Project Number: 2092-21220-002-052-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: May 1, 2020
End Date: Jun 30, 2023
Determine the effect of biological and genotypical differences in Stubby root nematode/Tobacco rattle virus populations from North Dakota and Washington State on insensitivity in Castle Russet and Payette Russet, using greenhouse and molecular diagnostic assays.
Objective 1: Greenhouse studies: Controlled greenhouse trials will be conducted in North Dakota and Washington State with North Dakota and Washington State stubby root nematodes/Tobacco rattle virus populations collected in 2020. To obtain these populations, soil will be collected from fields with known Tobacco rattle virus-infected stubby root nematodes. Stubby root nematodes will be extracted from the soil and added to soil used to grow tobacco plants to increase population size. Following successful increase, Russet Burbank, Castle Russet, Payette Russet, and tobacco (control) grown in loamy sand soil will be simultaneously infested with 300 stubby root nematodes per plant of either the North Dakota or Washington stubby root nematode, as well as the Washington State greenhouse stubby root nematode population (previously used in this project). Inoculations will occur when plants are ~3 inches tall, and plants will be challenged with the stubby root nematodes for ~2 months, or until plants senesce. At harvest, roots, tubers, stem/leaf tissue and 250cc of soil will be collected from each pot. Stubby root nematodes will be extracted from the soil and counted. Tubers will be cut in half longitudinally to determine internal Tobacco rattle virus incidence and severity. A sample will be taken from a single tuber with corky ringspot symptoms (or arbitrarily if no tuber symptoms are present) from each pot. Tuber samples, roots, and leaf tissue will be tested for Tobacco rattle virus using reverse transcription -polymerase chain reaction. Data gained from the side-by-side evaluations of the North Dakota and Washington State stubby root nematode/tobacco rattle virus populations at each greenhouse trial location will be compared and correlated to any genetic differences identified in stubby root nematode populations (Objective 2). This will help determine if biological differences exist between stubby root nematode populations, or if the stubby root nematode and/or virus directly cause the previously observed difference in insensitive reactions in Castle Russet and Payette Russet cultivars. Objective 2: Molecular analysis: The same stubby root nematode populations collected from Washington and North Dakota states during the 2020 growing season for Objective 1 will be used to complete Objective 2. Specimens will be compared using both microscopy analyses to identify morphological differences and molecular diagnostics of the three specific genomic regions analyzed previously in this project (the D2-D3 region of 28S rRNA gene, 18S rRNA and ITS1 rDNA regions). Additionally, other genes such as the cox1 gene and ITS2 rDNA region will be targeted to help improve the genetic analysis of each nematode population.