Malaria is a prevalent disease in several tropical and subtropical regions, including Brazil, where it remains a significant public health concern. Even though there have been substantial efforts to decrease the number of cases, the reoccurrence of epidemics in regions that have been free of cases for many years presents a significant challenge. Due to the multifaceted factors that influence the spread of malaria, influencing malaria risk factors were analyzed through regional outbreak cluster analysis and spatio-temporal models in the Brazilian Amazon, incorporating climate, land use/cover interactions, species richness, and number of endemic birds and amphibians. Results showed that high amphibian and bird richness and endemism correlated with a reduction in malaria risk. The presence of forest had a risk-increasing effect, but it depended on its juxtaposition with anthropic land uses. Biodiversity and landscape composition, rather than forest formation presence alone, modulated malaria risk in the period. Areas with low endemic species diversity and high human activity, predominantly anthropogenic landscapes, posed high malaria risk. This study underscores the importance of considering the broader ecological context in malaria control efforts.