Alfalfa and clover are two important forage crops of the family Fabaceae. More over they have useful effect on soil structure and have a fundamental role in feeding domesticated animals. As a consequence, they influence on meat and milk production, which influences on human nutrition. The powdery mildow of alfalfa and clover is one of the diseases in some regions of Iran . In 1385, twenty isolates of powdery mildews were collected from Isfahan province. Furthermore, one isolate of clover and one of alfalfa powdery mildow from Zanjan were used. Sixteen isolates were recognized as Oidium ., as they have no mature ascocarp or even without it. Two isolates recognized as Erysiphe pisi and three others recognized as E . trifolii var. trifolii . Isolate of Baghbahadoran's clover 1 was identified as E . trifolii var. desmanthi which is reported for first time from Iran . Five weeds were collected from inside of alfalfa and clover fields, to test the ability of their infectivity on alfalfa and clover, cross inoculation was done in greenhouse. Only isolates of clover and alfalfa could infect clover and alfalfa and no infection from weed sites to clover and alfalfa was observed. Considering these results powdery mildews of other collected hosts do not have any role in infection of alfalfa and clover plants. Six methods of DNA extraction have been tested and crushing of mycelia between two glass slides gave sufficient amount of DNA. ITS1-5.8s-ITS2 regions of rDNA was amplified using primer pairs of ITS5 and P3 and in nested PCR assay ITS1 and P3 primer pairs was used. A 522 bp fragment was amplified in E . pisi and 558 bp fragment was amplified in E . trifilii. PCR-RFLP profiles were obtained using Taq I, Alu I, Msp I, Hin fI restriction enzymes. The profiles from Alu I confirmed that different isolates are identical and the cluster analysis based on three other enzymes showed variability among different isolates. There was 75 percent similarity among different isolates. PCR-RFLP results was concordant with geographical distribution of isolates and could separated the two species E. pisi and E . trifolii .
