Prevalence and Antibiotic Resistance Profiles of Quinolone-Resistant Bacteria fromFreshwater Fish Farms in Southern Punjab, Pakistan

Quinolone resistance in bacteria poses a global health threat, and aquaculture environments may act as reservoirs for resistant strains and their genes.We investigated the prevalence, antimicrobial resistance profiles, and key genetic determinants of quinolone resistance by screening for associated resistance genes among 101 bacterial isolates from water and sediment at freshwater fish-farming sites in four districts of Southern Punjab, Pakistan (Bahawalpur, Lodhran, Multan, Muzaffargarh).Isolates were tested for susceptibility to enrofloxacin, ciprofloxacin, and norfloxacin via disk diffusion.Phenotypic prevalence and genetic associations were analyzed using Chi-square tests.Resistant isolates underwent PCR screening for plasmid-mediated quinolone resistance genes (qnrA, qnrB, qnrS) and mutations in the quinolone resistance-determining regions (QRDR) of gyrA and gyrB.Selected resistant isolates were identified by 16S rRNA sequencing.Phenotypic resistance was 30% (95% CI: 21-39) for norfloxacin, 27% (95% CI: 18-36) for enrofloxacin, and 18% (95% CI: 10-26) for ciprofloxacin.District-wise significant variations were observed for enrofloxacin (P=0.01) and ciprofloxacin (P<0.001)resistance.Among the three-plasmid mediated quinolone resistance (PMQR) genes (qnrA, qnrB, qnrS) screened, a significant association was found between the presence of these genes and phenotypic resistance to enrofloxacin (P=0.03),ciprofloxacin (P=0.05), and norfloxacin (P=0.01).Among all three PMQR genes, qnrS was the most frequently detected gene (37-40%), followed by qnrA (27-33%).District-level resistance to enrofloxacin was highest in Multan water (53%) and Bahawalpur sediment (44%), but low (0-10%) in Muzaffargarh and Lodhran.Ciprofloxacin resistance was predominately seen in Bahawalpur sediment (50%) and water from Multan (29%) and Muzaffargarh (25%), with none in Lodhran.Norfloxacin resistance peaked in Muzaffargarh water (50%), followed by Multan water (41%) and sediment (40%), with overall greater susceptibility in water isolates.Isolates had high multi-antibiotic resistance indices (~0.5), indicated anthropogenic pollution.Species identified included Pseudomonas aeruginosa and Achromobacter xylosoxidans.These findings highlight the dissemination of PMQR genes in multidrug-resistant opportunistic pathogens.We recommend strengthened antimicrobial stewardship, improved biosecurity, and routine AMR surveillance in aquaculture to protect animal, public, and environmental health.