Background: A leading malaria vaccine candidate, RTS,S/AS01, is based on immunogenic regions of Plasmodium falciparum circumsporozoite protein (CSP) from the 3D7 variant, and has shown modest efficacy against clinical disease in African children. It is unclear, however, what aspect(s) of the immune response elicited by this vaccine are protective. Better understanding of how diversity in the immunogenic regions of CSP (T-cell and B-cell epitopes) may relate to clinical immunity is needed to evaluate and improve the efficacy of vaccines based on CSP. Objectives: The goals of this study were to measure diversity in immunogenic regions of CSP in a natural population of parasites and identify associations between variation in amino acid sequences in CSP and the risk of infection and clinical disease caused by P. falciparum in African children. Methods: One hundred children were selected from those who had participated in a prospective cohort study designed to measure incidence of malaria infection in Bandiagara, Mali. DNA was extracted from 769 parasite-positive blood samples corresponding to both acute clinical malaria episodes and asymptomatic infections detected in monthly surveys and B- and T-cell epitope-encoding portions of the cs gene were sequenced. Non-synonymous SNP data were generated via 454, a next generation sequencing technology, for the T-cell epitopes and repeat length data was generated for the B-cell epitopes of the cs gene. Cox proportional hazards models were used to determine the effect of sequence variation in consecutive infections occurring within individuals on the time to new infection and new clinical malaria episode. Conclusions: Extensive diversity was found in the T-cell epitopes, but no associations were found between sequence variation in either the T-cell epitopes or the repeat region, and hazard of infection or clinical malaria, suggesting that naturally acquired immunity to CSP may not be allele-specific.