Genetic variation in the pistachio late blight fungus, Alternaria alternata, was investigated by restriction fragment length polymorphism (RFLP) in the rDNA region. Southern hybridization of EcoRI, HindIII, and XbaI digested fungal DNA with a RNA probe derived from Alt1, an rDNA clone isolated from a genomic library of the Japanese pear pathotype of A. alternata, revealed 34 different rDNA haplotypes among 56 isolates collected from four central valley locations in California. Analysis of molecular variation revealed a significant amount of genetic diversity within populations (85.8%), with only marginal variation accounting for differentiation among populations (14.2%, ϕST =0.142). All isolates examined were highly pathogenic. The identity of the four geographic populations sampled was not evident in both cluster and principal component analyses, probably indicating either the selectively neutral nature of rDNA variation or prevalence of widespread gene flow among populations combined with uniform host-selection.