Characterization of a new potyvirus infecting pepper crops in Ecuador

Sequencing 2,951 nucleotides of the 3' proximal region of the genome of a potyvirus isolate collected from Capsicum pubescens (rocoto) pepper in Ecuador revealed that this was the first representative of a new species tentatively named Ecuadorian rocoto virus (ERV). Phylogeny reconstruction showed that this isolate clustered with potato virus V (PVV), Peru tomato virus and wild potato mosaic virus into a monophyletic group, and was closest to PVV. The isolate was shown to be infectious in tobacco, tomato and, contrary to PVV, in pepper. The pvr2(1), pvr2(2), and Pvr4 genes present in many pepper cultivars conferred resistance toward this isolate and could help control ERV.     

Characterization of a new potyvirus isolated from Chinese artichoke in Japan

Summary A filamentous virus was isolated in Japan from virus-infected Chinese artichoke showing mosaic symptoms. This virus was assigned to the genus Potyvirus, based on particle morphology and serological properties. The virus could be transmitted to several test plants but not to Perilla frutesence, the host plant of Perilla mottle virus. cDNA corresponding to the 3-terminal 1675 nucleotides of the viral RNA, excluding the poly (A) tail, was cloned and sequenced. The amino acid sequence of the coat protein was different from those of 74 distinct potyviruses. Therefore, we propose that the new potyvirus should be designated as Chinese artichoke mosaic virus (ChAMV).Received January 15, 2003; accepted June 11, 2003 Published online August 7, 2003     

Molecular characterization of a new potyvirus infecting sunflower

Summary.  We have sequenced 1873 nucleotides from the 3′-end region of a sunflower potyvirus genome including the 3′-NIb protein coding region (813 nucleotides), the entire coat protein coding region (807 nucleotides) and 3′-NCR (253 nucleotides), excluding the poly (A) tail. Amino acids identity of the whole CP between the sunflower virus and Potyvirus members ranged from 49.5% (SCMV) to 81.5% (PVY-NsNr), and the core ranged from 55% (TVMV) to 87% (PVY-NsNr; PepMoV). The 3′-NCR nucleotides showed 38.7% homology to PeSMV and 61% to PepMoV-C. The sequence of 3′end region and analysis of phylogenetic relationships suggest this sunflower virus could belong to PVY subgroup and the name of “sunflower chlorotic mottle virus” (SuCMoV) is proposed. This is the first report on the partial nucleotide sequence of a potyvirus infecting sunflower. Received May 19, 2000/Accepted June 9, 2000     

Molecular characterization of a distinct potyvirus from whitegrass in China

Summary.  A potyvirus isolated from perennial whitegrass (Pennisetum centrasiaticum Tzvel.) in North China was characterized at the molecular level. The 3′ terminal nucleotide (nt) sequence of 1669 nt of the viral RNA genome has been determined, which covered the coding region of the C-terminal part of the large nuclear inclusion protein (NIb, RNA polymerase), capsid protein (CP) gene and the 3′ nontranslated region (NTR). The CP gene consisted of 909 nt (including the stop codon) encoding 302 amino acid residues, and the 3′ NTR was 241 nt in length excluding the polyadenylated tract. Sequence comparison of the amino acids of CPs showed that this virus was most closely related to Sorghum mosaic virus and Maize dwarf mosaic virus with percent identities of 77% to 78% while that of the 3′ NTRs suggested that it was most closely related to Zea mosaic virus with identity of 72%. This virus isolate was to some extent closely related to other members of the Sugarcane mosaic virus subgroup of potyviruses for the CP amino acid sequences. Phylogenetic analyses of the sequences indicated that this virus isolate represented a distinct potyvirus, and the name Pennisetum mosaic virus (PenMV) is proposed. Received November 22, 2002; accepted January 8, 2003 Published online March 21, 2003     

Characterization of a potyvirus isolated from Tradescantia fluminensis in northern Italy

Summary. We isolated a potyvirus from Tradescantia fluminensis that was causing leaf distortion and mild mosaic. We cloned and sequenced a 1500 bp cDNA obtained by RT-PCR corresponding to the 3′ proximal region of the genome. We determined the host range and tested a series of potyviral antisera against our tradescantia virus isolate by immuno-enzymatic methods. Based on our results, we suggest that our viral isolate could be considered a new potyvirus species named Tradescantia mild mosaic potyvirus. Phylogenetic analysis confirmed that Tradescantia mild mosaic virus belongs to the genus Potyvirus within the family Potyviridae, but the virus could not be assigned to any of the potyvirus groupings recently defined.     

Mutational analysis of the NIa protease from pepper vein banding potyvirus

Summary.  The nuclear inclusion protein a (NIa) protease plays an important role in the life cycle of potyviruses by processing the viral polyprotein into functional proteins. For functional characterization, the NIa protease from Pepper vein banding potyvirus (PVBV) was overexpressed in Escherichia coli and purified. Using a recombinant polyprotein substrate containing the nuclear inclusion protein b (NIb)-coat protein (CP) cleavage site, a trans-cleavage assay was developed for the NIa protease. The polyprotein substrate also possessed the cleavage site between NIa and NIb, in addition to the NIb-CP site. However, no trans-cleavage by the NIa protease between NIa and NIb was detected indicating that the cleavage between NIa and NIb under natural conditions would be by a cis-cleavage reaction. Site-specific mutations of the conserved residues D81, D90, C110, T146, C151 and H167 were performed to investigate their roles in the catalytic process of the protease. Such an analysis has revealed that D81 and C151 constitute two of the catalytic triad residues in the NIa protease, D90 and C110 are not essential for catalysis, and T146 and H167 are probably involved in binding to Gln at the P1 position of the substrate. Accepted June 16, 2000 Received April 7, 2000     

A potential new serotype of Riemerella anatipestifer isolated from ducks in Thailand

Eighty isolates of Riemerella anatipestifer representing 71 outbreaks of riemerellosis in Thailand between 1994 and 1999 were serotyped using the gel diffusion precipitin test. Based on the precipitation patterns, 25 serological profiles containing one to three antigenic determinants were recognized. Heat-stable antigens of the organism reacted with antisera raised against 16 known serotypes and an untypable strain 698/95. The most prevalent serotype appeared to be serotype 7, followed by serotypes 5, 10, 21 and 1. Further study demonstrated that the untypable strain probably represents a new serotype. Analysis of the polymerase chain reaction-amplified rrs genes for restriction fragment length polymorphisms verified the inclusion of strain 698/95 within the species R. anatipestifer and supported earlier work excluding strain 670/89, which had originally been designated the reference strain of serotype 20. Therefore, it is suggested that the strain 698/95 could be adopted as a replacement for the reference strain of serotype 20. Attention should be paid to strains with multiple antigenic factors as they may be useful for the preparation of vaccines.     

Molecular characterization of a new lytic bacteriophage isolated from cheese whey

In this study, we isolated and characterized a lytic Lactococcus lactis bacteriophage from the sera of a failed fermentation. The phage was isolated and cultured in L. lactis subsp. cremoris in M17 medium. The isolated bacteriophage was characterized by multiplex PCR, pulsed-field electrophoresis, DNA restriction digestion, analysis of the N-terminal sequence of the phage major structural protein, transmission electron microscopy and sequencing and analysis of a conserved fragment of its genome. Analysis of the viral genome indicates that its genome is composed of a DNA strand of approximately 48?kb in length, and PCR and microscopy confirmed that IL-P1 belongs to the group of 936-type phages in the family Siphoviridae, which is the most abundant type of lactococcal virus in dairy products worldwide. To our knowledge, this is the first report of a virus within this family that has a presumptive genome larger than 40?kb.     

Tomato necrotic ringspot virus, a new tospovirus isolated in Thailand

A new tospovirus isolated from naturally infected tomato plants grown in Nakhon Pathom province (Thailand) was characterized. Infected plants showed symptoms consisting of necrotic spots, necrotic ringspots and stem necrosis. This virus was detected using general antibodies that could recognize watermelon silver mottle virus (WSMoV), capsicum chlorosis virus (CaCV) and melon yellow spot virus (MYSV). However, it did not react with specific monoclonal antibodies (MAbs) to WSMoV and CaCV or a specific MAb to MYSV. The complete nucleotide sequences of S and M RNAs of the virus were determined. They were 3,023 and 4,716 nucleotides in length, respectively, and contained two ORFs in an ambisense arrangement. Sequence analysis indicated that amino acid sequence of the N protein shared 58.2%, 56.0% and 51.8% identity with those of CaCV, WSMoV and MYSV, respectively. The virus was experimentally transmitted by Thrips palmi and Ceratothripoides claratris. Based on our results, we conclude that this tospovirus isolate should be considered a member of a new species. The name tomato necrotic ringspot virus (TNRV) is proposed for this tospovirus.     

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 genome sequence of pepper yellow mosaic virus, a potyvirus, occurring in Brazil

The complete genomic sequence of pepper yellow mosaic virus (PepYMV), a member of the genus Potyvirus, was determined. The sequence was 9745 nucleotides long, excluding the 3' poly(A) tail. The genome contained a large open reading frame encoding a polyprotein of 3085 amino acids, which contained the typically conserved motifs found in members of the genus Potyvirus and an additional P3-PIPO (pretty interesting potyvirus ORF). In a pairwise comparison with other potyvirus sequences, the full genome of PepYMV shared a maximum of 63.84 % nucleotide sequence identity with pepper mottle virus (PepMoV), followed by verbena virus Y (VVY, 62.11 %), potato virus Y (PVY, 62.07 %) and Peru tomato mosaic virus (PTV, 62.00 %). Based upon a phylogenetic analysis, PepYMV was most closely related to PepMoV and PTV, within the PVY subgroup cluster, like most potyviruses isolated in solanaceous hosts in South America.     

A previously undescribed potyvirus isolated and characterized from arborescent Brugmansia

A suspected virus disease was identified from an arborescent Brugmansia x candida Pers. (syn. Datura candida Pers.) tree. The causal agent was aphid transmissible at low rates. Viral particles were purified from infected tobacco tissue, analyzed, and purified virions were inoculated into healthy tobacco plants to recreate the symptoms. The virions had a mean length of 720-729 nm, and infected cells contained inclusion bodies typical of potyvirus infections. Analysis of infected tissues and purified virions with a panel of potyvirus-specific antibodies confirmed identification as a potyvirus. Viral host range, dilution end point, thermal tolerance and aphid transmission characteristics were examined. The viral genome (9761 nt) is typical of potyviruses, with the closest related potyvirus being pepper mottle virus, at 72 % nt sequence identity. Based on conventions for naming novel potyviruses, the virus was determined to be a member of a previously undescribed species, tentatively named “Brugmansia mosaic virus” (BruMV).     

Molecular characterization of Anaplasma platys in a domestic cat from Thailand

A 5-year-old female cat was presented at Kamphaengsaen Animal Hospital for a complication of jaundice. Haematology revealed mild non-regenerative anaemia, lymphopenia, thrombocytopenia, hyper-proteinaemia and icteric plasma. The inclusion bodies in the platelets were detected in a stained blood smear. Partial sequences of the 16S rRNA gene from the isolate indicated that it had the closest relation to Anaplasma platys sequences from many dogs in Thailand and those obtained from GenBank. This is the first report of Anaplasma platys in a naturally infected domestic cat from Thailand.     

Molecular characterisation and evolutionary relationships of a potyvirus infecting Crotalaria in Brazil

Summary.  The 3′ terminal genomic region of a potyvirus causing mosaic disease in several Crotalaria species has been cloned and sequenced. Comparisons of the nucleotide and deduced amino acid (aa) sequences of the cloned cDNA with those from other potyviruses show that the Crotalaria-infecting virus (designated Crotalaria mosaic virus; CrMV) is closely related to Cowpea aphid-borne mosaic virus (CABMV). Maximum identity (95.4%) at the coat protein (CP) aa level was observed between CrMV and a Brazilian strain of CABMV. Phylogenetic analyses derived from the sequence alignments of the CP and 3′ untranslated region confirmed the identification of CrMV as a strain of CABMV and the name CABMV-Cr is suggested. Received April 24, 2001 Accepted October 5, 2001     

Molecular characterization of artemisia virus A, a new sobemovirus isolated from Artemisia annua

A distinct sobemovirus was isolated from diseased Artemisia annua plants grown in experimental culture plots in Switzerland. Electron microscopy performed on extracts of leaf and root samples of a diseased A. annua plant revealed icosahedral-30 nm viral particles. The complete nucleotide sequence of the viral genome was determined. The single positive-strand RNA of 4138 nt encodes four open reading frames with an organization similar to that described for sobemoviruses. Phylogenetic analysis revealed a close relationship to ryegrass mottle virus. The virus was efficiently acquired by healthy A. annua from contaminated soil samples. “Artemisia virus A” is tentatively proposed as a name for this new candidate member of the unassigned genus Sobemovirus.     

New serotypes of Riemerella anatipestifer isolated from ducks in Thailand

Thirty-two Riemerella anatipestifer isolates from ducks were serotyped by agar gel precepitin test using chicken antisera against serotypes 1 to 19 R. anatipestifer reference strains. The heat stable saline extracts from 29 field isolates reacted with the antisera of serotypes 1, 6, 7, 10, 11, 14, or 19. The isolates belonging to serotype 1 were the most prevalent (56.3%). Antigens from the remaining three isolates did not react with any of the available antisera. Additional investigations showed that they represent two new serotypes, serotypes 20 and 21.     

Identification and molecular characterization of a new potyvirus infecting Triteleia species

Plants of Triteleia hyacinthina, Triteleia ixioides Starlight, and Triteleia laxa Corina with severe mosaic and yellow vein-banding were found to be infected with a potyvirus. The 3′-terminal region of the virus was amplified by RT-PCR from total RNA using a potyvirus-specific degenerate primer (poty5P: 5′ GGN AAY AAY AGY GGN CAR CC 3′) and an oligo-dTprimer. The sequence generated included the 3′-NIb protein coding region (680 nucleotides), the entire coat protein coding region (840 nucleotides), and 3’-untranslated region (UTR) (253 nucleotides). Amino acid identity of the whole CP between the triteleia virus and potyvirus member ranged from 54% Apium virus Y (ApVY) to 67% Auraujia mosaic virus (ArjMV) and Twisted-stalk chlorotic streak virus (TSCSV) and the core ranged from 59% (ApVY) to 75% (ArjMV). The 3-UTR showed no significant homology with other known potyviruses. Phylogenetic relationships suggest this triteleia virus is a new member of the Potyvirus genus and the name of “Triteleia mosaic virus” (TriMV) is proposed. This is the first report of a potyvirus infecting triteleia.     

Molecular characterization of a plasmid isolated from Riemerella anatipestifer

Sixty strains of Riemerella anatipestifer were isolated from ducks and geese with infectious serositis in Taiwan. Sixty per cent of the isolates (36/60) contained a 3.9 b plasmid, 12% (7/60) contained 6.5 b and 16 b plasmids, 5% (3/60) contained 2.9, 16 and 18 b plasmids and 13% contained no plasmid (14/60). The 3.9 b plasmid (designated as pCFC1) was completely sequenced to determine if it encoded virulence factors. pCFC1 was 27% G-C and had four large open reading frames (ORF). Two of the ORFs (designated as VapD1 and VapD2) encoded proteins that shared 80, 83, 69 and 67 (VapD1) and 50, 48, 21 and 20% (VapD2) identity with virulence-associated proteins of Actinobacillus Actinomycetemcomitans, Dichelobcater nodosus, Haemophilus influenzae and Neisseria gonorrheae, respectively. pCFC1 also had an ORF (designated as RepAl) that encoded a protein with approximately 30% identity to the RepA proteins of Neisseria gonorrhoeae, Campylobactor hyointestinalis and Pseudomonas aeroginosa. The region upstream of the RepA ORF had an A-T-rich region that was followed by four 21 bp perfect and one 20 bp imperfect direct repeat. The fourth ORF (designated as RepA2) encoded a protein with a region that was 44% homologous to the Helicobactor pylori replication protein.