Development of a duplex PCR test system for differential diagnostics of wheat septoriosis

Abstract

Currently, the Septoria leaf spot is one of the most harmful and economically significant wheat diseases in the world, especially in countries with a temperate climate. Losses from the disease in epiphytotic years can reach 20-40%. The Septoria leaf spot is one of the dominant positions among fungal diseases of grain crops in Kazakhstan. Due to the unfavorable situation in the country for this type of disease, in order to timely identify phytopathogens, a duplex test system based on the polymerase chain reaction (PCR) method has been developed. This paper presents the results of studies obtained in the development of PCR test system for differential diagnosis of the Septoria leaf spot. Using the multiple nucleotide alignment method, the most conserved regions of the fungal genes Parastagonospora nodorum and Zymoseptoria tritici were determined, based on which species-specific primer pairs were constructed. The PCR parameters were optimized. The developed test system was tested for specificity and sensitivity using DNA of the fungi Zymoseptoria tritici, Parastagonospora nodorum, Pyrenophora tritici-repentis, Alternaria alternata, Alternaria infectoria, Fusarium, Ascomycota, Epicoccum. Laboratory test results showed that the test system works specifically, and the sensitivity is 5 picograms of DNA in one reaction. Key words: Parastagonospora nodorum, Zymoseptoria tritici, PCR, primers, DNA.