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Hantavirus cardiopulmonary syndrome (HCPS), which is caused by infection with Choclo virus, is uncommon in Panama, yet seropositivity among rural residents is as high as 60%. To clarify the environmental risk factors favoring rodent-to-human transmission, we tested serum from 3,067 rodents captured over a five-year period for antibodies against recombinant N protein of hantavirus by enzyme immunoassay and strip immunoblot. Among 220 seropositive rodents, Oligoryzomys fulvescens, the reservoir of Choclo virus, had the highest overall seroprevalence (23.5%); more abundant rodents (Zygodontomys brevicauda and Sigmodon hirsutus) had lower seroprevalences. In the mixed (combined modern and traditional) productive agroecosystem, the highest seroprevalence was among O. fulvescens captured in residences and in crops grown within 40 meters of a residence, with significantly lower seroprevalence in adjacent pasture and non-productive vegetation. Thus, crop habitats may serve as refugia for invasion into adjacent human residences and suggests several interventions to reduce human infection.