Complete sequence of the Pepino mosaic virus RNA genome

Abstract

Analysis of an RNA-seq strand-specific library revealed a complete ge-nome of Hardenbergia mosaic virus (HarMV) from RNA extracted from a native wiste-ria (Hardenbergia comptoniana) plant from southwest Australia. We compared it with eight other complete HarMV genomes. It most resembled (85.8% nucleotide identity) the genome of HarMV isolate MD4-D. I n July 2016, an apical leaf sample showing obvious mosaic and deformation symptoms was collected from a plant of the indigenous southwest Australian species native wisteria (Hardenbergia comptoniana; family Fabaceae). The plant sampled also exhibited symptoms typical of phytoplasma infection (leaf chlorosis, proliferation of axillary shoots, and stunting). It was growing on a fence line bordering a playing field area in the Victoria Park suburb of Perth in southwest Australia. Hardenbergia mosaic virus (HarMV; genus Potyvirus, family Potyviridae) causes a conspicuous disease in native wisteria plants, is spread nonpersistently by aphids, and, like native wisteria itself, is indigenous to the region (1-5). It invades introduced lupin species (Lupinus spp.) at the interface of the local ancient ecosystem and recent agroecosystem (6, 7). Analysis of polyadenylated transcripts derived from RNAseq-stranded libraries (8-17) prepared from RNA extracted from the collected sample (designated VPK) detected one complete HarMV genome. RNA was extracted from the VPK sample using a ZR Plant RNA MiniPrepTM kit (Zymo Research) and treated with RNase-free DNase (Invitrogen). The extract was subjected to library preparation using a TruSeq-stranded Ribo-Zero plant kit (Illumina, catalog no. RS-122-2401) and was subsequently subjected to quality control (8-17). The library was sent to Macrogen, Inc. (South Korea), where sequencing was done using the HiSeq 2500 platform with a TruSeq SBS version 4 kit (Illumina) with 151 cycles of paired-end reads. Reads were then assembled and genomes annotated using CLC Genomics Workbench version 6.5 (CLC bio) and Geneious version 8.1.7 (Biomatters) (18, 19). The VPK sample yielded 13,609,056 reads and, after trimming, 12,972,958 remained. De novo assembly generated 25 contigs and 622,179 reads mapped to the contig of interest with a coverage of 9,107. The complete genome obtained was named VPK-1. It consisted of 9,621 nucleotides (nt) and coded for 10 proteins, which is similar to other potyviruses (20, 21). There were eight other complete HarMV genomes already in GenBank (3, 7). A BLAST-based search (22) revealed that sequence VPK-1 most resembled the sequence of HarMV isolate MD4-D (KJ152157) with an 85.8% nt identity. In addition, the analysis revealed a partial sequence (6,002 nt in length) named VPK-2,

Differential Tomato Transcriptomic Responses Induced by Pepino Mosaic Virus Isolates with Differential Aggressiveness

“…Damage and economic losses caused by PepMV vary greatly and can at least partially be attributed to the differential symptomatology caused by different PepMV isolates (Hanssen et al, 2009(Hanssen et al, , 2010a. Currently, four PepMV genotypes are distinguished: the Peruvian genotype (LP), which was first isolated from Solanum peruvianum and is similar to the original pepino (S. muricatum) isolate (Ló pez et al, 2005;Pagán et al, 2006); the European tomato genotype (EU), which was first reported in greenhouse tomato production in Europe (Mumford and Metcalfe, 2001;Aguilar et al, 2002;Cotillon et al, 2002;Verhoeven et al, 2003;Pagán et al, 2006); the CH2 genotype, which was first isolated from tomato seeds from Chile (Ling, 2007); and the US1 genotype, which was first described in the United States (Maroon-Lango et al, 2005). In recent years, the CH2 genotype has largely replaced the EU genotype in commercial tomato production in several European countries (Hanssen et al, 2008;Gó mez et al, 2009).…”

Epidemiología y variabilidad patogénica del virus del mosaico del pepino dulce (Pepino mosaic virus). Nuevas enfermedades asociadas a su presencia (torrao o cribado)

“…En el gen del TGBp1 se observa un (región intergénica entre el TGB y la ORF5), se encuentra la última región del genoma del virus, que codifica para la proteína de la cápside (CP) de 25 kDa, y en la cual se encuentra la secuencia conservada de naturaleza antifática 5´KFAAFDFFDGVT 3´, responsable posiblemente de la unión del RNA viral a la CP mediante interacciones hidrofóbicas. En su extremo 3´, tras una serie de nucleótidos que no se traducen (3´UTR), se encuentra la cola poli-A (Aguilar et al, 2002;Cotillon et al, 2002;Maroon-Lango et al, 2005;Ling, 2007;Hasiów et al, 2007).…”

“…(Jordá et al, 2001b;Córdoba et al, 2004a) y una planta aromática (Ocimum basilicum L.) procedente de invernaderos de Silicia (Davino et al, 2009 Asimismo, en diversos estudios efectuados se han determinado experimentalmente una serie de especies hospedantes artificiales del virus, todas pertenecientes a la familia Solanaceae (Tabla 1.5), así como una serie de plantas habitualmente empleadas como plantas indicadoras que han resultado siempre no hospedantes (Tabla 1.6). Cabe destacar que existen dos especies, Chenopodium murale L. (Jones et al, 1980;Aguilar et al, 2002) y O. basilicum (Salomone y Roggero, 2002;Davino et al, 2009) Jordá et al, 2001a;Salomone y Rogero, 2002;Verhoeven et al, 2003 Nicotiana affinis T.Moore Solanaceae Pospieszny et al, 2008 N. benthamiana Domin. Solanaceae Jordá et al, 2001a;Aguilar et al, 2002;Salomone y Roggero, 2002;Davino et al, 2008;Pospieszny et al, 2008 N. bigelovii (Torr.)…”

“…Cabe destacar que existen dos especies, Chenopodium murale L. (Jones et al, 1980;Aguilar et al, 2002) y O. basilicum (Salomone y Roggero, 2002;Davino et al, 2009) Jordá et al, 2001a;Salomone y Rogero, 2002;Verhoeven et al, 2003 Nicotiana affinis T.Moore Solanaceae Pospieszny et al, 2008 N. benthamiana Domin. Solanaceae Jordá et al, 2001a;Aguilar et al, 2002;Salomone y Roggero, 2002;Davino et al, 2008;Pospieszny et al, 2008 N. bigelovii (Torr.) S. Wats Solanaceae Jones et al, 1980; N. clevelandii Gray Solanaceae Jordá et al, 2001a;Aguilar et al, 2002;Salomone y Roggero, 2002;Pospieszny et al, 2008 N. debneyi Domin.…”

“…Solanaceae Jordá et al, 2001a;Aguilar et al, 2002;Salomone y Roggero, 2002;Davino et al, 2008;Pospieszny et al, 2008 N. bigelovii (Torr.) S. Wats Solanaceae Jones et al, 1980; N. clevelandii Gray Solanaceae Jordá et al, 2001a;Aguilar et al, 2002;Salomone y Roggero, 2002;Pospieszny et al, 2008 N. debneyi Domin. Solanaceae Jones et al, 1980;Pospieszny et al, 2008 N. Verhoeven et al, 2003 N. tabacum L. cv.…”