Technical Abstract: Aflatoxins are toxic compounds produced by several Aspergillus species that contaminate food crops worldwide. A. flavus is the most common agent of aflatoxin contamination of corn, peanuts, cottonseed, figs and tree nuts in the US. Extensive studies have elucidated the biochemical and regulatory mechanisms of aflatoxin production, but basic knowledge of the evolutionary processes that maintain toxicity in A. flavus is lacking. We sequenced 21 intergenic regions in the aflatoxin gene cluster for a sample of 43 L-strain isolates of A. flavus representing the genetic diversity within a single peanut field in Georgia. Linkage disequilibrium analyses revealed six distinct recombination blocks that separate seven contiguous genes in the cluster (aflE, aflM, aflN, aflG, aflL, aflI, and aflO). This block-like organization is the result of recombination among haplotypes representing distinct vegetative compatibility groups. Subsequent screening of molecular sequence variation in these blocks in populations of A. flavus L-strain from Africa, Argentina, Australia, and India revealed shared haplotypes with the Georgia population as well as geographic-specific haplotypes. Although recombination blocks were not detected in the Australian and Indian populations, a similar block-like organization was found in other populations. We also detected significant geographic differentiation for two noncluster genes. Whether this isolation is consistent with restricted gene flow, or indicative of other demographic/ecological processes, is currently under investigation.