Objectives. To estimate the seroprevalence of leptospirosis in rice farmers of the Alto Mayo Valley in the Peruvian region of San Martín and to identify factors associated with infection. Materials and methods. A cross-sectional study was conducted between October and November of 2010 on a sample of 254 farmers from eleven irrigation committees in the Alto Mayo Valley. A blood sample was collected as well as sociodemographic, household and labor characteristics in the form of a questionnaire. The presence of antibodies against leptospira was determined using the microscopic agglutination test and the IgM ELISA. Results. The seroprevalence by agglutination was 64.6% (95% CI: 58.6-70.6). The IgM ELISA test was reactive in 15, 0% (95% CI: 10.5-19.5). The most frequent serovar were Icterohaemorrhagiae, Autumnalis, Australis, Panama and Grippotyphosa. Independently associated factors were handling rodents (adjusted OR (aOR): 7.9; 95% CI: 1.6-37.9), working barefoot (aOR: 2.9; 95% CI: 1.2-6.8) and male sex (aOR: 4.5; 95% CI: 1.3-15.3). Conclusions. The Alto Mayo Valley is a hyperendemic for leptospirosis. The most important factors were associated with working barefoot and handling rodents in crop fields. Consequently, it is recommended to implement interventions that affect these two factors in order to reduce the risk of infection in rice farmers.