Parvoviridae


The Parvoviridae are a family of small, rugged, genetically-compact DNA viruses, known collectively as parvoviruses. There are currently more than 100 species in the family, divided among 23 genera in three subfamilies. Parvoviridae is the sole taxon in the order Quintoviricetes.
Parvovirus B19 was the first pathogenic human parvovirus to be discovered and is best known for causing a childhood exanthem called "fifth disease", although it is also associated with other diseases including arthritis.
Parvoviruses can infect and may cause disease in many animals, from arthropods such as insects and shrimp, to echinoderms such as starfish, and to mammals including humans. Because most of these viruses require actively dividing cells to replicate, the type of tissue infected varies with the age of the animal. The gastrointestinal tract and lymphatic system can be affected at any age, leading to vomiting, diarrhea, and immunosuppression, but cerebellar hypoplasia is only seen in cats that were infected with feline parvovirus in the womb or at less than two weeks of age, and disease of the myocardium is seen in puppies infected with canine parvovirus 2 between the ages of three and eight weeks. Canine parvovirus causes a virulent and contagious disease in dogs. In cats, a parvovirus causes feline distemper.
Parvovirus can also be used specifically for members of one of the two Parvoviridae subfamilies: Parvovirinae, which infect vertebrate hosts, and Densovirinae, which infect invertebrate hosts, are more commonly referred to as densoviruses. In subfamily Densovirinae there are 5 genera and a total of 21 species.
Parvoviruses are linear, nonsegmented, single-stranded DNA viruses, with an average genome size of 5-6 kilo base pairs. They are classified as group II viruses in the Baltimore classification of viruses. Parvoviruses are among the smallest viruses and are 23–28 nm in diameter.

Structure

The viral capsid of a parvovirus is made up of 60 copies of two or more size variants of a single protein sequence, called VP1, VP2 etc., which form a resilient structure with T=1 icosahedral symmetry. These virions are typically resistant to dilute acids, bases, solvents, and temperatures up to 50°C. Parvoviruses do not have envelopes, thus are considered "naked" viruses. In addition, the shape of the virion is roughly spherical, with surface protrusions and canyons.
Inside the capsid is a linear, single-stranded DNA genome in the size range 4–6 kbp, so the small genome of parvovirus can encode only a few proteins. At the 5’ and 3’ ends of this genome are short complementary sequences of roughly 120 to 550 nucleotides that form secondary structures as hairpins for example inverted terminal repeats or unique sequences at the termini and are essential for viral genome replication mechanism called rolling-hairpin replication.
Parvovirus particles have a durable non-enveloped protein capsid ~20–30 nm in diameter that contains a single copy of the linear single-stranded ~ 5kb DNA genome, which terminates in small imperfect palindromes that fold into dynamic hairpin telomeres. These terminal hairpins are hallmarks of the family, giving rise to the viral origins of DNA replication and mediating multiple steps in the viral life cycle including genome amplification, packaging, and the establishment of transcription complexes. However, they are often refractory to detection by PCR amplification strategies since they tend to induce polymerase strand-switching. Many parvoviruses are exceptionally resistant to inactivation, remaining infectious for months or years after release into the environment.
Viruses in this family have small protein virions that exhibit T=1 icosahedral symmetry. Their capsid shells are assembled from 60 icosahedrally-ordered copies of a single core protein sequence, but some of these VP proteins also have N-terminal extensions that are not visible in X-ray structures. Biochemical and serological studies indicate that these extensions become successively exposed at the particle surface during virus maturation and cell entry, where they contribute to virion stability and mediate specific steps in cell trafficking. Parvoviruses appear to be unique in encoding a broad spectrum phospholipase A2 activity, typically in the N-terminus of the longest subset of their capsid proteins, which is deployed to mediate virion transfer across the lipid bilayer of host cells

Size

These viruses have small genomes, encoding just two genes, and must rely on the synthetic machinery of their host cell for their own preferential replication. This means that many parvoviruses require host cells to enter S-phase before viral DNA replication can initiate, but they do not encode any gene products that can drive this transition. Parvoviruses overcome this problem in various ways: viruses in many genera simply wait within the cell for it to enter S-phase under its own cell cycle control, which means that they can only infect actively-dividing cell populations.
In contrast, the so-called adeno-associated viruses from genus Dependoparvovirus must wait until the cell is co-infected by a helper DNA virus, commonly an adenovirus or herpes virus, which does encode gene products that can drive the cell into S-phase, allowing AAV infection to initiate and out-compete the helper virus. A third strategy is used by human bocavirus 1 from genus Bocaparvovirus, which appears to invoke a specific DNA-damage response in its host cell that ultimately supports viral DNA amplification and progeny virus production.

Genome

The viral genome is 4–6 kilobases in length and terminates in imperfectly-palindromic hairpin sequences of ~120–500 nucleotides that exhibit genus-specific secondary structures, and can either be identical at the two ends of the genome or can differ in size, sequence and predicted secondary structure. Homotelomeric viruses package DNA strands of both senses whereas heterotelomeric viruses generally package predominantly negative-sense DNA. All parvoviruses encode two major gene complexes: the non-structural gene that encodes the replication initiator protein, and the VP gene, which encodes a nested set of ~2–6 size variants derived from the C-terminus of the single VP protein sequence. Members of the Parvovirinae also encode a few small genus-specific ancillary proteins that are variably distributed throughout the genome, show little sequence homology to each other, and appear to serve an array of different functions in each genus. Viruses in most genera are mono-sense, meaning that both viral genes are transcribed in a single direction from open reading frames in the same DNA strand, but members of one genus of homotelomeric invertebrate viruses show ambisense organization, with the NS and capsid proteins being transcribed in opposite directions from the 5’-ends of the two complementary DNA strands.
The major non-structural protein, NS1, is a site- and strand-specific endonuclease belonging to the HuH protein superfamily, and also carries a AAA+ SF3 helicase domain. NS1 initiates and drives the viral “rolling hairpin” replication mechanism, which is a linear adaptation of the more-common “rolling-circle” replication strategy used by many small circular prokaryotic and viral replicons. RHR is a unidirectional mechanism that displaces a single, continuous DNA strand, which rapidly folds and refolds to generate a series of concatemeric duplex replication intermediates. Unit length genomes are then excised from these intermediates by the NS1 endonuclease, and packaged 3’-to-5’ into preformed empty capsids driven by the SF3 helicase activity of NS1/Rep.

Life cycle

Summary

Viral replication is nuclear. Entry into the host cell is achieved by attachment to host receptors, which mediate internalization via endocytosis. Capsids are metastable, undergoing a series of structural shifts during cell entry that sequentially expose peptides carrying PLA2 activity and trafficking signals. These signals ultimately mediate delivery of the intact virion into the cell nucleus, where genome uncoating allows the establishment of viral DNA replication and transcription complexes that rely predominantly upon the synthetic machinery of their host cell. Replication follows the unidirectional strand displacement mechanism discussed above. Packaged virions from viruses in at least two genera have mechanisms that allow mature virions to be trafficked out of viable host cells prior to cell lysis, but members of most other genera are only released into the environment following death and lysis of the infected cell. Natural animal hosts for parvoviruses include a wide range of vertebrates, arthropods and echinoderms. For viruses in the Parvovirinae transmission routes are commonly fecal-oral or respiratory.

Steps of life cycle

Attachment and entry

A virion attaches to receptors on the surface of a potential host cell. In the case of B19 virus the host cell is a red blood cell precursor and one of the receptors is the blood group P antigen. The virion enters the cell by endocytosis and is released from the endosome into the cytoplasm, where it associates with microtubules and is transported to a nuclear pore. With a diameter of 18–26 nm, the parvovirus virion is small enough to pass through a nuclear pore, unlike the herpesvirus nucleocapsid, though there is evidence that the virion must undergo some structural changes before it can be transported into the nucleus. Nuclear localization signals have been found in the capsid proteins of some parvoviruses.

Single-stranded DNA to double-stranded DNA

In the nucleus the single-stranded virus genome is converted to dsDNA by a cell DNA polymerase. The ends of the genome are double stranded as a result of base pairing, and at the 3' end the terminal nucleotides act as a primer to which the polymerase binds.

Transcription and translation

The cell's RNA polymerase II transcribes the virus genes and cell transcription factors play key roles. The primary transcript undergo various splicing events to produce several size classes of mRNA. The larger mRNAs encode the non-structural proteins and the smaller mRNAs encode the structural proteins. The non-structural proteins are phosphorylated and play roles in the control of gene expression and in DNA replication.

DNA replication

After conversion of the ssDNA genome to dsDNA, the DNA is replicated by a mechanism called rolling-hairpin replication. This is a leading strand mechanism that is an adaptation of rolling-circle replication, and sets parvoviruses apart from dsDNA viruses, which replicate their genomes through leading and lagging strand synthesis. Procapsids are constructed from the structural proteins and each is filled by a copy of the virus genome, either a DNA or a DNA as appropriate. The helicase function of one of the non-structural proteins serves as a motor to drive the packaging of displaced single-strands into the capsid.

Taxonomy

Group: ssDNA
The family is divided into three subfamilies: Parvovirinae, which infect vertebrates, Densovirinae, which infect invertebrates, and Hamaparvovirinae. Each subfamily has been subdivided into several genera.
Subfamily Densovirinae; genera:
Subfamily Hamaparvovirinae; genera:
Subfamily Parvovirinae:
To date, very few viruses from the Densovirinae have been studied and sequenced, so the taxonomy may poorly reflect the true diversity of this subfamily. Currently, members of genus Ambidensovirus, Brevidensovirus and Iterodensovirus typically infect insects, while the hepan- and penstyldensoviruses infect decapod shrimp. However, recently two viruses have been isolated that segregate phylogenetically with members of genus Ambidensovirus but infect non-insect hosts. One of these, cherax quadricarinatus densovirus, has been shown to infect and kill the freshwater crayfish Cherax quadricarinatus, subphylum Crustacea, while the other, called sea-star-associated densovirus, has been shown to replicate in and kill sea-stars and perhaps some other members of phylum Echinodermata.

Examples

Parvoviruses that infect vertebrate hosts make up the subfamily Parvovirinae, while those that infect invertabrates make up the subfamily Densovirinae. Prior to 2014, the name parvovirus was also applied to a genus within subfamily Parvovirinae, but this genus name has been amended to Protoparvovirus to avoid confusion between taxonomic levels.
Many mammalian species sustain infection by multiple parvoviruses. Parvoviruses tend to be specific about the species of animal they will infect, but this is a somewhat flexible characteristic. Thus, all isolates of canine parvovirus affect dogs, wolves, and foxes, but only some of them will infect cats.
Humans can be infected by viruses from five of the eight genera in the subfamily Parvovirinae: i) Bocaparvovirus Dependoparvovirus Erythroparvovirus Protoparvovirus Tetraparvovirus Amdoparvovirus Aveparvovirus Copiparvovirus.

Viruses infecting humans

Currently, viruses that infect humans are recognized in 5 genera: Bocaparvovirus, Dependoparvovirus, Erythroparvovirus, Protoparvovirus and Tetraparvovirus.

Dependoparvoviruses

Dependoparvoviruses, previously known as dependoviruses, require helper viruses to replicate. They are excellent vectors for experimental or clinical gene transduction and are now being used in patients for the therapeutic treatment of certain genetic diseases. The biggest advantage for such applications is that they are not known to cause any diseases.

Autonomous parvoviruses

Unlike dependoparvoviruse, autonomous parvoviruses do not require a helper virus. The autonomous parvovirus genome is a linear ssDNA with terminal palindromes, and several viruses have different sorts of palindromes at each end which are known to package the negative sense DNA strand for the most part. Most Autonomous Parvoviruses are known to encode only two NS structural proteins. Similar to RNA viruses, autonomous parvoviruses are highly effective when cells are in the S phase.
Autonomous Parvovirus Replication occurs through hairpins being formed by the palindrome at the 3′ end of the viral genome. This acts as a primer for the synthesis of complementary strands. Elongating strands then become covalently linked to hairpins at the 5' end of the template to create a linear duplex model that has covalent cross-links at the ends by DNA hairpins. Right end of the hairpin is nicked on the new strand by NS1. The genomic ssDNA is generated and packaged when the capsid is available. The RF structures create ssDNA but they are not linear duplex monomers. These Rf structures are potentially NS1/DNA complexes that are related to the structure of capsid.
Autonomous Parvoviruses have a very unique scheme of transcription. Several are known to utilize promoters at map regions 4 and 38 to create variable transcripts along with polyadenylation signals at the right end of the genome. Small introns exist between map positions 444 and 46, and large introns exist between map positions 10 and 39. All messengers in MVM have small introns spliced while alternative splice donors and acceptor sites can also be used. During genome replication, read-through of the internal poly S site occurs and the large transcript is translated into a capsid protein.
Protein synthesis is dependent on transcription. The earliest transcripts during the course of infection were the NS protein transcripts and they now are able to regulate gene expression. NS1 and NS2 are phosphorylated after translation, and NS1 is known to dimerize before nuclear localization. NS2 has been found to have profound impact on virus growth in a host-dependent manner.

Unclassified viruses

Since databases contain vast numbers of sequences that might be considered parvoviral in origin but are not real viruses, the Parvoviridae Study group cite the following criteria that must be established before a new viral sequence can be considered for recognition in the family:
"In order for an agent to be classified in the family Parvoviridae, it must be judged to be an authentic parvovirus on the basis of having been isolated and sequenced or, failing this, on the basis of having been sequenced in tissues, secretions, or excretions of unambiguous host origin, supported by evidence of its distribution in multiple individual hosts in a pattern that is compatible with dissemination by infection. The sequence must be in one piece, contain all the nonstructural and virus particle coding regions, and meet the size constraints and motif patterns typical of the family."
This means that partial coding sequences or sequences from a single host animal, with no evidence of virus exposure in the rest of the population, will not be considered sufficiently validated. Samples from feces are particularly problematic because they may be derived from food and so do not have an unequivocal host animal. Similarly, samples from aquatic animals that are also present in the surrounding environment are difficult to attribute until they can be shown to directly infect other members of the presumed host species.
Despite these caveats, every year many new viruses are identified that will likely merit recognition, so that the published taxonomy always trails studies in the field.
For example, a candidate parvovirus has recently been isolated by sequencing a histocytic sarcoma in a slow loris. The relationship between the virus and the sarcoma was not clear.

Additional genera

Possible new genera suggested in the literature include Chapparvovirus, but these candidate viruses are predicted to have structurally distinct capsids. At present it is not clear if their host range is restricted to vertebrates, which could make them candidates for inclusion in subfamily Parvovirinae, or if they also infect invertebrate hosts. Consideration of the taxon awaits further investigation and ultimately a decision by the ICTV.

Diseases

Parvovirus B19 which causes fifth disease in humans, is a member of species Primate erythroparvovirus 1 in the genus Erythroparvovirus. It infects red blood cell precursors and was the first parvovirus shown to cause human disease. Some infections do not result in visible infection, while some manifest with visible effects, such as fifth disease, which can give children a ‘slapped-cheek’ appearance.
Canine parvovirus is a member of species Carnivore protoparvovirus 1 in the genus Protoparvovirus. It causes a particularly deadly disease among young puppies, about 80% fatal, causing gastrointestinal tract damage and dehydration, as well as a cardiac syndrome in very young animals. It is spread by contact with an infected dog's feces. Symptoms include lethargy, severe diarrhea, fever, vomiting, loss of appetite, and dehydration.
However, mouse parvovirus 1, a member of species Rodent protoparvovirus 1, causes no symptoms but can contaminate immunology experiments in biological research laboratories.
Porcine parvovirus, a member of species Ungulate Protoparvovirus 1, causes a reproductive disease in swine known as SMEDI, which stands for stillbirth, mummification, embryonic death, and infertility.
Like canine parvovirus, feline panleukopenia, now more commonly known as feline parvovirus, is also a member of species Carnivore protoparvovirus 1 in the genus Protoparvovirus. This virus is common in kittens and causes fever, low white blood cell count, diarrhea, and death. Infection of the cat fetus and kittens less than two weeks old causes cerebellar hypoplasia.
Mink enteritis virus, also a member of species Carnivore protoparvovirus 1, is similar in effect to feline parvovirus, except that it does not cause cerebellar hypoplasia. A different parvovirus causes aleutian disease in mink and other mustelids, characterized by lymphadenopathy, splenomegaly, glomerulonephritis, anemia, and death.

Canine and feline

Canine parvovirus is a mutant strain of feline parvovirus. A very specific mutation is necessary for the virus to change species of infection. The mutation affects capsid proteins of feline parvovirus, giving it the ability to infect dogs. Both forms of the virus are very similar, so once the mutation has occurred, canine parvovirus is still able to infect cats. The canine parvovirus has the tradeoff of gaining the ability to infect canine cells, while becoming less effective at infecting feline cells. Both feline parvovirus and canine parvovirus bind to and infect the transferrin receptors, but both have different sequences in the cells and animals. Infection by both feline parvovirus and canine parvovirus are relatively quick, but because of constant mutation of canine parvovirus, canine parvovirus has a slower infection time than feline parvovirus. Studies of other strains of mutated canine parvovirus have revealed that changes in the viral capsid by just one protein can be fatal to the virus. Deleterious mutations have been noted to lead to inability to bind to transferrin receptors, bind to nonreceptive parts of the cell membrane, and identification of the virus by the host's antibody cells.
Dogs, cats, and swine can be vaccinated against parvovirus. In pet dogs and cats, this is a normal vaccine commonly administered when they are young. The dog vaccine that protects against parvovirus is DHPP, and the cat vaccine is FPV.

Replication as disease vector

To enter host cells, parvoviruses typically bind to a sialic acid-bearing cell surface receptor and penetration into the cytoplasm is mediated by a phospholipase A2 activity carried on the amino-terminal peptide of the capsid VP1 polypeptide. Once in the cytoplasm, the intact virus is translocated into the nucleus prior to uncoating. Transcription only initiates when the host cell enters S-phase under its own cell cycle control, when the cell's replication machinery converts the incoming single strand into a duplex transcription template, allowing synthesis of mRNAs encoding the nonstructural proteins, NS1 and NS2. The mRNAs are transported out of the nucleus into the cytoplasm, where the host ribosomes translate them into viral proteins. Viral DNA replication proceeds through a series of monomeric and concatemeric duplex intermediates by a unidirectional strand-displacement mechanism that is mediated by components of the host replication fork, aided and orchestrated by the viral NS1 polypeptide. NS1 also transactivates an internal transcriptional promoter that directs synthesis of the structural VP polypeptides. Once assembled capsids are available, replication shifts from synthesizing duplex DNA to displacement of progeny single strands, which are typically negative-sense and are packaged in a 3'-to-5' direction into formed particles within the nucleus. Mature virions may be released from infected cells prior to cell lysis, which promotes rapid transmission of the virus, but if this fails, then the virus is released at cell lysis.
Unlike most other DNA viruses, parvoviruses are unable to activate DNA synthesis in host cells. Thus, for viral replication to take place, the infected cells must typically be nonquiescent. Their inability to force host cells into S-phase means that parvoviruses are nontumorigenic. Indeed, they are commonly oncolytic, showing a strong tendency to replicate preferentially in cells with transformed phenotypes.

Management and therapy

Currently, no vaccine exists to prevent infection by all parvoviruses, but recently, the virus's capsid proteins, which are noninfectious molecules, have been suggested acting as antigens for improving of vaccines.
For pig vaccine, inactivated live, monovalent combined, most contain old PPV-1 strains to protect already positive sows. Vaccinate after 6 months, once or twice before mating, and repeat yearly.
Antivirals and human immunoglobulin-sourced treatments are usually for relief of symptoms. Using immunoglobulins is a logical solution for treatment as neutralizing antibodies because a majority of adults have been in danger from the parvoviruses, especially B19 virus.

Use of HeLa cells in parvovirus testing

Testing for how feline parvovirus and canine parvovirus infect cells and what pathways are taken, scientists used cat cells, mouse cells, cat and mouse hybrid cells, mink cells, dog cells, human cells, and HeLa cells. Both feline parvovirus and canine parvovirus enter their hosts, follow specific pathways, and infect at certain parts of cells before infecting major organs. Parvoviruses are specific viruses that are characterized in part by which receptors they attack. Testing found that parvovirus infects carnivorous animals through the oropharyngeal pathway. Parvovirus infects the oropharyngeal cells by binding to transferrin receptors, a glycoprotein, on the plasma membrane. The parvovirus plasmid is stored in a small non-enveloped capsid. Once oropharyngeal cells become infected the virus spreads to dividing lymph cells and continues to work to the bone marrow and spread to target organs through blood.
Testing of HeLa cells and human cells to exposure of both feline parvovirus and canine parvovirus resulted in infections of the cells at human transferrin receptors. When antibodies and parvovirus samples were added at the same time to human cells and HeLa cells, no infection was found to take place; both human cells and HeLa cells have transferrin receptors, but no evidence of humans contracting parvovirus was found.
Certain chromosomes in cells show more susceptibility to parvovirus than others. Testing of feline parvovirus on cat cells and cat mouse hybrid cells found cultures with cells having the highest concentrations of the C2 chromosome were the most highly infected cells. Slight mutations of binding sites were found to slow down or completely stop the infection of the given parvovirus, whereas cells that are naturally missing the receptors or are mutants lacking them cannot be mutated. Both feline parvovirus and canine parvovirus express plasticity during cellular infection. Although transferrin receptors may be limited on cell surfaces, the parvovirus will find available transferrin receptors and will use different pathways to gain entry to the cell. Unlike plasma membrane infection plasticity, all strains of parvovirus show related routes to the cell nucleus.

Etymology

Viruses of the family Parvoviridae take their name from the Latin parvum, meaning "small" or "tiny". The subfamily Densovirinae has its name from the Latin denso, meaning thick or compact.

History

Perhaps due to their extremely small size, the first parvoviruses were not discovered until the late 1950s. Parvovirus B19, the first known parvovirus to cause disease in humans, was discovered in London by Australian virologist Yvonne Cossart in 1974. Cossart and her group were focused on hepatitis B and were processing blood samples when they discovered a number of "false positives" later identified as parvovirus B19. The virus is named for the patient code of one of the blood-bank samples involved in the discovery.