High throughput sequencing-aided molecular characterization of resistance-breaking variants of Beet necrotic yellow vein virus in the United States.
Rhizomania, caused by Beet necrotic yellow vein virus (BNYVV), is an economically important disease of sugar beet industry globally. The disease is primarily managed by genetic resistance conferred by Rz1 and Rz2 genes. However, due to consistent disease pressure, the resistance has been overcome by resistance-breaking (RB) isolates worldwide. To characterize the molecular changes that occurred in the BNYVV genome that confer resistance breaking, we collected rhizomania-infested soil samples from sugarbeet production fields of California, Idaho, Minnesota, and North Dakota that were suspicious for RB ability. To recover the virus from the soil samples, soil-baiting was performed by growing sugar beet varieties possessing no known resistance to rhizomania (susceptible), with Rz1 alone, and with a combination of Rz1+Rz2 resistance genes in the test soils. Enzyme-linked immunosorbent assay (ELISA) was used to confirm the presence of BNYVV in the roots of the bait plants from susceptible, Rz1, and Rz1+Rz2 varieties. To gain insight at the nucleotide sequence level, high throughput sequencing (HTS) was applied to the total RNA isolated from the roots of Rz1+Rz2 bait plants to characterize the molecular changes associated with potential RB-strains. Among the various components of BNYVV genome, detailed analysis of the deduced amino acid sequence of P25 gene, which was previously implicated for RB pathotype, showed mutations at positions 67 and 68 on the hypervariable ‘tetrad’ amino acids (67-70). In addition, we observed scattered mutations in other area of the P25 gene. This study enhances our current understanding of the molecular changes that are associated with the developing RB population of BNYVV and underscoring the importance of alternative disease management strategies for the future.