A multiple single nucleotide polymorphisms interrogation assay for reliable Potato virus Y group and variant characterization

The complex Potato virus Y classification, including groups (PVYN and PVYO) and variants (PVYNTN and PVYN-W), is based mainly on biological properties of isolates. Published PVY detection tools targeting markers not associated with biological properties could fail to assign correctly isolates in the current classification. To improve PVY detection tools, a single nucleotide polymorphism (SNaPshot) detection assay was developed. The technique was adapted to target the T/C9259, A/C2271, G/C8573 and A/G2213 PVY polymorphic nucleotides. The “TAGA”, “CCCG”, “CACA” and “CAGA” four-digit codes associated with tested samples allowed identification of PVYN, PVYO, PVYN-W and PVYNTN isolates, respectively. The PVY SNaPshot procedure is efficient and reliable for PVY detection and characterization in samples containing as few as 102 viral RNA copies. Moreover, PVY group assignment is possible for fractions containing only 10 copies of a PVY RNA genome. Finally, the SNaPshot assay allows PVYN/PVYO dual characterization for mixed samples containing PVYN/PVYO quantity ratios in the range of 0.1–10. This innovative SNaPshot tool improved clearly PVY diagnostic assays described previously by targeting simultaneously major functional markers and sequence unlinked to biological properties used separately in PVY detection tools available currently.