Molecular characterization of the genome segments S4, S6 and S7 of rice gall dwarf virus

Summary Rice gall dwarf virus (RGDV) is a member of the genus Phytoreovirus within the family Reovirdae. Its genome has 12 segments of double-stranded RNA (dsRNA), of which the nucleotide sequences of segments S4, S6, and S7 were determined, providing the first complete genome sequence of RGDV. Each of the segments S4, S6, and S7 contained conserved terminal sequences conforming to the RGDV consensus, 5′-GGXA … UGAU-3′ (X = U or C). Each segment had a single predicted open reading frame encoding proteins with calculated molecular weights of 79.8, 58.6, and 53.3 kDa. These proteins appeared to be homologous to those encoded by the corresponding segments of rice dwarf virus and wound tumor virus, the other known members of the same genus, having about 20–30% amino acid identity to them. It is therefore likely that RGDV S4 and S6 encode non-structural proteins and S7 an inner core protein. Probable homologies between the segments of all known phytoreoviruses are summarized. Beyond these similarities, the RGDV proteins displayed no significant similarity to any other reported viral proteins.     

Molecular characterisation of segments 1 to 6 of Rice black-streaked dwarf virus from China provides the complete genome

Summary.  The nucleotide sequences of segments S1 to S6 of a Chinese isolate of Rice black-streaked dwarf virus (RBSDV) were determined. This provides the first complete sequence of a plant pathogenic member of the genus Fijivirus. The complete ten-segment genome has 29141 nucleotides, making it the largest reovirus genome so far reported. Each of the segments S1–S6 is predicted to encode a single major protein. Protein comparisons indicated that S1 encoded an RNA dependent RNA polymerase, with similarities to that encoded by S1 of Nilaparvata lugens reovirus (NLRV). S2 and S3 appeared to be homologous to S3 and S4 respectively of both Fiji disease virus (FDV) and NLRV. The protein encoded on S4 showed some similarity to that of NLRV S2. The proteins encoded on S5 and S6, though similar in size to those of NLRV S5 and S6, had no detectable homologies to them or to any other known protein. Received April 4, 2001 Accepted August 1, 2001     

Molecular characterization of the largest and smallest genome segments,S1 and S12, of <Emphasis Type="Italic">Rice gall dwarf virus</Emphasis>

The nucleotide sequences of segments S1 and S12 of a Chinese isolate of Rice gall dwarf virus (RGDV) were determined. This provides the first complete sequences of these segments. The complete sequence of S1, the largest genome segment of RGDV, was 4,505 nucleotides in length and was predicted to encode a large protein of 1,458 amino acids with a calculated molecular mass of nearly 166.2 kDa. The protein was related to that encoded by S1 of Rice dwarf virus (RDV; 50% identity and 67% similarity) and (to a lesser extent) to some large proteins of other reoviruses. It appears to be an RNA-dependent RNA polymerase (RdRp) and is probably present in particles as a minor core protein. S12, the smallest genome segment of RGDV, was 853 nucleotides in length, encoding a single major protein of 206 amino acids with a calculated molecular mass of nearly 23.6 kDa. This protein, though a little larger than those of RDV S11 and Wound tumor virus (WTV) S12 in size, showed some similarity to them, especially in the conserved N-terminal region and may have RNA-binding properties. Despite having a common host plant, RDV and RGDV were not more closely related to one another than either of them was to WTV. Phylogenetic analysis of the RdRp showed that members of the genus Phytoreovirus were more closely related to those of the genus Rotavirus than to any other genus within the family Reoviridae. The nucleotide sequence data reported in this article have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers DQ333946 and DQ494209.     

Sequence analysis of rice dwarf phytoreovirus genome segments S4, S5, and S6: comparison with the equivalent wound tumor virus segments

The complete nucleotide sequences of genome segments S4, S5, and S6 of rice dwarf phytoreovirus (RDV) were determined. S4 and S5 consist of 2468 and 2570 base pairs, respectively, S5 thus being larger in size than S4, contrary to the situation suggested by their relative migration in a polyacrylamide gel. S6 is 1699 nucleotides long. The individual segments have segment-specific inverted repeats adjacent to the conserved terminal sequences (5'GGUAAA---UGAU3' for S4, 5'GGCAAA---UGAU3' for S5 and S6). S4, S5, and S6 each have single long open reading frames encoding 727, 801, and 509 amino acids, respectively. A low level of amino acid sequence homology was observed between RDV S4 and wound tumor virus (WTV) S4 (22.4%), and between RDV S6 and WTV S6 (20.2%). On the other hand, RDV S5 and WTV S5 show 52.0% amino acid sequence similarity, indicating that S5 is much more conserved than any other segments of RDV and WTV reported so far. Further comparative analyses indicate that the RDV segment shows a greater frequency of usage of codons XYG and XYC, and much less frequent usage of codon XYA than the equivalent WTV segment, this codon preference bias being more conspicuous than expected from the base contents.     

Completion of the sequence of rice gall dwarf virus from Guangxi,China

The complete nucleotide sequences of segments S2, S3, S5, and S8-S11 of a Rice gall dwarf virus isolate from Guangxi Province, China, (RGDV-GX) were determined, completing the sequence of this isolate. The total genome (25,567nt) was similar in organization to a recently reported Thailand isolate (RGDV-T). A previously unreported second segment-specific repeat of 7 or 8nt was detected close to the 3'-end of segments S3, S5, and S8. The 3'-UTR of RGDV-GX S4 was 139nt shorter than that of RGDV-T; the insertion in RGDV-T contains a 34-nt inverted repeat, with the 3'-terminus probably abolishing the expected stem loop structure.     

Bluetongue virus evolution: sequence analyses of the genomic S1 segments and major core protein VP7.

The S1 segments, encoding the group-specific antigen, VP7, from the five United States prototype BTV serotypes were cloned as full-length entities. The nucleotide and deduced amino acid sequences of segment S1 of BTV-2 were determined and compared with BTV-10, -11, -13, and -17, completing the sequencing of this cognate gene segment from all five US BTV serotypes. Each segment is 1156 bp long and contains an open reading frame encoding the 349-amino acid VP7 protein. Most (greater than 94%) of the amino acids of VP7 among the serotypes are conserved, including the location (position 255) of a single lysine residue. Secondary structure analyses of VP7 predict a putative eight-stranded beta-barrel between amino acid positions 150 and 250, a structure similar to that observed in ssRNA viruses. The S1 genes are flanked by conserved 5' and 3' noncoding regions. Stem-loop structures are predicted at the 3' end of each gene (nucleotide positions 1058-1097). The S1 segments of BTV-2, -10, -11, and -17 have greater than 93% of the nucleotides conserved, while less than 80% of their bases are identical with BTV-13. Analyses of nucleotide mismatches in each codon position of the VP7 open reading frame, transition frequencies, and evolutionary distances show that of the five, BTV-13 is the most distantly related and that BTV-10 and -17 are the most closely related serotypes. Evolutionary distance calculations of segment L2 from BTV-10, -11, and -17 concur with these observations. Comparison of this relationship with hybridization data of segment M3, which codes for VP5, suggests that BTV-17 has evolved by a combination of genetic drift and genomic reassortment. The data also indicate that the five US BTV serotypes are derived from two distinct gene pools. Evolution distances were used to estimate an evolution rate of 2.2 x 10(-3) nucleotide substitution/site/year for BTV segment S1. This rate is similar to the genes of retroviruses and implies an absence of RNA polymerase proofreading activity for dsRNA viruses.     

Nucleotide sequence of segment S 9 of the rice dwarf virus genome

Summary The complete nucleotide sequence of genome segment S 9, which was reported as the ninth largest among the 12 genome segments of rice dwarf virus, was determined. The segment was 1,305 bp long and coded for a protein composed of 351 amino acids (M_r 38,930). The terminal sequences of the segment were found to consist of an imperfect 14 base pair inverted repeat located at positions 2–15 and 1289–1302, which is similar to that identified in other reoviruses.     

Nucleotide sequences of genome segments S6, S7 and S10 of Dendrolimus punctatus cypovirus 1

Summary. The nucleotide sequences of genome segments S6, S7 and S10 of Dendrolimus punctatus cypovirus 1 Hunan I (DpCPV-HN(I)) and DpCPV-HN(I)-Se₃ (DpCPV-HN(I) passed three times in Spodoptera exigua) were determined. Segment S10 was 944 nucleotides in length and encoded a polyhedrin of 248 amino acids (28 439Da). Only two nucleotide mutations were found between DpCPV-HN(I) S10 and DpCPV-HN(I)-Se₃ S10, and the deduced amino acid sequences of the polyhedrin proteins were identical. Segment S7, 1 501 nucleotides, encoded a protein of 448 amino acids (50kDa; p50). Thirty-one nucleotide mutations were found between DpCPV-HN(I) S7 and DpCPV-HN(I)-Se₃ S7, but these resulted in only four amino acid changes. DpCPV-HN(I) S6 encoded a protein of 561 amino acids (63 688Da; p64). The amino acid sequence of p64, had a high leucine content (10%), and contained a leucine zipper motif and one ATP/GTP-binding site motif.     

Nucleotide sequences of genome segments S6, S7 and S10 of Dendrolimus punctatus cypovirus 1

The nucleotide sequences of genome segments S6, S7 and S10 of Dendrolimus punctatus cypovirus 1 Hunan I (DpCPV-HN(I)) and DpCPV-HN(I)-Se(3) (DpCPV-HN(I) passed three times in Spodoptera exigua) were determined. Segment S10 was 944 nucleotides in length and encoded a polyhedrin of 248 amino acids (28,439 Da). Only two nucleotide mutations were found between DpCPV-HN(I) S10 and DpCPV-HN(I)-Se3 S10, and the deduced amino acid sequences of the polyhedrin proteins were identical. Segment S7, 1 501 nucleotides, encoded a protein of 448 amino acids ( approximately 50 kDa; p50). Thirty-one nucleotide mutations were found between DpCPV-HN(I) S7 and DpCPV-HN(I)-Se3 S7, but these resulted in only four amino acid changes. DpCPV-HN(I) S6 encoded a protein of 561 amino acids (63,688 Da; p64). The amino acid sequence of p64, had a high leucine content (10%), and contained a leucine zipper motif and one ATP/GTP-binding site motif.     

Complete genomic sequence of a Tobacco rattle virus isolate from Michigan-grown potatoes

Tobacco rattle virus (TRV) causes stem mottle on potato leaves and necrotic arcs and rings in potato tubers, known as corky ringspot disease. Recently, TRV was reported in Michigan potato tubers cv. FL1879 exhibiting corky ringspot disease. Sequence analysis of the RNA-1-encoded 16-kDa gene of the Michigan isolate, designated MI-1, revealed homology to TRV isolates from Florida and Washington. Here, we report the complete genomic sequence of RNA-1 (6,791 nt) and RNA-2 (3,685 nt) of TRV MI-1. RNA-1 is predicted to contain four open reading frames, and the genome structure and phylogenetic analyses of the RNA-1 nucleotide sequence revealed significant homologies to the known sequences of other TRV-1 isolates. The relationships based on the full-length nucleotide sequence were different from than those based on the 16-kDa gene encoded on genomic RNA-1 and reflect sequence variation within a 20–25-aa residue region of the 16-kDa protein. MI-1 RNA-2 is predicted to contain three ORFs, encoding the coat protein (CP), a 37.6-kDa protein (ORF 2b), and a 33.6-kDa protein (ORF 2c). In addition, it contains a region of similarity to the 3′ terminus of RNA-1, including a truncated portion of the 16-kDa cistron. Phylogenetic analysis of RNA-2, based on a comparison of nucleotide sequences with other members of the genus Tobravirus, indicates that TRV MI-1 and other North American isolates cluster as a distinct group. TRV M1-1 is only the second North American isolate for which there is a complete sequence of the genome, and it is distinct from the North American isolate TRV ORY. The relationship of the TRV MI-1 isolate to other tobravirus isolates is discussed.     

Complete nucleotide sequence and analysis of the putative polyprotein of maize dwarf mosaic virus genomic RNA (Bulgarian isolate)

Summary.  The complete nucleotide sequence of maize dwarf mosaic virus Bulgarian isolate (MDMV-Bg) was determined. The viral genome was 9515 nt and contained an open reading frame encoding 3042 amino acids, flanked by 3′- and 5′-UTRs of 139 and 250 nucleotides, respectively. MDMV-Bg was more conserved in the coding region (52.9%) than in the UTRs (45.8%) when compared to the 15 other potyviruses. Of ten putative gene products of MDMV-Bg, the P1 was the most variable protein (24.9%) while the NIb was the most conserved protein (67.3%). Several sequence variations were observed between MDMV-Bg and Johnson grass mosaic virus (JGMV), and more between MDMV-Bg and the dicot potyviruses. Phylogenetic analysis suggested that MDMV-Bg was the most closely related to JGMV. Received January 15, 1998 Accepted April 29, 1998     

Complete genome sequence of a Watermelon silver mottle virus isolate from China

The complete genome of a Watermelon silver mottle virus (WSMoV) (genus Tospovirus, family Bunyaviridae) isolate (WSMoV-GZ) from Guangdong province, China was sequenced. The genomes of WSMoV-GZ contained 3,603, 4,909, and 8,914 nt of small (S), medium (M), and large (L) RNA segments, respectively, and had a genomic organization characteristic of members of the genus Tospovirus. The amino acid sequence of the nucleocapsid (N) protein, S RNA-encoded nonstructural (NSs) protein, M RNA-encoded nonstructural (NSm) protein, Gn/Gc glycoprotein precursor, and RNA-dependent RNA polymerase (RdRp) protein showed 94.3–97.5 % identity with those of other WSMoV isolates. Phylogenetic analysis showed that the N protein of WSMoV-GZ was clustered together with those of the WSMoV isolates. The full sequence of WSMoV-GZ provides a reference genome for comparison with other tospoviruses.     

Nucleotide sequence of the genome of a citrus isolate of olive latent virus 1

Summary The 3699 nt genome of olive latent virus 1 (OLV-1), described years ago from Southern Italy as a putative sobemovirus, was completely sequenced. OLV-1 genomic RNA was not polyadenylated and had a structure virtually identical to that of species of theNecrovirus rather than theSobemovirus genus. Five open reading frames (ORFs) were identified, of which the 5-proximal encoded a 23 K protein and ended with an amber codon whose readthrough could yield a putative 82 K product. This polypeptide had extensive sequence similarity with polymerases of serotypes A and D of tobacco necrosis necrovirus (TNV-A and TNV-D) and species of the familyTombusviridae and related genera (Dianthovirus andMachlomovirus). Two small ORFs followed, which encoded polypeptides of 8 K and 6 K, respectively. The 6 K product had extensive homology with the comparable 6 K protein of TNV-A and was also related to the 11 K protein of shallot latent carlavirus, one of the triple block polypeptides involved in cell-to-cell virus movement. The 3-proximal ORF was in the same position as the coat protein (CP) cistron of necroviruses and encoded a 30 K product related to CP of both TNV-A and -D. Computer-assisted comparative analysis of structural and non-structural proteins of OLV-1, TNV-A and TNV-D disclosed an overall distant relationship between OLV-1 and TNV-D. OLV-1 genome appeared homologous to that of TNV-A, but differences from TNV-A were the absence of the small ORF downstream of the CP cistron and in the low degree of sequence identity in CP (39% aa identity). OLV-1 is serologically distantly related to TNV-A and even more distantly related to TNV-D. We propose that OLV-1 is a necrovirus species in its own right.The sequence data reported in this paper have been deposited in the EMBL database and given the accession number X85989.     

Molecular characterization of the full-length genome of a rabies virus isolate from India

Rabies is an important public health problem in South East Asia, with cases in this part of the world contributing to about 70% of the global burden. A large number of rabies cases occur in India, however, there is no organized system of surveillance and hence there is a lack of reliable data. Moreover, comprehensive molecular epidemiological studies have not been performed on Indian virus isolates. In this study, we determined the complete nucleotide and deduced amino acid sequence of a primary isolate of rabies virus obtained from the brain of an infected patient. Comparison of the genomic sequence with those of the ten fully sequenced rabies strains available in GenBank showed nucleotide homology ranging from 97% with AY956319 to 81% with AY705373. Amino acid homology of nucleoprotein ranged from 99.7% with AY352493 to 92% with DQ875051. In case of the glycoprotein gene, the homology ranged from 98.8% with AY956319 to 87.2 % with AY705373. An extensive nucleoprotein, glycoprotein, and full-length genome-based phylogenetic analysis was performed along with sequences available from the GenBank. Phylogenetic analysis of the complete genome sequence indicated that this isolate exhibited close homology with the ex Indian strain AY956319. Primer sequences and the scheme of amplification can be availed from the authors on request.     

Complete genome sequence of a dahlia common mosaic virus isolate from New Zealand

Dahlia mosaic disease of the ornamental flowering plant Dahlia is caused by two caulimoviruses, dahlia mosaic virus (DMV) and dahlia common mosaic virus (DCMV). We used a rolling-circle amplification method to amplify, clone and determine for the first time the full genome sequence of a DCMV isolate from New Zealand (DCMV-NZ). Within the 7949-bp circular double-stranded retro-transcribing DCMV-NZ DNA, we identified six putative open reading frames, typical of all genomes in the family Caulimoviridae. The availability of the complete DCMV sequence provides a reference genome against which all others can be compared.     

Complete genome sequence of a recombinant isolate of yambean mosaic virus from Canavalia ensiformis

Virus Genes - The complete genome sequence of a Brazilian isolate of yambean mosaic virus (YBMV) is presented. High-throughput sequencing (Illumina HiSeq) and Sanger sequencing revealed the...     

The complete nucleotide sequence of a capsicum chlorosis virus isolate from Lycopersicum esculentum in Thailand

Summary. The complete nucleotide sequence of a tospovirus isolated from Lycopersicum esculentum in Thailand was determined. The L RNA comprises of 8912 nt and codes for the RNA-dependent RNA-polymerase (RdRp) (2877 aa). Two ORFs are located on the M RNA (4823 nt) encoding the non-structural (NSm) protein (308 aa) and the viral glycoprotein precursors (Gn/Gc) (1121 aa) separated by an intergenic region of 433 nt. ORFs coding for the non-structural (NSs) and nucleocapsid (N) protein, 439 aa and 275 aa, respectively, were identified on the S RNA (3477 nt) separated by an intergenic region of 1202 nt. The N protein of the Thailand isolate was most closely related to that of capsicum chlorosis virus (CaCV), sharing an amino acid sequence identity of 92.7%. Additionally, multiple sequence analyses revealed significant similarities to tospoviruses of the species Watermelon silver mottle virus and to several putative tospovirus entries in GenBank. Based on these alignments it is proposed to refer to all these different viruses as isolates of CaCV.