Occurrence of aphidborne viruses in southernmost South American populations of Fragaria chiloensis ssp. chiloensis

Wild and cultivated Fragaria chiloensis ssp. chiloensis (Fcc) plants were collected at different locations in southern Chile in order to determine the current viral status of this native strawberry. The following aphidborne viruses (ABVs): Strawberry mild yellow edge virus (SMYEV), Strawberry mottle virus (SMoV), Strawberry crinkle virus (SCV) and Strawberry vein banding virus (SVBV), were found in wild and cultivated Fcc plants, but severe symptoms were not associated with viral infection. Furthermore, partial gene sequences of these ABV isolates were determined and displayed a high degree of conservation with virus isolates reported previously. In addition, partial gene sequences of SCV and SVBV from southernmost South American populations of Fcc are described for the first time. High‐throughput parallel sequencing (Illumina) of double‐stranded RNA was used to provide viral profiles of Fcc from different locations. Although strong evidence of novel viruses affecting Fcc was not found, it was confirmed that ABVs are the most frequent viruses affecting this subspecies. The information provided will help in the development of high‐quality molecular tools for virus detection and control in Fcc.     

Relationships of <Emphasis Type="Italic">Barley yellow dwarf virus</Emphasis>-PAV and <Emphasis Type="Italic">Cereal yellow dwarf virus</Emphasis>-RPV from Iran with viruses of the family <Emphasis Type="Italic">Luteoviridae</Emphasis>

Barley yellow dwarf disease is one of the most important problems confronting cereal production in Iran. Barley yellow dwarf virus-PAV (BYDV-PAV) and Cereal yellow dwarf virus-RPV (CYDV-RPV) are the predominant viruses associated with the disease. One isolate of BYDV-PAV from wheat (PAV-IR) and one isolate of CYDV-RPV from barley (RPV-IR) were selected for molecular characterisations. A genome segment of each isolate was amplified by PCR. The PAV-IR fragment (1264 nt) covered a region containing partial genes for coat protein (CP), read through protein (RTP) and movement protein (MP). PAV-IR showed a high sequence identity to PAV isolates from USA, France and Japan (96–97%). In a phylogenetic analysis it was placed into PAV group I together with PAV isolates from barley and oats. The fragment of RPV-IR (719 nt) contained partial genes for CP, RTP and MP. The sequence information confirmed its identity as CYDV. However, RPV-IR showed 90–91% identity with both RPV and Cereal yellow dwarf virus-RPS (CYDV-RPS). Phylogenetic analyses suggested that it was more closely related to RPS. These data comprise the first attempt to characterise BYD-causing viruses in Iran and southwest Asia. The nucleotide sequence data reported appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession numbers AY450425 and AY450454     

Biological and molecular variation amongst Australian Turnip mosaic virus isolates

Turnip mosaic virus (TuMV) causes crop losses worldwide. Eight Australian TuMV isolates originally obtained from five different species in two plant families were inoculated to 14 plant species belonging to four families to compare their host reactions. They differed considerably in virulence in Brassicaceae crop species and virus indicator hosts belonging to three other families. The isolates infected most Brassica species inoculated, but not Raphanus sativus, usually causing systemic mosaic symptoms, so they resembled TuMV biological host type [B]. Whole genome sequences of seven of the Australian isolates were obtained and had lengths of 9834 nucleotides (nt). When they were compared with 37 non‐recombinant TuMV genomes from other continents and another whole genome from Australia, six of them formed an Australian group within the overall world‐B phylogenetic grouping, while the remaining new genome sequence and the additional whole genome from Australia were part of the basal‐B grouping. When the seven new Australian genomes and the additional whole genome from Australia were subjected to recombination analysis, six different recombination events were found. Six genomes contained one or two recombination events each, but one was non‐recombinant. The non‐recombinant isolate was in the Australian grouping within the overall world‐B group while the remaining recombinant isolates were in the basal‐B and world‐B phylogenetic groups.     

Molecular and biological characterization of <Emphasis Type="Italic">Chrysanthemum stem necrosis virus</Emphasis> isolates from distinct regions in Japan

Three isolates of Chrysanthemum stem necrosis virus (CSNV) were obtained from chrysanthemum plants in distinct regions of Japan in 2006 and 2007. All the original host plants showed severe necrotic symptoms on the leaves and stems. Amino acid sequence data of the nucleocapsid protein genes of the three isolates (CbCh07A, TcCh07A, and GnCh07S) showed high identities with those of two other CSNV isolates, HiCh06A L1 from Japan and Chry1 from Brazil. Furthermore, for the first time the complete nucleotide sequence of the S RNA was determined for CSNV (isolate HiCh06A). In phylogenetic analysis based on the non-structural protein genes from the genus Tospovirus, HiCh06A L1 was placed in the same genetic group as Tomato spotted wilt virus (TSWV) and Impatiens necrotic spot virus. Host range examination for isolates HiCh06A L1 and CbCh07A showed that green pepper (cv. ‘Kyoyutaka’, ‘Saitamawase’, ‘Tosakatsura’, ‘L3 sarara’ and ‘L3 miogi’) and tomato (cv. ‘Sekaiichitomato’) were systemically susceptible hosts, whereas TSWV-resistant Solanaceae species, Capsicum chinense, Lycopersicon peruvianum and a TSWV-resistant cultivar of green pepper (cv. TSR miogi), were resistant.     

Susceptibility and resistance of <Emphasis Type="Italic">Beta vulgaris</Emphasis> subsp. <Emphasis Type="Italic">maritima</Emphasis> to foliar rub-inoculation with <Emphasis Type="Italic">Beet necrotic yellow vein virus</Emphasis>

Four lines (designated MR0, MR1, MR2, and M8) from 13 accessions of Beta vulgaris subsp. maritima were selected on the basis of phenotypes produced after foliar rub-inoculation with Beet necrotic yellow vein virus (BNYVV). The susceptible phenotype developed bright yellow local lesions, whereas the resistant phenotype had symptoms ranging from no visible lesions to necrotic lesions at the inoculation site. MR1 and MR2 lines had a resistant phenotype depending on the isolate and the MR0 line was susceptible to all isolates of BNYVV tested. The M8 line was highly susceptible; the virus spread systemically and caused severe stunting. These plant lines will be useful for distinguishing BNYVV isolates having different pathogenicities, especially those controlled by RNA3 and/or RNA5.     

Phylogeny of Egyptian isolates of <Emphasis Type="Italic">Cucumber mosaic virus</Emphasis> (CMV) and <Emphasis Type="Italic">Tomato mosaic virus</Emphasis> (ToMV) infecting <Emphasis Type="Italic">Solanum lycopersicum</Emphasis>

Four Cucumber mosaic virus (CMV) (CMV-HM 1–4) and nine Tomato mosaic virus (ToMV) (ToMV AH 1–9) isolates detected in tomato samples collected from different governorates in Egypt during 2014, were here characterized. According to the coat protein gene sequence and to the complete nucleotide sequence of total genomic RNA1, RNA2 and RNA3 of CMV-HM3 the new Egyptian isolates are related to members of the CMV subgroup IB. The nine ToMV Egyptian isolates were characterized by sequence analysis of the coat protein and the movement protein genes. All isolates were grouped within the same branch and showed high relatedness to all considered isolates (98–99%). Complete nucleotide sequence of total genomic RNA of ToMV AH4 isolate was obtained and its comparison showed a closer degree of relatedness to isolate 99–1 from the USA (99%). To our knowledge, this is the first report of CMV isolates from subgroup IB in Egypt and the first full length sequencing of an ToMV Egyptian isolate.     

<Emphasis Type="Italic">Turnip ringspot virus</Emphasis> recognised on Chinese cabbage in Russia

The nucleotide sequence of the 3′-terminal part of the RNA1 genome segment of the M12 isolate of comovirus Turnip ringspot virus (TuRSV) was established. This isolate originated in 1989 in Moscow (Russia) from Chinese cabbage with Radish mosaic virus-like symptoms. Comparison of the M12 RNA polymerase amino acid sequence with that of Radish mosaic virus (RaMV) revealed significant differences; these proteins are of different length and are only about 75% identical. On the other hand, the amino acid sequence of the M12 RNA polymerase was more than 94% identical with that of TuRSV recently described in Toledo (USA). We conclude that TuRSV occurs in Europe as well as in America and probably represents a new species of the genus Comovirus.     

Molecular characterisation of <Emphasis Type="Italic">Rice stripe necrosis virus</Emphasis> as a new species of the genus <Emphasis Type="Italic">Benyvirus</Emphasis>

The complete nucleotide sequences of RNAs 1 and 2 of Rice stripe necrosis virus (RSNV) were determined and compared to the corresponding genomes of all sequenced, rod-shaped plant viruses. The genome organisation of RSNV RNA1 and RNA2 is nearly identical to that of Beet necrotic yellow vein virus (BNYVV) and Beet soil-borne mosaic virus (BSBMV), definitive species of the genus Benyvirus. As demonstrated for BNYVV and BSBMV, the RNA1 of RSNV also encodes a single ORF with putative replicase-associated motifs, which distinguishes benyviruses from all other viruses possessing rod-shaped particles. As described for BNYVV, RNSV RNA-2 also contains six ORFs: the capsid protein gene, the read-through protein gene, a triple gene block gene that codes for three different proteins, and a 17 kDa cysteine-rich protein. RNAs 3 and 4 (or 5 in the case of BNYVV), identified in natural infections of BNYVV and BSBMV, were not detected in any of the 44 RSNV cDNA clones obtained in this investigation. Nevertheless, phylogenetic and amino comparative acid sequence analyses demonstrated that RSNV is more closely related to BNYVV and BSBMV than to any other rod-shaped plant virus characterised to date.     

Infectivity analysis of a soybean isolate of Mungbean yellow mosaic India virus by agroinoculation

Yellow mosaic disease (YMD) of legumes endemic to South Asia are caused by begomoviruses transmitted by whiteflies. Based on molecular characterization, two distinct viruses – Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV) – were found previously to be the etiological agents of YMD in legumes. Here, host range studies with a soybean isolate of MYMIV (MYMIV-[Sb]) were carried out by both whitefly transmission and agroinoculation. MYMIV-[Sb] was similar to a cowpea isolate of MYMIV (MYMIV-[Cp]) in its ability to infect cowpea, thus differing from blackgram (MYMIV) and mungbean (MYMIV-[Mg]) isolates, which do not infect cowpea. Genomic analysis of DNA A and DNA B components of these MYMIV isolates show characteristic differences in complete DNA B nucleotide sequence correlating with host range differences.     

Characterization of two distinct Polish isolates of Pepino mosaic virus

In Poland in 2002 and 2005 two different isolates of Pepino mosaic virus signed PepMV-SW and PepMV-PK were obtained. Both isolates were compared on the basis of their symptomatology on a series of plant species. In addition, the isolates were characterized by the nucleotide sequence analysis of the triple gene block, coat protein and a part of the polymerase genes. The studies showed that the Polish isolates differ from each other and belong to two strains. PepMV-SW was highly similar to European isolates, showing extensive sequence identity, ca. 99%. Pairwise comparisons of PepMV-PK with other PepMV isolates from the GenBank database showed that the highest nucleotide sequence identity was with two isolates: Ch2 from Chile and US2 from the USA.     

应用纳米磁珠实时荧光定量PCR方法检测番茄黄化曲叶病毒

In order to develop a rapid, sensitive and specific qPCR assay for detection and quantification of Tomato yellow leaf curl virus (TYLCV), a pair of primers and TaqMan probe were designed according to the conserved sequence of known TYLCV isolates. Combining with MNP technique, a novel MNP-qPCR detection method was established and verified based on specificity, sensitivity and reproducibility tests. The results indicated that the Ct value of plotted standard curve showed good linear relationship(R² =0.9994)with the log of copy number of template. The established method showed a high specificity for TYLCV detection without crossing reaction with Tomato severe leaf curl virus and Tomato yellow leaf curl Sadinia virus, and was 10-fold more sensitive than routine PCR. Both coefficients of variation were less than 2%, indicating a good reproducibility. We have provided a novel method for detection of TYLCV in plant samples rapidly and quantitatively.     

A novel putative member of the family Benyviridae is associated with soilborne wheat mosaic disease in Brazil

Soilborne wheat mosaic disease (SBWMD), originally attributed to infections by Soilborne wheat mosaic virus (SBWMV) and Wheat spindle streak mosaic virus (WSSMV), is one of the most frequent virus diseases and causes economic losses in wheat in southern Brazil. This study aimed to characterize molecularly the viral species associated with wheat plants showing mosaic symptoms in Brazil. Wheat leaves and stems displaying mosaic symptoms were collected from different wheat cultivars in Passo Fundo municipality, Rio Grande do Sul State, southern Brazil. Double-stranded RNA was extracted and submitted to cDNA library synthesis and next-generation sequencing. No sequences of SBWMV and WSSMV were detected but the complete genome sequence of a putative new member of the family Benyviridae was determined, for which the name wheat stripe mosaic virus (WhSMV) is proposed. WhSMV has a bipartite genome with RNA 1 and RNA 2 organization similar to that of viruses belonging to Benyviridae. WhSMV RNA 1 has a single open reading frame (ORF) encoding a polyprotein with putative viral replicase function. WhSMV RNA 2 has six ORFs encoding the coat protein, the major protein (read-through), triple gene block movement proteins (TGB 1, 2 and 3) and ORF 6 (hypothetical protein). In addition to the genomic organization and nucleotide and amino acid sequence identities, phylogenetic analyses also corroborated that WhSMV is a virus species of the Benyviridae. However, isolates of WhSMV formed a clade distinct from members of the genus Benyvirus. It was also demonstrated that the plasmodiophorid Polymyxa graminis is associated with wheat roots showing SBWMD symptoms and infected by WhSMV.     

Biological and molecular characterization of two closely related carlaviruses affecting brassica plants

Carlaviruses are plant-infecting viruses with flexuous filamentous particles of approximately 650 nm in length and a positive single-stranded hexacistronic RNA molecule as the genome. In this study, we analysed 14 samples of brassicas plants, reportedly affected by carlaviruses, that were collected in distant and edaphoclimatic distinct regions in Brazil. The genomes of four viral isolates detected in leaf kale (Brassica oleracea var. acephala) plants displayed the typical genomic organization of carlaviruses, subdivided into two contrasting identity profiles. Isolates T25, T107, and T110 showed a higher sequence identity among them (c.93%) than with T90 (c.67%), which had the highest nucleotide sequence identity (85.2%) with the only documented genomic fragment, approximately 1 kb of the RNA 3′-end, of cole latent virus (CoLV). Identity values among the other three isolates and CoLV were consistently lower than 78.6%. Nucleotide and amino acid sequence identity values of the replicase cistron of the isolates T25 and T90 are below the threshold for species demarcation in the genus Carlavirus. Upon mechanical transmission, these two isolates induced different symptoms in brassicas and solanaceous plants. Overall, data indicated that these viruses belong to two related but distinct species of carlaviruses. While T90 was recognized as an isolate of CoLV, the remaining isolates were considered members of a new tentative carlavirus named Cole mild mosaic virus (CoMMV). Current work provides convincing support for the taxonomic status of the species Cole latent virus, enlarges the known diversity of carlaviruses, and makes available new molecular tools to improve surveys of brassica-infecting viruses.