- Volume 76, Issue 5, 1995
Volume 76, Issue 5, 1995
- Animal
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Monoclonal antibodies to a peptide of human rhinovirus type 2 with different specificities recognize the same minimum sequence
More LessMonoclonal antibodies (MAbs) raised against a synthetic peptide including residues 156–170 of protein VP2 of human rhinovirus type 2 (HRV2) have previously been shown to be of differing specificities. The basis for these differences has now been examined in greater detail by ELISA, radioimmunoprecipitation and virus neutralization. Reactions with a panel of HRV2 mutant viruses indicated that substitution of some residues could enhance the apparent activity of one of the neutralizing anti-peptide MAbs. For one such substitution, VP2 P164H, there appeared to be a correlation between increased neutralizing activity and enhanced binding. Mapping experiments identified two overlapping neutralization epitopes (amino acids 156–163 and 160–165) and several non-neutralizing epitopes. Although some differences in antibody reactivity were due to epitope specificity alone, the explanation for others was less obvious. Significantly, the majority of MAbs that recognized, and in some cases neutralized, native virus had the same minimum binding sequence and critical residue requirement as others which recognized virus particles only after distortion. This demonstrates that factors other than the linear sequence of the peptide can be crucial in determining the fine specificity, and hence biological relevance, of peptide antigens.
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Comparison of bovine coronavirus isolates associated with neonatal calf diarrhoea and winter dysentery in adult dairy cattle in Québec
More LessCytopathic coronaviruses were isolated in HRT-18 cells from bloody faecal samples collected from cows in Québec dairy herds having experienced typical outbreaks of winter dysentery (WD). The formation of polykaryons in the infected cell cultures was found to be dependent on the presence of trypsin in the medium. The WD isolates differed from the prototype Mebus strain of bovine enteropathogenic coronavirus (BCV.Meb) in respect to haemagglutination inhibition (HI), haemagglutination patterns at 4 °C and 37 °C, and receptor destroying enzyme activity with rat erythrocytes. Other field strains of BCV associated with outbreaks of neonatal calf diarrhoea (NCD) also differed from the BCV.Meb strain by demonstrating differences in HI. In all cases, no differences were detected by virus neutralization and Western immunoblotting. Analysis and comparison of the nucleotide and deduced amino acid sequences of the PCR-amplified haemagglutinin esterase (HE) genes of one representative WD strain (BCQ.2590) and two highly cytopathic NCD strains (BCQ.3 and BCQ.571) revealed high degrees of similarities (nt and aa sequence homologies > 98%) with the BCV.Meb strain. The putative esterase active site FGDS was conserved among these four BCV strains, indicating that this domain is probably not a determinant for BCV virulence. Six amino acid substitutions occurred between the HE glycoproteins of BCV.Meb and BCQ.2590 strains; two proline substitutions occurred respectively in the signal peptide (at aa 5) and near the sequences of the putative esterase domain (at aa 53).
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- Plant
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Prunus necrotic ringspot ilarvirus: nucleotide sequence of RNA3 and the relationship to other ilarviruses based on coat protein comparison
More LessThe RNA3 of prunus necrotic ringspot ilarvirus (PNRSV) has been cloned and its entire sequence determined. The RNA3 consists of 1943 nucleotides (nt) and possesses two large open reading frames (ORFs) separated by an intergenic region of 74 nt. The 5′ proximal ORF is 855 nt in length and codes for a protein of molecular mass 31.4 kDa which has homologies with the putative movement protein of other members of the Bromoviridae. The 3′ proximal ORF of 675 nt is the cistron for the coat protein (CP) and has a predicted molecular mass of 24.9 kDa. The sequence of the 3′ noncoding region (NCR) of PNRSV RNA3 showed a high degree of similarity with those of tobacco streak virus (TSV), prune dwarf virus (PDV), apple mosaic virus (ApMV) and also alfalfa mosaic virus (AlMV). In addition it contained potential stem-loop structures with interspersed AUGC motifs characteristic for ilar- and alfamoviruses. This conserved primary and secondary structure in all 3′ NCRs may be responsible for the interaction with homologous and heterologous CPs and subsequent activation of genome replication. The CP gene of an ApMV isolate (ApMV-G) of 657 nt has also been cloned and sequenced. Although ApMV and PNRSV have a distant serological relationship, the deduced amino acid sequences of their CPs have an identity of only 51.8%. The N termini of PNRSV and ApMV CPs have in common a zinc-finger motif and the potential to form an amphipathic helix.
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Structural sites specific to citrus viroid groups
More LessSynthesis of cDNA probes by random-priming of a viroid template displays the unusual property of specificity to all members included within a single citrus viroid Group. The specificity of hybridization reactions was influenced by the structural conformation of the viroid RNA template, reaction conditions for reverse transcription and hybridization protocols. Mapping the loci for probe transcription from the CEVd, CVd-IIb, and CVd-IV genomes suggests that a similar structured conformation may be responsible for group specificity. A stem-loop configuration in the viroid template and hybridization target sites can be proposed to be responsible for the availability of the group-specific sequences.
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Characterization of cassava vein mosaic virus: a distinct plant pararetrovirus
More LessCassava vein mosaic virus (CVMV) was found to be widespread throughout the north-eastern region of Brazil. The complete sequence of CVMV was determined, and the genome was 8158 bp in size. A cytosolic initiator methionine tRNA (tRNAmet i)-binding site that probably acts as a primer for minus-strand synthesis was present. The genome contained five open reading frames that potentially encode proteins with predicted molecular masses of 186 kDa, 9 kDa, 77 kDa, 24 kDa and 26 kDa. The putative 186 kDa protein had regions with similarity to the zinc finger-like RNA-binding domain that is a common element in the capsid proteins and similarity to the intercellular transport domain of the plant pararetroviruses. The predicted 77 kDa protein had regions with similarity to aspartic proteases, reverse transcriptase and RNase H of pararetroviruses. This gene order was confirmed by the amplification of similar PCR products from total DNA extracted from CVMV-infected cassava plants. The genomic organization of CVMV was different from the organization of either the caulimoviruses or badnaviruses. In comparisons of the regions with the reverse transcriptase motif, CVMV was grouped between the caulimoviruses and badnaviruses. It appears that CVMV is distinct from the other well-characterized plant pararetroviruses.
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The nucleotide sequence of the RNA-2 of an isolate of the English serotype of tomato black ring virus: RNA recombination in the history of nepoviruses
More LessThe RNA-2 of a carrot isolate from the English serotype of tomato black ring nepovirus (TBRV-ED) has been sequenced. It is 4618 nucleotides long and contains one open reading frame encoding a polypeptide of 1344 amino acids. The 5′ non-coding region contains three repetitions of a stem-loop structure also conserved in TBRV-Scottish and grapevine chrome mosaic nepovirus (GCMV). The coat protein domain was mapped to the carboxy-terminal one-third of the polyprotein. Sequence comparisons indicate that TBRV-ED RNA-2 probably arose by an RNA recombination event that resulted in the exchange of the putative movement protein gene between TBRV and GCMV.
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Transfer of the 3′ non-translated region of grapevine chrome mosaic virus RNA-1 by recombination to tomato black ring virus RNA-2 in pseudorecombinant isolates
More LessIn grapevine chrome mosaic and tomato black ring viruses (GCMV and TBRV), as in many other nepoviruses, the 3′ non-translated regions (3′NTR) are identical between the two genomic RNAs. We have investigated the structure of the 3′NTR of two recombinant isolates which contain GCMV RNA-1 and TBRV RNA-2. In these isolates, the 3′NTR of RNA-1 was transferred to RNA-2, thus restoring the 3′ identity. The transfer occurred within three passages, and probably contributes to the spread of randomly appearing mutations from one genomic RNA to the other. The site of recombination is near the 3′ end of the open reading frame.
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Mutagenesis of the BC1 and BV1 genes of African cassava mosaic virus identifies conserved amino acids that are essential for spread
More LessThe products of three open reading frames encoded by the bipartite geminiviruses have been implicated in viral spread: AC2, BV1 and BC1. Alignment of the DNA B encoded gene products, BV1 and BC1, from African cassava mosaic virus (ACMV) with six other bipartite geminiviruses showed several highly conserved regions. Specific amino acids were selected for mutagenic studies to ascertain the tolerance of the virus to change and to identify the regions within these two proteins required for normal functioning. Various mutant DNA B constructs, and a wild-type construct, were inoculated onto three host plant species with an equivalent DNA A construct. Three of the mutant constructs were infectious on Nicotiana benthamiana and N. clevelandii, but only two induced ACMV disease symptoms on N. tabacum cv. Samsun. Sequencing of the viral DNA extracted from the sap of systemically infected plants confirmed the maintenance of introduced base changes. The amino acid at position 95 on the BV1 gene product was identified as non-essential for normal functioning of the protein. The alteration of the amino acid at position 145 in BC1 demonstrated the ability of the virus to tolerate a conservative change. The lack of tolerance to other changes in amino acids has given an indication of the importance of maintaining protein structure for these proteins to function normally.
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Sequence comparison of RNA2 of nematode-transmissible and nematode-non-transmissible isolates of pea early-browning virus suggests that the gene encoding the 29 kDa protein may be involved in nematode transmission
More LessA cDNA clone containing the complete coding region of RNA2 of the newly described TPA56 isolate of pea early-browning virus (English serotype, PEBV-E) has been obtained. A plasmid (pFLA56) containing this clone together with the 5′ and 3′ non-coding regions of PEBV isolate SP5 (the type isolate of PEBV-E) and flanked by the CaMV 35S promoter and NOS terminator is infectious when co-inoculated onto plants with pCaN1, an infectious clone of PEBV SP5 RNA1. Virus containing RNA2 derived from the cDNA clone of the TPA56 isolate is transmitted by Trichodorus primitivus nematodes, whereas virus containing RNA2 from a clone of the SP5 isolate is not transmitted. Sequencing revealed 11 differences out of 3374 nucleotides between the transmissible TPA56 and non-transmissible SP5 clones. However, only three of the base changes affected the amino acid sequences of virus gene products. A single, conservative amino acid change was present in the virus coat protein. Two non-conservative changes occurred in the protein of molecular mass 29.6 kDa expressed from an open reading frame located down-stream of the coat protein gene, suggesting that it has a function in nematode transmission and that changes in this protein prevent vector transmission of the SP5 isolate of PEBV.
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Volume 5 (1969)
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Volume 3 (1968)
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Volume 2 (1968)
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Volume 1 (1967)