1a.Objectives (from AD-416)
1. Sequence whole genomes for at least 3 PVY-N:O strains that do not cause tobacco vein necrosis, and analyze their sequences against at least 3 other PVY-N:O strains that do.
2. Sequence whole genomes for at least 3 PVY isolates that cause tuber necrosis in potato and analyze their sequences against at least 3 other PVY strains that do not.
3. Express PVYO and O5 capsid proteins in bacteria and correlate the monoclonal 1F5 reactivity with the point mutation in the PVY-O capsid protein.
1b.Approach (from AD-416)
Potato virus Y isolates collected by State Potato Seed Certification agencies and partially characterized by either State laboratories or the ARS laboratory at Ithaca, NY will be sent to the University of Idaho. Virus genomes will be amplified using a RT-PCR protocol and the products sequenced. Sequence information will be catalogued in a bioinformatics database that will be made available to all collaborators on the project. Virus protein fragments will be expressed in bacteria and used to characterize epitopes that recognize monoclonal antibodies.
Potato virus Y (PVY) strains were originally defined by interactions with different resistance genes in standard potato cultivars. In the most recent classification, five distinct strain groups are defined that cause a local and/or systemic hypersensitive response in the genetic background with a corresponding resistance (N) gene. The five strains were PVYO, PVYN, PVYC, PVYZ and PVYE. The nucleotide sequences of multiple isolates of PVYO and PVYN are known and differ from each other by about 8% along their genomes. Additionally, complete genome sequences of multiple recombinant isolates have been found to be composed of segments of parental PVYO and PVYN sequences. PVYC, PVYZ and PVYE are not known to occur in the US. We discovered a PVY isolate that is defined as belonging to the PVYZ strain, a group that has never been fully characterized. The complete genome sequence was determined and although the genome structure and sequence displays typical features of the recombinant PVYNTN strain known to occur in the US. PVYZ would not be recognized as a distinct strain using the current diagnostic tests employed by regulatory and seed certification agencies. Our complete characterization of PVYZ can now be used as a worldwide standard in the new classification of this group of viruses and we can develop modified diagnostic assays that can distinguish this new strain. Progress on the project is monitored by frequent conference calls and emails in addition to meetings at scientific conferences.