Species-specific primers derived from comparative genomics for PCR detection of Verticillium species

Verticillium is a small genus of ascomycete fungi containing ten species. Several of these species are destructive phytopathogens that cause vascular diseases on diverse crops including ornamentals in different regions of the world. Traditional methods of differentiating the species based on morphological characteristics can lead to misidentification. Although rapid PCR detection assays are available, there remains a lack of universal and accurate methods for the identification of the ten species. The aim of this research was to develop novel species-specific primers for the molecular detection of ten Verticillium species. The primers were designed based on screening of species-specific genomic regions. Among the ten primer sets tested, the detection limits ranged from approximately 0.1 ng to 1 ng of Verticillium DNA. The primer sets were further assessed on a collection of Verticillium strains from ten species. Additionally, DNA extracted from two plant species or soil samples infested with eight different species was employed to further validate the utility of these primers. These species-specific primers facilitate the rapid identification of Verticillium species.