Monitoring the Spread of Grapevine Viruses in Vineyards of Contrasting Agronomic Practices: A Metagenomic Investigation

SIMPLE SUMMARY: Grapevine red blotch virus (GRBV) and grapevine Pinot gris virus (GPGV) are two of the most concerning grapevine viruses in Canada. Currently, the secondary spreading of these viruses (spreading via an insect vector) in Canada is poorly documented. To monitor the spreading of these viruses, we deployed virus-free grapevines onto two virus-infected vineyards with different agronomic practices. One vineyard was organic while the other was conventional. Using two independent diagnostic tools, namely the polymerase chain reaction (PCR) and the high-throughput sequencing technique (HTS), we have found that GPGV is spreading in both vineyards. The spread of GPGV was noticeably faster in the organic vineyard than in the conventional one, possibly due to a higher insect population. There was no evidence of the spreading of GRBV. A small collection of plant species in and around the vineyard was also tested by the two methods above. None of the non-grape species harbored any grapevine viruses but a large portion of the wild grape found on the edge of the vineyard carried GPGV. This study is the first direct proof of insect-mediated spreading of GPGV in North America. ABSTRACT: This study investigated the transmission of grapevine viruses, specifically grapevine red blotch virus (GRBV) and grapevine Pinot gris virus (GPGV), in vineyards in Niagara Region, Ontario, Canada. Forty sentinel vines that were confirmed free of GRBV and GPGV by both high-throughput sequencing (HTS) and endpoint polymerase chain reaction (PCR) were introduced to two vineyards (one organic and one conventional) that were heavily infected with both GRBV and GPGV. Four months post-introduction, the sentinel vines were relocated to a phytotron. The HTS results from 15 months post-introduction revealed a widespread infection of GPGV among the sentinel vines but did not detect any GRBV. The GPGV infection rate of sentinel vines in the organic vineyard (13/18) was higher than in the conventional vineyard (1/19). The possibility of an alternative viral reservoir was assessed by testing the most abundant plants in between rows (Medicago sativa, Trifolium repens, Cirsium arvense and Taraxacum officinale), perennial plants in border areas (Fraxinus americana, Ulmus americana, Rhamnus cathartica) and wild grape (unknown Vitis sp.). The HTS result showed that cover crops and perennial plants did not harbor any grapevine viruses, while 4/5 wild grapes tested positive for GPGV but not GRBV. A pairwise sequence identity analysis revealed high similarities between the GPGV isolates found in the established vines on the vineyard and the newly contracted GPGV isolates in the sentinel vines, implicating a recent transmission event. This work provides novel insights into the spread of grapevine viruses in Niagara Region and is also the first direct proof of the spread of GPGV in natural vineyard conditions in North America.