Emergence of phenotypic and genotypic resistance in the intestinal microbiota of rainbow trout (Oncorhynchus mykiss) exposed long-term to sub-inhibitory concentrations of sulfamethoxazole.

Natural waters are contaminated globally with pharmaceuticals including many antibiotics. In this study, we assessed the acquisition of antimicrobial resistance in the culturable intestinal microbiota of rainbow trout (Oncorhynchus mykiss) exposed for 6 months to sub-inhibitory concentrations of sulfamethoxazole (SMX), one of the most prevalent antibiotics in natural waters. SMX was tested at three concentrations: 3000 µg/L, a concentration that had no observed effect (NOEC) on the in vitro growth of fish intestinal microbiota; 3 µg/L, a theoretical predicted no effect concentration (PNEC) for long-term studies in natural environments; and 0.3 µg/L, a concentration detected in many surveys of surface waters from various countries including the USA. In two independent experiments, the emergence of phenotypic resistance and an increased prevalence of bacteria carrying a sulfonamide-resistance gene (sul1) were observed in SMX-exposed fish. The emergence of phenotypic resistance to1000 mg/L SMX was significant in fish exposed to 3 µg/L SMX and was in large part independent of sul resistance genes. The prevalence of bacteria carrying the sul1 resistance gene increased significantly in the culturable intestinal microbiota of SMX-exposed fish, but the sul1-positive population was in large part susceptible to 1000 mg/L SMX, suggesting that the gene confers a lower resistance level or a growth advantage. The increased prevalence of sul1 bacteria was observed in all groups of SMX-exposed fish. Overall, this study suggests that fish exposed long-term to waters contaminated with low levels of antibiotics serve as reservoir of antimicrobial resistant genes and of resistant bacteria, a potential threat to public health.