Abstract
BackgroundGrapevine leafroll disease is one of the most economically important viral diseases affecting grape production worldwide. Grapevine leafroll-associated virus 4 (GLRaV-4, genus Ampelovirus, family Closteroviridae) is one of the six GLRaV species documented in grapevines (Vitis spp.). GLRaV-4 is made up of several distinct strains that were previously considered as putative species. Currently known strains of GLRaV-4 stand apart from other GLRaV species in lacking the minor coat protein.MethodsIn this study, the complete genome sequence of three strains of GLRaV-4 from Washington State vineyards was determined using a combination of high-throughput sequencing, Sanger sequencing and RACE. The genome sequence of these three strains was compared with corresponding sequences of GLRaV-4 strains reported from other grapevine-growing regions. Phylogenetic analysis and SimPlot and Recombination Detection Program (RDP) were used to identify putative recombination events among GLRaV-4 strains.ResultsThe genome size of GLRaV-4 strain 4 (isolate WAMR-4), strain 5 (isolate WASB-5) and strain 9 (isolate WALA-9) from Washington State vineyards was determined to be 13,824 nucleotides (nt), 13,820 nt, and 13,850 nt, respectively. Multiple sequence alignments showed that a 11-nt sequence (5′-GTAATCTTTTG-3′) towards 5′ terminus of the 5′ non-translated region (NTR) and a 10-nt sequence (5′-ATCCAGGACC-3′) towards 3′ end of the 3′ NTR are conserved among the currently known GLRaV-4 strains. LR-106 isolate of strain 4 and Estellat isolate of strain 6 were identified as recombinants due to putative recombination events involving divergent sequences in the ORF1a from strain 5 and strain Pr.ConclusionGenome-wide analyses showed for the first time that recombinantion can occur between distinct strains of GLRaV-4 resulting in the emergence of genetically stable and biologically successful chimeric viruses. Although the origin of recombinant strains of GLRaV-4 remains elusive, intra-species recombination could be playing an important role in shaping genetic diversity and evolution of the virus and modulating the biology and epidemiology of GLRaV-4 strains.
Highlights
Grapevine leafroll disease is one of the most economically important viral diseases affecting grape production worldwide
Internal gaps in viral genome sequence were filled by reverse transcription-polymerase chain reaction (RTPCR) using species-specific primers designed based on high-throughput sequencing (HTS) data and reference sequence corresponding to strain 4, 5, and 9 obtained from GenBank
Genome sequence analysis of three strains of Grapevine leafroll-associated virus (GLRaV)-4 from Washington vineyards After quality trimming, Illumina sequencing generated 29,859,206 paired-end 125 base-length reads from cv
Summary
Grapevine leafroll disease is one of the most economically important viral diseases affecting grape production worldwide. Grapevine leafroll-associated virus 4 (GLRaV-4, genus Ampelovirus, family Closteroviridae) is one of the six GLRaV species documented in grapevines (Vitis spp.). GLRaV-4 is made up of several distinct strains that were previously considered as putative species. Known strains of GLRaV-4 stand apart from other GLRaV species in lacking the minor coat protein. Grapevine leafroll-associated viruses (GLRaVs, family Closteroviridae) represent a group of highly complex and genetically distinct viruses infecting an agriculturally important perennial fruit crop [1]. The genus Ampelovirus contains higher number of GLRaVs compared to other genera in the family Closteroviridae. All GLRaVs are predominantly disseminated via plant propagation material, grapevine-infecting ampeloviruses are known to be transmitted by mealybugs (Pseudococcidae) and scale insects (Coccidae) in a semi-persistent manner [3]. GLRaV-7 was shown to be transmitted by the plant parasitic dodder, Cuscuta reflexa [4]
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