Dependovirus

Dependovirus: Description, Causes and Risk Factors:A genus of small defective single-stranded DNA viruses in the family Parvoviridae (A family of small viruses containing single-stranded DNA. Virions are 18-26 nm in diameter, are not enveloped, and are ether-resistant. Capsids are of cubic symmetry, with 32 capsomeres. Replication and assembly occur in the nucleus of infected cells. Three genera in the subfamily Parvovirinae are recognized: Parvovirus, Erythrovirus, and Dependovirus, which includes the adeno-associated virus. A second subfamily, Densovirinae, has 3 additional genera, all of which infect arthropods) that depend on adenoviruses for replication. DependovirusThe Dependovirus has an icosahedral shape, measure approximately 22 nm is composed of 60 wedge shaped proteins triangulation number equal to 1. Three virion proteins (VP1, VP2 and VP3) are present in each capsomere. Each capsid is made from 5 VP1, 5 VP2, and 50 VP3 proteins. The capsid does not have an envelope.The genome is a single molecule of single stranded with a length of 4.7 kilobases (kb). It has only two open reading frames. The 3' open reading frame is the structural capsid protein, cap, which can be spliced to form two RNAs, one for VP1 and the other goes on to eventually make VP2 and VP3. The second gene, rep, can be spliced into four different, nonstructural, regulatory proteins that all aid in the genome replication. These proteins are named Rep 78, Rep68, Rep 52, and Rep 40 based on their molecular weight.Due to inverted, complementary long terminal repeats (LTRs) at each end of the genome, a T shaped secondary structure is formed. The complementary areas leave a 3' hydroxyl group single stranded for the replication to begin. This 3' hydroxyl group is used as a primer for the leading strand synthesis. Both positive and negative sense strands of DNA are made. Double stranded intermediates are formed throughout the replication; this means the two strands, positive and negative sense, will be matched up. There is no known disease in humans associated with Dependovirus.RESEARCH:Electron microscopic particle counting of the defective dependovirus by use of pseudoreplication and negative staining with phosphotungstic acid was shown to be a reproducible quantitative assay procedure. Particles of satellite type 4 that were counted in fluids from infected cultures had the same morphology as particles that banded at a buoyant density of 1.43 g/cc in cesium chloride. Other satellite virus serotypes examined in the same manner had a buoyant density of 1.37 to 1.38 g/cc. A comparison of satellite titers obtained by complement fixation and by particle counting demonstrated that an increase in satellite particles resulted in a corresponding increase in CF titers; however, electron microscopy was at least 10 times more sensitive than complement fixation for detecting satellite virus. Growth cycle studies of satellite virus in cells co-infected with adenovirus, as assayed by particle counting, indicated that the kinetics of satellite virus production closely followed the kinetics of its helper adenovirus production, with an eclipse period of 12 to 16 hr. The eclipse period of the satellite remained the same when cultures were pre-infected with satellite 24 hr prior to adenovirus inoculation. However, when cultures were infected with adenovirus 12 hr before satellite virus, the eclipse period of the satellite was shortened to between 4 and 6 hr. Thus, satellite virus replication seems dependent upon a relatively late event in the adenovirus replication cycle. When cells were co-infected with adenovirus and its defective satellite, the yield of adenovirus was markedly reduced from that obtained in cells singly infected with adenovirus.
  • Adeno-associated satellite virus type 4 interferes with the replication of its helper adenovirus. No interferon-like soluble substance could be detected in satellite-infected cultures and other DNA- and RNA-containing viruses were not inhibited by co-infection with satellite virus under conditions which reduced adenovirus yields by more than 90% in monkey cells. Altering the concentration of adenovirus in the presence of constant amounts of satellite resulted in a constant degree of interference over a wide range of adenovirus inocula and suggested that adenovirus concentration was not a significant factor in the observed interference. The interference with adenovirus replication was abolished by pretreating satellite preparations with specific antiserum, ultraviolet light or heating at 80°C for 30 min. This suggested that infectious satellite virus mediated the interference. Satellite virus concentration was found to be a determinant of interference and studies indicated that the amount of interference with adenovirus was directly proportional to the concentration of satellite virus. 8 hr after adenovirus infection, the replication of adenovirus was no longer sensitive to satellite interference. This was true even though the satellite virus was enhanced as effectively as if the cells were infected simultaneously with both viruses. Interference with adenovirus infectivity was accompanied by reduced yields of complement-fixing antigen and of virus particles which suggested that satellite virus interfered with the formation and not the function of adenovirus products. When cells were infected either with adenovirus alone or with adenovirus plus satellite, the same proportion of cells plated as adenovirus infectious centers. However, the number of plaque-forming units of adenovirus formed per cell in the satellite-infected cultures was reduced by approximately 90%, the same magnitude of reduction noted in whole cultures coinfected with satellite and adenovirus. This suggested that all cells infected with the two viruses were producing a reduced quantity of adenovirus.
Symptoms of Dependovirus:There are no known symptoms.Diagnosis Dependovirus:Viruses ASV strain M, found in the stock culture of (infectious hepatitis virus [ICHV]). Four strains of ASV (types 1,2,3 and 4) were provided by M. ASV types 1, 2, 3 and 4 were grown in the culture of dog kidney cells, co-infected with ASV-free ICHV strain FD as a helper. Purification of the ASV strain M The culture fluid of DKC infected with the ASV strain M, associated with the helper ICHV strain Matsuda, was harvested when the cytopathic effect due to ICHV reached maximum. The fluid, frozen and thawed three times, was centrifuged differentially, at 11,000 g for 20 min and at 77,500 g for 90 min. The differential centrifugation was done several times. The resultant viral material of strain M was purified by banding three times in isopycnic CsCl density gradient ultracentrifugation at 120,000 g for 30 to 40 hours. Purification of ASV types 1, 2, 3 and 4 was carried out in the same way. The counting of the virus particles A drop of ASV samples, 0.01 ml in amount, was put on a carbon-coated collodion membrane grid. After the sample was dried, approximately 0.004 ml of 3 % sodium silicotungstate (pH 7.2) was placed on the same grid. After the staining solution was dried, the specimen was examined with a JEM-7 electron microscope at an instrumental magnification of 50,000 X. Virus particles were counted in 90 different fields (1 field = 1 mp2) and scored as the particles per mµ2.The immunization of a guinea pig against strain M 1 ml of the purified strain M, which contained 350 particles, per mµ2under the electron microscope at 50,000 X magnification, was mixed with an equal volume of complete Freund adjuvant. This mixture was injected into the foot pad and muscle of a guinea pig. Three weeks later, the same volume of the virus material without adjuvant was given as a booster. The serum was collected a week after the booster inoculation.Preparation of ASV antigens for the complement fixation (CF) test. Four units of the strain M antigen determined by theCF test using anti-strain guinea pig serum, contained 70-80 particles per mµ2under the electron microscope at 50,000 X magnification. Antigens of ASV types 1,2,3 and 4 were adjusted so as to contain an almost equal number of virus particles.The CF test was carried out according to the Microtiter technique.Treatment of Dependovirus:The Dependovirus is not a large enough virus to trigger an immune response; this makes it a good virus to use as a gene therapy tool. Gene therapy is a possible treatment for a variety of disorders and diseases that are genetic in origin. Viral vectors are currently being developed to transport genes into human cells. Since this virus does not stimulate an immune response it can be used multiple times effectively without being neutralized before infection. Another reason these viruses are reliable vectors is the known insertion point for the genome. This virus always inserts its contents into the same place on chromosome 19. This predictability can cut down on the chances of inserting into an important area that might disrupt normal gene function or increase the risk of developing cancer. At this time, one challenge using this virus as a therapy tool is the fact that the genome is fairly small. With less than 5kb in the genome the amount of genetic material that can fit into the capsid is limited. Work is currently being done to increase the amount of information this vector can deliver. This may be accomplished by the LTRs found at both the 5' and 3' end of the genome. Since the LTRs have the same sequence they will leave complementary strands exposed if they are removed. The complementary strands can undergo recombination and join two 5kb inserted fragments together.NOTE: The above information is educational purpose. The information provided herein should not be used during any medical emergency or for the diagnosis or treatment of any medical condition.DISCLAIMER: This information should not substitute for seeking responsible, professional medical care.

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