Cercospora leaf spot, caused by Cercospora beticola, is one of the most dangerous pathogenic fungi that caused large damage to sugar beet production in Egypt and across the world. The present study is a try to identify some Cercospora isolates collected from six districts in three Egyptian governorates using microsatellite DNA. The results confirmed a wide diversity among the six Cercospora isolates in their disease severity. The highest disease severity was recorded by El-Mansoura isolate, while El-Gemmeiza isolates showed the lowest disease severity among all tested isolates. The differences in the severity of the six isolates may be due to the wide genetic variation among these isolates. According to RAPD analysis a complete similarity between all isolations in OP-C3, OP-C15, and OP-O10 genes while the difference was complete between all isolations in both OP-A9 and OP- D1 genes. OP-A3 and OP-A5 genes in this study were associated with the high disease severity of Cercospora isolates while the absence of these two genes correlated with the low disease severity.With respect to DNA Polymorphic analysis, the results revealed that the polymorphic percentage in all tested isolates ranged between 83.33% to zero with a total polymorphic band reached 49%. The polymorphic bands exceeded 50% in the five primers OP-A3, OP-A5, OP-A9, OP-D1, and OP-O11 as clear evidence for the wide genetic diversity among tested isolates in these genes. The highest polymorphic band was showed in the OP-A5 gene (83.33%). As for Cluster analysis for RAPD, the result showed that the isolates were roughly grouped into two major groups according to their geographic origin. The first group consists of the two isolates collected from El-Mansoura, and Dekernes while the second group consists of the four isolates collected from Sidi-Salem, El-Reyad, and SendsesEl-Gemmeiza. The highest similarity between isolates was shownin the sub-sub group of between Sidi-Salem and El-Reyad isolates. In all cases, the similarity indices between all isolates were larger than 80% except for the two isolates collected from El-Mansoura, and Dekernes with Sendses isolates where the similarity index was less than 80%.