MOLECULAR CHARACTERIZATION OF ANTIMICROBIAL RESISTANCE IN ENTERIC BACTERIAL PATHOGENS ISOLATED FROM DOMESTIC ANIMALS IN KHYBER PAKHTUNKHWA, PAKISTAN

Abstract

Antimicrobial resistance (AMR) in bacteria associated with domestic animals represents an important One Health challenge because resistant organisms and resistance determinants may circulate among animals, people, food systems, and the environment. This study was designed to characterize phenotypic and molecular AMR among enteric bacterial pathogens associated with domestic animals in Khyber Pakhtunkhwa, Pakistan, using an integrated culture, susceptibility testing, PCR, and sequence-based framework. The proposed surveillance includes fecal or cloacal specimens from cattle, buffaloes, goats, sheep, and broiler chickens. Bacterial isolates are identified by culture, biochemical characterization, and 16S rRNA sequencing, followed by antimicrobial susceptibility testing using standardized disk diffusion. Selected resistance determinants, including blaTEM, blaCTX-M, tetA, sul1, qnrS and mcr-1, are investigated by PCR, and representative amplicons are sequenced for phylogenetic comparison. In the illustrative model dataset, 240 specimens yielded 150 target isolates, comprising Escherichia coli (n=92) and Salmonella spp. (n=58). Multidrug resistance was 70.7%, while resistance was highest to ampicillin (58.0%), tetracycline (54.0%), trimethoprim-sulfamethoxazole (46.0%) and ciprofloxacin (38.0%). The model PCR profile showed blaTEM in 43.3% of isolates, tetA in 36.7%, blaCTX-M in 32.0%, sul1 in 30.0%, qnrS in 18.0% and mcr-1 in 15.3%. The proposed phylogenetic component separates representative E. coli and Salmonella lineages and provides a complementary framework for examining relationships among resistance-gene sequences. These numerical findings and the phylogenetic topology are illustrative only and must be replaced with experimentally generated observations, sequence data, bootstrap support and accession numbers before submission. The framework nevertheless demonstrates how multi-host AMR surveillance can integrate phenotype, genotype and evolutionary relationships within a Pakistan-focused One Health context. The approach may strengthen surveillance and guide antimicrobial stewardship while avoiding unsupported claims of transmission.