Characterisation of different GLRaV-3 variant infections by determining virus concentration ratios and miRNA expression profiles

Grapevine leafroll disease (GLD) is present in all grape-growing regions of the world and is considered the most significant grapevine viral disease. Grapevine leafroll-associated virus 3 (GLRaV-3) is considered the primary cause of GLD and in South African vineyards five genetic variant groups (I, II, III, VI and VII) have been confirmed. Biological distinctions between GLRaV-3 variants have not been fully validated. By characterising virus concentration and stress-responsive microRNA expression in GLRaV-3 infected plants, this study aimed to glean a better understanding of the possible biological distinctions between GLRaV-3 variants. Quantitative reverse transcription PCR was utilised for virus concentration ratio (VCR) determination and miRNA quantitation in GLRaV-3 positive and negative grapevines grown under greenhouse and field conditions. This study found statistically significant differences in VCRs in plants singly infected with different GLRaV-3 variants. Interestingly, no difference in mean VCRs were observed between data sets, despite notable differences in plant age, duration of GLRaV-3 infection, scion/rootstock combination and growing conditions. Several miRNAs showed statistically significant expression modulation between infected and healthy samples. miRNA expression between data sets varied substantially and a greater overall miRNA response was observed in plants with more established GLRaV-3 infections. The lack of significant differences in mean VCRs between data sets, coupled with the consistent modulation of certain miRNAs in plants that have likely been infected for longer is a promising result. This finding could indicate that successful inhibition of further virus replication by plant defence mechanisms occurred, and that these miRNAs are implicated in this response.