Complete nucleotide sequence of the Toledo isolate of turnip ringspot virus

The complete genomic sequence of the Toledo isolate of the comovirus, turnip ringspot virus (TuRSV), was found to consist of 2 polyadenylated RNAs. RNA 1 is 6082 nucleotides long and encodes a single predicted polypeptide of 1860 amino acids. The predicted RNA 1 polyprotein contains the polypeptides for viral replication and proteolytic processing. RNA 2, that is 3985 nucleotides long, codes for a single predicted 1095 amino acid polypeptide containing the movement and coat proteins. Phylogenetic analysis indicates that TuRSV is most closely related to radish mosaic virus, and these crucifer-infecting pathogens form a distinct clade within the comoviruses.     

Complete sequence of RNA1 of grapevine Anatolian ringspot virus

The nucleotide sequence of RNA1 of grapevine Anatolian ringspot virus (GARSV), a nepovirus of subgroup B, was determined from cDNA clones. It is 7,288 nucleotides in length excluding the 3′ terminal poly(A) tail and contains a large open reading frame (ORF), extending from nucleotides 272 to 7001, encoding a polypeptide of 2,243 amino acids with a predicted molecular mass of 250 kDa. The primary structure of the polyprotein, compared with that of other viral polyproteins, revealed the presence of all the characteristic domains of members of the order Picornavirales, i.e., the NTP-binding protein (1B^Hel), the viral genome-linked protein (1C^VPg), the proteinase (1D^Prot), the RNA-dependent RNA polymerase (1E^Pol), and of the protease cofactor (1A^Pro-cof) shared by members of the subfamily Comovirinae within the family Secoviridae. The cleavage sites predicted within the polyprotein were found to be in agreement with those previously reported for nepoviruses of subgroup B, processing from 1A to 1E proteins of 67, 64, 3, 23 and 92 kDa, respectively. The RNA1-encoded polyprotein (p1) shared the highest amino acid sequence identity (66 %) with tomato black ring virus (TBRV) and beet ringspot virus (BRSV). The 5′- and 3′-noncoding regions (NCRs) of GARSV-RNA1 shared 89 % and 95 % nucleotide sequence identity respectively with the corresponding regions in RNA2. Phylogenetic analysis confirmed the close relationship of GARSV to members of subgroup B of the genus Nepovirus.     

Complete nucleotide sequence of an Argentinean isolate of sweet potato virus G

Sweet potato virus G belongs to the largest plant virus genus Potyvirus. This virus was detected for the first time in Argentina and then sequenced using the method of next-generation pyrosequencing. The complete genome was found to be 10,798 nucleotides excluding the poly-A tail with a predicted genome organization typical for a member of the genus Potyvirus. This is the first report of the complete genomic sequence of a SPVG isolated from South America.     

Complete nucleotide sequence of RNA-2 of Olive latent ringspot virus

Summary.  The complete nucleotide sequence of Olive latent ringspot virus (OLRSV) RNA-2 was determined. This RNA is 3969 nucleotides in length and contains a single open reading frame of 3448 nt, that encodes a polypeptide of 1146 amino acids, with a calculated M_r of 126,044. OLRSV RNA-2 has a structural organization typical of nepoviruses, with the coat protein (CP) cistron located in the C-terminal and the putative movement protein (MP) in the N-terminal regions of the polyprotein. Computer-assisted comparison of coat proteins of OLRSV and other nepoviruses disclosed relationships that tally with subgrouping based onphysicochemical properties. Received June 5, 2000 Accepted July 18, 2000     

Complete nucleotide sequence of a maize chlorotic mottle virus isolate from Nebraska

The complete genome of a maize chlorotic mottle virus isolate from Nebraska (MCMV-NE) was cloned and sequenced. The MCMV-NE genome consists of 4,436 nucleotides and shares 99.5 nucleotide sequence identity with an MCMV isolate from Kansas (MCMV-KS). Of 22 polymorphic sites, most resulted from transition with a clear bias for U to C and C to U substitutions. The MCMV-NE genome was assembled into a single plasmid insert and used as a template to transcribe RNA in vitro. As RNA transcribed from the cloned MCMV-NE genome was infectious to maize plants, sequence differences between MCMV-NE and MCMV-KS are most likely neutral with respect to pathogenicity and virulence.     

Sequence analysis of grapevine isolates of Raspberry ringspot nepovirus

Summary. The nucleotide sequences of RNAs 1 and 2 of a German isolate of Raspberry ringspot virus (RpRSV) infecting grapevine (RpRSV-Grapevine), as well as partial sequences of another grapevine isolate from Switzerland (RAC815) were determined. The sequences of the protease-polymerase region encoded by RNA1, and the movement protein and coat protein genes encoded by RNA 2, of these isolates were compared with those of other isolates available in databases. The coat proteins of the grapevine isolates formed a sister group to all those from other RpRSV isolates, but whether this resulted from divergence or recombination was uncertain.     

Complete nucleotide sequence and genome organisation of grapevine Bulgarian latent virus

The complete genome sequence of grapevine Bulgarian latent virus (GBLV) has been determined. RNA-1 (7,452 nt in length) contains a single ORF of 6,285 nt, encoding a polyprotein with conserved motifs characteristic of the viral protease cofactor (Prot-cofact), the NTP-binding protein (NTP), the cysteine-like protease (Cyst-Prot) and the RNA-dependent RNA polymerase (RdRp) of members of the order Picornavirales and show high aa sequence identity with blackcurrant reversion virus (BRV, 64%). RNA-2 (5,821 nt) contains a single ORF of 4,500 nt, encoding a polyprotein in which the conserved motifs of the movement protein (MP) and coat protein (CP) have been identified. The GBLV CP aa sequence shows highest homology with that of blueberry leaf mottle virus (BLMoV, 68%). Both RNAs have a poly(A) tail and a NCR at the 3' and 5' termini, respectively. The results of this study confirm the classification of GBLV as a member of a distinct species in subgroup C of the genus Nepovirus.     

Complete nucleotide sequence of Velvet tobacco mottle virus isolate K1

Velvet tobacco mottle virus (VTMoV) infects the native Australian plant Nicotiana velutina, which is endemic to central Australia. This virus is included in the genus Sobemovirus based on virion morphology and serological relationships. We report here the full genome sequence of VTMoV, attained using a genome-walking strategy with both degenerate and specific primers. This sequence confirms that VTMoV is a sobemovirus, with the same open reading frame (ORF) organisation as other described sobemoviruses. The VTMoV sequence is closest to those sobemoviruses isolated from monocotyledonous plants, although the narrow host range of VTMoV is limited to dicotyledonous plants.     

Complete nucleotide sequence of Cherry leaf roll virus (CLRV), a subgroup C nepovirus

The complete nucleotide sequence of both genomic (+)ss RNAs of a rhubarb isolate of Cherry leaf roll virus (CLRV) was determined. The larger RNA1 is 7918 nucleotides and the shorter RNA2 6360 nucleotides in size, each genome component comprising a single open reading frame (ORF). The RNA1-encoded polyprotein (P1) is 2112 amino acids long (235.6 kDa) containing domains characteristic for a proteinase-cofactor (PCo), nucleotide-binding helicase (Hel), genome-linked protein (VPg), proteinase (Pro), and an RNA-dependent RNA polymerase (Pol). The RNA2-encoded polyprotein (P2) has a molecular mass of 174.9 kDa (1589 aa) encoding the putative movement protein (MP) and the coat protein (CP) of CLRV. The genome region upstream of the MP has a coding capacity of 77 kDa, however processing of P2 by the putative virus-encoded proteinase and protein-function encoded by this region is unknown. Furthermore, it could be demonstrated that the 5′-termini including the N-terminal region (208 aa) of P1 and P2 of the rhubarb isolate of CLRV are nearly identical among the two genome segments.The taxonomic position of CLRV as member of the genus Nepovirus was confirmed by phylogenetic analyses employing the amino acid sequences of the conserved Pro-Pol region of RNA1, the complete P2, and the CP. However, clustering of Nepovirus-species according to allocated subgroups was inconsistent and depended on the compared genome fragment.     

Complete genome sequence of turnip ringspot virus

Here we present the complete genome sequences of two TuRSV isolates. They are 90–100% identical in distinct genes, but reasonably less identical with RaMV isolates. Regarding the CPs, TuRSV and RaMV have an aa sequence identity of 72–74% among all isolates and the proposed cut-off level is 75%. For the proteinase–polymerase region, the average value between the two isolates of TuRSV and three isolates of RaMV is 79.8% and the cut-off level is 80%. At the moment, TuRSV and RaMV are the two identified species most closely related within the genus Comovirus.     

Complete nucleotide sequence of a severe isolate of cucumber vein yellowing virus from Jordan

The complete genome of a severe isolate of Cucumber vein yellowing virus (CVYV) from Jordan was sequenced. Comparison with the genome of a Spanish CVYV isolate inducing very mild symptoms in cucumber cultivars revealed a nucleotide identity of 94% for the complete genome and an amino acid identity of 96% for the coding region. Comparison of synonymous and non-synonymous substitutions suggested a negative selection at amino acid and nucleotide levels with different degrees depending on the different coding regions. Finally, specific amino acid changes in the zinc finger domain of P1b and in the P1-P3 proteolytic site were found which could be involved in the virulence of CVYV.     

Complete nucleotide sequence of a South African isolate of Grapevine fanleaf virus

The complete sequences of RNA1 and RNA2 have been determined for a South African isolate of Grapevine fanleaf virus (GFLV-SAPCS3). The two RNAs are, respectively, 7,342 and 3,817 nucleotides in length, excluding the poly(A) tails. RNA1 has a large open reading frame (ORF) of 6,852 nucleotides and a 5′-UTR and a 3′-UTR of 243 and 244 nucleotides, respectively. RNA2 encodes for an ORF of 3,330 nucleotides and has the highest nucleotide identity (90.4 %) with GFLV-F13. The full length nucleotide sequence of GFLV-SAPCS3 RNA1 had the highest nucleotide identity (86.5 %) to the French isolate GFLV-F13. The 5′- and 3′-UTRs of GFLV-SAPCS3 RNA2 are 272 nucleotides and 212 nucleotides (nt) in length, respectively. The GFLV-SAPCS3 RNA2 5′-UTR is 32–53 nt longer compared to other GFLV isolates. The GFLV-SAPCS3 RNA2 5′-UTR is also more closely related to GFLV-GHu and Arabis mosaic virus (ArMV) isolates than to other GFLV isolates. Putative intra- and interspecies recombination events between GFLV and ArMV isolates involving GFLV-SAPCS3 RNA1 and RNA2 were investigated. Recombination analysis software has indicated that the GFLV-SAPCS3 5′-UTR might have evolved from a recombinational event between GFLV-F13-type and ArMV-Ta-type isolate.     

Complete nucleotide sequence of a new isolate of passion fruit woodiness virus from Western Australia

We determined the complete genome sequence of the passion fruit woodiness virus Gld-1 isolate (PWV-Gld-1) from Australia and compared it with that of PWV-MU-2, another Australian isolate of PWV. The genomes shared high sequence identity in both the complete nucleotide sequence and the ORF amino acid sequence. All of the cleavage sites of each protein were identical to those of MU-2, and the sequence identity for the individual proteins ranged from 97.2 % to 100.0 %. However, the 5′ untranslated region (5′UTR) of the Gld-1 isolate shared only 46.8 % sequence identity with that of PWV-MU-2 and was 177 nucleotides shorter. Re-sequencing of the 5′UTR of MU-2 revealed that the 5′ end of the original sequence includes an artifact generated by deep sequencing.     

The complete nucleotide sequence of hydrangea ringspot virus

Summary. The complete nucleotide sequence of the genome of the potexvirus, hydrangea ringspot virus, has been determined. The sequence is 6,185 nt in length, excluding the poly (A) tail, and contains six ORFs coding for proteins of 156, 26, 12, 8, 24, and 16 kDa, respectively. ORF 6 is contained within ORF 5 and in this respect the virus is similar to the potexviruses CsCMV, NMV, and SMYEV. Phylogenetic analysis of the putative products of the ORFs and signature motifs contained within these products shows the virus to be most closely related to CsCMV. A similar analysis of data for the coat proteins of potexviruses did not support the previously reported serological relationships between HdRSV and other potexviruses. This is the first complete sequence published for the genome of a potexvirus infecting a dicotyledonous, temperate, deciduous woody species.     

Polypeptide synthesis induced inNicotiana clevelandii protoplasts by infection with raspberry ringspot nepovirus

Summary Infection ofNicotiana clevelandii protoplasts by raspberry ringspot nepovirus resulted in the accumulation of about 24 polypeptides that differed in M_r and pI from polypeptides accumulating in mock-inoculated protoplasts. Similar polypeptides accumulated in protoplasts infected with the S and E strains of RRV but different infection-specific polypeptides were detected in protoplasts infected with tobacco ringspot nepovirus. The M_r of RRV-specific polypeptides ranged from 210 000 to 18 000 and most are presumed to be derived from others by proteolytic cleavage. No evidence was found for marked changes in polypeptide abundance with time after inoculation or for any virus-specific polypeptide becoming disproportionately abundant in the medium during culture.     

The complete nucleotide sequence of RNA 3 of a peach isolate of Prunus necrotic ringspot virus

The complete nucleotide sequence of RNA 3 of the PE-5 peach isolate of Prunus necrotic ringspot ilarvirus (PNRSV) was obtained from cloned cDNA. The RNA sequence is 1941 nucleotides and contains two open reading frames (ORFs). ORF 1 consisted of 284 amino acids with a calculated molecular weight of 31,729 Da and ORF 2 contained 224 amino acids with a calculated molecular weight of 25,018 Da. ORF 2 corresponds to the coat protein gene. Expression of ORF 2 engineered into a pTrcHis vector in Escherichia coli results in a fusion polypeptide of approximately 28 kDa which cross-reacts with PNRSV polyclonal antiserum. Analysis of the coat protein amino acid sequence reveals a putative "zinc-finger" domain at the amino-terminal portion of the protein. Two tetranucleotide AUGC motifs occur in the 3'-UTR of the RNA and may function in coat protein binding and genome activation. ORF 1 homologies to other ilarviruses and alfalfa mosaic virus are confined to limited regions of conserved amino acids. The translated amino acid sequence of the coat protein gene shows 92% similarity to one isolate of apple mosaic virus, a closely related member of the ilarvirus group of plant viruses, but only 66% similarity to the amino acid sequence of the coat protein gene of a second isolate. These relationships are also reflected at the nucleotide sequence level. These results in one instance confirm the close similarities observed at the biophysical and serological levels between these two viruses, but on the other hand call into question the nomenclature used to describe these viruses.