Fluoroquinolones (FQs) and ciprofloxacin (Cp) are important antimicrobials that pollute the environment in trace amounts. Although Cp has been recommended as prophylaxis for patients undergoing leech therapy to prevent infections by the leech gut symbiont Aeromonas, a puzzling rise in Cp-resistant (Cp r) Aeromonas infections has been reported. We report on the effects of subtherapeutic FQ concentrations on bacteria in an environmental reservoir, the medicinal leech, and describe the presence of multiple antibiotic resistance mutations and a gain-offunction resistance gene. We link the rise of Cp r Aeromonas isolates to exposure of the leech microbiota to very low levels of Cp (0.01 to 0.04 g/ml), Ͻ1/100 of the clinical resistance breakpoint for Aeromonas. Using competition experiments and comparative genomics of 37 strains, we determined the mechanisms of resistance in clinical and leech-derived Aeromonas isolates, traced their origin, and determined that the presence of merely 0.01 g/ml Cp provides a strong competitive advantage for Cp r strains. Deep-sequencing the Cp r-conferring region of gyrA enabled tracing of the mutation-harboring Aeromonas population in archived gut samples, and an increase in the frequency of the Cp r-conferring mutation in 2011 coincides with the initial reports of Cp r Aeromonas infections in patients receiving leech therapy.