Introduction: Escherichia coli is one of the most prevalent bacterial species which cause gastrointestinal and digestive tract infections in humans and livestock. This study examined genotypic diversity of the E. coli isolates taken from fecal specimens in Zabol using random amplification of polymorphic DNA (RAPD) method and phylogenetic background.Materials and Methods: In this study, 100 isolates were collected from human samples and identified by the common biochemical tests. The isolates were categorized into four main phylogenetic groups including group A (28 isolates), group B1 (7 isolates), group B2 (48 isolates), and group D (17 isolates). Two primers (H1 & H2) were used to study the genetic variation of E. coli and the electrophoresis band pattern was analyzed by the NTSYS.Results: The analysis of the difference in isolates using the RAPD technique showed a genetic similarity between 14% and 100%.Conclusion: The phylogenetic groups B2 and A were more frequent in fecal isolates. In addition, the number of isolates related to phylogenetic groups B1 and D was significantly lower than that of the other groups.