Implementation of a new method to identify Verticillium isolates affecting potato cultivars in Alberta

Potato (Solanum tuberosum L.) is the most important non-cereal crop in the world, with 300 million tons produced every year. Despite its economic importance, potato plants are susceptible to numerous diseases caused by fungi, bacteria, and viruses, that lead to tremendous financial losses for the industry. A common potato disease called Verticillium wilt, or potato early dying, is caused by fungi from the genus Verticillium. The two species that affect potato cultivars in Alberta are V. dahliae and V. albo-atrum. These soil-borne fungi affect xylem tissues preventing water intake causing dehydration in plants. This results in necrosis of the tissues and thus, leads to early plant mortality. Unfortunately, once a crop has developed disease symptoms, there is little that can be done to prevent harvest losses that year. This is why a fast and reliable method for early detection of the fungi is necessary. Traditionally, fungal identification is performed by observing morphological characteristics of mycelia on solid media with a microscope; however, it is time consuming, requires specialized skill set, and species identification can only be achieved by genome sequencing. We are currently implementing a new method developed by Inderbitzin et al. (2013) to identify the species of Verticillium isolates by detecting the presence of species-specific nucleotide sequences1. After extracting fungal DNA, a polymerization chain reaction (PCR) is conducted with species-specific primers for both V. dahlia and V. albo-atrum. By visualizing the presence of an amplified fragment on agarose gel, it is possible to determine the Verticillium species. We are testing and optimizing this method on eight non-identified pure Verticillium cultures isolated from fields in Alberta to determine their species. We are also working on the isolation of new Vertcillium cultures from seven potato plant samples collected during the past growing season across Southern Alberta that showed symptoms of Verticillium wilt. This will allow us to build a collection of Verticillium isolates for future genetic analysis. Overall, this new method will be a very powerful tool to detect the presence of Verticillium fungi in soil and plants before the emergence of any disease symptoms, which allows potato grower to control this disease in a timely manner preventing harvest losses. 1Inderbitzin P, Davis RM, Bostock RM, Subbarao KV (2013) Identification and Differentiation of Verticillium Species and V. longisporum Lineages by Simplex and Multiplex PCR Assays. PLoS ONE 8:e65990 *Indicates presenter

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