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. 2019 Jun 29;11(7):592.
doi: 10.3390/v11070592.

High-Throughput Sequencing Assists Studies in Genomic Variability and Epidemiology of Little Cherry Virus 1 and 2 infecting Prunus spp. in Belgium

Rachid Tahzima  1   2 , Yoika Foucart  1 , Gertie Peusens  3 , Tim Beliën  3 , Sébastien Massart  2 , Kris De Jonghe  4
Affiliations

High-Throughput Sequencing Assists Studies in Genomic Variability and Epidemiology of Little Cherry Virus 1 and 2 infecting Prunus spp. in Belgium

Rachid Tahzima et al. Viruses. .

Abstract

Little cherry disease, caused by little cherry virus 1 (LChV-1) and little cherry virus 2 (LChV-2), which are both members of the family Closteroviridae, severely affects sweet (Prunus avium L.) and sour cherry (P. cerasus L.) orchards lifelong production worldwide. An intensive survey was conducted across different geographic regions of Belgium to study the disease presence on these perennial woody plants and related species. Symptomatic as well as non-symptomatic Prunus spp. trees tested positive via RT-PCR for LChV-1 and -2 in single or mixed infections, with a slightly higher incidence for LChV-1. Both viruses were widespread and highly prevalent in nearly all Prunus production areas as well as in private gardens and urban lane trees. The genetic diversity of Belgian LChV-1 and -2 isolates was assessed by Sanger sequencing of partial genomic regions. A total RNA High-Throughput Sequencing (HTS) approach confirmed the presence of both viruses, and revealed the occurrence of other Prunus-associated viruses, namely cherry virus A (CVA), prune dwarf virus (PDV) and prunus virus F (PrVF). The phylogenetic inference from full-length genomes revealed well-defined evolutionary phylogroups with high genetic variability and diversity for LChV-1 and LChV-2 Belgian isolates, yet with little or no correlation with planting area or cultivated varieties. The global diversity and the prevalence in horticultural areas of LChV-1 and -2 variants, in association with other recently described fruit tree viruses, are of particular concern. Future epidemiological implications as well as new investigation avenues are exhaustively discussed.

Keywords: Ampelovirus; Closteroviridae; HTS; RNA virus phylogeny; Velarivirus; fruit trees and plant viruses; genetic variability; molecular evolution; total RNA.

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Conflict of interest statement

The authors declare no conflict of interest. The founding sponsors had no direct role in the design of the study; in the collection, analyses, or interpretation of the data; in the writing of the manuscript, and in the decision to publish the results.

Figures

Figure 1
Figure 1
Maximum likelihood phylogenetic tree inferred from partial RdRp (left) and CP (right) nucleotide sequences of the LChV-1 Belgian (purple dot) and other LChV-1 isolates. The GenBank accession numbers of the sequences are indicated together with the isolate name, host plant and cultivar. MEGA 7.0 analysis included most of the available LChV-1 sequences. The phylogenetic clusters are delineated with vertical bars. Branch lengths on the phylogenetic tree represent the genetic distance the numbers at the branches represent the percentage of replicates in which the topology of the branch was observed after 1000 bootstrap replicates.
Figure 2
Figure 2
Maximum likelihood phylogenetic tree inferred from partial RdRp (left) and CP (right) nucleotide sequences of the LChV-2 Belgian (purple dot) and other LChV-2 isolates. The GenBank accession numbers of the sequences are indicated together with the isolate name, host plant and cultivar. MEGA 7.0 analysis included most of the available LChV-2 sequences. The phylogenetic clusters are delineated with vertical bars. Branch lengths on the phylogenetic tree represent the genetic distance the numbers at the branches represent the percentage of replicates in which the topology of the branch was observed after 1000 bootstrap replicates.
Figure 3
Figure 3
Maximum likelihood phylogenetic trees inferred from the full genome nucleotide sequences of LChV-1 (left) and LChV-2 (right) isolates. All isolates from Genbank are reported with their accession numbers followed by their names. Belgian isolates are indicated with a purple dot. The numbers above or below each branch are the nonparametric bootstrap (NPB) values given as percentages of 1000 replicates.
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