Complete nucleotide sequence of rose yellow mosaic virus, a novel member of the family Potyviridae

The complete genomic sequence of rose yellow mosaic virus (RoYMV) was determined and found to have all the features that are characteristic of members of the family Potyviridae. The RoYMV genome is 9508 nucleotides long excluding the 3′-poly-(A) tail and contains a single open reading frame encoding a polyprotein of 3067 amino acids. The RoYMV P3 and CI cistrons are shorter than those of other members of the family Potyviridae, and the 6K1 cistron is completely absent. Comparative sequence analysis revealed that RoYMV had highest amino acid sequence identity across the entire genome sequence to brome streak mosaic virus (33 %) and to turnip mosaic virus (30 %) at the coat protein level. Based on its low sequence similarity to known members of the family Potyviridae and phylogenetic analysis, RoYMV appears to be a distinct, previously undescribed, member of this family.     

Complete sequence and genetic characterization of Raspberry latent virus, a novel member of the family Reoviridae

A new virus isolated from red raspberry plants and detected in the main production areas in northern Washington State, USA and British Columbia, Canada was fully sequenced and found to be a novel member of the family Reoviridae. The virus was designated as Raspberry latent virus (RpLV) based on the fact that it is symptomless when present in single infections in several Rubus virus indicators and commercial raspberry cultivars. RpLV genome is 26,128 nucleotides (nt) divided into 10 dsRNA segments. The length of the genomic segments (S) was similar to those of other reoviruses ranging from 3948 nt (S1) to 1141 nt (S10). All of the segments, except S8, have the conserved terminal sequences 5'-AGUU----GAAUAC-3'. A point mutation at each terminus of S8 resulted in the sequences 5'-AGUA----GAUUAC-3'. Inverted repeats adjacent to each conserved terminus as well as stem loops and extended pan handles were identified by analyses of secondary structures of the non-coding sequences. All segments, except S3 and S10, contained a single open reading frame (ORF) on the positive sense RNAs. Two out-of-frame overlapping ORFs were identified in segments S3 (ORF S3a and S3b) and S10 (ORF S10a and S10b). Amino acid (aa) alignments of the putative proteins encoded by the main ORF in each segment revealed a high identity to several proteins encoded by reoviruses from different genera including Oryzavirus, Cypovirus, and Dinovernavirus. Alignments of the polymerase, the most conserved protein among reoviruses, revealed a 36% aa identity between RpLV and Rice ragged stunt virus (RRSV), the type member of the genus Oryzavirus, indicating that these two viruses are closely related. Phylogenetic analyses showed that RpLV clusters with members of the genera Oryzavirus, Cypovirus, Dinovernavirus and Fijivirus. These genera belong to the subfamily Spinareovirinae which includes reoviruses with spiked core particles ('turreted' reoviruses). In addition, two nucleotide binding motifs, regarded as 'signature' sequences among turreted reoviruses, were also found in RpLV P8, suggesting that RpLV is a novel dicot-infecting reovirus in the subfamily Spinareovirinae.     

Complete nucleotide sequence of Nootka lupine vein-clearing virus

The complete genome sequence of Nootka lupine vein-clearing virus (NLVCV) was determined to be 4,172 nucleotides in length containing four open reading frames (ORFs) with a similar genetic organization of virus species in the genus Carmovirus, family Tombusviridae. The order and gene product size, starting from the 5'-proximal ORF consisted of: (1) polymerase/replicase gene, ORF1 (p27) and ORF1RT (readthrough) (p87), (2) movement proteins ORF2 (p7) and ORF3 (p9), and, (3) the 3'-proximal coat protein ORF4, (p37). The genomic 5'- and 3'-proximal termini contained a short (59 nt) and a relatively longer 405 nt untranslated region, respectively. The longer replicase gene product contained the GDD motif common to RNA-dependent RNA polymerases. Phylogenetically, NLVCV formed a subgroup with the following four carmoviruses when separately comparing the amino acids of the coat protein or replicase protein: Angelonia flower break virus (AnFBV), Carnation mottle virus (CarMV), Pelargonium flower break virus (PFBV), and Saguaro cactus virus (SgCV). Whole genome nucleotide analysis (percent identities) among the carmoviruses with NLVCV suggested a similar pattern. The species demarcation criteria in the genus Carmovirus for the amino acid sequence identity of the polymerase (<52%) and coat (<41%) protein genes restricted NLVCV as a distinct species, and instead, placed it as a tentative strain of CarMV, PFBV, or SgCV when both the polymerase and CP were used as the determining factors. In contrast, the species criteria that included different host ranges with no overlap and lack of serology relatedness between NLVCV and the carmoviruses, suggested that NLVCV was a distinct species. The relatively low cutoff percentages allowed for the polymerase and CP genes to dictate the inclusion/exclusion of a distinct carmovirus species should be reevaluated. Therefore, at this time we have concluded that NLVCV should be classified as a tentative new species in the genus Carmovirus, family Tombusviridae.     

Complete nucleotide sequence of rose yellow vein virus, a member of the family Caulimoviridae having a novel genome organization

This report describes the complete nucleotide sequence and genome organization of rose yellow vein virus (RYVV), a proposed new member of the family Caulimoviridae. The RYVV genome is 9314 bp in size and contains eight open reading frames (ORFs). ORFs 1, 2, and 3 have 22–38 % amino acid sequence similarity to known members of the family Caulimoviridae. The remaining ORFs have no significant amino acid sequence similarity to known viruses. Based on differences in its genome organization, its low sequence similarity to known members of the family Caulimoviridae, and the results of phylogenetic analysis, RYVV appears to be a distinct new member of this family.     

Complete nucleotide sequence of rose yellow leaf virus,a new member of the family Tombusviridae

The genome of the rose yellow leaf virus (RYLV) has been determined to be 3918 nucleotides long and to contain seven open reading frames (ORFs). ORF1 encodes a 27-kDa peptide (p27). ORF2 shares a common start codon with ORF1 and continues through the amber stop codon of p27 to encode an 87-kDa (p87) protein that has amino acid similarity to the RNA-dependent RNA polymerase (RdRp) of members of the family Tombusviridae. ORFs 3 and 4 have no significant amino acid similarity to known functional viral ORFs. ORF5 encodes a 6-kDa (p6) protein that has similarity to movement proteins of members of the Tombusviridae. ORF5A has no conventional start codon and overlaps with p6. A putative +1 frameshift mechanism allows p6 translation to continue through the stop codon and results in a 12-kDa protein that has high homology to the carmovirus p13 movement protein. The 37-kDa protein encoded by ORF6 has amino acid sequence similarity to coat proteins (CP) of members of the Tombusviridae. ORF7 has no significant amino acid similarity to known viral ORFs. Phylogenetic analysis of the RdRp amino acid sequences grouped RYLV together with the unclassified Rosa rugosa leaf distortion virus (RrLDV), pelargonium line pattern virus (PLPV), and pelargonium chlorotic ring pattern virus (PCRPV) in a distinct subgroup of the family Tombusviridae.     

Complete sequence and genetic characterization of pigeon avian nephritis virus, a member of the family Astroviridae

In the current study, the complete genome sequence of a member of the family Astroviridae isolated from pigeons was determined through genetic characterization and phylogeny analysis. The isolated genome sequence was proposed to be that of pigeon avian nephritis virus (ANV), whose genome structure and characteristics were similar to previously reported avian astroviruses. The sequenced ssRNA genome comprises 6928 nucleotides, excluding the poly(A) tail, and contains three open reading frames. Phylogenetic analysis using a partial nucleotide sequence of the polymerase gene and the entire amino acid sequence of the full-length capsid protein revealed that pigeon avian nephritis virus is closely related to the previously published ANV, especially to the Japanese G-4260 and Chinese strains. This investigation provides information on the sequence and genetic characteristics of this virus and contributes to a better understanding of pigeon ANV and the possible occurrence of astrovirus transmission between chickens and pigeons.     

Complete nucleotide sequence of Rosa rugosa leaf distortion virus, a new member of the family Tombusviridae

This report describes the complete nucleotide sequence and genome organization of Rosa rugosa leaf distortion virus (RrLDV), the causal agent of a previously undescribed virus disease of Rosa rugosa. The RrLDV genome is a positive-sense ssRNA, 3971 nucleotides in length, containing five open reading frames (ORFs). ORF1 encodes a 27-kDa peptide (p27). ORF2 shares a common start codon with ORF1 and continues through the amber stop codon of p27 to produce an 87-kDa protein (p87) with amino acid sequence similarity to the RNA-dependent RNA polymerases (RdRp) of members of the family Tombusviridae. ORF3 encodes a protein of 8 kDa with no significant similarity to known viral sequences. ORF4 encodes a 6-kDa protein (p6) with similarity to the p13 movement proteins of members of the family Tombusviridae. ORF5 has no conventional start codon and overlaps with p6. A putative +1 frame shift mechanism allows p6 translation to continue through the stop codon and results in a 12-kDa protein with high homology to the carmovirus p13 movement protein. The 37-kDa protein encoded by ORF6 has amino acid sequence similarity to coat proteins (CPs) of members of the family Tombusviridae. Phylogenetic analyses of the RdRp and CP amino acid sequences placed RrLDV in a subgroup close to members of the genus Carmovirus of the family Tombusviridae.     

Full-length genome sequence of Mossman virus, a novel paramyxovirus isolated from rodents in Australia

Mossman virus (MoV) was isolated on two occasions from wild rats trapped in Queensland, Australia, during the early 1970s. Together with Nariva virus and J-virus MoV belongs to a group of novel paramyxoviruses isolated from rodents during the last 40 years, none of which had been characterized at the molecular level until now. cDNA subtraction strategies used to isolate virus-specific cDNA derived from both MoV-infected cells and crude MoV pellets were pivotal steps in rapid characterization of the complete genome sequence. Analysis of the full-length genome and its encoded proteins confirmed that MoV is a novel member of the subfamily Paramyxovirinae which cannot be assigned to an existing genus. MoV appears to be more closely related to another unclassified paramyxovirus Tupaia paramyxovirus (TPMV), isolated from the tree shrew Tupaia belangeri. Together with Salem virus (SalV), a further unclassified paramyxovirus that was isolated from a horse, MoV and TPMV make up a new collection of paramyxoviruses situated evolutionally between the genus Morbillivirus and the newly established genus Henipavirus.     

Complete genome sequence of Tunisvirus,a new member of the proposed family Marseilleviridae

Marseillevirus is the founding member of the proposed family Marseilleviridae, which is the second discovered family of giant viruses that infect amoebae. These viruses have been recovered from environmental water samples and, more recently, from humans. Tunisvirus was isolated from fountain water in Tunis, Tunisia, by culturing on Acanthamoeba spp. and is a new marseillevirus. We describe here its 380,011 base-pair genome. A total of 484 proteins were identified, among which 320 and 358 have an ortholog in Marseillevirus and Lausannevirus (e-value <1e-2), respectively, and 259 and 299 have best reciprocal hits with a Marseillevirus and a Lausannevirus protein, respectively. In addition, a significant hit was found in organisms other than marseilleviruses for 144 Tunisvirus proteins, indicating extensive lateral gene transfers, as has been demonstrated previously for Marseillevirus. Finally, a total of 21 ORFans were identified. Phylogeny reconstructions and analysis of the gene repertoires of marseilleviruses, including the proportion of orthologs and the mean amino acid identity between genes in pairs, suggest that the proposed family Marseilleviridae encompasses three lineages. Lineage A is composed of Marseillevirus, Cannes 8 virus and Senegalvirus; lineage B is represented by Lausannevirus alone; and lineage C has Tunisvirus as its first member. Taken together, these findings suggest that the marseilleviruses display a substantial level of diversity.     

Detection of potentially pathogenic bacteria in the drinking water distribution system of a hospital in Hungary

The drinking water distribution system of a hospital was investigated using standard cultivation techniques, taxon-specific PCRs targeting pathogenic bacteria, denaturing gradient gel electrophoresis, cloning and sequencing. The results obtained verify the higher sensitivity of PCR compared to cultivation for detecting Legionella and Pseudomonas aeruginosa . Moreover, several other opportunistic pathogenic bacteria, such as Escherichia albertii , Acinetobacter lwoffi and Corynebacterium tuberculostrearicum , were detected, emphasizing that drinking water systems, especially those with stagnant water sections, could be the source of nosocomial infections.     

Morphology and genome organization of the virus PSV of the hyperthermophilic archaeal genera Pyrobaculum and Thermoproteus: a novel virus family, the Globuloviridae

A novel virus, termed Pyrobaculum spherical virus (PSV), is described that infects anaerobic hyperthermophilic archaea of the genera Pyrobaculum and Thermoproteus. Spherical enveloped virions, about 100 nm in diameter, contain a major multimeric 33-kDa protein and host-derived lipids. A viral envelope encases a superhelical nucleoprotein core containing linear double-stranded DNA. The PSV infection cycle does not cause lysis of host cells. The viral genome was sequenced and contains 28337 bp. The genome is unique for known archaeal viruses in that none of the genes, including that encoding the major structural protein, show any significant sequence matches to genes in public sequence databases. Exceptionally for an archaeal double-stranded DNA virus, almost all the recognizable genes are located on one DNA strand. The ends of the genome consist of 190-bp inverted repeats that contain multiple copies of short direct repeats. The two DNA strands are probably covalently linked at their termini. On the basis of the unusual morphological and genomic properties of this DNA virus, we propose to assign PSV to a new viral family, the Globuloviridae.     

Mild variable Noonan syndrome in a family with a novel PTPN11 mutation

Noonan syndrome (OMIM 163950) is a common genetic condition with variable clinical expression and genetic heterogeneity. About half of the cases can be accounted to activating mutations in the PTPN11 gene encoding SHP-2. We report on a family with mild, variable expression of Noonan syndrome in five individuals. Clinical manifestations included short stature, craniofacial anomalies and thorax deformity, but none of the affected family members had a heart defect. Sequencing of the entire coding region of PTPN11 revealed a novel mutation c.1226G-->C in exon 11 predicting the amino acid exchange G409A. This mutation is not located in the previously known mutation clusters. Our observation and the recent report of a mutation affecting a neighbouring residue (T411M) in a family with a variable phenotype suggest that mutations in this particular region of SHP-2 may have effects on the protein that differ from those of the classical mutations.     

Complete nucleotide sequence of the temperate bacteriophage LBR48, a new member of the family Myoviridae

The complete genomic sequence of LBR48, a temperate bacteriophage induced from a lysogenic strain of Lactobacillus brevis, was found to be 48,211 nucleotides long and to contain 90 putative open reading frames. Based on structural characteristics obtained from microscopic analysis and nucleic acid sequence determination, phage LBR48 can be classified as a member of the family Myoviridae. Analysis of the genome showed the conserved gene order of previously reported phages of the family Siphoviridae from lactic acid bacteria, despite low nucleotide sequence similarity. Analysis of the attachment sites revealed 15-nucleotide-long core sequences.     

Deep sequencing analysis of RNAs from a grapevine showing Syrah decline symptoms reveals a multiple virus infection that includes a novel virus

In a search for viruses associated with decline symptoms of Syrah grapevines, we have undertaken an analysis of total plant RNA sequences using Life Sciences 454 high-throughput sequencing. 67.5 megabases of sequence data were derived from reverse-transcribed cDNA fragments, and screened for sequences of viral or viroid origin. The data revealed that a vine showing decline symptoms supported a mixed infection that included seven different RNA genomes. Fragments identified as derived from viruses or viroids spanned a ∼ ten thousand fold range in relative prevalence, from 48,278 fragments derived from Rupestris stem pitting-associated virus to 4 fragments from Australian grapevine viroid. 1527 fragments were identified as derived from an unknown marafivirus. Its complete genome was sequenced and characterized, and an RT-PCR test was developed to analyze its field distribution and to demonstrate its presence in leafhoppers (vector for marafiviruses) collected from diseased vines. Initial surveys detected a limited presence of the virus in grape-growing regions of California.