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PubMed1979

Sudden death in young dogs with myocarditis caused by parvovirus.

Sudden death of pups in the 4- to 6-week age range has recently been occurring in western Canada as a result of severe, primary, nonsuppurative myocarditis. At necropsy, the prominent macroscopic lesion was pulmonary edema, and microscopically, characteristic intranuclear inclusion bodies were found within cardiac myofibers in association with myocarditis. Ultrastructurally, numerous small particles resembling parvoviruses were found within the intranuclear inclusion bodies, which were positive by direct fluorescent antibody test for canine parvovirus. Of three pups inoculated with homogenate from affected myocardium, one developed lesions resembling canine parvoviral enteritis.

Animals↗
PubMed2026

Nanopore-based, long-range Parvovirus B19 amplicon sequencing for near-whole genome characterization.

BACKGROUND: Whole-Genome Sequencing (WGS) enables monitoring of genomic variation and evaluation of diagnostic PCR assays. However, WGS data for Parvovirus B19 (B19V) remains limited despite its relevance for clinical care and transfusion safety. To increase the availability of high-quality B19V genomic data, a near-WGS protocol was developed and validated. METHOD: The protocol combines long-range PCR to generate a 4.6-kb amplicon, covering ∼82% of the B19V genome, with Oxford Nanopore sequencing. Validation was performed using six reference samples and nineteen B19V-positive donor plasma samples. RESULTS: After quality control, samples achieved a median sequencing depth of 152x. Sequences generated from the six reference samples showed 100% concordance with previously published data. Genomic analysis of donor samples explained atypical amplification profiles observed during routine PCR screening. CONCLUSION: The newly developed protocol provides a scalable method for B19V genome characterization, enabling assessment of oligonucleotide-binding regions for PCR assay monitoring and facilitating the generation of genomic data for future epidemiological investigations.

PCR assay monitoring↗
PubMed2024

N6-methyladenosine modification of a parvovirus-encoded small noncoding RNA facilitates viral DNA replication through recruiting Y-family DNA polymerases.

Human bocavirus 1 (HBoV1) is a human parvovirus that causes lower respiratory tract infections in young children. It contains a single-stranded (ss) DNA genome of ~5.5 kb that encodes a small noncoding RNA of 140 nucleotides known as bocavirus-encoded small RNA (BocaSR), in addition to viral proteins. Here, we determined the secondary structure of BocaSR in vivo by using DMS-MaPseq. Our findings reveal that BocaSR undergoes N6-methyladenosine (m6A) modification at multiple sites, which is critical for viral DNA replication in both dividing HEK293 cells and nondividing cells of the human airway epithelium. Mechanistically, we found that m6A-modified BocaSR serves as a mediator for recruiting Y-family DNA repair DNA polymerase (Pol) η and Pol κ likely through a direct interaction between BocaSR and the viral DNA replication origin at the right terminus of the viral genome. Thus, this report represents direct involvement of a viral small noncoding RNA in viral DNA replication through m6A modification.

Humans↗
PubMed1979

Nucleotide sequence of the self-priming 3' terminus of the single-stranded DNA extracted from the parvovirus Kilham rat virus.

The parvovirus genome is a linear, single-stranded DNA molecule with double-stranded hairpin termini. The 3' terminus can serve in vitro as a self-primer for the synthesis of a double-stranded viral DNA intermediate. We have sequenced the nucleotides in the 3' terminus and propose a model for the secondary structure of the terminus and the in vitro origin of replication for the complementary viral DNA strand.

Base Sequence↗
PubMed1976

The pathogenesis of parvovirus-induced cerebellar hypoplasia in the Syrian hamster, Mesocricetus auratus. Fluorescent antibody, foliation, cytoarchitectonic, Golgi and electron microscopic studies.

Cerebellar histogenesis was studied in hamsters infected at birth with a parvovirus, rat virus strain PRE 308. Cerebellar granule cell precursors in these animals were selectively infected and lysed in the external germinal layer before their migration to form the internal granular layer. The effects of the absence of granule cells on cerebellar development and especially on the development of the Purkinje cells and their dendrites was analyzed using fluorescent antibody. Golgi, conventional paraffin, and electron microscopic methods. This study represents the first Golgi and ultrastructural study of the pathogenesis of rat virus infections in the cerebellum. The destruction of the granule cell precursors resulted in a dysplastic cerebellar hypoplasia with total disruption of normal cerebellar stratification and cytoarchitectonics. The Purkinje cells developed misshapen, progressively disoriented dendritic stems lacking tertiary dendrites and studded with numerous spines, devoid of afferent synaptic contacts (naked spines) and encased by glial processes. These developmental studies, together with the mouse mutant studies, demonstrated that the spines of the Purkinje cells were elaborated in the absence of both tertiary dendrites and afferent parallel fiber contacts. Such data suggested that spine formation, once triggered, was intrinsically programmed rather than being dependent on the development of parallel fiber contacts. Despite the loss of a major interneuronal component and disintegration of normal cytoarchitectonic relationships, synapses in the cerebellar cortex developed normally as long as both the pre- and post-synaptic elements were present. Thus synaptic specificity is maintained in the face of gross disruption of cytoarchitectonic relationships. If either the pre- or post-synaptic portion of a contact was absent, then glial processes isolated the persisting element or aberrant contacts formed. In addition to glial encasement of naked spines, there were dendrodendritic articulations between Purkinje cell dendrites, some of which were joined by septate, plaque-like junctions. Aberrant synaptic contacts between mossy and climbing fiber glomeruli and the smooth surface of the Purkinje cell somata were found rarely. In addition to these contacts which also occur in the hypoplastic cerebella produced by other methods, previously undescribed non-synaptic spine-articulations between Purkinje cell dendrites were seen. The role played by granule cells and their axons in Purkinje cell development appeared to be two-fold. First, the development of the orderly array of parallel fibers in the normal animal played a role in orienting and flattening the dendritic trees of Purkinje cells. Second, the formation of tertiary dendritic branches appeared to depend primarily upon the presence of an external germinal layer throughout this stage of Purkinje cell development. By contrast, dendritic spines developed and persisted in the absence of granule cells.

Animals↗
PubMed1977

Embryonal carcinoma cells (and their somatic cell hybrids) are resistant to infection by the murine parvovirus MVM, which does infect other teratocarcinoma-derived cell lines.

Minute virus of mice (MVM), a non-defective parvovirus, has been shown to infect cultures of non-pluripotent differentiated teratocarcinoma-derived cells, but pluripotent (and "nullipotent") embryonal carcinoma cells derived from the same teratocarcinoma resist MVN infection. Somatic cell hybrids between an embryonal carcinoma line and Friend erythroblastic leukemia cells are also resistant to MVM, even though Friend cells are susceptible. Among three blastocyst-derived lines tested, only one, a parietal yolk sac cell line, resists MVM infection. These results suggest that teratocarcinoma cultures may provide useful systems in which to study the cellular factors which mediate susceptibility to this teratogenic and oncolytic virus.

Animals↗
PubMed1976

Observations on the pathogenesis of porcine parvovirus infection.

Differences in the pathogenesis of porcine parvovirus (PPV) were shown when pregnant gilts were infected by the oral and intramuscular (i.m.) routes. By the oral route, PPV took 23-32 days to cross the placenta following infection of the dam, as compared to 15 days by the i.m. route, Successful transplacental infection occurred following oral infection of dams only in the second third of gestation, whilst i.m. infection resulted in infection of foetuses in both first and second thirds of gestation. Foetal infection resulted in death and mummification only where infection of foetuses occurred before onset of immune competence--estimated at 70 days gestation. Infected foetuses either died before onset of immune competence, or survived to mount an immune response with subsequent death or survival to farrowing. It is suggested in discussion that reproductive failure due to PPV, characterised by mummification or occasional stillbirth, is associated in nature with oral infection, and occurs only when dams are infected in the first part of the midthird of gestation.

Animals↗
PubMed1976

Infection of newborn and fetal hamsters induced by inoculation of LuIII parvovirus.

The LuIII parvovirus was adapted to the newborn hamster and produced a systemic infection with massive intestinal hemorrhage. Inoculation of pregnant hamsters lead to transplacental infection of the fetuses and abortion. Most fetal deaths were observed in animals inoculated on days 8 and 10 of gestation. Virus was recovered from dead fetuses, placentas, and viable fetuses. Histological lesions were found in the heart, liver, kidney and CNS of infected fetuses.

Animals↗
PubMed1979

Subfractionation of CsCl-purified H-1 parvovirus on metrizamide gradients.

The different density classes of H-l parvovirus, collected within 30 hr of infection of par-asynchronous cultures, following the standard CsCl purification step, have been shown to be heterogeneous. Rebanding of the denser form (HF, p = 1.46 g/cm3) and the less dense form (LF, p = 1.42 g/cm3) of infectious virus in the nonionic density generating solute, metrizamide, showed that both HF and LF virus bands were heterogeneous in density. The infectivity banded with isotopically labeled virus protein and DNA at 1.32 g/cm3 for both HF and LF virus. Amounts of protein and DNA which varied from preparation to preparation, but which were greater from the HF virus band, were distributed throughout the rest of the gradient, but predominated in a peak at a density of 1.2 g/cm3. The protein in this peak was without hemagglutinating activity but had the molecular weights and proportions of the H-l virion proteins (VPl, VP2', and VPZ). The DNA was of the same size as H-l DNA monomers and its proportion to the protein was similar to that of the infectious peak. The DNA was susceptible to micrococcal nuclease digestion. The nature of this noninfectious viral material thus seemed to be incompletely assembled virus. Radiolabeled H-l virus collected after 72 hr of infection formed a discrete single peak in both CsCl (p = 1.42 g/cm3), and metrizamide gradients (p = 1.32 g/cm3). There was no significant amount of the 1.20 g/cm3 viral protein-DNA complex in these mature preparations.

Centrifugation, Density Gradient↗
PubMed2026

Development of a recombinant goose parvovirus VP2 neutralizing epitope-containing region vaccine adjuvanted with IL-2 and FliC for enhanced immune responses and protection against challenge.

Gosling plague (GP), caused by goose parvovirus (GPV), is a highly contagious and fatal viral disease. Vaccination is essential for disease prevention; however, conventional attenuated and inactivated vaccines have several limitations. Genetically engineered vaccines based on defined antigenic regions represent a promising alternative strategy. This study aimed to identify neutralizing epitope-containing regions within the GPV VP2 protein and develop effective recombinant vaccines. The GPV VP2 protein was divided into 11 overlapping fragments, and the anchored periplasmic expression (APEx) bacterial display system combined with flow cytometry (FCM) was used for antigenic region screening. GPV VP2-specific single-domain antibodies (VHHs) were further applied to identify neutralizing epitope-containing regions. Six neutralizing epitope-containing regions were identified and linked together to construct the VP2M recombinant antigen. The VP, VP2M, interleukin-2 (IL-2), and flagellin (FliC) genes were inserted into prokaryotic and eukaryotic expression vectors to generate protein and DNA vaccines. Three-day-old goslings were randomly assigned into 15 experimental groups for immunization. Immune responses were evaluated by measuring anti-GPV antibody levels, IgG, IgM, and IgA production, IFN-γ levels, immune-related gene expression, splenocyte proliferation, neutralizing activity, and protective efficacy against GPV challenge. The results showed that vaccines containing neutralizing epitope-containing regions induced stronger immune responses than control vaccines. Vaccinated groups exhibited increased anti-GPV antibody levels, IgG, IgM, IgA production, IFN-γ levels, immune-related gene expression, and splenocyte proliferation. Following GPV challenge, VP2M-based vaccines significantly reduced viral genome copies in the bursa of Fabricius, spleen, thymus, and intestinal tissues, accompanied by decreased histopathological lesions based on semi-quantitative scoring. Furthermore, the protective efficacy exceeded 50% in vaccines without adjuvants and reached 90% in groups containing combined IL-2 and FliC adjuvants. In conclusion, this study identifies novel neutralizing epitope-containing regions within GPV VP2 and provides a potential strategy for developing safe and effective recombinant vaccines against GP infection.

GP↗
PubMed1975

Parvovirus-like particles in human sera.

A parvovirus-like antigen has been found in sera of nine healthy blood-donors and two patients. Its pathogenicity is unknown, but 30% of adults possess specific antibody. The new agent can be confused with hepatitis-B antigen both morphologically and serologically.

Adolescent↗
PubMed1976

Rolling hairpin model for replication of parvovirus and linear chromosomal DNA.

A novel, quasicircular scheme is proposed for the replication of parvovirus DNA. Daughter strands are initiated after the copying and rearrangement of a terminal palindromic sequence, a process termed 'hairpin transfer'. Such a process may be involved in the replication of other viruses and host cell DNA.

Base Sequence↗
PubMed1979

Covalent association of protein with replicative form DNA of parvovirus H-1.

The double-stranded replicative form (RF) DNA of the autonomous parvovirus H-1 can be isolated from infected cells in a covalent complex with protein. The protein is present on most or all of the RF DNA, including actively replicating molecules, and is associated with the 5'-terminal endonuclease Hae III fragments of both the viral and complementary strands of RF. The size of the protein is estimated to be 60,000-70,000 daltons from its effect on buoyant density of DNA. DNA with covalently bound protein has not been found in H-1 virions.

Centrifugation, Isopycnic↗
PubMed1979

Mechanism for circularization of linear DNAs: circular parvovirus MVM DNA is formed by a "noose" sliding in a "lasso"-like DNA structure.

During an electron-microscopic survey with the aim of identifying the parvovirus MVM transcription template, we observed previously unidentified structures of MVM DNA in lysates of virus-infected cells. These included double-stranded "lasso"-like structures and relaxed circles. Both structures were of unit length MVM DNA, indicating that they were not intermediates formed during replication; they each represented about 5% of the total nuclear MVM DNA. The proportion of these structures was unchanged after digestion with sodium dodecyl sulfate/Pronase and RNase and after mild denaturation treatment. Cleavage of the "lasso" structures with EcoRI restriction endonuclease indicated that the "noose" part of the "lasso" structure is located on the 5' side of the genomic single-stranded MVM DNA. A model is presented for the molecular nature of the circularization process of MVM DNA in which the "lasso" structures are identified as intermediates during circle formation. This model proposes a mechanism for circularization of linear DNAs.

DNA Replication↗
PubMed1977

Serological responses in pigs vaccinated with inactivated porcine parvovirus.

The safety and immunogenicity of inactivated porcine parvovirus (PPV) vaccines were investigated. Both beta-propiolactone and formalin successfully inactivated virus without destroying immunogenicity, which was considerably enhanced by incorporation of a gel adjuvant in the vaccine. Using the formalised-gel vaccine, initial antibody responses were demonstrated in susceptible piglets and adult pigs at 7 days after vaccination. These antibody responses persist at significant levels for at least 6 months after vaccination. Antibody levels increased up to 16 fold when revaccination was carried out. Vaccination of gilts with low level (passive) immunity resulted in antibody responses comparable to those recorded in susceptible pigs. The vaccine was safe as determined by absence of residual virus in the vaccine, absence of viraemia and excretion in vaccinted stock, and absence of effect on litters of sows vaccinated at different gestational ages. Vaccine stored at 4 degrees C for 6 months was as immunogenic as fresh vaccine.

Animals↗
PubMed1978

Serological procedures to determine time of infection of pigs with porcine parvovirus.

It was found possible to correlate serological responses to porcine parvovirus (PPV) with the time after infection. Two procedures were used for PPV antibody measurement: an immunodiffusion technique to measure precipitating antibody and a haemagglutination inhibition (HI) procedure to measure concentration of 2-mercaptoethanol (2-ME) sensitive antibody. Both procedures successfully related antibody titres to time post-infection, but it was considered that a 2 M 2-ME HI procedure showed the greater promise for field investigations.

Animals↗
PubMed1978

Experimental infection of 35, 50 and 60 day old pig foetuses with porcine parvovirus.

Foetuses of six seronegative gilts, two of which each respectively 35, 50 and 60 days pregnant, were inoculated intrauterinely with porcine parvovirus (PPV) and examined 7 and 11 days after inoculation. HI antibody was not detected in any of the foetuses although all but one gilt developed low levels of antibody. All but one of the foetuses inoculated with PPV died in utero prior to examination at 11 days after inoculation. Infection also spread to non-inoculated litter mates. Histological changes were mild in the gilts but there was widespread tissue necrosis in infected foetuses, and intranuclear inclusion bodies were observed in cells of the liver, lung, kidney and cerebellum. The increased survival of foetuses infected at later stages of gestation appeared to be related to increased numbers of mononuclear cells then present in many tissues.

Animals↗
PubMed1977

Isolation and characterization of a parvovirus of rabbits.

The isolation and characterization of a new virus from rabbit stool are described. The virus replicated in rabbit kidney cell cultures and agglutinated human group O erythrocytes at 4 degrees C. It was stable at acid pH and resistant to chloroform and heat treatment. The growth of the virus was inhibited by 5-iodo-2-deoxyuridine, and virions were stained red with acridine orange, suggesting that they contain single-stranded deoxyribonucleic acid. The density of virions was 1.41 to 1.44 g/ml in CsCl, and the sedimentation value was 137S in sucrose at 4 degrees C. The infectious particles had cubic symmetry and were 27 to 28 nm in diameter by electron microscopy. By these properties this virus can be classified as a member of the parvovirus group. Antibody response was demonstrated in the rabbit from which this virus was recovered. A number of rabbits from a commercial source were found to contain hemagglutination-inhibiting antibody to this virus.

Acridines↗