High-Throughput Sequencing Identified Distinct Bipartite and Monopartite Begomovirus Variants Associated with DNA-Satellites from Tomato and Muskmelon Plants in Saudi Arabia

Abstract

The studies on the prevalence and genetic diversity of begomoviruses in Saudi Arabia are minimal. In this study, field-grown symptomatic tomato and muskmelon plants were collected, and initially, begomovirus infection was confirmed by the core coat protein sequences. Four tomato and two muskmelon plants with viral infections were further evaluated for Illumina MiSeq sequencing, and twelve sequences (2.7-2.8 kb) equivalent to the full-length DNA-A or DNA-B components of begomoviruses were obtained along with eight sequences (~1.3-1.4 kb) equivalent to the begomovirus-associated DNA-satellite components. Four begomovirus sequences obtained from tomato plants were variants of tomato yellow leaf curl virus (TYLCV) with nt sequence identities of 95.3-100%. Additionally, two tomato plants showed a mixed infection of TYLCV and cotton leaf curl Gezira virus (CLCuGeV), okra yellow crinkle Cameroon alphasatellite (OYCrCMA), and okra leaf curl Oman betasatellite (OLCuOMB). Meanwhile, from muskmelon plants, two sequences were closely related (99-99.6%) to the tomato leaf curl Palampur virus (ToLCPalV) DNA-A, whereas two other sequences showed 97.9-100% sequence identities to DNA-B of ToLCPalV, respectively. Complete genome sequences of CLCuGeV and associated DNA-satellites were also obtained from these muskmelon plants. The nt sequence identities of the CLCuGeV, OYCrCMA, and OLCuOMB isolates obtained were 98.3-100%, 99.5-100%, and 95.6-99.7% with their respective available variants. The recombination was only detected in TYLCV and OLCuOMB isolates. To our knowledge, this is the first identification of a mixed infection of bipartite and monopartite begomoviruses associated with DNA-satellites from tomato and muskmelon in Saudi Arabia. The begomovirus variants reported in this study were clustered with Iranian isolates of respective begomovirus components in the phylogenetic dendrogram. Thus, the Iranian agroecological route can be a possible introduction of these begomoviruses and/or their associated DNA-satellites into Saudi Arabia.

Keywords: DNA-satellites; Illumina MiSeq; Saudi Arabia; begomovirus; mixed infection; muskmelon; tomato.

Figure 3

Maximum likelihood phylogenic dendrograms showing the evolutionary relatedness of (A) full-length sequences of identified okra yellow crinkle Cameroon alphasatellite (OYCrCMA) and (B) okra leaf curl Oman betasatellite (OLCuOMB). In each case, the full-length sequences of the most closely related DNA-satellite sequences were retrieved from the NCBI GenBank database for comparison with the identified respective DNA-satellite. The host plant species, geographical origins, and database accession numbers of each isolate used in this tree have been provided. The DNA-satellite isolates identified in this study are written in bold text and indicated with a red star. The trees were constructed with percentage bootstrap values set at 1000 replicates. The guidelines suggested by Briddon et al. [6] and Briddon et al. [31] were followed for taxonomic abbreviations of alphasatellite and betasatellites. Abbreviations used for alphasatellites were; Ageratum yellow vein Singapore alphasatellite (AYVSGA), bhendi yellow vein alphasatellite (BYVA), Cassava mosaic Madagascar alphasatellite (CMMGA), cotton leaf curl Egypt alphasatellite (CLCuEGA), cotton leaf curl Gezira alphasatellite (CLCuGeA), cotton leaf curl Multan alphasatellite (CLCuMuA), cotton leaf curl Saudi Arabia alphasatellite (CLCuSAA), Gossypium mustelinum symptomless alphasatellite (GMusSLA), Hollyhock yellow vein alphasatellite (HoYVA), Malvastrum yellow mosaic Cameroon alphasatellite (MaYMCMA), okra enation leaf curl alphasatellite (OEnLCuA), okra yellow crinkle Cameroon alphasatellite (OYCrCMA), and tomato yellow spot alphasatellite (ToYSA). Abbreviations used for betasatellite species were chili leaf curl betasatellite (ChLCuB), cotton leaf curl Gezira betasatellite (CLCuGeB), okra leaf curl Oman betasatellite (OLCuOMB), papaya leaf curl betasatellite (PaLCB), and tomato leaf curl betasatellite (ToLCB).

Figure 1

Virus infection symptoms on plants from Al-Ahsa region, Saudi Arabia; (A) symptomatic tomato plants showing leaf curling and yellowing; and (B) muskmelon plants showing leaf yellowing and mosaic symptoms.

Figure 2

Maximum likelihood phylogenic dendrograms showing the evolutionary relatedness of (A) full-length genomes of identified cotton leaf curl Gezira virus (CLCuGeV), tomato yellow leaf curl virus (TYLCV), tomato leaf curl Palampur virus (ToLCPalV) DNA-A, and (B) ToLCPalV DNA-B. In each case, the full-length sequences of the most closely related begomvovirus genomes were retrieved from the NCBI GenBank database for comparison with the identified respective begomovirus genomes. The host plant species, geographical origins, and database accession numbers of each isolate used in this tree have been included. The begomovirus isolates identified in this study are written in bold text and indicated with a red star. The trees were constructed with percentage bootstrap values set at 1000 replicates. For taxonomic abbreviations of begomovirus species, the guidelines by Zerbini et al. [1] were followed. The ToLCPalV isolates reported from the Indo–Pak subcontinent were merged. Abbreviations used for begomovirus species were cotton leaf curl Gezira virus (CLCuGeV), tomato yellow leaf curl virus (TYLCV), tomato leaf curl Palampur virus (ToLCPalV), and African cassava mosaic virus (ACMV).

Figure 4

Detection of recombination events using Recombination Detection Program (RDP5.0). (A) Recombination analysis of the potential recombinant begomovirus and (B) betasatellite isolates. The graphical representation of begomovirus and betasatellite genomes is followed by GARD and RDP analyses, respectively.