- Volume 40, Issue 2, 1978
Volume 40, Issue 2, 1978
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Studies on Cotia Virus — an Unclassified Poxvirus
More LessSUMMARYThis paper is a report of studies on Cotia virus; this had been first isolated in 1965 in Brazil and was subsequently shown to be a poxvirus.
Cotia virus grew in a wide range of cell cultures and on the chick chorioallantois (CAM). Its growth characteristics are similar to those of other poxviruses. Microscopy showed virus factories or type B inclusions appearing before infectious progeny virus could be demonstrated. Type A inclusions appeared later, after development of progeny virus; these were shown by electron microscopy to differ from the type A inclusions of cowpox and other poxviruses and they have been termed Cotia bodies. Immunofluorescent staining also showed ring structures which appeared before the development of Cotia bodies. The growth of Cotia virus in human embryo lung (HEL) cells was sensitive to inhibitors of DNA and protein synthesis but was resistant to a concentration of rifampicin which inhibited vaccinia virus. Sharing of antigens between the Cotia virus and vaccinia virus was shown by gel precipitation tests and immunofluorescent staining. There was no cross neutralization between Cotia virus and vaccinia virus nor did anti-Cotia sera neutralize representatives of other poxvirus groups.
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Electron Microscopic Observation of African Swine Fever Virus Development in Vero Cells
More LessSUMMARYAfrican swine fever virus emerges from infected Vero cells either from areas of smooth cell surface or from microvilli. The two patterns may occur at different sites on the same cell and are unique for this virus. The scanning electron micrographs supplement regular thin section views.
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Observations on the Fine Structure of Lymphocystis Virus from European Flounders and Plaice
More LessSUMMARYLymphocystis virus has been classified with the icosahedral cytoplasmic deoxyriboviruses (ICDVs) but a detailed structure of the virion has not yet been established. Virus from tumours of European plaice and flounders was allowed to decay and this revealed details of its structure. Freshly prepared virus has a hexagonal outline which breaks down to reveal surface subunits 4.5 nm in diam. and a spheroidal core with tubular material 13 nm in diam. inside it. This tubular material appears to have a periodicity suggesting a helical conformation and may be a deoxyribonucleoprotein. No triangular or pentagonal facets, as found with some other ICDVs, were seen. A possible construction of the virus is proposed.
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Physical and Chemical Properties of Cynosurus Mottle Virus
More LessSUMMARYParticles of cynosurus mottle virus (CyMV) purified from systemically infected oat leaves have a sedimentation coefficient of 113S, buoyant density of 1.399 g/ml in CsCl and 1.332 in Cs2SO4, diffusion coefficient of 0.130 × 10−10 m2/s and E 0.1% 260 of 7.3. Virus particles are isometric with a diameter of 27 to 29 nm in negatively stained preparations; the hydrodynamic diameter was calculated from the D 20, w as 32.8 nm. The particle mol. wt. was calculated as 6.86 × 106. CyMV particles contain 22.5% single-stranded RNA of mol. wt. approx. 1.5 × 106 and a molar nucleotide composition of: A, 24.2%; G, 24.3%; C, 26.2% and U, 25.3%; RNA purified by the phenol-SDS method was infectious. The melting profile of native CyMV-RNA was typical of a single stranded RNA with secondary structure; the hyperchromicity was 33% and the Tm 61 °C. The protein mol. wt. was 30500. The completed cryptogram for CyMV is R/1:1.5/22.5:S/S:S/Ve/Ap.
CyMV particles were degraded by 0.02 to 0.05% SDS at pH 7.0 into a component with a sedimentation coefficient of 83S and by 0.1% SDS into the RNA and protein constituents; virus dissociation was enhanced by EDTA and reduced by MgCl2. At pH 5.0, the virus was not affected by SDS concentrations of up to 8%. The virus was unstable in EDTA and NaCl at pH 8.25.
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Photochemical Inactivation of DNA and RNA Viruses by Psoralen Derivatives
More LessSUMMARYWestern equine encephalitis virus, and RNA virus, and herpes simplex virus type 1, a DNA virus, were efficiently inactivated in less than 1 min by exposure to long-wave ultraviolet light (320 to 380 nm) in the presence of several psoralen derivatives. The psoralen photochemical reaction was chosen for study due to its known specificity for nucleic acids. Neither the light nor any of the drugs alone caused appreciable inactivation. The inactivation kinetics and dependence on light intensity and on different derivatives of psoralen were studied. The high solubility of a new aminomethyl psoralen derivative was found to be advantageous in the photochemical inactivation of the RNA virus, but was not in the case of the more easily inactivated DNA virus. Within its limited solubility range trimethylpsoralen was superior to its aminomethyl derivative on a molar basis for the inactivation of both types of viruses under most of the conditions studied.
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The Use of the Enzyme Linked Immunosorbent Assay to Detect a Nuclear Polyhedrosis Virus in Heliothis armigera Larvae
More LessSUMMARYThe enzyme linked immunosorbent assay (ELISA) has been shown to be a specific and sensitive method to detect a nuclear polyhedrosis virus in Heliothis armigera larvae. 1 ng/ml of purified virus particle antigen could be detected. The presence of larval extracts did not interfere in the assay. The growth of the virus in Heliothis armigera larvae was analysed by ELISA and the conventional counting of virus polyhedra and this showed that ELISA was a superior detection method.
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Defective Herpes Simplex Virus DNA: Circular and Circular-linear Molecules Resembling Rolling Circles
SUMMARYThe formation of defective herpes simplex virus (HSV) in BSC-1 cells and the synthesis of defective virus DNA was studied. The fourth consecutive passage of undiluted virus yielded defective DNA that was 0.008 g/ml more dense than wild type (w.t.) virus DNA. The amount of defective DNA increased at passage 6 concomitantly with the decrease in infectious virus progeny. The synthesis of defective DNA was always accompanied by w.t. virus DNA synthesis. Defective DNA from both infected nuclei and defective virions had a mol. wt. of 100 × 106 and was linear as determined by electron microscopy. Electron microscopy of defective virus DNA at passage 6 revealed circular molecules varying in size in addition to linear DNA molecules with the length of intact virion DNA. The circular DNA molecules had contour lengths of 10, 5, 2.5 and less than 2.5 µm. The smallest circular DNA molecules had a contour length of 0.3 µm, possibly one virus gene. In addition, circular-linear DNA molecules were observed in which both the circular and the linear components varied in length. Most of these DNA molecules had circular components of either 2.5 or 5.0 µm, and linear components varying in length from less than 1 to 50 µm. Based on the present study, it is proposed that the S component of w.t. virus DNA is fragmented into small circular molecules that serve as templates for DNA synthesis, possibly by the rolling circle mechanism.
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Increase by Calcium in Production of Interferon by L929 Cells Induced with Polyriboinosinate-Polyribocytidylate Complex
More LessSUMMARYCalcium chloride (5 to 20 mm) potentiated interferon production induced by rIn:rCn in L929 mouse fibroblasts up to a thousand-fold. Higher concentrations of calcium (20 to 65 mm) mixed with rIn:rCn were associated with increased cytotoxicity and a more acidic medium, but were effective in enhancing interferon production if preparations were adjusted to a uniform pH. Although calcium increased cellular binding of 3H-rCn:rIn, only a partial correlation between binding and interferon production was observed.
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ELISA in the Study of Homologous and Heterologous Reactions of Plant Viruses
More LessSUMMARYSerological relationships between plant viruses within six different taxonomic groups were studied by means of the double antibody sandwich form of enzyme-linked immunosorbent assay (ELISA). This technique was very sensitive not only for the detection of low concentrations of viruses, but also for the differentiation of closely related antigens. With some viruses, e.g. Andean potato latent virus, the specificity of the test was so great that conjugates prepared to one strain failed to detect other serologically closely related strains. With other viruses and with host proteins the cross-reactivity was broader, but distant and even intermediate serological relationships between viruses were usually not detected. The great specificity of ELISA is mainly caused by the behaviour of the conjugated antibodies. The binding of conjugated antibodies was completely inhibited by simultaneously added native antibodies. Antibodies from very early bleedings were not suitable for ELISA.
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Replication of Herpesviruses in Human Cells Transformed by Cytomegalovirus
More LessSUMMARYThe replication of human cytomegalovirus (CMV) and herpes simplex virus (HSV) was studied in three human embryo cell lines (CMV-Mj-HEL-1, CMV-Mj-HEL-2, and CMV-Mj-HEL-2,T-1) transformed in vitro by human CMV. Growth studies revealed that these cells were completely resistant to infection by CMV strains AD169 and Mj and partially resistant to HSV types 1 and 2. Neither virus DNA nor virus proteins were synthesized in the transformed cells infected with CMV AD169. The HSV production in CMV-transformed human embryo lung (HEL) cells was delayed when compared to the virus production in normal HEL cells and spread of HSV c.p.e. was slower in the transformed cells. The treatment of normal HEL cells with a crude extract of CMV-transformed HEL cells also resulted in inhibition of the spread of c.p.e. of HSV types 1 and 2. The inhibitory effect was not due to interferon since vesicular stomatitis virus replication was not affected and several experiments showed that it was not due to mycoplasma. The presence of virus inhibitor molecules in CMV-transformed cells absent in normal HEL cells is postulated.
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Complement-mediated Cytolysis of HSV-1 and HSV-2 Infected Cells: Plasma Membrane Antigens Reactive with Type-specific and Cross-reactive Antibody
More LessSUMMARYSurface antigens of BHK-21 cells infected with HSV-1 or HSV-2 were radio-iodinated (125I) with lactoperoxidase, immune precipitated and analysed by polyacrylamide gel electrophoresis (PAGE). Experiments using antiserum to HSV-1 or HSV-2, absorbed with appropriate homotypic or heterotypic antigens, revealed that both type-specific (homotypic) and cross-reactive antibody combined with surface glycoproteins to form a single large radioactive peak. This peak, which constituted the major glycoprotein region (region a) observed in electropherograms, represented a range in mol. wt. from 115000 to 130000. Sensitization of cells to complement lysis, neutralization of infectious virus and immune precipitation of surface glycoproteins (region a) were found to be generally correlated properties of all the antibody preparations analysed, including antibody prepared specifically against region a antigens. These findings suggest a major immunological role for the surface glycoproteins migrating in PAGE region a.
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Characterization of Kawino Virus, an Entero-like Virus Isolated from the Mosquito Mansonia uniformis (Diptera: Culicidae)
More LessSUMMARYSome properties of a small RNA virus isolated from Mansonia uniformis are described. The virus particles have a sedimentation coefficient of about 165S, buoyant density in CsCl of 1.33 g/ml and diameter of 28 nm. They contain a single-stranded RNA which probably lacks poly (A) but is infective. Four polypeptides with mol. wt. of 33, 30, 27 and about 7 × 103 are present in the virus particle.
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Metabolism of Myoinositol in Avian and Mammalian Cells Infected with Naked and Enveloped DNA and RNA Viruses
More LessSUMMARYThe uptake of 3H-inositol into the pool of free inositol and its incorporation into the lipid phosphatidylinositol have been studied in various avian and mammalian cells infected by different viruses. In all the virus-cell systems investigated, virus infection results in a drastically reduced amount of free 3H-inositol about 3 to 5 h post-infection, demonstrable in the infected cells as compared to the mock-infected controls. In contrast, the incorporation of 3H-inositol into lipid can be enhanced, reduced, or not influenced at all, depending on the virus-cell system under observation.
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Conformational Studies on Particles of Turnip Yellow Mosaic Virus
More LessSUMMARYCircular dichroism studies (CD) of turnip yellow mosaic virus (TYMV) nucleoprotein and of its isolated RNA and capsid revealed that: (i) the nucleic acid structure, which comprises a considerable amount of base pairing and/or stacking, remains essentially unchanged irrespective of whether the RNA is encapsidated or free; (ii) the secondary structure of the protein component is mainly accounted for by β- and irregular forms without appreciable amounts of α-helix; (iii) the interaction of capsid protein and RNA induces some conformational changes in the protein probably involving a decrease of β-structure and a perturbation of the microenvironment of some aromatic residues. The influence of temperature on the CD spectra of virus nucleoprotein, RNA and capsid was also investigated. The results are discussed in connection with particle stability.
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Biochemical Study of Certain Enzymes and Metabolites of the Carbohydrate Metabolism in the Skeletal Muscle of the Dengue Virus-Infected Mice
More LessSUMMARYChanges in enzymes and metabolites of the carbohydrate metabolism in skeletal muscles were studied in mice after intracerebral inoculation of dengue type 2 virus. It was noted that lactic dehydrogenase, aldolase, phosphogluco-isomerase, phosphoglucomutase, GO-T and GP-T activity were enhanced initially by two- to three-fold, reaching a peak on day 5. As the illness appeared in mice, all the enzyme activities were lowered and were about three times less in the paralytic stage on the 8th day as compared to controls. Fructose-1,6-diphosphatase activity was increased on the 4th and 5th days but decreased later. Acid phosphatase increased abruptly from the 6th day while alkaline phosphatase activity was irregular. Creatine increased on the 4th and 5th days but diminished later. Glycogen decreased from the beginning and was lowest on the 5th day, but the levels increased later and were maximum in paralysed muscles. On the other hand, lactic acid began accumulating in the muscles and was maximum on the 5th day, then declined. Dengue virus was detected in the muscles from the 2nd day but higher titres were seen from the 6th day. Changes similar to the preparalytic stage of mice may occur in human beings, causing myalgia.
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Potentiation of the Neutralization of Feline Leukaemia Virus in Hypotonic Medium
More LessSUMMARYThe neutralization of feline leukaemia virus could be enhanced by performing the reaction in hypotonic medium. Absorption of antibody by virus specific antigens was also made more sensitive under these conditions.
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Sensitivity of the Transforming and Replicative Functions of Epstein-Barr Virus to Inhibition by Phosphonoacetate
More LessSUMMARYDisodium, phosphonoacetate (PA), at concentrations of 50 to 200 µg/ml, which still allowed continued growth of the EB virus-transformed B95-8 cell line on a routine culture regimen, was able to inhibit the production of virus capsid antigen and of virus particles by these cells down to very low but finite levels which persisted despite prolonged treatment. Further experiments measured the effects of these same drug concentrations on the EB virus-induced in vitro transformation of foetal cord blood lymphocytes and on the colony forming ability of already established EB virus-transformed foetal cell lines; in both types of culture, doses of PA up to and including 50 µg/ml did not affect cell growth within the 8-week observation period, whereas doses of 100 µg/ml and above were increasingly inhibitory. The cell lines established by EB virus-induced transformation in the continual presence of PA at 50 to 150 µg/ml contained multiple copies of the virus genome per cell just as did the corresponding cell lines established in control medium. The results argue against the existence of any PA-sensitive event unique to the EB virus-induced transformation process.
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In Vitro Infectivity of the Autographa californica Nuclear Polyhedrosis Virus Under Various Conditions
H. A. Wood and G. SpeyerSUMMARYThe stability of the in vitro infectivity of the non-occluded virions (NOV) of the Autographa californica nuclear polyhedrosis virus (Ac-NPV) was tested. Exposure of Ac-NPV NOV to solutions ranging from pH 7.5 to 11.4 with molarities of 0.1 to 0.001 showed that infectivity was most stable in 0.001 m, pH 7.5, tris(hydroxymethyl)aminomethane (tris)-HCl buffer. Infectious NOV could be concentrated by precipitation with 50% saturated ammonium sulphate or 9% (w/w) polyethylene glycol 4000. Treatment of concentrated preparations with 1 m-urea did not affect infectivity or the centrifugal properties of the NOV. NOV were partially purified by centrifugation through a sucrose underlay.
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Development of Early Nuclear Antigen in Cytomegalovirus Infected Cells in the Presence of RNA and Protein Synthesis Inhibitors
More LessSUMMARYHuman cytomegaloviruses (HCMV) induce nuclear antigens as early as 1 h after infection as detected by immunofluorescence. Early antigen development occurs in the presence of DNA, RNA, and protein synthesis inhibitors and is dependent on intact viral DNA in the inoculum.
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Infection with BMV of Mesophyll Protoplasts Isolated from Five Plant Species
I. Furusawa and T. OkunoSUMMARYMesophyll protoplasts isolated from plants of wheat, oat, maize (a local lesion host) and Japanese radish (normally a non-host) were efficiently infected with brome mosaic virus (BMV) without using poly-l-ornithine, and the optimum conditions for infection were very similar to those found using barley protoplasts. These results suggest that the host ranges of viruses can be extended by using protoplasts and that the conditions optimal for infection of protoplasts with BMV are determined mainly by the virus.
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A Semi-continuous System for the Production of Human Interferon in Lymphoblastoid Cell Cultures
More LessSUMMARYThe simultaneous occurrence of virus replication and of interferon production in Namalva lymphoblastoid cell cultures infected with measles virus has allowed the development of a semi-continuous production system yielding about 104 units of interferon for each 106 cells.
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Attachment of Herpes Simplex Virus to Neurons and Glial Cells
More LessSUMMARYCells of brain tissue of rabbits, rats and mice were dissociated and glial cells, neuronal perikarya and synaptosomes were separated by centrifugation on discontinuous Ficoll gradients. HSV was shown to attach well to rat and rabbit glial cells and synaptosomes but not to neuronal perikarya. Of intracerebrally infected mice the fractions with glial cells contained the infective virus. The interactions between HSV and neuronal cells and the implication of the observations on the HSV infection of the nervous system are discussed.
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Comparative Protein Analysis of Non-salmonid Fish Rhabdoviruses
More LessSUMMARYOn the basis of gel precipitation, reactions with rabbit anti-pike fry rhabdovirus sera, a recent virus isolate (V75/94) from pike fry, and the grass carp rhabdovirus were found to be indistinguishable from pike fry rhabdovirus. The proteins of these viruses were compared to those of spring viraemia of carp virus. In ‘rocket’ immunoelectrophoresis the pike fry rhabdoviruses and the grass carp rhabdovirus showed up to 4 lines with homologous antiserum in which the precipitates formed by the G and M proteins were fused. With this antiserum, spring viraemia of carp virus gave only one precipitation are which was formed by the N protein. All the viruses are very similar to vesicular stomatitis virus with respect to the mol. wt., localization and phosphorylation of structural proteins. In polyacrylamide-SDS slab gels the viruses showed minor differences in the migration of their respective NS proteins.
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Cytological Alterations in Tissues of Gomphrena globosa Plants Systemically Infected with Tomato Bushy Stunt Virus
More LessSUMMARYSpecial environmental conditions were used to induce systemic infection by tomato bushy stunt virus (TBSV) in Gomphrena globosa. Severe mottle symptoms were followed by the formation of necrotic lesions. The mottled tissue consisted of small patches of necrotic, chlorotic and green tissue closely intermingled. The ultrastructure of such systemically infected G. globosa leaves was investigated. In the mesophyll and bundle sheath cells of the chlorotic patches the chloroplasts were severely altered and often connected to multivesicular (MV) bodies, whose vesicles and vacuoles contained fibrillar material. The MV bodies are thought to originate from chloroplasts and may be involved in TBSV replication. Virus appeared in the cytoplasm and central vacuole as scattered particles or small paracrystalline aggregates. Dark spots, probably proteinaceous material, were present in the cytoplasm. The cells of the necrotic patches appeared completely collapsed and contained virus particles. The necrotic lesions were cytologically quite similar to those produced in hypersensitive conditions.
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Improved Detection of Virus-Specific IgM Antibodies. Elimination of Non-Specific IgM Binding
More LessSUMMARYA non-specific reaction between human IgM and cytoplasmic structures of virus infected cells can often be observed if IgM antibodies to virus antigens are detected by indirect immunofluorescence or by immuno enzyme assays. Formaldehyde selectively inactivates the cytoplasmic receptors for human IgM without affecting the virus structural proteins. Alternatively, receptor-free antigens can be obtained by isolation of nuclei from virus infected cells. Due to reduced back-ground, a more specific and more sensitive detection of IgM antibodies to Epstein-Barr virus, cytomegalovirus or central European encephalitis virus is possible.
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Neutralization and Enhancement of Infectivity of Non-salmonid Fish Rhabdoviruses by Rabbit and Pike Immune Sera
More LessSUMMARYPlaque neutralization assays were carried out using the rhabdoviruses of pike fry (2 isolates), spring viraemia of carp and grass carp. When rabbit anti-pike fry rhabdovirus sera were used, no neutralizing activity was observed but plaque counts increased with heat-inactivated sera or sera that had been stored at -20 °C for prolonged periods. Antibody binding to the virus was demonstrated by gradient centrifugation experiments. The use of immune serum alone or in combination with anti-rabbit gamma globulin serum resulted in aggregation of radioactive virus. Aggregation was also observed when anti-rabbit gamma globulin serum was added under neutralization test conditions. Addition of non-inactivated guinea-pig serum to mixtures of pike fry rhabdovirus and rabbit antiserum to spring viraemia of carp virus resulted in strong neutralization, whereas this antiserum alone or in the presence of inactivated guinea-pig serum gave an increase in plaque numbers. The increase in plaque number and the complement-dependent neutralization phenomena were weak or absent in the homologous spring viraemia of carp virus system.
Immunization of pike with pike fry rhabdovirus resulted in an increase in virus neutralizing activity which could be attributed to the macroglobulin fraction of the serum. After immunizations over a period of 6 months with virus antigens, no further increase in neutralizing activity was observed. Pike sera showed 50% plaque reduction at concentrations between 0.017 and 0.3%. Using these sera it could be demonstrated that the grass carp virus and the V 75/94 isolate from pike fry were indistinguishable from the original pike fry rhabdovirus, whereas spring viraemia of carp virus was different.
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Properties of the Viruses Selected during Persistent Infection of L Cells with VSV
More LessSUMMARYTwo virus clones, VSV-mp and VSV-sp, were isolated from L cells persistently infected with VSV (New Jersey serotype). Both clones were more temperature sensitive than the parent virus, VSV-o, and grew more slowly, gave smaller plaques, less c.p.e. and lower virus yields in L cells. Unlike the parent virus, both persistent viruses induced interferon production in L cells. Stable carrier cultures could be obtained from L cells infected with VSV-sp at low multiplicities without pretreatment with interferon. This may be related to the fact that VSV-sp is more sensitive to interferon than either VSV-mp or VSV-o.
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Volume 105 (2024)
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