Official reprint from UpToDate

www.uptodate.com 2014 UpToDate

Author
Jeanne A Jordan, PhD
Section Editors
Martin S Hirsch, MD
Morven S Edwards, MD
Deputy Editor
Allyson Bloom, MD
Epidemiology and laboratory diagnosis of parvovirus B19 infection
All topics are updated as new evidence becomes available and our peer review process is complete.
Literature review current through: Jul 2014. | This topic last updated: Nov 21, 2013.
INTRODUCTION The Parvoviridae family contains two subfamilies: Parvovirinae, which infect mammals and
birds, and Densovirinae, which infect arthropods. The Parvovirinae subfamily has been subdivided into five genera:
Erythrovirus, Dependovirus, Parvovirus, Amdovirus, and Bocavirus. The focus here will be limited to Erythrovirus
genera, and mainly to human parvovirus B19 (B19 prototype strain); only brief coverage will be given to the less
common genotype 2 (prototype strain: LaLi) and genotype 3 (prototype strain: V9) genera within this subfamily [1].
There are three genotypes (genotypes 1, 2 and 3) within the Erythrovirus genera. Genotype 1 (Prototype stain; Au)
circulates worldwide, whereas genotypes 2 and 3 are geographically more restricted and found at lower
prevalences. The nucleotide divergence is significantly greater when comparing genotype 1 to either genotypes 2 or
3 (>11 percent), with slightly less divergence between genotypes 2 and 3 (~8 percent). Not surprisingly, the
divergence at the amino acid level among the three genotypes is significantly less than that seen at the nucleotide
level.
Unless otherwise specified, B19 will be the strain referred to when describing the spectrum of disease
manifestations, epidemiologic patterns and diagnostic testing related to Erythrovirus infections. Compared with
genotype 1, much less has been published on the transmission and epidemiology of genotype 2 and genotype 3.
(See "Clinical manifestations and pathogenesis of human parvovirus B19 infection" and "Treatment and prevention of
parvovirus B19 infection".)
One of the hallmarks of Erythroviruses is their extremely limited host range. Productive infections occur only in
human erythroid precursor cells. B19 is directly cytotoxic to colony-forming units (CFU-E) and burst-forming units
(BFU-E) of the erythroid series [2,3].
B19 is a relatively simple, small nonenveloped DNA virus consisting of a linear single-stranded 5.6 kb genome
encoding for the following proteins:
B19 is among the smallest of the DNA animal viruses, with a virion capsid diameter of between 18 to 26 nm. Both
its small size and its lack of a lipid envelope make B19 extremely difficult to inactivate or remove from blood
products. This is an important issue in the blood banking industry, because many individuals infected with B19 are
asymptomatic, yet are circulating very high viral loads (10(9) to 10(12) viral particles/mL of blood) [6,7].
EPIDEMIOLOGY Parvovirus B19 infection occurs worldwide. Cases can be sporadic or can occur in clustered
outbreaks. The infection is easily transmitted from person to person via the respiratory route. In the United States,

Two viral capsid proteins, VP1 (83 kDa) and VP2 (58 kDa) are expressed during productive infection; the
smaller VP2 protein constitutes 95 percent of the capsid while the larger VP1 protein makes up only 5
percent.

The nonstructural protein, NS1 (71 or 77 kDa), is involved in transregulation of its P6 promoter, and has
helicase activity. It also plays a role in inducing cellular apoptosis in both permissive and non-permissive cell
types [4,5].

B19 infection occurs more frequently between late winter and early summer. Where reportable, communities have
documented not only a seasonality to B19 infections, but also cycles of local epidemics with case numbers that
can peak every four to 10 years [8-10].
The genetic variants of the Erythrovirus genera mentioned above, genotypes 2 and 3, have primarily been detected
in Northern European countries (eg, Denmark, Finland, Sweden, and France), while they are uncommon in the
United States. These genotypes are also identified among patients with underlying immune deficiencies [11-15].
Prevalence Parvovirus B19 infection is common throughout the world. The percentage of people with
measurable levels of B19-specific IgG increases with increasing age, with most individuals becoming infected during
their school years. During school outbreaks, 25 to 50 percent of students and 20 percent or more of susceptible
staff may become infected. More than 70 percent of adults have measurable B19-specific IgG antibodies [16,17].
When considering women as a separate group, approximately 30 to 40 percent of pregnant women lack
measurable IgG to B19 and are therefore presumed to be susceptible to B19 infection, which then places their
infant at risk.
Incubation period and infectivity Infected patients are most contagious during the phase of active viral
replication and viral shedding, which occurs approximately 5 to 10 days after the exposure. During this phase,
patients can be asymptomatic or present with non-specific flu-like illness. It is not until the second phase of
parvovirus infection that a person can present with specific symptoms or signs (eg, arthralgia, arthritis, and/or an
exanthem) of B19 infection. This is also the phase in which B19-specific antibody production and B19 antigen-
antibody immune complex formations occur. Individuals are no longer infectious when exhibiting these clinical
characteristics.
Transmission There are three documented modes of transmitting B19:
LABORATORY DIAGNOSIS
B19 immune status The B19 immune status varies with acute infection, previous infection, and reactivation of
infection.
Respiratory transmission is the most common way an individual acquires this virus. Although B19 is primarily
associated with nonrespiratory symptoms, it is nevertheless transmitted through close person-to-person
contact, fomites, and respiratory secretions and/or saliva [18]. Young children are the main source of
respiratory acquired B19. Therefore, individuals at highest risk for acquiring the virus include household
contacts, daycare workers and those in a crowded environment.
This scenario places pregnant women with young children at home at high risk of becoming infected. This can
be problematic if she is not immune to the virus [19-21]. Healthcare workers may or may not be at increased
risk of transmission depending upon the degree of contact that they have with infected patients [22-24].

Vertical transmission can occur if a susceptible woman becomes infected during her pregnancy [25]. The risk
of a poor outcome for the fetus is greatest when the congenital infection occurs within the first 20 weeks of
gestation.

Hematogenous transmission can occur from administration of blood or blood products containing B19 [26-31].
Individuals requiring regular infusions of blood product(s) that are made from large plasma pools are at
greatest risk for acquiring the virus compared to those individuals receiving single units [28,29].
In contrast to B19 infections, PARV4 infections are most frequently detected in injection drug users,
particularly those who are coinfected with HIV-1. One study demonstrated prevalent exposure to a novel
human parvovirus (PARV4) among hemophiliacs and injection drug users. Serum from persons at risk showed
frequent reactivity in ELISA testing using VP2 as an antigen, while uniform negativity was noted among adult
control subjects at low risk of parenteral exposures [32].

Acute infection The choice of diagnostic testing for acute B19 infection will depend on the patient's immune
status. In immunocompetent individuals, the preferred method is serologic testing to detect circulating B19-specific
IgM and IgG antibodies (see 'B19-specific assays' below). However, in the immunosuppressed patient, or for
congenital infections, nucleic acid amplification testing (NAAT) is recommended. (See 'Nucleic acid detection'
below.)
Analyzing sera for the presence of B19-specific IgM antibodies is a practical and common approach for diagnosing
acute infection in the immunocompetent individual [33]. Detectable levels of B19-specific IgM can be found within 7
to 10 days of virus exposure and remain measurable for several months before diminishing [34]. In some patients,
B19-specific IgM antibodies can persist for six months or more. Therefore, the presence of these antibodies,
especially at low titers, is suggestive but not conclusive proof of recent infection.
Acute infection can also be diagnosed by demonstrating a fourfold or greater rise in serum B19-specific IgG
antibody titers. However, since this procedure requires two separate time points for sample collection, it is
considered impractical in most clinical situations. Today, most serology testing will include concurrent analyses for
both B19-specific IgM and IgG antibodies from a single blood sample.
NAAT is frequently used today to detect B19 DNA. At present, NAAT is considered the most sensitive approach for
direct detection of virus within a specimen, and is the test of choice for diagnosing acute infection in the
immunocompromised or immunosuppressed patient, including the fetus and neonate.
Previous infection Previous infection is best confirmed by serologic testing that detects B19-specific IgG
antibodies. Documenting previous infection, which infers immunity, is a common practice in obstetrics when the
clinician is concerned about the immune status of a pregnant woman who has a history of exposure to an individual
infected with B19, especially if the contact is her own school-aged or younger children.
Reactivation of infection Reactivation of persistent infection occurs in immunocompetent and, more often,
immunocompromised individuals [35]. These infections are confirmed by demonstrating the presence of the virus for
a prolonged period; this is most often accomplished by NAAT to detect B19 DNA within a specimen. Individuals
with persistent infection may also have measurable levels of B19-specific IgM antibodies over time if they are
healthy enough to produce immunoglobulin.
B19-specific assays A thorough serologic analysis of B19-specific antibody status should include testing for
both IgM and IgG antibodies recognizing viral capsid antigen(s) (VP1 and/or VP2). Many clinical laboratories
complement B19 serology testing with NAAT to detect B19 DNA directly. Specifics on these assays are discussed
below:
IgM antibody assays Detecting B19-specific IgM antibodies to viral capsid antigen(s) is the cornerstone in
determining whether immunocompetent individuals are actively infected. Several B19-specific IgM enzyme
immunoassays (EIAs) are commercially available. However, the analytical parameters of these assays can, vary
considerably both in sensitivity (70 to 100 percent) and specificity (76 to 100 percent) [36-38]. In a study that was
designed to compare correlations between B19 viral DNA loads and antibody responses to viral antigens VP1 and
VP2, the anti-VP2 EIA (Biotrin, Ltd, Dublin, Ireland, recently acquired by Diasorin S.p.A., Saluggia, Italy)
demonstrated significantly better sensitivity compared to the anti-VP1 immunofluorescence assay (91 versus 66
percent); both assays had good specificity (94 and 97 percent, respectively) [39].
B19-specific IgM assay performance and thus its accuracy is greatly enhanced when the serologic method
includes a step in its protocol to deplete or remove IgG antibodies from the serum sample [33,38]. Because IgG
antibodies are present in significantly higher concentrations than IgM antibodies, assays lacking this design are
subject to higher rates of false negative results due to antibody competition. The mu capture format helps minimize
false negative IgM results resulting in excellent sensitivity and specificity [37,40,41]. The presence of rheumatoid
factor, anti-nuclear antibodies, and Epstein-Barr virus IgM in a specimen can also generate false positive IgM
results.
IgG antibody assays Several EIA kits are commercially available for B19-specific IgG analysis, many of
which have excellent sensitivity and specificity. The different kits vary in their antigen composition: some contain
recombinant VP2 alone, while others consist of a combination of VP1 and VP2 antigens. Kits containing
conformational B19 antigens have improved performance over kits containing only linear B19 antigens [36].
The choice of antigen(s) is critical when selecting a diagnostic serology assay [42,43]. Commercial companies
cannot rely on native virus as a source of antigen for their kits, since B19 isn't readily propagated in a tissue culture
system. As a result, recombinant viral antigens are used. These antigens have been produced both in a prokaryotic
expression vector (E. coli) and a eukaryotic expression system (Baculovirus) [44,45]. The viral antigens produced in
the baculovirus system can self assemble into empty capsids as demonstrated by electron microscopy studies,
and as such are more similar to native virions than the linear antigens produced in E. coli.
Unlike NAAT, serology testing does not appear to suffer from recognition issues between genotypes 1, 2 and 3 as
the level of divergence among the strains at the amino acid level is significantly less than that seen at the
nucleotide level.
In addition to serology tests that detect circulating antibodies to viral capsid proteins, other assays have been
described that detect B19-specific IgM and IgG antibodies to the nonstructural protein NS1. Including these, in
addition to detecting viral capsid proteins (VP1 and/or VP2), can improve the accuracy of diagnosing acute B19
infection [46-49]. However, NS1 specific serologic reagents are not yet commercially available.
Nucleic acid detection Detecting B19 DNA using NAAT is now routinely performed in many clinical
laboratories, and has been shown to be much more sensitive than antigen-based detection systems [50-52].
Analyte-specific reagents are available from several manufacturers for detecting B19 DNA. Numerous PCR-based
primer and probe sets have been described for detecting B19-specific DNA target(s). Most of these oligonucleotides
were designed to detect genotype 1 DNA, but not genotypes 2 or 3, which can lead to failure in detecting non-
genotype 1 erythrovirus infections [11,12,14,15]. The lightCycler-Parvovirus B19 quantitative assay (Roche
Diagnostics, Indianapolis, IN) is highly sensitive for genotype 1 but is not suitable for detecting genotypes 2 or 3.
RealArt Parvo B19 LC PCR (Qiagen, Hamburg, GmbH) has been reported to detect all three genotypes by some
investigators, but not by others [53,54].
Appropriate clinical specimens for NAAT analysis include serum, plasma, bone marrow, amniotic fluid, placental
and fetal tissues. To ensure diagnostic testing accuracy, it is critical that the laboratory has adequately assessed
the analytical performance characteristics (eg, analytical sensitivity, analytical specificity, reproducibility, and limit
of detection) of the B19 NAAT prior to its clinical implementation on each sample type that will be analyzed. As
with any diagnostic test, NAAT can produce false positive and/or false negative results.
A false negative NAAT result can occur if the individual's infection is due to the less common genotypes 2 or 3
if the primer and probe sequences used are located in a non-homologous gene set being used targets a
divergent region [12,14,15].
A false negative NAAT result can also occur when specimens contain inhibitor(s). This is especially true when
using a crude cell lysate rather than purified nucleic acid in the NAAT assay. An internal control should be
included in each run when analyzing samples so that significant levels of inhibitors can be detected. This
approach allows one to be more confident that the negative result is due to lack of detectable target and not to
inhibitors in the specimen.

A false positive NAAT result can occur because of contamination of either carry-over of genomic DNA during
processing of a specimen containing a high viral load, or from high levels of the amplified target generated from
a previous run. This is especially true in laboratories using a nested PCR assay for B19 DNA amplification
and those do not incorporate pre-amplification contamination control steps. Risk of contamination can be
reduced significantly if real-time PCR testing is used. In this approach, amplification and detection are
performed in a closed, self-contained vessel that is not opened during the analysis, which eliminates post
amplification handling of the amplicons. Another strength of the real-time PCR assay is its ability to be

Several real-time PCR assays have been described for detecting B19 DNA, although none have been cleared by the
Food and Drug Administration, though some NAAT kits have been CE marked (ie, Roche Molecular Diagnostics,
Abbott Molecular, Altona Diagnostics) [57-61]. Several manufacturers now offer analyte specific reagents for
detecting B19 DNA; some have been designed to detect and differentiate between all three genotypes while others
do not [54].
As noted above, B19 transmission can occur via the administration of blood products [26-30]; the risk is greatest in
those products prepared from large sized plasma pools [28,29]. As a result, there is heightened interest both in
Europe and North America for establishing a maximum cutoff for an acceptable viral load in certain blood products.
Observations in healthy volunteers suggest that acute B19 infection can be acquired from administration of blood
components that contain greater than 10 (log 7) genome equivalents/mL of B19 DNA. In contrast, patients receiving
greater than 10 (log 4) genome equivalents/mL have not shown evidence of virus transmission [62-64]. The presence
of neutralizing activity in all these pools may help to explain the lack of infectivity with exposure to lower viral loads
[26,62]. With the introduction of a World Health Organization International Standard for B19 DNA (NIBSC 99/800),
quantitative PCR assay standardization is possible [65].
Despite the fact that contaminated blood products can transmit B19 infection, there are at present no regulations in
the United States governing B19 contamination of pooled plasma or blood products. However, many manufacturers
voluntarily test plasma minipools for B19 DNA viral load and eliminate those with high levels of B19 virus. Because
NAAT can detect not only intact B19 genomes, but also fragments of degraded B19 DNA, it is important to be
cautious in interpreting NAAT data from filtered blood products [66]. The issue of providing blood products, which
have been screened for B19 DNA, to special high-risk populations such as pregnant women, immunocompromised
patients, and individuals with chronic anemia may also be addressed.
Antigen detection Immunohistochemical (IHC) techniques can be used to detect B19 antigens in a variety of
tissues, especially fetal and placental tissues [67-69]. There are commercially available sources of monoclonal or
polyclonal B19 specific antibodies that recognize B19 capsid proteins VP1 and VP2. Antibodies recognizing NS1
protein have been reported, but are not yet commercially available. Although IHC techniques allow for direct
visualization of virus within a tissue, it suffers from suboptimal sensitivity compared with NAAT and if used alone for
diagnosing B19 infections will miss B19-positive cases [25].
Virus isolation Freshly harvested bone marrow or fetal cord blood, or several continuous cell lines can support
low level B19 replication in vitro. However, these in vitro systems have not been used for clinical applications
[70,71].
SUMMARY AND RECOMMENDATIONS
quantitative. However, at this time it does not appear that determining viral load has a prognostic value outside
of the blood banking industry [55].
A false positive result can be seen in immunocompromised patients where B19-specific DNA has been
described to be found circulating for months or even years after the infection. This is especially true when
testing bone marrow or synovial fluid [6,35,56]. Thus, detection of B19 DNA, especially at very low levels,
does not necessarily indicate an acute or recent infection.

Parvovirus B19 infection occurs worldwide as sporadic cases or within clustered outbreaks. (See
'Epidemiology' above.)

Parvovirus B19 infection is highly prevalent. More than 70 percent of adults have evidence of B19-specific IgG
antibodies. (See 'Prevalence' above.)

Infected patients are most contagious during the phase of active viral replication and viral shedding, which
occurs approximately 5 to 10 days after exposure. During this initial phase, patients may present with a non-
specific flu-like illness or may be asymptomatic. During the second phase of illness, more specific symptoms
of parvovirus infection include arthralgia, arthritis, and/or rash. Patients are no longer infectious when

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Topic 8274 Version 7.0
exhibiting these clinical characteristics. (See 'Incubation period and infectivity' above.)
The three documented modes of transmitting parvovirus B19 include respiratory spread, vertical transmission
during pregnancy and hematogenous transmission through blood products. (See 'Transmission' above.)

The choice of diagnostic testing for acute parvovirus B19 infection will depend on the patient's immune status.
In immunocompetent individuals, the preferred method is serologic testing to detect circulating B19-specific
IgM and IgG antibodies. Among immunocompromised patients or infants, nucleic acid amplification testing
(NAAT) is preferred. (See 'Acute infection' above.)

Previous infection is best confirmed through serologic testing for B19-specific IgG antibodies, which infer
immunity. Such testing is helpful in determining susceptibility in a pregnant woman with a history of parvovirus
exposure. (See 'Previous infection' above.)

Reactivation of infection occurs in immunocompetent and, more often, among immunocompromised

individuals with serologic evidence of past infection. Reactivation is confirmed by demonstrating the continued
presence of parvovirus B19 DNA over time through nucleic acid amplification testing. IgM-specific parvovirus
antibodies are often detectable during reactivation among those patients with sufficient humoral immunity.
(See 'Reactivation of infection' above.)

Acute infection is demonstrated through serologic detection of B19-specific IgM antibodies to viral capsid
antigen. Commercial assays vary both in sensitivity (70 to 100 percent) and specificity (76 to 100 percent).
The presence of rheumatoid factor, anti-nuclear antibodies, and Epstein-Barr virus IgM in a specimen can lead
to false positive IgM results. (See 'IgM antibody assays' above.)

Serologic testing for past infection using parvovirus B19-specific IgG antibodies has excellent sensitivity and
specificity. (See 'IgG antibody assays' above.)

Detecting B19 DNA using nucleic acid amplification testing (NAAT) is more sensitive than antigen-based
detection systems. Appropriate clinical specimens for NAAT analysis include serum, plasma, bone marrow,
amniotic fluid, placental and fetal tissues. (See 'Nucleic acid detection' above.)

Viral isolation is usually only performed in research laboratories and is not generally available for patient
management. (See 'Virus isolation' above.)