Eimeria species from poultry breeder farms without previous exposure to anticoccidial vaccines in five distinct geographical regions of Iran were examined for genetic relatedness by the random amplified polymorphic DNA (RAPD) assay. Eight different oligonucleotide decamers with arbitrary DNA sequences were tested as primers to amplify DNA from five isolates of each E. acervulina, E. tenella, and E. maxima. Depending on the species/isolate-primer combination, between 1 and 14 DNA fragments ranging in size from 240 to 3000 bp were amplified. The two isolates originated from Northeast and North parts of Iran showed minor differences and two isolates originated from Northeast and Southwest of Iran showed major differences in their amplified DNA patterns. The intra-specific similarity coefficient within five isolates of each species of, E. acervulina, E. tenella, E. maxima was 74, 82 and 72%, respectively. The distance indices observed between species were greater than those found between isolates (80-90%) with all examined primers. The inferred phylogenetic tree on the fingerprinting of all species revealed that the RAPD-PCR can easily differentiate within and between species and could be a useful and valuable tool in future epidemiological studies, designing and developing of vaccines against avian coccidosis, here in Iran and neighboring countries.