OBJECTIVE:The aquaculture sector in Pakistan remains underdeveloped, as aquaculture has not yet received adequate attention. Studies focusing on the health status of fish in the natural water bodies within Pakistan are rare. METHODS:This study isolated and characterized bacteria from naturally infected cyprinid species across Khyber Pakhtunkhwa province. In total, 56 bacterial species were identified from 100 different naturally infected freshwater fish samples, with both internal and external organs examined. RESULTS:Molecular characterization showed that among the nine families characterized in total, the dominant families were Enterobacteriaceae and Aeromonadaceae, and the dominant species were Enterobacter spp., Aeromonas spp., Klebsiella spp., Symbiopectobacterium spp., and Citrobacter spp. This study also found bacterial species for the first time in Pakistan that are rarely reported in global aquaculture. These included Rahnella contaminans and species of Rahnella, Yokenella, and Morganella in the gills, liver, and muscle of Common Carp Cyprinus carpio, Rohu Labeo rohita, and Grass Carp Ctenopharyngodon idella. Additionally, human bacterial pathogens were also detected, which may indicate a potential health risk for fish consumers. CONCLUSIONS:Based on the level of bacterial infections observed, there is a critical need for continuous monitoring of fish populations to prevent potential disease outbreaks in aquaculture and thus avert economic losses.