BACKGROUND: The Norway rat (Rattus norvegicus) is the principal reservoir for leptospirosis in many urban settings. Few studies have identified markers for rat infestation in slum environments while none have evaluated the association between household rat infestation and Leptospira infection in humans or the use of infestation markers as a predictive model to stratify risk for leptospirosis. METHODOLOGY/PRINCIPAL FINDINGS: We enrolled a cohort of 2,003 urban slum residents from Salvador, Brazil in 2004, and followed the cohort during four annual serosurveys to identify serologic evidence for Leptospira infection. In 2007, we performed rodent infestation and environmental surveys of 80 case households, in which resided at least one individual with Leptospira infection, and 109 control households. In the case-control study, signs of rodent infestation were identified in 78% and 42% of the households, respectively. Regression modeling identified the presence of R. norvegicus feces (OR, 4.95; 95% CI, 2.13-11.47), rodent burrows (2.80; 1.06-7.36), access to water (2.79; 1.28-6.09), and un-plastered walls (2.71; 1.21-6.04) as independent risk factors associated with Leptospira infection in a household. We developed a predictive model for infection, based on assigning scores to each of the rodent infestation risk factors. Receiver operating characteristic curve analysis found that the prediction score produced a good/excellent fit based on an area under the curve of 0.78 (0.71-0.84). CONCLUSIONS/SIGNIFICANCE: Our study found that a high proportion of slum households were infested with R. norvegicus and that rat infestation was significantly associated with the risk of Leptospira infection, indicating that high level transmission occurs among slum households. We developed an easily applicable prediction score based on rat infestation markers, which identified households with highest infection risk. The use of the prediction score in community-based screening may therefore be an effective risk stratification strategy for targeting control measures in slum settings of high leptospirosis transmission.