct resistance gene combinations were identified, including combinations involving ESBL, carbapenemase and colistin resistance determinants. CONCLUSIONS: Poultry-associated K. pneumoniae showed a substantial burden of multidrug resistance and diverse combinations of clinically important resistance determinants, particularly among isolates from commercial farms. The findings support strengthened antimicrobial stewardship, farm-level biosecurity and integrated molecular surveillance within poultry production systems as components of a One Health approach to AMR control.