A Multiplex PCR Assay to Characterize Potato virus Y Isolates and Identify Strain Mixtures

Abstract

Potato virus Y (PVY) has become a serious problem for the seed potato industry, with increased incidence and rejection of seed lots submitted for certification. New PVY strains and strain variants have emerged in recent decades in Europe and North America, including the PVYN strain that causes veinal necrosis in tobacco, and strain variants that represent one or three recombination events between the common strain (PVYO) and PVYN. Several reverse transcription-polymerase chain reaction (RT-PCR) assays have been described that characterize PVY isolates as to strain type, but they are limited in their ability to detect some combinations of mixed strain infections. We report here the development of a single multiplex RT-PCR assay that can assign PVY strain type and detect mixed infections with respect to the major strain types. Validation of this assay was achieved using 119 archived PVY isolates, which had been previously characterized by serology and bioassay and/or previously published RT-PCR assays. Results for single-strain isolates were comparable to previous results in most cases. Interestingly, 16 mixed infections were distinguished that had previously gone undetected. The new multiplex RT-PCR assay will be useful for researchers and seed production specialists interested in determining PVY infection type using a single assay.

Complete nucleotide sequence of a potato isolate of strain group C of Potato virus Y from 1938

“…PVY is one of the most common viruses of potato (Solanum tuberosum L.) and causes significant economic losses worldwide, especially in the production of seed potatoes. [12,16,24,25].PVY O and PVY C can be distinguished in potato cultivars bearing the resistance genes Ny tbr and Nc [7,25]. PVY C isolates are defined by the typical hypersensitive response they induce in potato cultivars carrying the Nc gene.…”

“…PVY is one of the most common viruses of potato (Solanum tuberosum L.) and causes significant economic losses worldwide, especially in the production of seed potatoes. [12,16,24,25].…”

“…More recently, new PVY strains have been found worldwide that appear to be genetic recombinants bearing genetic material of the PVY O and PVY N strains. Examples of such strains are PVY NTN and PVY N-Wi [12,16,24,25].…”

“…Many RT-PCR tests have been described for PVY detection, including tests for detection of recombinant strains [16,21,24], but none of them can distinguish PVY C isolates easily.…”

Epidemiología y variabilidad patogénica del virus del mosaico del pepino dulce (Pepino mosaic virus). Nuevas enfermedades asociadas a su presencia (torrao o cribado)

“…This diagnostic method allows the simultaneous detection and identification of different viruses with less time and cost waste. Furthermore, a multiplex RT-PCR assay has been developed to identify different strain types of a single virus (Lorenzen et al, 2006;Ratti et al, 2005;Rigotti and Gugerli, 2007) and even different species of a virus (Martínez-Culebras et al, 2001). In the present work we have developed a one-step multiplex RT-PCR reaction plus a restriction analysis that permits the simultaneous detection and identification of five different PepMV isolates (here after RT-PCR-SacI).…”