Showing papers in "Journal of General Virology in 1988"

The complete DNA sequence of the long unique region in the genome of herpes simplex virus type 1.

Abstract: We have determined the DNA sequence of the long unique region (UL) in the genome of herpes simplex virus type 1 (HSV-1) strain 17. The UL sequence contained 107,943 residues and had a base composition of 66.9% G + C. Together with our previous work, this completes the sequence of HSV-1 DNA, giving a total genome length of 152,260 residues of base composition 68.3% G + C. Genes in the UL region were located by the use of published mapping analyses, transcript structures and sequence data, and by examination of DNA sequence characteristics. Fifty-six genes were identified, accounting for most of the sequence. Some 28 of these are at present of unknown function. The gene layout for UL was found to be very similar to that for the corresponding part of the genome of varicella-zoster virus, the only other completely sequenced alphaherpesvirus, and the amino acid sequences of equivalent proteins showed a range of similarities. In the whole genome of HSV-1 we now recognize 72 genes which encode 70 distinct proteins.

Typing Hepatitis B Virus by Homology in Nucleotide Sequence: Comparison of Surface Antigen Subtypes

Abstract: The complete nucleotide sequences of the DNA of three hepatitis B virus (HBV) genomes of subtype adw, cloned from plasma samples of asymptomatic carriers living in the mainland and Okinawa Prefecture of Japan and Indonesia were determined. All three comprised 3215 bp and differed in sequence by only 3.9 to 5.6%. When these isolates were compared with the reported sequences of two HBV genomes of the same subtype derived from American carriers, however, the differences were greater (8.3 to 9.3% to an extent comparable with the nucleotide divergence between an HBV genome of subtype adw and that of a heterotypic subtype, such as adr, ayw or ayr. A total of 18 HBV genomes of various subtypes, including the three described here, 10 reported previously and five unpublished ones, were classified into four groups based on an inter-group divergence in nucleotide sequence of 8% or greater: group A (two adw genomes), group B (four adw), group C (three adw, four adr and one ayr) and group D (four ayw). Thus, the nine genomes of HBV subtype adw were distributed into three groups with considerably different sequences. These results indicate that the four major antigenically defined subtypes of envelope polypeptide do not reflect true genotypic variation of HBV. The fact that d to y, as well as w to r, subtypic change can be induced by an A----G point mutation at nucleotides 365 and 479 in the S gene, respectively, supports this view.

Coronaviruses: structure and genome expression.

Abstract: Introduction. Progress in coronavirology is illustrated by the number of workshops convened and reviews written. International meetings have been held in Germany (1980), the Netherlands (1983) and the U.S.A. (1986), and the Fourth Coronavirus Symposium will be organized by one of us (D.C.) in Cambridge, U.K. in July 1989. In addition, reviews have appeared which highlighted particularly interesting characteristics of the family, e.g. the replication strategy (Lai, 1986) and the glycoproteins (Sturman & Holmes, 1985). As the last general accounts were published some 5 years ago (Siddell et al., 1983; Sturman & Holmes, 1983) an update is timely. The present article is based on the large amount of sequence data accumulated in these years and focuses on the viral nucleic acids and proteins and their function. Coronaviruses cause infections in man, other mammals and birds. Most experimental data have been obtained from studies of mouse hepatitis virus (MHV) and infectious bronchitis virus of chickens (IBV).

Epstein-Barr virus gene expression in nasopharyngeal carcinoma.

Abstract: Summary Epstein—Barr virus (EBV), an agent with growth transforming potential for human B cells, is associated with certain B cell lymphomas in man and also with an epithelial tumour, undifferentiated nasopharyngeal carcinoma (NPC). Since B cell growth transformation is associated with the constitutive expression of a small number of EBV-coded latent proteins, the nuclear antigens EBNA 1, EBNA 2, EBNA 3 and EBNA-LP and the latent membrane protein (LMP), the present work sought to determine whether this same pattern of virus gene expression occurred in NPC. Tumour biopsies were taken from NPC patients from three areas of differing tumour incidence (Kenya, Algeria, Britain) and immediately snap-frozen, as were biopsies of non-EBV-related carcinomas for controls. Immunoblotting of PAGE-separated proteins with selected human sera identified 24 NPC biopsies clearly expressing EBNA 1. When the analysis was extended using selected human sera with antibodies against the other EBNAs, there was no detectable expression of EBNA 2, EBNA 3 or EBNA-LP in any of these 24 biopsies; their EBNA 2-negative status was confirmed using a monoclonal antibody (MAb) PE2 which was reactive in immunoblotting and in immunoprecipitation with EBNA 2A and EBNA 2B proteins. Similar experiments with two different LMP-specific MAbs, CS1 to 4 and S12, revealed heterogeneity between NPC biopsies; 9/24 biopsies were demonstrably LMP-positive, the degree of expression varying considerably between individual tumours in a manner which was not related to the level of EBNA 1 expression. None of the 24 NPC biopsies expressed detectable amounts of EBV lytic cycle antigens. A nude mouse-passaged NPC cell line, C15, likewise expressed EBNA 1 and LMP but none of the other EBV latent proteins nor lytic cycle antigens. This work identifies a novel type of EBV-cell interaction in NPC cells which is distinct from that seen in in vitro transformed B cell lines and from that seen to date in EBV-positive B cell lymphomas.

Biochemistry and Immunology of Infectious Bursal Disease Virus

Abstract: Introduction. The aetiological agent of infectious bursal disease (IBD), IBD virus (IBDV), belongs to a new group of viruses referred to as ‘birnaviruses’ (Dobos et al., 1979), which has been characterized only recently (Brown, 1986). There are excellent reviews dealing with the clinical, pathological, serological and epidemiological aspects of IBDV infection (Faragher, 1972; Becht, 1980; Okoye, 1984; Cummings et al., 1986). The molecular biology of birnaviruses has also been reviewed (Dobos & Roberts, 1983) but with an emphasis on infectious pancreatic necrosis virus (IPNV), the birnavirus genus prototype. The purpose of the present review is to compile information on structural and immunological aspects of IBDV. These are subjects of much recent interest, and have great relevance to the control of IBD in chickens. IBD is a highly contagious viral disease of young chickens which is characterized by destruction of the lymphoid cells in the bursa of Fabricius; other lymphoid organs are also affected but to a lesser degree (Cheville, 1967).

The N and C termini of the coat proteins of potyviruses are surface-located and the N terminus contains the major virus-specific epitopes.

Abstract: Summary Mild proteolysis by trypsin of particles of six potyviruses (bean yellow mosaic virus, clover yellow vein virus, Johnson grass mosaic virus, passion-fruit woodiness virus, potato virus Y and watermelon mosaic virus II) revealed that the N- and C-terminal regions of their coat protein are exposed on the particles' surfaces. The enzyme treatment removed the N-terminal region (30 to 67 amino acids long, depending on the virus) and 18 to 20 amino acids from the C terminus of the coat proteins, leaving a fully assembled virus particle composed of coat protein cores consisting of 216 or 218 amino acid residues. These core particles were indistinguishable from untreated native particles in an electron microscope and were still infectious. The core particles lacked the virus-specific surface epitopes that are recognized by the bulk of the polyclonal antibodies raised against the whole virus particles. Epitopes thought to be group-specific were located in the trypsin-resistant core protein region. The implications of these findings are discussed in relation to the similar surface location of the N- and C-terminal regions of the coat protein of other rod-shaped plant viruses and the observed common structural features displayed by isometric plant and animal viruses.

Amino Acid Sequence Homology of Coat Proteins as a Basis for Identification and Classification of the Potyvirus Group

Abstract: Summary Analysis of the 136 possible pairings of the coat protein amino acid sequences from 17 strains of eight distinct potyviruses revealed a bimodal distribution of sequence homology. Distinct members of the group exhibited sequence homologies ranging from 38 to 71% (average 54%) with major differences in the length and sequence of their N termini and high sequence homology in the C-terminal half of the coat proteins. In contrast strains of individual viruses exhibited sequence homologies of 90 to 99% (average 95%) and had very similar N-terminal sequences. These findings cast doubt on the currently held ‘continuum’ hypothesis proposed to explain the unsatisfactory taxonomy of the potyvirus group. The coat protein sequence data, in combination with information on the nature of the potyvirus particle assembly, can be used to develop rationally designed, simple serological techniques that appear to be more useful and more easily applied than those properties previously used for potyvirus identification and classification.

Nucleotide and complete amino acid sequences of Kunjin virus: definitive gene order and characteristics of the virus-specified proteins.

Abstract: Summary A Kunjin (KUN) virus cDNA sequence of 10664 nucleotides was obtained and it encoded a single open reading frame for 3433 amino acids. Partial N-terminal amino acid analyses of KUN virus-specified proteins identified the polyprotein cleavage sites and the definitive gene order. The gene order relative to that proposed for yellow fever (YF) virus is as follows: KUN 5′-C·GP20·E·GP44·P19·P10·P71·(?)·P21·P98-3′ YF 5′-C·prM·E·NS1·ns2a·ns2b·NS3·ns4a·ns4b·NS5-3′. The order of putative signal sequences and stop transfer sequences indicated that KUN NS1, NS2A and NS4B are probably cleaved in the lumen of the endoplasmic reticulum, at a consensus site Val-X-Ala↓ where X is an uncharged residue, and NS2B, NS3 and NS5 are cleaved in the cytosol at the site Lys-Arg↓Gly. Comparisons with the complete amino acid sequences of YF and West Nile (WN) viruses showed that KUN virus shared 93% homology with WN virus, but only 46% homology with YF virus. Comparisons among individual gene products of six flaviviruses showed that E, NS1, NS3 and NS5 tended to be the most highly conserved, and C among the least conserved. Homologous cleavage sites were evident, and six domains in NS5, a total of over 170 residues, shared at least 85% homology. Comparisons with the KUN C to NS2B sequence defined a gradient of relationships of all gene products in decreasing order WN > Murray Valley > Japanese encephalitis > St Louis encephalitis viruses within this closely related serological complex. A non-coding 5′ sequence (75 nucleotides) of KUN virus shared 95% homology with WN virus and a shorter imperfect match with Murray Valley encephalitis virus (15 of 18 nucleotides). The KUN non-coding 3′ sequence of 290 nucleotides contained several short and imperfectly matched sequences, and shared 87% homology over the distal region of 191 nucleotides with the corresponding region of WN virus RNA.

The complete nucleotide sequence of potato virus X and its homologies at the amino acid level with various plus-stranded RNA viruses.

Abstract: Summary Double-stranded cDNA of potato virus X (PVX) genomic RNA has been cloned and sequenced. The sequence [6435 nucleotides excluding the poly(A) tract] revealed five open reading frames (ORFs) which were numbered one to five starting at the 5′ terminus of the RNA. They encoded proteins of M r 165588 (166K), 24622 (25K), 12324 (12K), 7595 (8K) and 25080 (coat protein), respectively. ORFs 1 and 2 were inphase coding regions. The ORF 1 product contained domains of homology with the tobacco mosaic virus 126K and 183K products. The ORF 2 and 3 products showed homologies with the barley stripe mosaic virus 58K and 14K proteins, the beet necrotic yellow vein virus 42K and 13K products and the white clover mosaic virus 26K and 13K products, respectively. The significance of these homologies with respect to putative functions of the PVX-encoded proteins are discussed.

Membrane fusion by peptide analogues of influenza virus haemagglutinin.

Abstract: We have studied the interactions of synthetic peptides corresponding to the sequence of the amino terminus of the HA2 subunit of influenza virus haemagglutinin with artificial lipid membranes. The peptides could fuse cholesterol-free liposomes at neutral as well as acid pH; however, liposomes containing cholesterol could only be fused below pH 6. The fusion process caused leakage of aqueous liposomal contents. Peptides with amino acid substitutions had fusion properties similar to whole haemagglutinin molecules with the corresponding sequence changes. Non-fusogenic peptides still interacted with the membrane but did not cause leakage of liposomal contents. A correlation between the alpha-helical content of peptide and its fusogenicity was noted, but this was not absolute. The results reported here support suggestions for a role of the amino terminus of HA2 in virus-endosome fusion.

Comparative studies on structural and antigenic properties of two serotypes of infectious bursal disease virus.

Abstract: The electrophoretic mobilities of the two genome segments and the structural polypeptides of the chicken strain Cu-1 (serotype I) and the turkey isolate 23/82 (serotype II) of infectious bursal disease virus were compared. There is a close antigenic relationship between the smaller of the two major structural proteins (32K) of both strains. Neutralizing monoclonal antibodies are induced by the larger protein (40K in Cu-1) which differentiates between the two serotypes. The 40K structural protein also has epitopes which do not induce neutralizing antibodies and which are common to both strains. There is evidence that the antigenic region responsible for the production of neutralizing antibodies is highly conformation-dependent. Passively administered neutralizing antibodies directed against the 40K structural polypeptide of Cu-1 confer protective immunity to susceptible chickens, whereas antibodies directed against the 32K structural protein do not have any protective effect.

Synergistic Interactions of Anti-NS1 Monoclonal Antibodies Protect Passively Immunized Mice from Lethal Challenge with Dengue 2 Virus

Abstract: Summary Non-neutralizing, serotype-specific anti-NS1 monoclonal antibodies partially protected passively immunized mice from lethal dengue 2 virus intracerebral challenge. There was no apparent correlation between complement-fixing activity and protective capacity among individual anti-NS1 monoclonal antibodies. Immunization with specific combinations of non-protective or partially protective antibodies resulted in prolonged survival or reduced mortality. Solid protection, equal to that achieved after immunization with neutralizing polyclonal antibody, was achieved only with an antibody pair which individually fixed complement to high titre with homologous virus. Some groups of mice had increased morbidity after immunization with combinations of protective monoclonal antibodies that bind to overlapping epitopes. These results may affect the design of recombinant dengue vaccines which may require the inclusion of serotype-specific antigenic domains.

Functional studies on the p10 gene of Autographa californica nuclear polyhedrosis virus using a recombinant expressing a p10-beta-galactosidase fusion gene.

Abstract: SUMMARY The fl-galactosidase gene (lacZ) of Escherichia coli was inserted in phase with the coding sequence of the Autographa californica nuclear polyhedrosis virus (AcMNPV) late-expressed Mr 10000 (pl0) gene. The fusion gene was inserted into the AcMNPV genome by cotransfection of a recombinant plasmid pAcR159Z, consisting of the EcoRI P fragment-containing pBR325-derived plasmid pAcR159 and the lacZ insert inthe p 10 gene, and wild-type AcMNPV DNA. Infection of Spodopterafrugiperda cells by the resulting recombinant AcMNPV/p 10Z-2 showed high level expression of a p 10lacZ fusion protein, but no synthesis of pl0. Therefore, the pl0 gene is dispensable for virus replication and the pl0 promoter is effective in driving the expression of foreign genes. Cells infected with AcMNPV/pl0Z recombinants resembled those infected with wild-type AcMNPV in the amounts of polyhedrin synthesized and polyhedra formed, although pl0 was absent. The nucleus and cytoplasm of AcMNPV/pl0Z-2infected cells lacked the fibrous structures that are associated with pl0 in wild-type AcMNPV-infected cells. Instead, large granular structures were observed that were found by immunogold labelling to contain the lacZ gene product. The electron-dense 'spacers', thought to be precursors of the polyhedron membrane, were absent from cells infected by the recombinant virus and the polyhedra did not have a membrane. The recombinant AcMNPV/pl0Z-2 was at least twice as virulent for second instar S. exigua larvae than was wild-type AcMNPV. The increased virulence of the recombinant is an important property for the control of insects.

Excretion of non-infectious virus particles lacking glycoprotein H by a temperature-sensitive mutant of herpes simplex virus type 1: evidence that gH is essential for virion infectivity.

Abstract: Summary A temperature-sensitive mutant of herpes simplex virus type 1, tsQ26, was shown to contain an amino acid substitution in glycoprotein H (gH). The mutant entered cells efficiently at the non-permissive temperature and replicated to give nearly normal yields of intracellular infectivity. The intracellular virions contained, predominantly, an immature form of gH and no gH was found on the surface of infected cells. Excreted virions were devoid of gH and were not infectious. Virions excreted at the permissive temperature were infectious and contained gH and no loss of gH resulted from incubation of these virions at the non-permissive temperature. The temperature-sensitive phenotype apparently results from the loss of gH from virions during their transport to the cell surface, and since loss of gH is accompanied by loss of infectivity we conclude that gH is an essential component of the infectious virion.

The DNA sequences of the long repeat region and adjoining parts of the long unique region in the genome of herpes simplex virus type 1

Abstract: We have determined the DNA sequence of the long repeat region (RL) in the genome of herpes simplex virus type 1 (HSV-1) strain 17, as 9215 bp of composition 71.6% G + C. In addition, the sequences of parts of the long unique region (UL) adjacent to the terminal (TRL) and internal (IRL) copies of RL were determined (2611 and 3836 bp, respectively). Gene organization in these regions of UL was deduced from the sequences and other available data. It was proposed that the region of UL sequenced, adjacent to TRL, contains three complete genes, none with significant previous characterization, and that the region of UL adjacent to IRL also contains three genes, one encoding the immediate early protein IE63. The RL sequence contains one well characterized gene, for the protein IE110, whose organization we have described previously. Between the downstream end of the IE110 gene and UL there is a 3500 bp segment of RL in which we did not find convincing protein-coding sequences, and which thus remains of obscure functionality. Upstream of the IE110 gene is a region previously proposed by others to contain a gene. However, our sequence data are not compatible with their interpretation. We do consider it possible that the region is protein-coding, but regard gene organization here as still unresolved.

Gene sequence and mapping data from Marek's disease virus and herpesvirus of turkeys: implications for herpesvirus classification.

Abstract: Purified DNAs from Marek's disease virus (MDV) and the herpesvirus of turkeys (HVT) were randomly sheared and cloned into the M13 bacteriophage. Two-hundred and ten MDV and 130 HVT clones were sequenced to give representative samples of the genome sequences. The predicted amino acid sequences from these gammaherpes-viruses were compared to known sequences from other herpesviruses using computer analysis. Thirty-five MDV and 24 HVT genes were identified by comparison with varicella-zoster virus (VZV), an alphaherpesvirus. However, only 14 MDV and seven HVT genes, giving generally lower homology scores, were found by comparison with Epstein-Barr virus (EBV), a gammaherpesvirus, indicating that MDV and HVT sequences bear greater similarity to VZV than to EBV sequences. A number of sequences were mapped by hybridizing labelled M13 clones to Southern blots of restriction fragments of MDV or HVT DNA. The results were consistent with the MDV and HVT genomes being collinear with VZV.

Neutralizing monoclonal antibodies to bovine viral diarrhoea virus bind to the 56K to 58K glycoprotein.

Abstract: A panel of murine monoclonal antibodies (MAbs) against the two major glycoproteins of bovine viral diarrhoea virus (BDV) was produced and assayed by serum neutralization, radioimmunoprecipitation (RIP) and immunoblotting. Based on their viral polypeptide specificity and on their ability to neutralize viral infectivity, the MAbs in the panel were divided into three classes: Class 1 MAbs reacted with the 56K to 58K glycoprotein and neutralized the virus, class 2 MAbs recognized the 56K to 58K glycoprotein but were not neutralizing, and class 3 MAbs reacted with the 48K glycoprotein and did not neutralize the virus. These results identify the 56K to 58K protein as one of the envelope glycoproteins of BDV. Evidence was obtained indicating that it is responsible for the induction of neutralizing antibodies. No large uncleaved precursors of the 56K to 58K protein could be identified unequivocally by RIP of infected cell extracts, suggesting that this polypeptide is proteolytically processed cotranslationally. A subset of MAbs that reacted with BDV isolates of the noncytopathic biotypes yielded similar results, indicating that these findings are applicable to both biotypes of BDV.

Poliovirus protein 3CD is the active protease for processing of the precursor protein P1 in vitro.

Abstract: A transcription/translation system for generating poliovirus proteins in vitro has been used to assess the proteolytic activity of various polypeptides containing the virus-coded 3C region towards the poliovirus precursor protein P1. Plasmids containing a phage T7 promoter followed by either the complete poliovirus P1 sequence or various sequences containing the 3C region were used for this purpose. We showed that all except one of the 3C-containing polypeptides had a very restricted activity towards P1, generating only a small amount of VP1 and no VP0 or VP3. The only polypeptide capable of fully processing P1 into VP0, VP3 and VP1 in vitro was protein 3CD, consisting of the complete 3C and 3D sequences.

Comparisons of the pestivirus bovine viral diarrhoea virus with members of the flaviviridae.

Abstract: Summary The molecular features of bovine viral diarrhoea virus (BVDV), a member of the Pestivirus genus currently classified in the Togaviridae, were examined for characteristics resembling those of the Flaviviridae family. Like flaviviruses, BVDV possesses a single-stranded RNA genome (approx. 4.3 × 106 M r) deficient in a 3′ poly(A) tract. This RNA has a single open reading frame spanning the length of the genome in the viral RNA sense (positive polarity), implying an expression strategy involving the processing of a precursor polyprotein. With the exception of several short but significant stretches of identical amino acids within two non-structural proteins, no extended regions of nucleotide or amino acid sequence homology between BVDV and representatives of three serological subgroups of mosquito-borne flaviviruses were noted. However, comparison of the organization of protein-coding domains along the genomes and the hydropathic profiles of amino acid sequences revealed pronounced similarities. It is proposed that Pestivirus, of which BVDV is the prototype member, should no longer be grouped in the Togaviridae family, but rather be considered a genus of non-arthropod-borne viruses within the Flaviviridae.

The global spread and replacement of canine parvovirus strains.

Abstract: Canine parvovirus type 2 (CPV-2) became widespread during 1978 and was reported in many countries during 1978 and 1979. Earlier studies showed that CPV-2 was replaced in the U.S.A. around 1980 by an antigenically and genetically variant virus (CPV-2a). Here we show that CPV-2 was present in the U.S.A., Japan, Belgium and Australia prior to 1980, but that between 1979 and 1982 CPV-2 was replaced by CPV-2a in all of those countries as well as in France and Denmark. Examination of sera collected between 1979 and 1984 from wild coyotes (Canis latrans) in the U.S.A. by an agar gel precipitin assay indicated that the coyotes were originally infected by CPV-2, but that after 1980 the juvenile coyotes were being infected with CPV-2a. The natural global replacement of CPV-2 by CPV-2a over a period of 2 to 3 years indicates that CPV-2a has a strong epidemiological advantage over CPV-2, although the mechanism involved remains to be defined.

Molecular Cloning and Complete Nucleotide Sequence of an Adult T Cell Leukaemia Virus/Human T Cell Leukaemia Virus Type I (ATLV/HTLV-I) Isolate of Caribbean Origin: Relationship to Other Members of the ATLV/HTLV-I Subgroup

Abstract: Summary We report the first complete nucleotide sequence of an adult T cell leukaemia virus/human T cell leukaemia virus type I (ATLV/HTLV-I) isolate from a British patient of Caribbean origin. Sequence comparisons of our proviral clone (HS-35) with other molecular clones are shown. We note the strong sequence conservation between isolates of Caribbean and Japanese origin (2.3% divergence), but demonstrate the higher homologies existing between isolates originating from similar geographical areas (approximately 1% divergence). Implications for the origin, evolution and dissemination of the ATLV/HTLV-I subgroup are discussed. Analysis of defective proviral clones isolated from the same genomic library is also reported, and suggests a pattern of proviral sequence deletions during the biogenesis of defective proviruses.

Studies on the structure of the influenza virus haemagglutinin at the pH of membrane fusion.

Abstract: At the pH required to trigger the membrane fusion activity of the influenza virus haemagglutinin (HA) the soluble ectodomain of the molecule, BHA, which is released from virus by bromelain digestion, aggregates into rosettes. Analyses of soluble proteolytic fragments derived from the rosettes indicated that aggregation is mediated by association of the conserved hydrophobic amino-terminal region of BHA2, the smaller glycopolypeptide component of each BHA subunit. Further analyses of the structure of the soluble fragments and of HA in its low pH conformation by electron microscopy, spectroscopy and in crosslinking experiments showed that, although the membrane distal globular domains lose their trimer structure at the pH of fusion, the central fibrous stem of the molecule remains trimeric and assumes a more stable conformation. The increase in length of BHA2 at low pH observed microscopically appears to result from movement of the amino-terminal region to the membrane proximal end of the molecule and in virus incubated at low pH the amino terminus may insert into the virus membrane. The consequences of these possibilities for the mechanism of membrane fusion are discussed.

A Definition of Citrus Viroid Groups and Their Relationship to the Exocortis Disease

Abstract: Summary Nucleic acid extracts from citrons (Citrus medica cv. Etrog) displaying mild and moderate symptoms associated with the exocortis disease were analysed by sequential and denaturing PAGE which revealed the presence of several viroids. A comparison was made of electrophoretic patterns displaying one or more distinct citrus viroids from field isolates of citrus with exocortis. Citrus viroids were characterized by the physical parameters of electrophoretic mobility, chromatography on CF-11 cellulose and hybridization to cDNA probes of the well characterized citrus viroids, citrus exocortis viroid, CV-Ib from the ‘citron variable viroid’ isolate, and citrus cachexia viroid. These characteristic properties combined with biological distinctions in the host range and symptom expression suggested a scheme for the organization of the citrus viroids into five major groups. The association of the symptoms induced by these citrus viroids in citron cv. Etrog, their organization into individual viroid groups and their presumed relationship to the exocortis disease of citrus are discussed.

Ribonucleotide reductase encoded by herpes simplex virus is a determinant of the pathogenicity of the virus in mice and a valid antiviral target.

Abstract: Summary The role of the herpes simplex virus (HSV)-encoded ribonucleotide reductase (RR) in the pathogenicity of the virus has been examined by use of mutants with lesions in either the large or small subunit of the enzyme. The virulence of the mutants in mice was reduced by about 106-fold when compared with that of the parental virus (HSV type 1 strain 17), while the virulence of a revertant of one of the mutants was restored to within about 100-fold of that of the parent virus. These experiments demonstrate that activity of the HSV RR is essential for virus pathogenicity in mice and suggests that the enzyme is a valid target for specific antiviral compounds.

Aetiological agent of enterically transmitted non-A, non-B hepatitis.

Abstract: Summary Virus-like particles (VLPs) with a mean diameter of 32 nm were recovered from the stools of three acute phase cases of enterically transmitted non-A, non-B hepatitis (ET-NANBH) occurring in the Soviet Union, North Africa and North America. VLPs from two of these cases were studied in detail and were shown to react specifically with antibody in acute phase sera obtained from other cases of ET-NANBH in Asia, the Soviet Union, North Africa and North America. Partially purified VLPs were found to sediment at 183S in sucrose gradients and to cross-react with antibody in acute phase sera from geographically isolated cases of ET-NANBH. The latter virus preparations were also used to document the seroconversion of experimentally ET-NANBH-infected cynomolgus macaques to 32 nm VLPs. Our findings indicate that one virus or class of viruses is responsible for the majority of ET-NANBH.

The fusion glycoproteins of human respiratory syncytial virus of subgroups A and B: sequence conservation provides a structural basis for antigenic relatedness.

Abstract: Summary Two major antigenic subgroups (A and B) have been described for human respiratory syncytial virus. The complete nucleotide sequence was determined for the fusion (F) mRNA of the subgroup B strain 18537 and the amino acid sequence of the F protein deduced, for comparison with the previously described sequences for the A2 strain of subgroup A. The F proteins (excluding the cleaved signal peptide) were 91% identical between the subgroups, consistent with the previously described high degree of antigenic relatedness. The greatest divergence occurred within the F2 subunit immediately preceding the cleavage activation site.

The Complete Nucleotide Sequence of the Genome of a Hepatitis B Virus Isolated from a Naturally Infected Chimpanzee

Abstract: The complete nucleotide sequence of a strain of hepatitis B virus, originally isolated from a naturally infected chimpanzee, has been determined. Interesting features of the sequence include the presence of an in-phase stop codon in the 'pre-core' region of the core antigen open reading frame. The sequence shows approximately 10% nucleotide divergence from all of the other hepatitis B virus sequences previously published and the possibility that this divergence is the result of passage through chimpanzees is discussed.

Effects of Mutagenesis in vitro on the Ability of Cloned Tomato Golden Mosaic Virus DNA to Infect Nicotiana benthamiana Plants

Abstract: Summary Insertion mutations introduced in vitro into cloned DNA of tomato golden mosaic virus that considerably shortened the length of the open reading frames (ORFs) AL1, AL2/AL3, BL1 or BR1, abolished the ability of the DNA to infect Nicotiana benthamiana seedlings. Mutants in which ORF AR1 was similarly shortened by an insertion or a 28 bp deletion were infectious, showing that the formation of coat protein or virions is not required for replication and systemic spread of virus DNA, although the appearance of symptoms was delayed in infections with the deletion mutant. Mutants with larger deletions (178 bp to 603 bp) in ORF AR1 were not infectious. Infections could be initiated with mixtures of AL1 and AL2/AL3 mutants, or BL1 and BR1 mutants, primarily as a result of complementation, although a low proportion of wild-type DNA A molecules regenerated by recombination or reversion was detected in the progeny of infection with the DNA A mutants.

The effect of sequences in the 5' non-coding region on the replication of polioviruses in the human gut.

Abstract: The genomic sequences of a portion of the 5' non-coding region of polioviruses excreted by recipients of the Sabin vaccine strains of poliovirus were examined. It was found that in all three types a residue changed in the course of excretion to give a sequence representing the consensus of poliovirus sequences. For type 1 this residue was at position 481, which altered in half the vaccines, for type 2 it was at position 480, which altered in all vaccinees and for type 3 it was at position 472 in all vaccinees, as previously reported. Other vaccine strains unrelated to the Sabin strains but used in limited vaccination programmes did not have such changing residues. It was concluded that these regions indicated sequences that were strongly selected during replication in the human gut. Such sequences were further defined by examination of strains of poliovirus unrelated to the vaccine strains, the sequences of which may be expected to vary at less stringently defined regions.

Gene Mapping and Positive Identification of the Non-structural Proteins NS2A, NS2B, NS3, NS4B and NS5 of the Flavivirus Kunjin and Their Cleavage Sites

Abstract: Partial N-terminal amino acid analyses of five radiolabelled non-structural (ns) proteins specified by Kunjin (KUN) virus provided positive identification of NS3, NS5 and three previously hypothetical ns proteins of flaviviruses, ns2a, ns2b and ns4b. Their correct gene order was obtained from their deduced amino acid sequences. Thus the gene order for KUN virus relative to that proposed for yellow fever (YF) virus was as follows: KUN 5'...GP44.P19.P10.P71.(?).P21.P98-3', YF 5'...NS1.ns2a.ns2b.NS3.ns4a.ns4b.NS5 -3'. The identity of GP44 as NS1 was assumed from the known nucleotide and deduced amino acid sequences; ns4a was not identified. The cleavage sites in the polyprotein for KUN NS2B, NS3 and NS5 were identical, Lys-Arg decreased Gly, similar in form to the sequence Arg-Arg decreased Ser defined at the cleavage sites of YF NS3 and NS5. A new consensus cleavage site for NS1, NS2A and NS4B in the form Val-X-Ala decreased, where X is any one of several uncharged amino acids, was found at corresponding sites homologous to those of KUN virus in all published flavivirus sequences (a total of 18 sites). NS1 and NS4B, but not NS2A, were preceded by a putative signal sequence.