Project number:
GenUS GP 20-01
Project leader:
Pietersen, G
Institution:
Department of Genetics, Stellenbosch University
Start date:
2020
EXECUTIVE SUMMARY
Objectives and Rationale
Certification schemes prevent the spread of pathogens via planting material. In the Vine Improvement Association (VIA) Vitis certification scheme of South Africa, only a few viruses are tested for and certified free of, while more than 80+ viruses occur on grapevine. In this project, we determine the total virus and viroid status of nuclear grapevine material in 97 accessions from three Plant Improvement Organisations.
Methods
Nuclear plant accessions from three plant improvement organisations were selected for analysis. Total nucleic acid from these accessions was extracted at multiple time points and from various tissues and pooled. The pooled nucleic acid was subjected to next-generation sequencing, and the presence of viruses and viroids was determined using bioinformatic techniques. The viruses or viroids found were confirmed by specific PCR tests.
Key Results
Only seventeen of the 97 nuclear accessions tested had no detectable viruses or viroids. Amongst the remaining nuclear accessions, five viruses and four viroids or viroid-like entities were identified. The virus infections were rare, being found in only 8 accessions, while viroid infections were common, being found in 76 of the 97 accessions. Viruses observed included Rupestris stem pitting-associated virus (GRSPaV), Grapevine fleck virus (GFkV), Grapevine syrah virus 1 (GSyV-1), Grapevine leafroll-associated virus 4 (GLRaV-4) and Grapevine Rupestris vein feathering virus (GRVFV). Viroids found were hop stunt viroid (HSVd), grapevine yellow speckle viroid-1 (GYSVd-1), grapevine yellow speckle viroid-2 (GYSVd-2), Australian grapevine viroid (AGHVd) and Grapevine hammerhead viroid-like RNA in various combinations.
Conclusion and Discussion / Recommendation
South African vine nuclear material appears remarkably free of grapevine viruses despite only a few of the more than 80 known viruses being tested for locally. This is probably due to the mandatory application of heat therapy and meristem tip culture to create nuclear plants. All remaining and future nuclear plants should be subjected to NGS analysis to confirm their virus/viroid status. Those found to be virus-infected need to be subjected to secondary virus elimination processes. The common presence of viroids suggests that these agents are not eliminated by current elimination processes or spread exceptionally rapidly within the nuclear block facilities. A risk assessment as to the importance of these agents must be made prior to deciding to eliminate these pathogens from nuclear material, and whether to manage these agents.