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C R Pringle

Publications and source records attributed to C R Pringle.

At least 91 records · Page 5Linked to original sources
PubMed1982

Mutant identifying a third recombination group in a bunyavirus.

Only two recombination groups have been reported in genetic analyses of ts mutants of 10 different bunyaviruses from the Bunyamwera and California encephalitis serogroups, although three groups are expected from the tripartite structure of the genome of all members of the family Bunyaviridae. We describe now a ts mutant of Maguari virus, MAGts23(III), which recombined in both vertebrate (BHK-21) and invertebrate (Aedes albopictus) cells with mutants representing recombination groups I and II of this Bunyamwera serogroup virus. In addition, MAGts23(III) recombined with two mutants MAGts20 and MAGts21, provisionally identified as double mutants by their failure to recombine with group I or group II mutants, Mutant MAGts23(III) therefore represents a third bunyavirus recombination group. Mutant MAGts23(III) differed phenotypically from other bunyavirus mutants by growth restriction in BS-C-1 cells. Wild-type recombinants were obtained in the heterologous cross of MAGts23(III) and a group II mutant of Bunyamwera virus, but not in a cross with a group I mutant. The recombinants had the G protein of the Maguari virus parent and the N protein of the Bunyamwera virus parent. Analysis of the phenotypes of clones isolated at permissive temperature from the progeny of the other cross [MAGts23(III) and a group I mutant of Bunyamwera virus] indicated that recombination occurred in this cross, but that the possible recombinant phenotypes were not recovered with equal frequency. As a consequence, it has not been possible to obtain a gene assignment for group III from genetic data alone.

Bunyamwera virus↗
PubMed1981

Acute fatal pneumonia in calves due to respiratory syncytial virus.

An acute pneumonia developed in 28 calves which had been housed together from one to two weeks of age. The clinical signs included pyrexia, tachypnoea, respiratory distress and coughing. Some of the calves died. The pneumonia was characterised by an alveolitis with multinucleated syncytia, alveolar epithelial hyperplasia and bronchiolitis. Interstitial emphysema was also present. Fifteen of 19 calves examined serologically had rising neutralising antibody titres to respiratory syncytial virus; in nine calves the rise was fourfold or greater. Respiratory syncytial virus was not isolated from the calves. There was no evidence of parainfluenza type 3 virus involvement. The adult cows being sucked by the calves remained clinically normal throughout the incident. Six calves examined six weeks after the outbreak started had a chronic cuffing pneumonia characterised by lymphocytic bronchiolitis; some of the calves also had bronchiolitis obliterans. Mycoplasma dispar was found in two of them.

Acute Disease↗
PubMed1981

Antigen and polypeptide synthesis by temperature-sensitive mutants of respiratory syncytial virus.

A revised nomenclature for the polypeptides of respiratory syncytial (RS) virus has been devised on the basis of comparison of the Long, A2 and RSN-2 strains by slab-gel electrophoresis. Seven polypeptides, now designated VP200, VGP48, VPN41, VPP32, VPM27, VP25 and VP10, were observed in preparations of all three strains of RS virus, irrespective of the host cell of origin. In addition, a slowly migrating glycopolypeptide GP1 was prominent in partially purified RS virus of the Long and A2 strains obtained from Hep-2 cells, and to a lesser extent from BS-C-1 cells. In the case of the RSN-2 strain, this polypeptide was only resolved clearly in virus obtained from Hep-2 cells. GP1 was an atypical glycopolypeptide in that 35S-methionine incorporation was poor relative to 3H-glucosamine incorporation. The ts mutants of RS virus exhibited four distinct phenotypes with respect to intracellular polypeptide synthesis and antigen production of 39 degrees C. Mutants ts 17 (complementation group B') and ts 19 (group E) were almost completely restricted, suggesting defective early functions. Mutants ts A1 (group A), ts A7 (group C) and ts 1 (group D) synthesized antigen and polypeptides normally, but the amount of antigen at the cell surface was reduced, suggesting maturation defects. In addition, the VPP32 of ts 1 (group D) exhibited an aberrant mobility, confirming its viral specificity. The remaining mutants, representing groups B, F and G exhibited generally impaired synthesis at 39 degrees C. Absence of surface filaments in ts mutant-infected cells at 39 degrees C confirmed their virus-specific nature.

Antigens, Viral↗
PubMed1981

Genetic interactions among viruses of the Bunyamwera complex.

Seventy-seven temperature-sensitive (ts) mutants belonging to three antigenically distinct and geographically isolated members of the Bunyamwera complex--Batai virus, Bunyamwera virus, and Maguari virus--have been isolated after 5-fluorouracil treatment. High-frequency recombination was observed, and the mutants of each virus were classified into two groups, which were shown to be equivalent by heterologous recombination experiments. In most combinations heterologous recombination was less efficient than homologous recombination, but all crosses of group I and II mutants yielded viable recombinants. Recombination was an early event. Analysis by polyacrylamide gel electrophoresis of the proteins of the wild-type viruses and recombinant clones obtained from the six possible heterologous combinations of group I and II mutants indicated that recombination occurred by reassortment of genome subunits. Group I appeared to correspond to the genome subunit coding for the N protein, and group II corresponded to the G1/G2 determinant. The G1 (or G2 or both) protein was associated with neutralization specificity and plaque diameter, and the N protein was associated with plaque opacity. Complementation was observed between two nonrecombining mutants of Maguari virus belonging to group I, which may indicate that the N genome subunit codes for an additional protein. There appeared to be no genetic barrier to exchange of genetic material between Batai, Bunyamwera, and Maguari viruses in vitro, and it is concluded that the Bunyamwera complex is potentially a single gene pool if geographical and ecological constraints are discounted.

Bunyamwera virus↗
PubMed1981

Enhanced mutability associated with a temperature-sensitive mutant of vesicular stomatitis virus.

Temperature-sensitive (ts) mutant tsD1 of vesicular stomatitis virus, New Jersey serotype, is the sole representative of complementation group D. Clones derived from this mutant exhibited three different phenotypes with respect to electrophoretic mobility of the G and N polypeptides of the virion in sodium dodecyl sulfate-polyacrylamide gel. Analysis of non-ts pseudorevertants showed that none of the three phenotypes was associated with the temperature sensitivity of mutant tsD1. Additional phenotypes, some also involving the NS polypeptide, appeared during sequential cloning, indicating that mutations were generated at high frequency during replication of tsD1. Furthermore, mutations altering the electrophoretic mobility of the G, N, NS, and M polypeptides were induced in heterologous viruses multiplying in the same cells as tsD1. These heterologous viruses included another complementing ts mutant of vesicular stomatitis virus New Jersey and ts mutants of vesicular stomatitis virus Indiana and Chandipura virus. Complete or incomplete virions of tsD1 appeared to be equally efficient inducers of mutations in heterologous viruses. Analysis of the progeny of a mixed infection of two complementing ts mutants of vesicular stomatitis virus New Jersey with electrophoretically distinguishable G, N, NS, and M proteins yielded no recombinants and excluded recombination as a factor in the generation of the electrophoretic mobility variants. In vitro translation of total cytoplasmic RNA from BHK cells indicated that post-translational processing was not responsible for the aberrant electrophoretic mobility of the N, NS, and M protein mutants. Aberrant glycosylation could account for three of four G protein mutants, however. Some clones of tsD1 had an N polypeptide which migrated faster in sodium dodecyl sulfate-polyacrylamide gel than did the wild type, suggesting that the polypeptide might be shorter by about 10 amino acids. Determination of the nucleotide sequence to about 200 residues from each terminus of the N gene of one of these clones, a revertant, and the wild-type parent revealed no changes compatible with synthesis of a shorter polypeptide by premature termination or late initiation of translation. The sequence data indicated, however, that the N-protein mutant and its revertant differed from the parental wild type in two of the 399 nucleotides determined. These sequencing results and the phenomenon of enhanced mutability associated with mutant tsD1 reveal that rapid and extensive evolution of the viral genome can occur during the course of normal cytolytic infection of cultured cells.

Animals↗
PubMed1980

Temperature-sensitive mutants of Chandipura virus. I. Inter- and intragroup complementation.

Fifty temperature-sensitive (ts) mutants of Chandipura virus, a human rhabdovirus, have been classified into six complementation groups, designated ChI, ChII, ChII, ChIV, ChV, and ChVI and containing 44, 2, 1, 1, 1, and 1 mutants, respectively. Weak complementation was observed within group ChI, allowing the division of the group into subgroups ChIA and ChIB. Intragroup complementation was most extensive within subgroup ChIB, and one mutant in this subgroup complemented all but one (ts Ch598) of the mutants in group ChI. If ts Ch598 had been omitted from the analysis the number of complementation groups would have been increased to seven. Consequently, in circumstances where intragenic and intergenic complementation cannot be clearly distinguished, the number of complementation groups identified in rhabdoviruses could be overestimated. The identification of six complementation groups in three different rhabdoviruses need not imply the existence of an as yet unidentified sixth virus-specified polypeptide. The extensive intragroup complementation observed in Chandipura virus suggests that the functional form of one at least of the virion proteins of Chandipura virus is a multimer.

Genetic Complementation Test↗
PubMed1980

Temperature-sensitive mutants of Chandipura virus. II. Phenotypic characteristics of the six complementation groups.

Fifty temperature-sensitive (ts) mutants of the rhabdovirus Chandipura virus have been classified into six complementation groups designated ChI to ChVI. Group ChI contains 44 mutants, group ChII contains 2 mutants, and the remaining groups have 1 mutant each. Mutants in groups ChI, ChIII, ChIV, and ChVI had RNA-negative phenotypes in experiments measuring amplification of RNA synthesis at restrictive temperature. The two mutants in group ChII had RNA-positive phenotypes, and the virions were thermolabile. Mutant ts Ch851 of group ChV was also RNA positive, and the M polypeptide of this mutant appeared to be unstable in cells incubated at restrictive temperature. It is likely, therefore, that complementation groups ChII and ChV represent the genes coding for the two viral proteins of the virion envelope. No precise assignment can be made in the case of the four RNA-negative groups, since all the mutants examined showed some polymerase activity in vitro at restrictive temperature. An attempt to obtain polymerase mutants by screening for sensitivity to rifampin was not successful. Six temperature-dependent host range mutants (the tdCE phenotype) of Chandipura virus failed to multiply in chicken embryo cells at restrictive temperature, but otherwise they differed in their host range properties from similar mutants of vesicular stomatitis virus.

Animals↗
PubMed1980

Comparisons of nucleotide sequences in the genomes of the New Jersey and Indiana serotypes of vesicular stomatitis virus.

Nucleotide sequences of around 200 residues were determined adjacent to the 3' terminus of the genome RNA of vesicular stomatitis virus, New Jersey serotype, and adjacent to the 3'-terminal polyadenylic acid tract of the N protein mRNA of the same virus. These sequences were compared with the corresponding sequences previously determined for the Indiana serotype of vesicular stomatitis virus. The sequences obtained for the two strains were readily aligned, showing 70.8% homology overall. Examination of the sequences allowed identification of the translation initiation and termination codons for the N mRNA of each serotype. The deduced N-terminal and C-terminal amino acid sequences of the two N polypeptides were each similar, and most of the differences between them consisted of substitution by a clearly homologous amino acid. It was proposed that these nucleotide sequences, within limits imposed by their functions, comprise reasonably representative measures of the extent of sequence homology between the genomes of the two serotypes, and that this is higher than previously estimated, but with little exact homology over extended regions.

Amino Acid Sequence↗
PubMed1979

Feline syncytium-forming virus: identification of a virion associated reverse transcriptase and electron microscopical observations of infected cells.

The maturation of feline syncytium-forming virus (FSFV), a member of the foamy virus sub-family (Spumavirinae), has been studied by electron microscopy of thin sections of infected feline embryo (FEA) cells. The initial event observed was formation of crescent-shaped nucleoids at the plasma membrane. As budding progressed, the nucleoid became circular in outline with an electron-lucent centre in fully mature extracellular particles. These observations suggested that the maturation of FSFV in fully permissive FEA cells resembled that of C-type RNA tumour viruses, rather thant the B-type mouse mammary tumour virus. In this respect FSFV may be distinct from other foamy viruses. However, like other foamy viruses FSFV possessed reverse transcriptase activity. Polymerase activity co-sedimented with infectivity in an equilibrium density gradient and exhibited a preference for poly(rA).oligo(dT)10 over poly(dA).oligo(dT)10 as exogenous template.

Animals↗
PubMed1979

Feline syncytium-forming virus: DNA provirus size and structure.

An infectious DNA assay has been used to investigate the size and structure of the genome of feline syncytium-forming virus (FSFV). The dose response between DNA extracted from FSFV-infected cells and plaque number on feline embryo cells followed two-hit kinetics and the mol. wt. of the proviral DNA was estimated as approx. 6 x 10(6).

Animals↗
PubMed1979

Pneumoviruses: the cell surface of lytically and persistently infected cells.

Human embryonic lung (MRC-5), feline embryo (FEA), mink lung (Mv1Lu) and monkey kidney (BSC-1) cells infected by respiratory syncytial virus showed characteristic morphological changes when viewed by scanning electron microscopy. The surfaces of respiratory syncytial virus-infected cells developed a profusion of slender filaments after 48 h incubation at 31 degrees C. Similar changes in surface morphology were observed in BSC-1 cells infected by murine pneumonia virus. Filament production therefore appears to be a common property of pneumo-viruses. Filaments were not observed in cells infected with either syncytial and non-syncytial herpes simplex virus, the cytocidal vesicular stomatitis and Batai (Bunyaviridae) viruses, or the focus-inducing rabbit fibroma virus. Filament production was not observed in cells infected with ts mutants of respiratory syncytial (RS) virus during incubation at the restrictive temperature, or in a persistently infected culture of BSC-1 cells at 37 degrees C. The persistently infected cells (the RS ts 1/BSC-1 line) had some of the characteristics of cells transformed by oncogenic viruses, namely ability to overlap adjacent cells and agglutination by a low concentration of concanavalin A. The pseudo-transformed phenotype was temperature-dependent, however, and suppressed by raising the temperature of incubation to 39 degrees C. The presence of virus antigen at the cell surface was similarly temperature-dependent in these cells, diminished at high temperature (39 degrees C) and enhanced at low temperature (31 degrees C), suggesting that the changes in the host cell were the result of insertion of virus protein into the cell membrane. Evidently, persistent infection by a cytoplasmic virus can produce alterations in the host cell usually associated with transformation by nuclear viruses.

Animals↗
PubMed1979

Effect of temperature-sensitive mutation on activity of the RNA transcriptase of vesicular stomatitis virus New Jersey.

The virion-associated RNA transcriptase activity of vesicular stomatitis virus New Jersey temperature-sensitive (ts) mutants was assayed in vitro at the permissive (31 degrees C) and restrictive (39 degrees C) temperatures. RNA synthesis at 39 degrees C by the RNA-negative ts A1 and the RNA-positive ts C1 and ts D1 mutants was similar to that of wild-type virus. The RNA-negative ts B1 synthesized only small amounts of RNA in vitro at 39 degrees C. The three mutants of complementation group E were dissimilar in the amounts of RNA they synthesized at 39 degrees C: ts E1 synthesized very little RNA, ts E2 synthesized moderate amounts, and RNA synthesis by ts E3 was not inhibited. The two mutants of group F were also dissimilar, since ts F1 synthesized very little RNA at 39 degrees C, whereas ts F2 synthesized as much RNA as wild-type virus. The revertant clones ts B1/R1, ts E1/R1, and ts F1/R1 synthesized RNA at 39 degrees C in amounts comparable to wild-type virus, indicating that the heat sensitivity of the transcriptase activity of the mutants ts B1, ts E1, and ts F1 was associated with temperature sensitivity. Similar heat sensitivities were observed when transcribing nucleoprotein complexes were used in the assays, showing that the mutated polypeptides were part of the viral core. The heat stability of the mutant ts B1 was similar to that of wild-type virus, and in vitro RNA synthesis was fully restored when the temperature was lowered to 31 degrees C after 30 min of preincubation at 39 degrees C, showing that the inhibition was due to reversible configurational change of the mutated polypeptide. When virions of the mutant ts E1 were heated for 5 h at 39 degrees C, their infectivity and transcriptase activity were as stable as those of the wild-type virus, whereas transcriptase activity became very heat labile after disruption of the viral coat with a neutral detergent. This suggests an interaction between the mutated polypeptide and a coat polypeptide which stabilizes the activity of the transcriptase. The RNA transcriptase activity of the mutant ts F1 was also heat labile, although to a lesser extent than that of ts E1. Thus, the defects in transcriptase activity of groups B, E, and F suggest that all three polypeptides of the virus core, polypeptides L, N, and NS, are involved in the transcription. In addition, we postulate that the mutated gene products of groups E and F are multifunctional, being required both in transcription and replication, and that the gene product of group E may also be involved in some late stage of virus development.

DNA-Directed RNA Polymerases↗
PubMed1979

Temperature-sensitive mutants of complementation group E of vesicular stomatitis virus New Jersey serotype possess altered NS polypeptides.

In vesicular stomatitis virus New Jersey serotype polyacrylamide gel electrophoresis was unable to distinguish the polypeptides of the temperature-sensitive (ts) mutants of complementation groups A, B, C, and F from those of the wild-type virus. However, the NS polypeptide of the representative mutant of group E, ts E1, had a significantly greater electrophoretic mobility than that of the wild-type virus NS polypeptide. The electrophoretic mobilities of the NS polypeptides of the three mutants of complementation group E varied, being greatest in the case of ts E1, slightly less for ts E2, and only a little greater than that of wild-type virus NS polypeptide in the case of ts E3. Since the NS polypeptides of the revertant clones ts E1/R1 and ts E3/R1 have mobilities identical to that of wild-type NS polypeptide, the observed altered mobilities of the group E mutants are almost certainly the direct result of the ts mutations in the E locus. The electrophoretic mobilities of the intracellular NS polypeptides of the group E mutants were indistinguishable from those of their virion NS polypeptides. The electrophoretic mobilities of the NS polypeptides of the group E mutants synthesized in vitro using mRNA synthesized in vitro by TNP were identical to those of the NS polypeptides of their purified virions. The NS polypeptides of all three mutants were labeled with (32)P(i) to approximately the same extent as wild-type virus NS polypeptide, indicating that gross differences in phosphorylation of this polypeptide are unlikely to account for the altered mobilities. We propose a model in which the NS polypeptide consists of at least three loops held in this configuration by hydrophobic or ionic forces or both and stabilized by phosphodiester bridges. If a mutation affects one of the amino acids to which the phosphate is covalently linked, the phosphodiester bridge cannot be formed, and, as a result, in the presence of sodium dodecyl sulfate the affected loop opens and thus the NS polypeptide migrates further into the gel. Such a configuration may also explain the multifunctional nature of the NS polypeptide.

Genetic Complementation Test↗
PubMed1979

Rhabdoviridae. Report of the Rhabdovirus Study Group, International Committee on Taxonomy of Viruses.

The family Rhabdoviridae comprises approximately 75 viruses infecting vertebrates, invertebrates and plants. The main characteristics of the member viruses are: (i) the viruses infecting vertebrates and invertebrates are bullet-shaped and the viruses infecting plants are usually bacilliform; (ii) the viruses have particle lengths varying from 130 to 380 nm and widths varying from 60 to 95 nm; (iii) the viruses possess unit-membrane envelopes from which protrude spikes 5 to 10 nm long; (iv) the viruses have precisely coiled helical nuecleocapsids with a diameter of approx. 50 nm; (v) most of the viruses which have been studied contain 5 proteins; the prototype, vesicular stomatitis virus, contains proteins designated L (large), G (glycoprotein), N (nucleoprotein), NS (nonstructural) and M (matrix); N or NS is phosphorylated in most members which have been studied; (vi) the viruses contain single-stranded RNA which is transcribed into several messenger RNA species with sizes corresponding to the structural proteins; (vii) the nucleocapsid contains the RNA-dependent RNA polymerase and is infectious; and (viii) many of the viruses produce morphologically distinct defective-interfering (T) particles.

RNA, Viral↗