The complete sequence of the genome of Cocksfoot streak virus (CSV), a grass infecting Potyvirus

Summary.  The complete nucleotide sequence of Cocksfoot streak virus (CSV) has been determined. The viral genome comprises 9663 nucleotides, excluding a 3′-terminal poly(A) sequence. The genome of CSV has a 133 nt 5′-non coding and a 260 nt 3′-non coding region. The RNA of CSV encodes a single polyprotein of 3089 amino acid residues and has a deduced genome organization typical for a member of the family Potyviridae. CSV is transmissible by aphids and has a narrow host range in the Gramineae. It was compared to two potyviruses having monocotyledonous hosts (monocot potyviruses) and several potyviruses infecting dicotyledonous plants (dicot potyviruses). CSV is most closely related to other monocot potyviruses like Maize dwarf mosaic virus (MDMV) and Johnson grass mosaic virus (JGMV), but also closely related to the dicot potyviruses. On the other hand, CSV is less related to monocot viruses from the other genera (Rymovirus, Tritimovirus) within the Potyviridae. Specific motifs, described for potyviral polyproteins, are also present in the polyprotein of CSV. Only two motifs in the HC-Pro – motif involved in long distance movement and motif for HC-Pro self-interaction – were different in comparison to most of the dicot potyviruses. Received September 10, 2001; accepted March 4, 2002 Published online May 24, 2002     

The complete genome sequence of a Passion fruit woodiness virus isolate from Australia determined using deep sequencing, and its relationship to other potyviruses

The complete genome sequence (9,858 nucleotides) of the Passion fruit woodiness virus isolate MU-2 was determined using Illumina sequencing. The large open reading frame (ORF) encodes a polyprotein containing 3,086 amino acids, with an AUG start codon and UAA stop codon. The polyprotein yielded 11 proteins (P1, HC-Pro, P3, PIPO, 6K1, CI, 6K2, NIa-VPg, NIa-Pro, NIb and CP). Putative cleavage sites between them were identified by sequence comparison to those of other known potyviruses. Accuracy of the genome sequence information was provided by 42-1691-fold sequence coverage, and viral RNA accounted for 7.38% of total polyadenylated RNA from the host plant.     

Complete nucleotide sequence of the genomic RNA of a Japanese yam mosaic virus, a new potyvirus in Japan

Summary.  We determined the complete nucleotide sequence of a potyvirus purified from a Japanese yam plant. The genomic RNA of this virus is 9 757 nucleotides (nts) in length, excluding the 3′-terminal poly(A) tail. It contains a single open reading frame (ORF) encoding a polyprotein of 3130 amino acids (aa) with a calculated Mr of 356,793. The genomic organization of this potyvirus is similar to that of other members of the genus Potyvirus and nine potential cleavage sites for the viral proteinase were found by comparison of its sequence with those available for other potyviruses. The nucleotide sequence and genome characteristics show that this isolate is a new potyvirus species. Its polyprotein differs substantially from Yam mosaic virus (YMV) (50% amino acid sequence identity) and fourteen other potyvirus species examined (44–59% identity). Although this potyvirus has been classified as YMV, our results suggest that the potyvirus infectious to the Japanese yam plant in Japan is distinct from YMV. Therefore, we propose that the Japanese yam potyvirus should be designated as Japanese yam mosaic virus (JYMV). September 14, 1998 Received August 3, 1998     

The complete genome sequence of a Brazilian isolate of yam mild mosaic virus

In this study, the complete genome of an isolate of yam mild mosaic virus (YMMV) from Brazil was sequenced, and the predicted amino acid sequence was analyzed. The YMMV RNA genome consists of 9538 nt without the poly(A) tail, encoding a putative typical potyvirus polyprotein of 3084 amino acids. Furthermore, the small overlapping ORF (PIPO) in the P3 gene was also deduced, and the cleavage sites of the polyprotein were predicted. Multiple alignment with other potyviruses showed a maximum nucleotide sequence identity of 64 % to wild tomato mosaic virus. A phylogenetic tree showed that YMMV clustered with Asian potyviruses that mainly infect solanaceous plants.     

Complete nucleotide sequence of Sesbania mosaic virus: a new virus species of the genus Sobemovirus

Summary.  The complete nucleotide sequence of the Sesbania mosaic virus (SeMV) genomic RNA was determined by sequencing overlapping cDNA clones. The SeMV genome is 4149 nucleotides in length and encodes four potential overlapping open reading frames (ORFs). Comparison of the nucleotide sequence and the deduced amino acid sequence of the four ORFs of SeMV with that of other sobemoviruses revealed that SeMV was closest to southern bean mosaic virus Arkansas isolate (SBMV-Ark, 73% identity). The 5′ non-coding regions of SeMV, SBMV and southern cowpea mosaic virus (SCPMV) are nearly identical. However ORF1 of SeMV which encodes for a putative movement protein of M_r 18370 has only 34% identity with SBMV-Ark. ORF 2 encodes a polyprotein containing the serine protease, genome linked viral protein (VPg) and RNA dependent RNA polymerase domains and shows 78% identity with SBMV-Ark. The N-terminal amino acid sequence of VPg was found to be TLPPELSIIEIP, which mapped to the region 326–337 of ORF2 product and the cleavage site between the protease domain and VPg was identified to be E³²⁵-T³²⁶. The cleavage site between VPg and RNA dependent RNA polymerase was predicted to be E⁴⁴⁵-T⁴⁴⁶ based on the amino acid sequence analysis of the polyprotein from different sobemoviruses. ORF3 is nested within ORF2 in a − 1 reading frame. The potential ribosomal frame shift signal and the downstream stem-loop structure found in other sobemoviruses are also conserved in SeMV RNA sequence, indicating that ORF3 might be expressed via − 1 frame shifting mechanism. ORF4 encodes the coat protein of SeMV, which shows 76 and 66% identity with SBMV-Ark and SCPMV, respectively. Thus the comparison of the non-coding regions and the ORFs of SeMV with other sobemoviruses clearly revealed that it is not a strain of SBMV. Phylogenetic analysis of six different sobemoviruses, including SeMV, suggests that recombination event is not frequent in this group and that SeMV is a distinct member of the genus sobemovirus. The analysis also shows sobemoviruses infecting monocotyledons and dicotyledons fall into two distinct clusters. Received April 20, 2000 Accepted August 28, 2000     

The complete nucleotide sequence,genome organization,and specific detection of <Emphasis Type="Italic">Beet mosaic virus</Emphasis>

Summary. Beet mosaic virus (BtMV) was identified almost five decades ago but has not been fully characterized at the molecular level. In this study, we have determined for the first time the complete nucleotide sequence of BtMV genomic RNA and have developed a specific molecular means for its diagnosis. The viral genome of BtMV comprises 9591 nucleotides, excluding the 3 terminal poly (A) sequence, and contains a single open reading frame (ORF) that begins at nt 166 and terminates at nt 9423, encoding a single polyprotein of 3086 amino acid residues. A 3 untranslated region of 168 nucleotides follows the ORF. The deduced genome organization is typical for a member of the family Potyviridae and includes 10 proteins: P1, HC-Pro, P3, 6K1, CI, 6K2, NIa-VPg, NIa-Pro, NIb and coat protein (CP). Nine putative protease cleavage sites were predicted computationally and by analogy with genome arrangements of other potyviruses. Conserved sequence motifs of homologous proteins of other potyviruses were found in corresponding positions of BtMV. BtMV is a distinct species of the genus Potyvirus with the most closely related species being Peanut mottle virus (55% amino acid identity). Based on the nucleotide sequence obtained, we have developed a virus-specific RT-PCR assay for accurate diagnosis and differentiation of BtMV.     

Divergence and conservation of the genomic RNAs of Taiwan and Hawaii strains of papaya ringspot potyvirus

Summary. The complete nucleotide sequence of the genome of a Taiwan isolate of papaya ringspot potyvirus (PRSV YK) was determined from three overlapping cDNA clones and by direct RNA sequencing. Comparison was made with the reported Hawaii isolate of PRSV HA. Both genomes are 10 326 nucleotides long, excluding the poly(A)-tail. They encode a polyprotein of 3 344 amino acids with a 5′ leader of 85 nucleotides and a 3′ non-translated region of 209 nucleotides. The two genomes share an overall nucleotide identity of 83.4% and an amino acid identity of 90.6%. The 3′ non-translated regions show 92.3% identity. The first 23 nucleotides of the leaders are identical, while the remaining parts of the leaders only show 51.6% identity. The P1 protein genes of the two isolates are very different, with 70.9% nucleotide identity and 66.7% encoded amino acids identity. However, the other viral proteins of the two virus isolates are similar, with a 82.5–89.8% nucleotide identity of their genes and 91.2–97.6% amino acid identity, indicating that they are strains of the same potyvirus. Analysis of the ratios of nucleotide differences to the actual amino acid changes revealed that there are only 2.63 nucleotide changes for each amino acid change in the P1 protein, whereas for the other proteins 4.0–16.4 nucleotide changes are required for each amino acid replacement. The P1 protein has 58% of all the differences of polyprotein. The unusual variation in the leader sequences and the P1 proteins suggests that the two PRSV strains were derived from different evolutionary pathways in different geographic areas. Received February 5, 1996 Accepted September 17, 1996     

Nucleotide sequence analysis of the coat protein genes of two Korean isolates of sweet potato feathery mottle potyvirus

Summary.  The coat protein (CP) genes of the genomic RNA of two Korean isolates of sweet potato feathery mottle potyvirus (SPFMV), SPFMV-K1 and SPFMV-K2, were cloned and their complete nucleotide sequences were determined. Sequence comparisons of the two Korean isolates showed 97.8% amino acid identity in the CP cistron, and 79.9% to 99.0% identity with those of 6 other known SPFMV strains. Of 74 amino acid changes totally among the SPFMV strains, 39 changes were located at the N-terminal region. Pairwise amino acid sequence comparison revealed sequence similarities of 48.6 to 70.2% between SPFMV and 20 other potyviruses, indicating SPFMV to be a quite distinct species. Multiple alignment of the CP cistrons from other potyviruses showed that most of the conserved amino acid residues of the genus Potyvirus are well preserved in the corresponding locations. Accepted November 13, 1997 Received September 1, 1997     

The genomic sequence of cowpea aphid-borne mosaic virus and its similarities with other potyviruses

Summary.  The genomic sequence of a Zimbabwe isolate of Cowpea aphid-borne mosaic virus (CABMV-Z) was determined by sequencing overlapping viral cDNA clones generated by RT-PCR using degenerate and/or specific primers. The sequence is 9465 nucleotides in length excluding the 3′ terminal poly (A) tail and contains a single open reading frame (ORF) of 9159 nucleotides encoding a large polyprotein of 3 053 amino acids and predicted Mr of 348. The size of the genome and the encoded polyprotein is in agreement with other potyviruses and contains nine putative proteolytic cleavage sites and motifs conserved in homologous proteins of other potyviruses. The P1 and P3 were the most variable proteins while CI, NIb and CP were the most conserved. Received August 2, 2001; accepted January 15, 2002     

Characterization of the potyviral HC-pro autoproteolytic cleavage site.

The helper component-proteinase (HC-Pro) encoded by potyviruses functions to cleave the viral polyprotein by an autoproteolytic mechanism at the HC-Pro C-terminus. This protein belongs to a group of viral cysteine-type proteinases and has been shown previously to catalyze proteolysis between a Gly-Gly dipeptide. The amino acid sequence requirements surrounding the HC-Pro C-terminal cleavage site of the tobacco etch virus polyprotein have been investigated using site-directed mutagenesis and in vitro expression systems. A total of 51 polyprotein derivatives, each differing by the substitution of a single amino acid between the P5 and P2' positions, were tested for autoproteolytic activity. Substitutions of Tyr (P4), Val (P2), Gly (P1), and Gly (P1') were found to eliminate or nearly eliminate proteolysis. Substitutions of Thr (P5), Asn (P3), and Met (P2'), on the other hand, were permissive for proteolysis, although the apparent processing rates of some polyproteins containing these alterations were reduced. These results suggest that auto-recognition by HC-Pro involves the interaction of the enzymatic binding site with four amino acids surrounding the cleavage site. Comparison of the homologous sequences of five potyviral polyproteins revealed that the residues essential for processing are strictly conserved, whereas the nonessential residues are divergent. The relationship between HC-Pro and other viral and cellular cysteine-type proteinases is discussed.     

Analyses of the complete sequence of the genome of wheat Eqlid mosaic virus,a novel species in the genus <Emphasis Type="Italic">Tritimovirus</Emphasis>

The complete nucleotide sequence of the genome of wheat Eqlid mosaic virus (WEqMV) (excluding the poly A tail) comprised 9636 nucleotides including 5' and 3' noncoding regions of 137 and 172 nt, respectively. It contained a single ORF coding for a polyprotein of 3,109 amino acid residues and had a deduced genome organization typical of members of the family Potyviridae and with proteinase cleavage sites very similar to those of the members of the genus Tritimovirus. Pairwise and multiple alignments and phylogenetic analysis showed that WEqMV is a distinct species in the genus Tritimovirus. WEqMV and Wheat streak mosaic virus (WSMV) shared the greatest nucleotide sequence identity in the NIb and HC-Pro cistrons (63.2% and 60.8%, respectively) and the lowest sequence identity in the P1 and CP cistrons (51.2% and 51.1%, respectively). Sequence identity for the complete genome of WEqMV and WSMV was 56.8% at the nucleotide level and 50.7% at the amino acid level. WEqMV had 57.2% nucleotide identity and 50.6% amino acid identity with Oat necrotic mottle virus and 52.5% nucleotide identity and 45.5% amino acid identity with Brome streak mosaic virus. The relationship of WEqMV with other members of the family Potyviridae was more distant. Structural analysis of WEqMV protein showed presence of potential transmembrane helices in 6k1, 6k2, and P3 proteins.     

Identification and molecular characterization of a new potyvirus from Panax notoginseng

A potyvirus causing distortion and mosaic symptoms in the herbal plant Sanqi (Panax notoginseng) was isolated from Yunnan province, China, and the complete nucleotide sequence of one isolate and the partial sequences of two other isolates were determined. The viral RNA genome comprised 9,750 nt excluding the 3′-terminal poly(A) tail, with the capacity to encode a single polyprotein of 3,089 amino acids. Phylogenetic analysis with other completely sequenced potyviruses revealed that the virus in this study was most closely related to plum pox virus, with 56.3% nt identity in the genomic RNA sequence and 53.3% aa identity in the polyprotein. However, the most closely related 3′-terminal sequences were from four partially sequenced potyviruses infecting plants of the family Apiaceae (67.7–75.3% nt identity and 73.8–76.7% aa identity in their coat protein cistrons), especially Angelica virus Y. These results suggest that this virus isolate should be designated a member of a new species in the genus Potyvirus, which is tentatively named Panax virus Y (PanVY).     

Complete nucleotide sequence of wheat yellow mosaic bymovirus genomic RNAs

Summary.  The complete sequences of wheat yellow mosaic bymovirus (WYMV) RNA1 and RNA2 were determined. RNA1 is 7 636 nucleotides long [excluding the 3′-poly(A)], and codes for a 269 kDa polyprotein of 2 404 amino acids which contains the capsid protein (CP) at the C terminus and seven putative non-structural proteins. RNA2 is 3 659 nucleotides long and codes for a polyprotein of 904 amino acids which contains a 28 kDa putative proteinase and a 73 kDa polypeptide. These functional proteins are arranged as in RNA1 and RNA2 of barley yellow mosaic bymovirus (BaYMV). Comparisons with the sequence reported for the 3′ half of RNA1 of wheat spindle streak mosaic bymovirus (WSSMV) from Southern France show that WYMV and WSSMV have a similar genetic organization. However, WYMV and WSSMV share only 77% amino acid sequence identity in their deduced CPs in spite of their close serological relationship, and 74% nucleotide sequence identity in their 3′ non-coding regions. Thus, the sequence data indicate that WYMV and WSSMV are not strains of the same virus, which has long been suggested, but are distinct virus species within the genus Bymovirus of the family Potyviridae. Accepted December 19, 1997 Received November 14, 1997     

The complete nucleotide sequence and genome organization of barley mild mosaic virus (Na1 strain)

Summary The 5-terminal half of RNA 1 and whole of RNA 2 of barley mild mosaic bymovirus Na1 strain (BaMMV-Na1) were sequenced to give together with the published data its complete genomic sequence. BaMMV-Na1 RNA 1 and RNA 2 consist of 7 263 and 3 516 nucleotides, excluding the 3 poly A tails, respectively. RNA 1 encodes a single large polyprotein of 2 258 amino acids (M_r 256 K), containing eight putative functional proteins, and RNA 2 also encodes one polyprotein of 891 amino acids (M_r 98 K), containing two functional proteins. These functional proteins are arranged in the same manner as in RNA 1 and RNA 2 of barley yellow mosaic bymovirus (BaYMV), and significant amino acid sequence homology (25–58%) exists between the proteins of the two viruses. The BaMMV-Na1 proteins show less amino acid sequence homology (18–32%) with the corresponding proteins of potyviruses or rymoviruses than with those of Ba YMV. Comparisons of the BaMMV-Na1 proteins with the corresponding proteins of other partially sequenced BaMMV isolates show 87–98% amino acid sequence identity. There is 91–94% nucleotide sequence identity between the 3 non-coding regions (NCRs) in RNA 1 or RNA 2 of BaMMV-Na1 and other BaMMV isolates, but only 68–72% identity between the 5 NCRs in RNA 2 of BaMMV-Na1 and other BaMMV isolates.The nucleotide sequence data described in this paper will appear in the DDBJ, EMBL and GenBank nucleotide databases under the accession numbers D83408 (RNA 1) and D83409 (RNA 2).     

Complete genome sequence of the original Taiwanese isolate of sweet potato latent virus and its relationship to other potyviruses infecting sweet potato

The complete genome of sweet potato latent virus (SPLV) was determined to be 10081 nucleotides long excluding the 3’ poly (A) tail. The genome contains a single large open reading frame encoding a polyprotein of 3247 amino acids. Its genomic organization is typical of potyviruses and contains motifs conserved in members of the genus Potyvirus. Pairwise comparisons show that SPLV shares identities of 50.0 %-56.3 % to other potyviruses at the genomic sequence level. Phylogenetic analysis shows that SPLV is closely related to four other sweet potato potyviruses in the sweet potato feathery mottle virus lineage, but it lacks the unique PISPO in the P1 region of those viruses. The genome analyses confirm that SPLV is a distinct sweet potato virus in the genus Potyvirus.     

The complete nucleotide sequence of bean yellow mosaic potyvirus RNA

Summary The complete nucleotide sequence of an Australian strain of bean yellow mosaic virus (BYMV-S) has been determined from cloned viral cDNAs. The BYMV-S genome is 9 547 nucleotides in length excluding a poly(A) tail. Computer analysis of the sequence revealed a single long open reading frame (ORF) of 9 168 nucleotides, commencing at position 206 and terminating with UAG at position 9 374–6. The ORF potentially encodes a polyprotein of 3 056 amino acids with a deduced Mr of 347 409. The 5 and 3 untranslated regions are 205 and 174 nucleotides in length respectively. Alignment of the amino acid sequence of the BYMV-S polyprotein with those of other potyviruses identified nine putative proteolytic cleavage sites. The predicted consensus cleavage site of the BYMV NIa protease was found to differ from that described for other potyviruses. Processing of the BYMV polyprotein at the designated proteolytic cleavage sites would result in a typical potyviral genome arrangement. The amino acid sequences of the putative BYMV encoded proteins were compared to the homologous gene products of twelve individual potyviruses to identify overall and specific regions of amino acid sequence homology.The nucleotide sequence data reported in this paper has been submitted to GenBank nucleotide sequence database and has been assigned the accession number U47033.     

Complete genome sequence of Brugmansia suaveolens mottle virus, a potyvirus from an ornamental shrub

Brugmansia suaveolens mottle virus (BsMoV) was the first potyvirus isolated from “angel trumpet” (Brugmansia suaveolens), described in Brazil. In this study, the complete nucleotide (nt) genome sequence of BsMoV was determined, and the deduced amino acid (aa) sequence was analyzed. The BsMoV RNA genome consists of 9870 nt without a poly-A tail, encoding a putative typical potyviral polyprotein of 3090 aa. Pairwise comparisons of the complete BsMoV genome with those of the most closely related potyviruses revealed a maximum nucleotide identity of 63.7% with pepper mottle virus. These results and phylogenetic analyses based on the complete genome sequence of the most closely related potyviruses confirmed that BsMoV should be considered a member of a distinct species of the genus Potyvirus.     

The complete genomic sequence of <Emphasis Type="Italic">Tobacco vein banding mosaic virus</Emphasis> and its similarities with other potyviruses

The complete genomic sequence of an isolate of Tobacco vein banding mosaic virus (TVBMV-YND) from Yunnan, China was determined by sequencing overlapping cDNA fragments obtained by RT-PCR with degenerate and/or specific primers. The genome is composed of 9,570 nucleotides (nt) excluding the 3′-terminal poly (A) tail and contains one single open reading frame of 9,240 nt encoding a large polyprotein of 3,079 amino acids with predicted Mr of 348.6 kDa. Phylogenetic analysis of complete genomic sequences confirmed that TVBMV is a distinct species of the genus Potyvirus. Different parts of TVBMV-YND genome shared different levels of identity with other species of potyviruses, while most parts showed greatest identity with Chilli veinal mottle virus among the potyviruses with available complete genomic sequences. TVBMV-YND had a rare Q/N cleavage site for NIb/CP and uncommon RITC motif in HC-Pro that is crucial for aphid transmission of potyviruses. Xiao-Qing Yu and Yu-Fei Lan contributed equally to this research     

Complete nucleotide sequence of the geonomic RNA of a mild strain of Japanese yam mosaic potyvirus

Summary.  We determined the complete nucleotide sequence of a mild strain of Japanese yam mosaic potyvirus (JYMV-M) and compared it with the published sequence of severe strain of JYMV (JYMV-J1). The genomic RNA of JYMV-M is 9,760 nucleotides (nts) in length, excluding the poly (A) tail, and encodes a polyprotein of 3,132 amino acids. Among nine potential cleavage sites, only the P1 and NIa recognition sites (between 6K1 and CI) had different sequences from those of JYMV-J1. The data confirm the strain status of these two viruses with 91.1% sequence identity for the polyprotein and ∼94–97% identities for HC, CI, NIa, NIb and CP. The most divergent products P1 and P3 had 62% and 90% sequence identities respectively. Accepted September 27, 1999