Molecular identification, genetic diversity, and antibiotic resistance of pathogenic bacteria in finfish aquaculture systems of southwestern Bangladesh

Abstract

Bacterial disease outbreaks continue to threaten finfish aquaculture in southwestern Bangladesh, resulting in significant economic losses and heightened food security concerns. To address the absence of robust molecular diagnostics and genetic diversity assessment for bacterial pathogens, a comprehensive five-year study (2018-2023) was conducted to evaluate pathogen prevalence, genetic diversity, and antibiotic resistance, with the goal of informing region-specific management strategies. A total of 340 fish, representing 17 species from 10 districts, were systematically collected from farms, hatcheries, nurseries, and markets. Specimens underwent phenotypic and biochemical characterization, followed by DNA extraction and quantification under stringent quality control protocols. Quantitative PCR (qPCR) utilizing species-specific primers, including the 16S rRNA housekeeping gene, enabled sensitive detection and quantification of bacterial loads. Statistical analyses included one-way ANOVA to compare mean log10 copy numbers and Ct values across districts, fish species, and tissue types, with post hoc t-tests identifying significant differences. Random Amplified Polymorphic DNA (RAPD) and cluster analyses were employed to assess genetic similarity and diversity among 75 representative isolates. Regression analysis identified nutrient concentration as a significant predictor of bacterial growth (p = 0.0461), with optimal culture conditions determined at 1-2 % nutrients, pH 6.5-7, and temperature 25-37 °C. High bacterial loads were detected in gill tissues, with Edwardsiella ictaluri (Ct 19.8 ± 2.1) and Aeromonas hydrophila (Ct 21.4 ± 1.9) being most prevalent, and the highest E. ictaluri load (8.5 ± 0.3 log10 copies) observed in Pangasius hypophthalmus from Bagerhat (p < 0.001). The lowest pathogen prevalence was recorded in Magura district. RAPD analysis demonstrated significant genetic similarity among isolates from Khulna, Satkhira, and Bagerhat. Antibiotic susceptibility testing revealed resistance to Tetracycline but susceptibility to Ciprofloxacin and Gentamicin. These findings highlight pronounced spatial variation in pathogen prevalence, genetic diversity, and antibiotic resistance, emphasizing the critical need for integrated molecular diagnostics, targeted surveillance, and sustainable management practices to protect aquaculture health and productivity in Bangladesh.

Keywords: Antibiotic resistance; Aquaculture health; Bacterial pathogens; Genetic diversity; PCR and qPCR analysis.