Comparison of the coat protein genes of Lettuce big-vein associated virus isolates from Australia with those of isolates from other continents

The complete coat protein (CP) nucleotide sequences of seven Lettuce big-vein associated virus (LBVaV) isolates from Australia were compared to those of 22 other LBVaV and five tobacco stunt virus (TStV) isolates. On phylogenetic analysis, clade I contained only LBVaV isolates from Europe, sub-clade IIa only Australian LBVaV isolates, IIb only Japanese LBVaV isolates, and IIc only TStV isolates from Japan. In the amino acid sequences deduced, the central region of the gene was most divergent. Mean Dn/Ds ratios were 0.283 and 0.124 for clades I and II, respectively. The suggestion that TStV is a strain of LBVaV was supported.     

Molecular analysis of coat protein coding region of tobacco stunt virus shows that it is a strain of <Emphasis Type="Italic">Lettuce big-vein virus</Emphasis> in the genus <Emphasis Type="Italic">Varicosavirus</Emphasis>

Summary. To evaluate the relationship between tobacco stunt virus (TStV) and Lettuce big-vein virus (LBVV), we determined nucleotide sequences of the coat protein (CP) coding region of five TStV and three LBVV isolates and compared them with those of one Japanese and four Spanish isolates of LBVV. CP coding regions were identical in size and the nucleotide and amino acid sequence identities between TStV and LBVV were 95.6–96.5% and 97.2–98.7%, respectively. Phylogenetic analysis of nucleotide sequences indicated that TStV was very closely related to LBVV and a strain of LBVV rather than a distinct species.     

Comparison of the coat protein genes of Mirafiori lettuce big-vein virus isolates from Australia with those of isolates from other continents

The complete coat protein nucleotide encoding sequences of 13 Mirafiori lettuce big-vein virus isolates from Australia were compared to those of 23 other isolates, including one from Australia. On phylogenetic analysis, sub-clade A1 contained isolates from Australia (13), Europe and Japan, A2 contained isolates from Australia (1), Europe and South America, and B1 and B2 contained only European isolates. In the amino acid sequences deduced, the N-terminus and central regions varied considerably between clades A and B. Mean Dn/Ds ratios were 0.112, 0.076, 0.187 and 0.063 for all isolates, Australian isolates, clade A isolates and clade B isolates, respectively.     

An Ophiovirus isolated from lettuce with big-vein symptoms

Summary.  Big-vein is a widespread and damaging disease of lettuce, transmitted through soil by the chytrid fungus Olpidium brassicae, and generally supposed to be caused by Lettuce big-vein virus (LBVV; genus Varicosavirus). This virus is reported to have rigid rod-shaped particles, a divided double-stranded RNA genome, and one capsid protein of 48 kD, but has not been isolated or rigorously shown to cause the disease. We provide evidence that a totally different virus, here named Mirafiori lettuce virus (MiLV), is also very frequently associated with lettuce showing big-vein symptoms. MiLV was mechanically transmissible from lettuce to Chenopodium quinoa and to several other herbaceous test plants. The virus was partially purified, and an antiserum prepared, which did not react with LBVV particles in decoration tests. As reported for LBVV, MiLV was labile, soil-transmitted and had a single capsid protein of 48 kD, but the particles morphologically resembled those of ophioviruses, and like these, MiLV had a genome of three RNA segments approximately 8.5, 1.9 and 1.7 kb in size. MiLV preparations reacted strongly in Western blots and in ISEM with antiserum to Tulip mild mottle mosaic virus, an ophiovirus from Japan also apparently Olpidium-transmitted. They reacted weakly but clearly in Western blots with antiserum to Ranunculus white mottle virus, another ophiovirus. When lettuce seedlings were mechanically inoculated with crude or partially purified extracts from MiLV-infected test plants, many became systemically infected with MiLV and some developed big-vein symptoms. Such plants did not react in ELISA using an LBVV antiserum or an antiserum to tobacco stunt virus, and varicosavirus-like particles were never seen in them in the EM after negative staining. We conclude that MiLV is a hitherto undescribed virus assignable to the genus Ophiovirus. The cause or causes of lettuce big-vein disease and the properties of LBVV may need to be re-evaluated in light of our results. Accepted May 2, 2000 Received January 18, 2000     

Konjak mosaic virus: the complete nucleotide sequence of the genomic RNA and its comparison with other potyviruses

Summary. Konjak mosaic virus (KoMV) belongs to the genus Potyvirus, family Potyviridae. The complete nucleotide sequence of KoMV F isolate (KoMV F) was determined. The genome is 9,544 nucleotides long excluding the 3′ terminal poly A tail and encodes a typical potyviral 350-kDa polyprotein of 3,087 amino acids. Phylogenetic analysis using known potyvirus polyproteins shows that KoMV constitutes a branch with yam mosaic virus, close to another branch including Japanese yam mosaic virus, turnip mosaic virus, scallion mosaic virus and lettuce mosaic virus. The 3′ terminal 1,842 nucleotides of a different isolate of KoMV, K-2, was also determined, covering the C-terminal 292 amino acids of the nuclear inclusion protein b (NIb), coat protein (CP), and the 3′ untranslated region. The amino acid sequences of the KoMV F CP and the nucleotide sequences of the KoMV F 3′ untranslated region showed 92.5 and 90.5% identity to the corresponding genes of K-2, 88.7–96.8 and 92.7–94.4% to those of Zantedeschia mosaic virus (ZaMV) isolates, 87.5–89.7% and 85.5–90.3% to those of Japanese hornwort mosaic virus (JHMV) isolates. These results showed that KoMV is a distinct potyvirus and that KoMV, ZaMV, and JHMV are members of the same potyvirus species. Considering that KoMV was the first of these to be described, ZaMV and JHMV may be considered isolates of KoMV.     

Sequence Divergence of Four Soilborne Sugarbeet-Infecting Viruses

Soilborne viruses are among the most harmful pathogens of sugarbeet (Beta vulgaris L.ssp. vulgaris) but most of them lack information on genetic variability due to paucity of sequence data. Only one isolate of Beet soil borne virus (BSBV; genus Pomovirus), Beet virus Q (BVQ; genus Pomovirus) and Beet soil borne mosaic virus (BSBMV; genus Benyvirus) has been characterised for the coat protein (CP) gene. In this study, the CP gene sequences of three isolates each of BSBV and Beet necrotic yellow vein virus (BNYVV; genus Benyvirus) (France, Germany and USA), two isolates of BVQ (France and Germany), and one isolate of BSBMV (USA) were determined. Phylogenetic analyses including sequences from databanks indicated that the French BNYVV isolate of this study belongs to so-called P-type, the American isolate to A-type and the German isolate to B-type. The CP genes of the three BSBV isolates characterised in this study and the one available from databank were highly identical (98.4–99.0% at nucleotide level; one variable amino acid). The BSBMV isolate studied here differed from the previously characterised isolate for five nucleotides and four amino acids in the CP region. The two BVQ isolates characterised in this study contained three additional nucleotides resulting in an additional amino acid residue (arginine) at CP position 86, as compared to the only isolate available in databank.     

Complete genome sequences of Maize dwarf mosaic and Sugarcane mosaic virus isolates coinfecting maize in Spain

Summary The genomes of Spanish isolates of Maize dwarf mosaic virus (MDMV-Sp) and Sugarcane mosaic virus (SCMV-Sp) were completely sequenced. Nucleotide sequence identities of SCMV-Sp to those of other SCMV isolates ranged from 79 to 90%. MDMV-Sp shared 85% nucleotide identity with the only other fully sequenced isolate of MDMV. MDMV-Sp and SCMV-Sp differed from each other by 31% in their nucleotide sequences. Phylogenetic analyses showed that SCMV isolates group by host rather than by geographical location. Two significant recombination signals were identified in the NIa and NIb regions of the SCMV-Sp genome.     

Genetic variation of coat protein gene among the isolates of Rice tungro spherical virus from tungro-endemic states of the India

Rice tungro disease, one of the major constraints to rice production in South and Southeast Asia, is caused by a combination of two viruses: Rice tungro spherical virus (RTSV) and Rice tungro bacilliform virus (RTBV). The present study was undertaken to determine the genetic variation of RTSV population present in tungro endemic states of Indian subcontinent. Phylogenetic analysis based on coat protein sequences showed distinct divergence of Indian RTSV isolates into two groups; one consisted isolates from Hyderabad (Andhra Pradesh), Cuttack (Orissa), and Puducherry and another from West Bengal, Coimbatore (Tamil Nadu), and Kanyakumari (Tamil Nadu). The results obtained from phylogenetic study were further supported with the SNPs (single nucleotide polymorphism), INDELs (insertion and deletion) and evolutionary distance analysis. In addition, sequence difference count matrix revealed 2–68 nucleotides differences among all the Indian RTSV isolates taken in this study. However, at the protein level these differences were not significant as revealed by Ka/Ks ratio calculation. Sequence identity at nucleotide and amino acid level was 92–100% and 97–100%, respectively, among Indian isolates of RTSV. Understanding of the population structure of RTSV from tungro endemic regions of India would potentially provide insights into the molecular diversification of this virus.     

Genetic variability and phylogenetic analysis of hosta virus X

Triple gene block 1 (TGB1) and coat protein (CP) sequences of 30 hosta virus X (HVX) isolates from Tennessee (TN), USA, were determined and compared with available sequences in GenBank. The CPs of all known HVX isolates, including those from TN, shared 98.3–100% and 98.2–100% nucleotide and amino acid sequence identity, respectively, whereas TGB1 shared 97.4–100% nucleotide and 97–100% amino acid sequence identity. TGB1 of TN isolates were all longer by one codon from that of a Korean isolate, which is the only sequence publicly available. Phylogenetic analysis of nucleotide and amino acid sequences of TGB1 and CP of all known HVX isolates, separately or combined, revealed a close relationship, suggesting that all of them are derived from a common ancestor. Phylogenetic analysis with the type member of each genus of the family Flexiviridae confirmed that HVX is a member of a distinct species of the genus Potexvirus.     

Biological and molecular variability among geographically diverse isolates of sweet potato virus 2

Summary. Sweet potato virus 2 (SPV2) is a tentative member of the genus Potyvirus, family Potyviridae. In addition to the type isolate of SPV2 recently characterised in greater detail, twelve additional isolates of this virus were obtained from sweet potato clones originating from China, Portugal, South Africa and Zambia. Sequences of the coat protein (CP) gene and 3′ non-translated region (NTR) were determined. Comparisons of the CP gene sequences of these isolates revealed nucleotide and amino acid sequence identities ranging from 81 to 99% and from 86 to 99%, respectively. Phylogenetic analysis of sequences distinguished several groups, which partially correlated with the geographic origin of the isolates, and indicated that some isolates from South Africa and a Zambian isolate are most distinct both in CP and 3′NTR sequences. Host range studies of a selected number of isolates revealed some differences in test plant reactions, which appeared to correlate to some extent with the geographic origin and molecular distinctness of the SPV2 isolates. The results strongly suggest the occurrence of biologically and genetically diverse strains of SPV2.     

Distribution and molecular diversity of three cucurbit-infecting poleroviruses in China

Cucurbit aphid-borne yellows virus (CABYV) and Melon aphid-borne yellows virus (MABYV) have been found to be associated with cucurbit yellowing disease in China. Our report identifies for the first time a third distinct polerovirus, tentatively named Suakwa aphid-borne yellows virus (SABYV), infecting Suakwa vegetable sponge. To better understand the distribution and molecular diversity of these three poleroviruses infecting cucurbits, a total of 214 cucurbitaceous crop samples were collected from 25 provinces in China, and were investigated by RT-PCR and sequencing. Of these, 108 samples tested positive for CABYV, while 40 samples from five provinces were positive for MABYV, and SABYV was detected in only 4 samples which were collected in the southern part of China. Forty-one PCR-amplified fragments containing a portion of the RdRp gene, intergenic NCR and CP gene were cloned and sequenced. Sequence comparisons showed that CABYV isolates shared 78.0–79.2% nucleotide sequence identity with MABYV isolates, and 69.7–70.8% with SABYV. Sequence identity between MABYV and SABYV was 73.3–76.5%. In contrast, the nucleotide identities within each species were 93.2–98.7% (CABYV), 98.1–99.9% (MABYV), and 96.1–98.6% (SABYV). Phylogenetic analyses revealed that the polerovirus isolates fit into three distinct groups, corresponding to the three species. The CABYV group could be further divided into two subgroups: the Asia subgroup and the Mediterranean subgroup, based on CP gene and partial RdRp gene sequences. Recombination analysis suggested that MABYV may be a recombinant virus.     

Evidence that <Emphasis Type="Italic">Cymbidium mosaic virus</Emphasis> (CymMV) isolates divide into two subgroups based on nucleotide diversity of coat protein and replicase genes

Summary The genetic diversity of Cymbidium mosaic virus (CymMV, family Flexiviridae) was assessed by analysing the nucleotide sequences of coat protein (CP) and partial RNA-dependent RNA polymerase (RdRp) genes. Thirty CymMV sequences from vanilla isolates, obtained in this work by direct sequencing of RT-PCR products, were compared to the sequences from ornamental orchid isolates available in GenBank. The CymMV population exhibited overall low genetic diversity (π = 0.054 and π = 0.053 for CP and RdRp genes, respectively). Phylogenetic analyses of the 85 CP and 37 RdRp sequences revealed the segregation of the isolates into two congruent monophyletic clusters; however these two subgroups did not cluster in amino sequence analysis because most of the nt mutations were synonymous. Nevertheless, the subgrouping was confirmed by highly significant Kst tests for the CP and RdRp genes. Analysis of population genetic parameters and distribution of synonymous and nonsynonymous mutations revealed that both genes were under negative selection with no recombination events. These results suggested that the CymMV isolates found in cultivated orchids worldwide have a dual origin and are expanding as if following bottlenecks.     

Analysis of the biological and molecular variability of Watermelon mosaic virus isolates from Iran

Watermelon mosaic virus (WMV) is one of the most important viruses that causes different symptoms in Cucurbitaceae. WMV is a potyvirus with a worldwide distribution, but occurs most commonly in temperate and Mediterranean regions. Cucurbit species grown in Yazd, Esfahan, West Azerbaijan, Hormozgan, and Kerman provinces were surveyed for the relative incidence of WMV in 2004–2005. A total of 757 symptomatic cucurbit and 31 weed species were collected and assayed for infection with WMV. Of 788 leaf samples from cucurbit and weed plants, 190 samples were positive by double antibody sandwich ELISA (DAS-ELISA) using specific polyclonal antibody. Among the weed species tested only colocynth (Citrullus colocynthis) was found to be infected with WMV. The coat protein (CP) gene from 18 representative isolates was PCR amplified, cloned, sequenced, and compared with the sequences available in GeneBank. Phylogenetic analysis using 778 nucleotide long sequences of the coat protein gene showed that these isolates fell into two; groups I and II. Only one isolates (KER.JI.1) was classified in the group II. This isolate had a wider host range and infected Nicotiana debneyii and Datura metel. None of the other 17 isolates could infect these two species. Members of group I were divided into three subgroups; A, B, and C. The subgroup I_B appears to be a new subgroup comprising only of the Iranian isolates. Phylogenetic analysis based on 200 nucleotides coding for the N-terminal segment of the CP showed that all Iranian isolates except KER.JI.1 clustered with the previously reported WMV strains. All Iranian isolates had a DAG amino acid triplet which is involved in aphid transmissibility. This is the first report on sequence analysis of the nearly full-length CP cDNA clones of WMV isolates from Iran.     

Phylogenetic analysis of <Emphasis Type="Italic">Beet necrotic yellow vein virus</Emphasis> isolates from China

A survey detected Beet necrotic yellow vein virus (BNYVV) infection in six Chinese sugar-beet-growing regions. To study the diversity of virus isolates among the regions, nucleotide sequences of four proteins, namely CP, p25, p31, and p26, were determined and the amino acid sequences thus deduced were analyzed using sequence alignments and the phylogenetic method, respectively. Amino acid sequence analysis of CP revealed A-type isolates in Harbin, Hohhot, Baotou, Wuwei, and Jiuquan and B-type isolates in Hohhot and Changji; no Chinese isolate was found to cluster with European P-type isolates. Chinese p25 proteins clustered into three groups with seven tetrad motifs (positions 67–70). Of the seven, the tetrad ASHG has not been reported previously. Most Chinese p31 proteins clustered in p31-2 group, diverging from European p31 proteins. Isolates containing RNA 5 occurred in most of the investigated regions and were associated with both A-and B-types. Phylogenetic analyses of these four proteins showed complex patterns of genetic diversity among these Chinese isolates and indicated that the isolates of China and Japan were more closely related and may have a common origin. The nucleotide sequence data reported in this article have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers EF473089–EF473126.     

Nucleotide sequence of a Japanese isolate of Squash mosaic virus. Brief report

Summary.  The complete nucleotide sequence of the RNA-1 of Squash mosaic virus (SqMV) was determined using a Japanese isolate (Y-SqMV). The sequence consisted of 5865 nucleotides excluding the poly (A) at the 3′ terminus and contained a single long open reading frame with a coding capacity for a protein of M_r209971. Analysis of the deduced amino acid sequence suggested a genomic organization typical of comoviruses. The nucleotide sequence of the RNA-2 of Y-SqMV was also determined and compared with the SqMV isolates from the United States. The larger and smaller capsid protein (CP) coding region was compared to those of K-SqMV and Z-SqMV, which represent two subgroups of SqMV. The larger CP gene of Y-SqMV showed 93.0% and 88.0% identities with those of K-SqMV and Z-SqMV, respectively at the nucleotide level. The smaller CP gene of Y-SqMV was 94.1% and 88.4% identical with those of K-SqMV and Z-SqMV. The results suggested that the Japanese SqMV isolate (Y-SqMV) is distinct from those in the United States, and might represent a third subgroup. Received May 7, 2001 Accepted August 1, 2001     

Sequence diversity of readthrough proteins of <Emphasis Type="Italic">Soybean dwarf virus</Emphasis> isolates from the Midwestern United States

The amino acid sequence diversity of readthrough proteins (RTPs) of 24 dwarfing isolates of Soybean dwarf virus (SbDV) from Wisconsin and Illinois was analyzed. The RTP, a minor component of viral capsids, has a significant role in specificity of aphid transmission of luteovirids. Among the isolates, nucleotide sequence identities ranged from 95 to 100%. The predicted amino acid sequences differed at 56 amino acid positions in the 54 kDa RTD compared to only five positions in the 22 kDa CP. Phylogenetic analysis of both amino acid and nucleotide sequences showed three distinct clusters of SbDV isolates. The GenBank accession numbers of the sequence reported in this paper are EU095846, EU095847, and EU419570–EU419584.     

The complete genome sequence for a Turkish isolate of <Emphasis Type="Italic">Wheat dwarf virus</Emphasis> (WDV) from barley confirms the presence of two distinct WDV strains

The complete genome for a barley isolate of Wheat dwarf virus (WDV) from Tekirdağ, Turkey, WDV-Bar[TR], was isolated and sequenced. The genome was found to be 2739 nucleotides long, which is shorter than wheat-infecting WDV isolates, and with a genome organization typical for mastreviruses. The complete genome of WDV-Bar[TR] showed 83–84% nucleotide identity to wheat isolates of WDV, with the non-coding regions SIR and LIR least conserved (72–74% identity). The deduced amino acid sequences for Rep and RepA were most conserved (92–93%), while CP and MP were less conserved (87% and 79–80%, respectively). The identity to other mastrevirus species was significantly lower. In phylogenetic analyses, the WDV isolates formed a distinct clade, well separated from the other mastreviruses with the wheat isolates grouping closely together. Phylogenetic analyses of WDV-Bar[TR], the partial sequence for another Turkish barley isolate (WDV-Bar[TR2]) and published WDV sequences further supported the division of WDV into two distinct strains. The barley strain could also be divided into three subtypes based on relationships and geographic origin. This study shows the first complete published sequence for a barley isolate of WDV. The nucleotide sequence data reported appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession numbers AJ783960 and AJ811960.     

Low genetic diversity of Squash vein yellowing virus in wild and cultivated cucurbits in the U.S. suggests a recent introduction

Squash vein yellowing virus (SqVYV) isolates were collected from cultivated and weedy cucurbits representing major hosts and locations in the U.S. and analyzed to better understand the diversity and population structure. No differences in symptoms were observed in field-collected isolate source plants or subsequently inoculated greenhouse plants, and the complete genome of an SqVYV isolate from a wild cucurbit host (smellmelon, Cucumis melo var. dudaim) was highly similar (99.4% nucleotide identity, 99.3% amino acid identity) to the previously published type isolate from squash. Although analysis of the coat protein (CP) and two serine proteases (P1a and P1b) sequences for 41 isolates showed little diversity across seven years of sampling, it revealed two distinct groups of SqVYV isolates with low intra-group diversity. Our analyses also suggested that recombination had occurred between SqVYV isolates, similar to other ipomoviruses. Selection pressures on the genome regions analyzed were negative indicating purifying selection was occurring. The magnitude of negative selection in SqVYV was consistent with what has been reported for other ipomoviruses, and was greatest for the CP and least for the P1b. The observed genetic diversity was similar to that reported for Cucumber vein yellowing virus but less than that reported for Sweet potato mild mottle virus, Cassava brown streak virus and Ugandan cassava brown streak virus. Collectively, these results indicate that the current U.S. population of SqVYV has undergone a recent genetic bottleneck and was introduced from elsewhere.