Taxonomic Structure of the Family
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Baculoviridae |
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One or two virion phenotypes may be involved in baculovirus infections. The virion phenotype that initiates infection in the gut epithelium is occluded in a crystalline protein matrix which may be (1) polyhedral in shape, range in size from 0.15 to 15 µm and contain many virions (genus Nucleopolyhedrovirus), or (2) ovicylindrical (about 0.3 0.5 µm) and contain only one, or rarely two or more virions (genus Granulovirus). Virions within occlusions consist of one or more rod-shaped nucleocapsids that have a distinct structural polarity and are enclosed within an envelope. For occluded virions, nucleocapsid envelopment occurs within the nucleus (genus Nucleopolyhedrovirus) or in the nuclear-cytoplasmic milieu after rupture of the nuclear membrane (genus Granulovirus). Nucleocapsids average 30-60 nm in diameter and 250-300 nm in length. Spike-like structures have not been reported on envelopes of the occlusion derived virions (ODV). If infection is not restricted to the gut epithelium, a second virion phenotype is required for infection of other tissues. Virions of the second phenotype (termed budded virions or BV) are generated when nucleocapsids bud through the plasma membrane at the surface of infected cells. BV typically contain a single nucleocapsid. Envelopes of the BV are derived from the cellular plasma membrane and characteristically appear as a loose-fitting membrane that contains terminal peplomers 14-15 nm in length composed of a glycoprotein (Fig. 1).
Physicochemical and Physical Properties
ODV buoyant density in CsCl is 1.18-1.25 g/cm3, and that of the nucleocapsid is 1.47 g/cm3. BV buoyant density in sucrose is 1.17-1.18 g/cm3. Virions of both phenotypes are sensitive to organic solvents and detergents. BV is marginally sensitive to heat and pH 8-12, inactivated by pH 3.0, and stable in Mg++ (10-1 M to 10-5 M).
Nucleocapsids contain a single molecule of circular supercoiled dsDNA, 80-180 kbp in size.
Genomic analyses suggest that baculoviruses encode 100-200 proteins. Virions may contain from 12 to 20 or more different polypeptides. Nucleocapsids from both virion phenotypes (ODV and BV) contain a major capsid protein, a basic DNA binding protein that is complexed with the viral genome, and at least 2-3 additional proteins. BV contains a major envelope glycoprotein (the peplomer protein) that serves as a membrane fusion protein. Several ODV envelope proteins have been identified. The major protein of the occlusion body matrix is a viral encoded polypeptide, Mr = 25-33 103. This protein is called polyhedrin for nucleopolyhedroviruses and granulin for granuloviruses. The occlusion body is surrounded by an envelope that contains at least one major protein.
Lipids are present in the envelopes of ODV and BV. Lipid composition differs between the two virion phenotypes.
Carbohydrates are present as glycoproteins and glycolipids.
Genome Organization and Replication
Circular genomic DNA is infectious suggesting that after cellular entry and uncoating, no virion-associated proteins are essential for infection. Transcription of baculovirus genes is temporally regulated, and two main classes of genes are recognized: early and late. Late genes may be further subdivided as late and very late. Gene classes (early, late, and very late) are not clustered on the baculovirus genome, and both strands of the genome are involved in coding functions. Early genes are transcribed by host RNA polymerase II, while late and very late genes are transcribed by an alpha-amanitin resistant RNA polymerase activity. RNA splicing occurs, but appears to be rare since only one spliced gene has been identified. Transient early and late gene transcription and DNA replication studies suggest that at least three virus encoded proteins regulate early gene transcription, while approximately 19 viral encoded proteins known as Late Expression Factors (LEFs) are necessary for late gene transcription. Of the approximately 19 LEFs, 9-10 appear to be involved in DNA replication. Late gene transcription initiates within or near a highly conserved 5-TAAG-3 sequence, which appears to be an essential core element of the baculovirus late promoter. Putative replication origins consist of repeated sequences found at multiple locations within the baculovirus genome. These sequences, termed homologous repeat (hr) regions, do not appear to be highly conserved between different baculovirus species. Single copy, non-hr putative replication origins have also been identified. DNA replication is required for late gene transcription. Most structural proteins of the virions are encoded by late genes. While transcription of late and very late genes appears to begin immediately after DNA replication, some very late genes that encode occlusion-specific proteins are transcribed at extremely high levels at a later time. BV production occurs primarily during the late phase, and occlusion production occurs during the very late phase (Fig. 2).
In infected animals, viral replication begins in the midgut (insects) or digestive gland epithelium (shrimp) of the arthropod host. Following ingestion, occlusions are solubilized in the gut lumen releasing the enveloped virions which are thought to enter the target epithelial cells via fusion with the plasma membrane at the cell surface. In lepidopteran insects, viral entry into midgut cells occurs in an alkaline environment, up to pH 12. Infection of the midgut is required for initiation of infection in the animal. In most cases, the virus is believed to undergo one round of replication in the midgut epithelium prior to transmission of infection to secondary tissues within the hemocoel. A mechanism for direct movement from the midgut to the hemocoel has also been proposed. DNA replication takes place in the nucleus. In granulovirus-infected cells, the integrity of the nuclear membrane is lost during the replication process. With some baculoviruses, replication is restricted to the gut epithelium and progeny virions become enveloped and occluded within these cells, and may be shed into the gut lumen with sloughed epithelium, or released upon death of the host. In other baculoviruses, the infection is transmitted to internal organs and tissues. The second virion phenotype, BV, which buds from the basolateral membrane of infected gut cells is required for transmission of the infection into the hemocoel. In secondarily infected tissues, BV is produced during the late phase and occluded virus during the very late phase of the infection. Infected fat body cells are the primary location of occluded virus production in lepidopteran insects. Occluded virus matures within nuclei of infected cells for nucleopolyhedroviruses and within the nuclear-cytoplasmic milieu for granuloviruses. Occlusion bodies containing infectious ODV virions are released upon death, and usually liquification, of the host.
Antigenic determinants that cross-react between different baculoviruses exist on virion proteins and on the major occlusion body polypeptide: polyhedrin or granulin. Neutralizing antibodies react with the major surface glycoprotein of BV.
Baculoviruses have been isolated from arthropods only; primarily from insects of the order Lepidoptera, but also Hymenoptera, and Diptera as well as from the crustacean order Decapoda (shrimp). Transmission: (1) natural-horizontal transmission by contamination of food, egg surface, etc. with occlusion bodies; (2) vertical transmission within the egg has been reported; (3) experimental-by injection of intact hosts with BV or by infection or transfection of cell cultures. Typically the infection process in a permissive insect host requires approximately one week, and as an end result, the diseased insect liquifies releasing infectious occlusion bodies into the environment. Occlusion bodies represent an environmentally stable form of the virus with increased resistance to inactivation by light.
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