Comparison of the full-length genome sequence of avian metapneumovirus subtype C with other paramyxoviruses

We determined the nucleotide (nt) sequence of the small hydrophobic (SH), attachment glycoprotein (G), and RNA polymerase (L) genes, plus the leader and trailer regions of the Colorado strain of Avian metapneumovirus subtype C (aMPV/C) in order to complete the genome sequencing. The complete genome comprised of 13,134 nucleotides, with a 40 nt leader at its 3' end and a 45 nt trailer at its 5' end. The aMPV/C L gene was the largest with 6173 nt and consisting of a single open reading frame encoding a 2005 amino acids (aa) protein. Comparison of the aMPV/C SH, G, and L nt and predicted aa sequences with those of Human metapneumoviruses (hMPV) revealed higher nt and aa sequence identities than the sequence identities between the aMPV subtypes A, B, C, and D, supporting earlier finding that aMPV/C was closer evolutionary to hMPV than the other aMPV subtypes.     

Complete genome sequence of highly virulent neurotropic Newcastle disease virus strain Texas GB

Newcastle disease virus (NDV) strain Texas GB is a highly virulent neurotropic virus that is used as a standard vaccine challenge virus in the U.S. In this study, the complete genome sequence of strain Texas GB was determined and compared with the complete genome sequences of other NDV strains. The genome is 15,186 nucleotides (nt) long and consists of six genes in the order of 3′leader-N-P-M-F-HN-L-5′trailer. The genome contains a 55-nt leader sequence at the 3′ end and a 114-nt trailer sequence at the 5′ end. The intergenic sequences are 2, 1, 1, 31, and 47 nt between N/P, P/M, M/F, F/HN, and HN/L genes, respectively. The putative cleavage site of fusion protein showed amino acid sequence of R-R-Q-K-R↓F in position 112 to 117, which corresponds to those of virulent NDV strains. The phylogenetic analysis showed that strain Texas GB is closely related to the neurovirulent mesogenic strain Beaudette C (BC) and to NDV viruses isolated in China and Egypt than to other strains of NDV.     

Genomic characterization of two avian paramyxovirus type 2 isolates from chickens in China

The complete genome sequences were determined for avian paramyxovirus type 2 (APMV-2) strains F8 and NK isolated from chickens in China. Both strains had a genome of 14,904 nucleotides (nt) in length, which followed the “rule of six”. Each genome consisted of six genes in the order 3′-N-P-M-F-HN-L-5′, with a 55-nt leader at the 3′ end and a 154-nt trailer at the 5′ end. Sequence alignment and phylogenetic analysis showed that APMV-2 strains F8 and NK shared the highest sequence identity with APMV-2 prototype strain Yucaipa, being classified in the same subgroup as strains Yucaipa, England and Kenya, while strain Bangor represented another subgroup of APMV-2. Among the APMVs, APMV-2 strains F8 and NK exhibited a closer evolutionary relationship with APMV-7 and APMV-8 representative strains.     

Complete genome sequence of a virulent Newcastle disease virus isolated from an outbreak in chickens in Egypt

The complete genome sequence of a virulent Newcastle disease virus (NDV) isolated from chickens in Egypt was determined and compared to the sequence of NDV strains isolated from different parts of the world. The genome is 15,186 nucleotides (nt) long and consists of 6 genes in the order of 3′-N-P-M-F-HN-L-5′. The genome contains a 55-nt leader region at the 3′ end and a 114-nt trailer region at the 5′ end. Interestingly, the phylogenetic analysis showed that strain Egypt is closely related with the NDV strains isolated in China. In addition, the sequence of the fusion protein cleavage site of strain Egypt was identical to that of the NDV strain recently isolated in Mali. Determination of complete genome sequences of additional NDV strains from Africa is necessary to understand the epidemiology of currently circulating viruses in Africa.     

Subtype B avian metapneumovirus resembles subtype A more closely than subtype C or human metapneumovirus with respect to the phosphoprotein,and second matrix and small hydrophobic proteins

Avian metapneumovirus (aMPV) subtype B (aMPV/B) nucleotide sequences were obtained for the phosphoprotein (P), second matrix protein (M2), and small hydrophobic protein (SH) genes. By comparison with sequences from other metapneumoviruses, aMPV/B was most similar to subtype A aMPV (aMPV/A) relative to the US subtype C isolates (aMPV/C) and human metapneumovirus (hMPV). Strictly conserved residues common to all members of the Pneumovirinae were identified in the predicted amino acid sequences of the P and M2 protein-predicted amino acid sequences. The Cys(3)-His(1) motif, thought to be important for binding zinc, was also present in the aMPV M2 predicted protein sequences. For both the P and M2-1 protein-predicted amino acid sequences, aMPV/B was most similar to aMPV/A (72 and 89% identity, respectively), having only approximately 52 and 70% identity, respectively, relative to aMPV/C and hMPV. Differences were more marked in the M2-2 proteins, subtype B having 64% identity with subtype A but < or = 25% identity with subtype C and hMPV. The A and B subtypes of aMPV had predicted amino acid sequence identities for the SH protein of 47%, and less than 20% with that of hMPV. An SH gene was not detected in the aMPV/C. Phylogenetically, aMPV/B clustered with aMPV/A, while aMPV/C grouped with hMPV.     

Comparison of the complete sequences of five different isolates of Potato virus A (PVA), genus Potyvirus

Summary.  The complete nucleotide (nt) and deduced amino acid (aa) sequences of isolates Ali, U, Her (from potato, Solanum tuberosum) and TamMV (from tamarillo, Solanum betacea) of Potato virus A (PVA, genus Potyvirus) were determined and compared with the previously reported sequence of PVA isolate B11. Most parts (proteins) of the polyprotein showed over 95% aa sequence similarity. The cylindrical inclusion (CI) protein and the 6K 1 protein were the most conserved proteins among the five isolates. TamMV was the most different isolate. Sequence similarity between TamMV and the other isolates was the lowest in regions close to the 5′-end [5′-non-translated region (NTR) and P1 region] and 3′-end (N-terminus of coat protein) of the genome. However, the termini of the genome (the first 60 nt of the 5′-NTR and the entire 3′-NTR) were highly similar in all five isolates. A frameshift region in the replicase (NIb) was identified the PVA isolates Ali, B11, Her and U, as compared to TamMV and other potyviruses. Received May 25, 1999/Accepted July 23, 1999     

Erratum to: Genomic characterization of the first class I Newcastle disease virus isolated from the mainland of China

The complete genomic sequence of Newcastle disease virus (NDV) strain NDV08-004, isolated from domestic ducks in China, was determined in this study. The genome is 15198 nucleotides (nt) in length, follows the “rule of six” and contains a 55-nt leader sequence at the 3′ end and a 114-nt trailer sequence at the 5′ end. Compared with the full genome sequences of Class II NDV strains, the NDV08-004 isolate has a 12-nt insertion (TGGGAGACGGGG) in the phosphoprotein gene between nucleotides 2381 and 2382 of the genome (numbered according to the genomic sequence of the La Sota strain, which consists of 15186 nt). Strain NDV08-004 has the motif ¹¹²E-Q-Q-E-R-L¹¹⁷ at the cleavage site of the fusion protein, which is typical of lentogenic NDV strains, and this is in agreement with the results of pathogenic tests based on the mean death time (MDT) and the intracerebral pathogenicity index (ICPI). Phylogenetic analysis based on the full genome revealed that all the NDV strains studied could be divided into two distinct clades, namely class I and class II, and the NDV08-004 isolate characterized in this study was grouped in class I. Further phylogenetic analysis based on a 374-bp fragment of the F gene in class I strains of NDV demonstrated that NDV08-004 belongs to genotype 3, and should be therefore similar to strains obtained from live bird markets in Hong Kong in recent years.     

Citrus viroid Ia is a derivative of citrus bent leaf viroid (CVd-Ib) by partial sequence duplications in the right terminal region

Summary.  Nucleotide sequences of group I citrus viroids Ia (CVd-Ia) and citrus bent leaf viroid (CBLVd, formerly designated CVd-Ib) isolated from citrus plants in Japan, the Philippines and China have been determined. Citrus samples in Japan and the Philippines contained CVd-Ia, which consists of 328 nucleotides(nt). Although 10 nt longer than the type CBLVd-225A in Israel they share 94% identity in overall nucleotide sequence. The Philippines sample also contained a 329-nt long CVd-Ia sequence variant, in which one base insertion and three substitutions were observed. A citrus in China contained CBLVd, which consists of 318 nt and shares 98% identity to CBLVd-225A. CVd-Ia was clearly separated from CBLVd by two 5-nt insertions located in upper (5′-AGCUG-3′) and the lower (5′-CUUCU-3′) strand of the right terminal region (which is also designated T2 domain) in rod-like secondary structure. Since both of the additional 5-nt sequences are similar to the adjacent sequences (5′-AGUUG-3′ and 5′-CUUCU-3′), we hypothesize that CVd-Ia is a derivative of CBLVd caused by partial sequence duplications and substitutions taking place in the right terminal region. Accepted December 11, 1997 October 14, 1997     

Genome comparison of a novel classical swine fever virus isolated in China in 2004 with other CSFV strains

The genome of a novel classical swine fever virus (CSFV), SWH/CA/2004, isolated from a hog pen in Henan Province, central China, is 12296 nucleotides (nt) in length. It is composed of a 373-nt 5′ terminal non-translated region (NTR), a 11697-nt open reading frame (ORF) encoding a polyprotein of 3898 amino acids (aa), and a 226-nt 3′-NTR. Genome comparison of the SWH/CA/2004 isolate (GenBank Accession: DQ127910) with other known CSFV isolates was performed and analyzed. Corresponding segments from SWH/CA/2004 and other reported strains shared 80.4–99.8% identity at the nucleotide level and 89.5–99.8% identity at the amino acid level. From an evolutionary point of view, isolate SWH/CA/2004 is closely related to the highly virulent isolate cF114/CA/2001, with a pairwise distance of 0.013; and distantly related to the moderately virulent isolate GXWZ02/CA/2003, with pairwise distance 0.170. The phylogenetic trees of the full-length genome and the following region E^rns, E1, E2, and NS5B-based neighbor-joining (NJ) method were constructed and approximately divided into different genetic groups according to avirulence, moderate virulence and high virulence, while other region-based NJ trees demonstrated sequence conservation between these groups. The four genomic regions may constitute important criteria for genetic typing of diverse CSFV isolates. Based on these analyses, isolate SWH/CA/2004 was deduced to belong to the highly virulent isolate group. However, SWH/CA/2004 also contains a 14-U deletion in the 3′-NTR that is characteristic of avirulent isolates. These analyses constitute a comprehensive study of the phylogenetics of CSF based on distinct regions of the genome and may provide the basis for future molecular epidemiology research to identify virulent strain outbreaks and trigger implementation of appropriate control measures. Xiang-Min Li and Zhou-Fei Xu contributed equally to this work     

Complete genomic sequence analysis of a highly virulent isolate revealed a novel strain of <Emphasis Type="Italic">Sugarcane mosaic virus</Emphasis>

Sugarcane mosaic virus (SCMV) is the most prevalent virus causing maize dwarf mosaic disease in northern China. A SCMV isolate, BD8, was obtained from the maize showing dwarf and mosaic symptoms in Baoding, China. The complete genomic sequence of BD8 is 9,576 nucleotides (nt) excluding the poly(A) tail. It contains one single open reading frame of 9,192 nt and encodes a large polyprotein of 3,063 amino acids (aa), flanked by a 5′-untranslated region (UTR) of 148 nt and a 3′-UTR of 236 nt. The entire genomic sequence of BD8 shares identities of 79.1–80.8% with those of other 13 SCMV isolates available in the GenBank at nt level, while their CP genes share identities of 76.9–82.6 and 82.8–86.9% at nt and aa levels, respectively. Phylogenetic analysis of the complete genomic sequences reveals that SCMV can be clustered to four groups: group I includes isolates from maize, group II consists of isolates from sugarcane or maize, groups III and IV contain single isolate of AU-A (AJ278405) and BD8, respectively. Thus, BD8 represents a new strain of SCMV. Furthermore analysis of the CP gene sequences of more isolates shows that BD8 is clustered to a group with the isolates from Thailand and Vietnam, which implies that isolates of this strain have been distributed in South Asia. In the greenhouse, BD8 can cause severe symptoms in all the 12 maize varieties tested with high incidence, indicating that BD8 is highly virulent.     

Nucleotide and Predicted Amino Acid Sequences of All Genes Encoded by the 3′ Genomic Portion (9.5 kb) of Respiratory Bovine Coronaviruses and Comparisons Among Respiratory and Enteric Coronaviruses

The 3′-ends of the genomes (9538 bp) of two wild-type respiratory bovine coronavirus (RBCV) isolates LSU and OK were obtained by cDNA sequencing. In addition, the 3′-end of the genome (9545) of the wild-type enteric bovine coronavirus (EBCV) strain LY-138 was assembled from available sequences and by cDNA sequencing of unknown genomic regions. Comparative analyses of RBCV and EBCV nucleotide and deduced amino acid sequences revealed that RBCV-specific nucleotide and amino acid differences were disproportionally concentrated within the S gene and the genomic region between the S and E genes. Comparisons among virulent and avirulent BCV strains revealed that virulence-specific nucleotide and amino acid changes were located within the S and E genes, and the 32 kDa open reading frame. This revised version was published online in July 2006 with corrections to the Cover Date.     

The readthrough region of Potato mop-top virus (PMTV) coat protein encoding RNA, the second largest RNA of PMTV genome, undergoes structural changes in naturally infected and experimentally inoculated plants

Summary.  Molecular data on Potato mop-top virus (PMTV), genus Pomovirus, is currently mostly based on analysis of two Scottish isolates, PMTV-S and PMTV-T. Here we report the complete sequence of “the coat protein (CP) encoding RNA” of an isolate of PMTV obtained from the field in Sweden. Our data show that this RNA (3134 nt) is the second largest of the three RNA species in the tripartite PMTV genome, and it should, therefore, be referred to as RNA 2. This nomenclature is consistent with other pomoviruses. The sequence of the readthrough domain (RT) of RNA 2 was determined also in two additional field isolates of PMTV from Finland and Denmark. All three isolates contained a novel, 109 nucleotides long sequence at the 3′-end of the RT, which has not been found in PMTV-S and PMTV-T. Hence, our data suggest that the RNA 2 sequences previously described for the isolates PMTV-T and PMTV-S may represent deletion derivatives. The C-proximal half of RT contained many amino acid (aa) differences among the isolates, in contrast to only few aa differences in the N-proximal part of RT. Deletion variants of RNA 2 were generated from the Nordic isolates in potato tubers infected in the field, and in the mechanically inoculated test plants. All deletions started within a short region (18 nt) and removed 558–940 nt from the 3′-end of RT region. This study for the first time describes the full-length sequence of the “CP-encoding RNA” (RNA2) of PMTV, and reveals considerable aa variability and occurrence of deletion variants of RT in the field isolates of PMTV. Received February 15, 2000 Accepted September 26, 2000     

Complete genomic characterization of a European type 1 porcine reproductive and respiratory syndrome virus isolate in Korea

Porcine reproductive and respiratory syndrome virus (PRRSV) isolates belonging to the European genotype 1 have recently emerged in South Korea, suggesting potential problems for disease control. In the present study, we attempted to determine the complete nucleotide sequence of the first Korean type 1 PRRSV isolate, designated KNU-07. The full-length genome of KNU-07 was found to be 15,038 nucleotides in length, which was 60 nucleotides shorter than the type 1 prototype strain Lelystad due to a notable 60-bp deletion within the nonstructural protein 2 (NSP2). The KNU-07 genome was shown to consist of a 221-nucleotide (nt) 5′ untranslated region (UTR), a 14,703-nt protein-coding region, and a 114-nt 3′ UTR, followed by a 42-73-bp poly(A) tail. A nucleotide sequence comparison of the KNU-07 genome with 20 complete PRRSV genomes revealed a 10.5–13.3% and 39.5–40.3% divergence from type 1 and type 2 strains, respectively, at the genome level, indicating a high similarity to the virus strains commonly identified as the European genotype. In order to investigate genetic variation and to understand the molecular evolution of the type 1 isolate in Korea, extensive phylogenetic analyses were performed using the ORF5 and ORF7 nucleotide sequences of published type 1 PRRSV isolates. The data further indicated that the newly emerging type 1 isolate KNU-07 belongs to the recently proposed pan-European subtype 1. Taken together, the results of this study describe the genomic characterization of the type 1 PRRSV isolated in South Korea, suggesting a recent introduction of the virus typical for this genotype that has commonly appeared worldwide.     

Phylogenetic analysis of the Potyviridae with emphasis on legume-infecting potyviruses

Summary.  The 3′-terminal nucleotide sequences of thirteen authenticated strains of bean common mosaic virus (BCMV) and one strain of bean common mosaic necrosis virus (BCMNV) were obtained. The regions sequenced included the coat protein coding sequence and 3′-end non-coding region. These data, combined with sequence information from other legume-infecting potyviruses and the Potyviridae were used for phylogenetic analysis. Evidence is provided for delineation of BCMNV as distinct from BCMV and the inclusion of azuki mosaic, dendrobium mosaic, blackeye cowpea mosaic, and peanut stripe viruses as strains of BCMV. This relationship defines the members of the BCMV and BCMNV subgroups. These data also provide a basis upon which to define virus strains, in combination with biological data. Other aspects and implications of legume-infecting potyvirus phylogenetics are discussed. Received December 24, 1996 Accepted June 3, 1997     

Molecular epidemiology of porcine epidemic diarrhea virus in China

Since early 2006, porcine epidemic diarrhea virus (PEDV) has been reemerging in immunized swine herds. Open reading frame 3 (ORF3) is the only accessory gene in the PEDV genome. The entire ORF3 genes of 12 PEDV field strains and one vaccine strain were sequenced. The ORF3 genes of Chinese PEDV field strains (excluding CH/GSJIII/07) contain a single 672- or 675-nucleotide (nt) ORF, which encodes a 223- or 224-aa-long peptide. However, the CV777 vaccine strain and CH/GSJIII/07 contain a 276-nt ORF because of a 49-nt deletion at nt 245–293. The Chinese PEDV field strains and PEDV reference strains are divided into three groups based on the phylogenetic relationship of their ORF3 genes. Chinese PEDV field strains (excluding CH/GSJIII/07) have a close phylogenetic relationship to Korean strains and are genetically different from the PEDV vaccine strains. However, CH/GSJIII/07 has a close phylogenetic relationship to two vaccine strains, suggesting that it might have evolved from a live vaccine strain. Chinese PEDV field strains (excluding CH/GSJIII/07) can be differentiated from PEDV vaccine strains by a nested RT-PCR method.     

Molecular relationships between type Asia 1 new strain from China and type O Panasia strains of foot-and-mouth-disease virus

The complete genome of Asia 1/HNK/CHA/05 strain of foot-and-mouth disease virus (FMDV) was sequenced, which was isolated from Chinese Hongkong in 2005. It is 8187 nt long in size and contains 5′-UTR, polyprotein region, and 3′-UTR. Polyprotein region can be divided into four parts of L, P1, P2 and P3. In this report, these six parts of the whole genome of the strain were compared with 12 reference strains using DNAStar and Simplot softwares. The comparison of P1 confirmed that Asia 1/HNK/CHA/05 has a high identity with nine type Asia 1 reference sequences from 85.9 to 92.6% (Ind/491/97 strain is the highest) but from 69.6 to 69.7% with three type O Panasia sequences. The identities of 5′-UTR, L, P2, P3 and 3′-UTR with three Panasia strains are from 89.0 to 90.6%, 92.5 to 93.4%, 94.8 to 95.5%, 96.0 to 96.7% and 90.7 to 92.5% separately, but with nine type Asia 1 strains are from 83.5 to 85.9%, 87.7 to 90.7%, 87.0 to 91.6%, 91.6 to 92.8% and 86.0 to 100% separately, which illuminates the closer relationship between Asia 1/HNK/CHA/05 and Panasia strains in 5′-UTR, L, nonstructural region and 3′-UTR although they do not belong to the same serotype. The SimPlot software was used to examine the authentic relationships of Asia 1/HNK/CHA/05 with 12 reference sequences. It was found that Asia 1/HNK/CHA/05 strain has a highest similarity with three Panasia strains especially Tibet/CHA/99 in 5′-UTR, L, nonstructural region and 3′-UTR but has a highest similarity with Asia 1/Ind/491/97 strain in P1 region, which suggested that the gene recombination had occurred around nucleotide position 1811 and 3971in the polyprotein region between Tibet/CHA/99 and Ind/491/97 to recombine the Asia 1/HNK/CHA/05 strain. The nucleotide sequence data of Asia 1/HNK/CHA/05 strain reported in this article has been submitted to the Genbank nucleotide sequence database and has been assigned the accession number EF149010.     

Genetic diversity of the Junin virus in Argentina: geographic and temporal patterns

RNA was purified from 39 strains of cell-cultured Junin virus (JUN) from central Argentina, which included both human- and rodent-derived isolates (a total of 26 and 13, respectively), as well as from 2 laboratory JUN strains, XJ Cl3 and XJ #44. JUN-specific primers were used to amplify a 511-nucleotide (nt) fragment of the nucleocapsid protein gene and a 495-nt fragment of the glycoprotein 1 (GP1) gene. Genetic diversity among JUN strains studied was up to 13% at the nt level and up to 9% at the amino acid (aa) level for the GP1 gene and up to 9% (nt) and 4% (aa) for the NP gene. Phylogenetic analyses of both genes revealed three distinct clades. The first clade was composed of the JUN strains from the center of the endemic area and included the majority of JUN strains analyzed in the current study. The second clade contained 4 JUN strains isolated between 1963 and 1971 from Cordoba Province, the western-most edge of the known endemic area. The third clade contained 4 JUN strains that originated from Calomys musculinus trapped in Zarate, the northeastern edge of the known endemic area. Certain JUN sequences, which were obtained from GenBank and identified as XJ, XJ #44, and Candid #1 strains, appeared to form a separate clade. Over 400 nt of the GP1 and GP2 genes were additionally sequenced for 7 JUN strains derived from patients with different clinical presentations and outcomes of Argentine hemorrhagic fever. Analysis of the corresponding aa sequences did not allow us to attribute any particular genetic marker to the changing severity or clinical form of the human disease.     

Molecular cloning of an Australian isolate of hepatitis C virus

Summary.  The genomic sequence of an Australian isolate of hepatitis C virus (HCV) was determined from overlapping cDNA clones obtained from a small amount (1.2 ml) of serum from a single individual with hepatitis C. The isolate (HCV-A) comprises 9 379 nucleotides (nt) including 324 nt of a 5′ untranslated region (5′UTR), a single long open reading frame of 9 033 nt encoding a polyprotein of 3 010 amino acids (aa), and 22 nt of a 3′ untranslated region (3′UTR). Sequence analysis of a 251 nt region within the 5′UTR and a 222 nt region within NS5B showed the genotype of HCV-A to be subtype 1b. A striking difference in the amino acid sequence of the hypervariable region 1 (HVR-1), and not in the surrounding sequence, was seen in cDNA clones synthesised from serum taken 52 weeks after the initial sample, indicating a significant population diversity of HCV genomes. Accepted October 17, 1977 Received July 7, 1997