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ADNA virus was purified from the salivary gland affected by a large proliferation in the Diptera Merodon equestris. Structure, nucleic acid as well as development and intracellular cycle of the virus have been characterized.
BACKGROUND: Japanese encephalitis virus (JEV) is a zoonotic mosquito-borne Orthoflavivirus that circulates primarily in birds and pigs. Previous observations of vector-free transmission between pigs indicates the possibility of single-host cycling in swine. Therefore, the aim of this work was to investigate the evolutionary pressure of single host cycling using a relevant primary cell culture model. METHODS: To investigate whether such single-host cycles affect viral infectivity, fitness and genomic adaptations, two strains and a reverse genetic cDNA-derived clone of JEV were serially passaged 12 times in primary porcine monocyte-derived macrophages (MDMs), in Aedes albopictus-derived C6/36 cells, and alternately between both cell types. Next-generation sequencing analysis was used to identify selected single nucleotide variants (SNVs) and haplotypes. Phenotype-to-genotype connections were confirmed using reverse genetics. RESULTS: For all viruses, serial passaging in MDMs - but not in C6/36 cells - led to a rapid increase in relative infectivity toward MDMs, accompanied by reduced plaque sizes in porcine endothelial cells. In contrast to C6/36 cells, MDM imposed a strong selective pressure, rapidly favoring selection of many SNVs and viral haplotypes. In addition, we identified a dominant selection of mutants with glutamic acid to lysine substitutions at positions 49 or 138 in the E protein, which explained the small plaque phenotype and caused viral sensitivity to heparin-mediated inhibition of attachment, indicating enhanced virus binding to glycosaminoglycans (GAG). The E138K mutant also explained the increased relative infectivity for MDM. CONCLUSION: This work demonstrates a high evolutionary pressure on JEV in MDM causing rapid selections of minor haplotypes. Furthermore, the efficient selection of E49K and E138K SNV, which were responsible for the phenotype, are likely caused by a selective pressure for GAG binding, observed in vitro with other mammalian cells.
Four baculoviruses produced visible plaques in the Spodoptera frugiperda cell line when cell monolayers were infected and overlayed with a simplified, agarose-nutrient formulation. Macroscopic plaques were first detected 4 days postinoculation, and by 10 days plaques ranging from 0.5 to 3 mm in diameter were seen. Dose-response experiments indicated that a single particle initiated the formation of a plaque, because a linear response was demonstrated with increased dosage.
The behaviour of uncloned, low-passage Tahyna virus in an Aedes albopictus (AA) cell line at 28 and 20 degrees C was studied in the course of 10 passages. The virus multiplied at both temperatures without any apparent effect on the host cell. At 28 and 20 degrees C reduction of plaque size, decrease of peripheral virulence and weakening of thermostability were observed. Differences between both temperatures were only in the intensity of these changes.
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Maturation of mosquito densonucleosis virus (MDV) was studied in its natural host, Aedes aegypti L. larvae. First ultrastructural changes were observed in the cytoplasm of virus-infected cells. They consisted in the formation of paracrystalline structures containing particles 18--20 nm in diameter as well as microtubules 20 nm in diameter. Virogenic stromas and paracrystalline virion arrays were found in the nuclei of virus-infected cells.
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N-polyhedrin of inclusion bodies of two nucleopolyhedrosis viruses of Orgyia pseudotsugata was characterized. Alkali-dissolved N-polyhedrin from both virus strains was of similar size and consisted of 12S molecule of 209 000 daltons. Eight subunits of approximately 26 000 daltons were found to form the 12S molecules. N-polyhedrin from both viruses showed two main antigens by immunodiffusion. The subunits appear to possess one antigen and, upon formation of the 12S molecule, a new antigen is created. Both the subunit and 12S antigens from the two virus strains were shown to be antigenically related. The 12S molecule of both viruses also appears to possess a minor antigen unique to each virus.
The photometric method of quantitative determination of dry mass by electron microscopy has been applied to the study of various types of viruses: animal, plant, insect, and bacterial. The method is applicable to all viruses having a mass of 1 x 10-18g or greater. The molecular weight of viruses can be calculated from the mass value by multiplying it by Avogadro's number. In comparison to other methods of determining the molecular weight of viruses, sedimentation and diffusion, sedimentation equilibrium, light scattering, and electron microscopy counting, the method of quantitative electron microscopy is competitive. In some ways quantitative electron microscopy is superior to other methods for the determination of molecular weight: There is no limitation to the size of the virus, no experimental time involved and no concentration and purity of virus preparations required, and finally it is independent of the geometry of the virion. In one important aspect it is unique when compared to other methods; namely, it affords one the capacity to analyse individual virus particles.
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An iridovirus, Apis iridescent virus (AIV), isolated from sick adult specimens of Apis cerana (Hymenoptera) from Kashmir, closely resembles iridescent viruses from Tipula and Sericesthis spp. (TIV and SIV). However, AIV is only distantly related serologically to TIV and SIV and is even more remotely related to several other similar viruses that were tested in tube precipitation tests with intact particles. AIV multiplies in Apis mellifera, forming cytoplasmic iridescent crystalline aggregates in several tissues, but unlike all the other iridoviruses tested, it failed to multiply in Galleria mellonella.
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