DETECTION OF TRICHOTHECENS CHEMOTYPE PRODUCED BY FUSARIUM GRAMINEARUM ISOLATED FROM GRAINS USING SPECIFIC PCR ASSAYS

Abstract

The present study aims at using several techniques in detecting some isolates of the Fusarium fungi and testing their ability to produce tricothecencs toxin and detection of the genes responsible for production of trichothecenes (Nivalenol) in Fusarium graminearum by using the PCR and Real time PCR. The eight isolates of Fusarium spp. are isolated from clinical and environmental sources. The clinical isolates are isolated from a cows lung, while the environmental isolates are isolated from dry grains of wheat and barley. These fungi are cultured on Sabouraud Dextrose agar (SDA). The isolates are identified depending on their morphological characteristics (cultural and microscopical). Concerning the molecular part of the study, genomic DNA in extracted from all Fusarium graminearum isolates. These genomic DNA samples are found to have suitable concentration and purity for Real time PCR technique. The genes sites are amplified (Tri4, Tri6, Tri10) in the targeted DNA by using Real time PCR. One the first hand, the results of macroscopic and microscopic examination of the Fusarium graminearum showed that colonies had a cottony appearance at the beginning of growth and then whitely turned into irradiated which was floral and had the ability to form abundant conidial structures with a large banana shape in microscopic examination . On the other hand, the average yield of DNA was in the range of (96.9-181.2) ng/μl with purity of (1.6-1.8). Three structural genes have been observed in the present study; Tri4, Tri6 and Tri10. These genes are involved in the biosynthesis of trichothecens (Nivalenol), the regulating gene that plays a major role in the production of trichothecens.