Verticillum wilt disease, caused by Verticillium dahliae Kelb, is one of most important olive diseases worldwide. The pathogen is a well known soil borne pathogen causing vascular wilt in more than 200 host plants, in a varity of economically important plants worldwide. The fungus survives for a long time in soil as microsclerotia and in plant material as mycelia. Symptoms are similar in many plant and reduce crop quantity and quality, causes wilt. Microsclerotia provide pathoge survival in the absence of host for 15 years. Several factors might account for the increasing spread of the disease, including pathogen population densities in soil, highly virulent pathogen, host susceptibility and environmental conditions. Integrated disease management system (IDM) increases plant resistance against pathogen, but it's not disease complete cure and remove from the tree, thus prevention of disease through the use of healthy plants and new planting in non-infested soils is the best method of control. The purpose of this investigation was to achieve an accurate and sensitive method in shortest possible time for pathogen detection from infested soils and plant tissuses. In this study, sixteen isolates of V. dahliae were obtained from soil and roots of olive and apricot trees, after isolation, purification and confirmation using species specific primers, various methods of detection viz Nested PCR method, trapping plant(using twig pepper) and semi-selective medium were used to determine best pathogen detection method from soil and root. Trap plant method isn't sensitive and accurate and also time, is time consuming. Varkup dish method is able to detect V. dahliae in soil but inlong term isn't appropriate. but nested PCR technique was evaluated better than other methods for pathogen detection because of high sensitivity and accuracy in the shortest possible time. Research of the deal initial inoculums and the required time pathogen detection showed that soil temperature has major role in pathogen detection. Loamy soils contain 2% organic matter in the absence of host , increased the initial inoculums and reduced required time pathogen detection, although soil temperature had fluctuatation but fungus maintained viable in along time. DNA concentration in contaminated soil is detectable more dilution 1.2 mg/ng. In order to verify the accuracy of pathogen detection V. dahliae from soil and root,Nested PCR product was sequenced. Comparison of the sequenced showed highly similarity to recognized isolate of V. dahlia e. According to results, it seems that Nested PCR is an efficient method for detection of V. dahliae from soil.
