ABSTRACT Background This retrospective case-control study aimed to investigate the factors associated with the occurrence of human visceral leishmaniasis (VL) in the municipality of Rondonópolis, which is a highly endemic area for VL in Central-Western Brazil. Methods All VL cases reported in Rondonópolis from 2011 to 2016 were included. For each case, two neighborhood- and age-matched controls with the absence of past and present history of VL and negative rk39-based immunochromatographic test, were recruited. Data were retrospectively collected through interviews that assessed socioeconomic aspects, housing features, backyard characteristics, presence of animals and daily habits. A multivariate conditional logistic regression analysis was used to identify factors associated with human VL. Results During the study period, 81 VL cases were reported in Rondonópolis, of which 37 (45.7%) were included in this study. Seventy controls were also included. The variables associated with VL occurrence were the outdoor location of a bathroom and/or kitchen separated from the rest of the house (OR 7.8; 95% CI 2.1 to 29.5), the presence of a vegetable garden (OR 5.3; 95% CI 1.3 to 21.2) and the presence of decaying fruits (OR 10.7; 95% CI 2.0 to 52.1) in the backyard. Conclusions Our findings suggest an association between backyard characteristics and VL. This may be useful in guiding preventive and control measures against zoonotic VL worldwide.