Amanda Corcoran

Impaired Gamma Interferon Responses against Parvovirus B19 by Recently Infected Children

ABSTRACT Parvovirus B19 is the causative agent of “fifth disease” of childhood. It has been implicated in a variety of conditions, including unsuccessful pregnancy and rheumatoid arthritis, and is a potential contaminant of blood products. There has been little study of immunity to parvovirus B19, and the exact nature of the protective humoral and cell-mediated immune response is unclear. Immune responses to purified virus capsid proteins, VP1 and VP2, were examined from a cohort of recently infected children and compared with responses from long-term convalescent volunteers. The results demonstrate that antibody reactivity is primarily maintained against conformational epitopes in VP1 and VP2. The unique region of VP1 appears to be a major target for cell-mediated immune responses, particularly in recently infected individuals. We confirm that antibody reactivity against linear epitopes of VP2 is lost shortly after infection but find no evidence of the proposed phenotypic switch in either the subclass of parvovirus B19-specific antibody or the pattern of cytokine production by antigen-specific T cells. The dominant subclass of specific antibody detected from both children and adults was immunoglobulin G1. No evidence was found for interleukin 4 (IL-4) or IL-5 production by isolated lymphocytes from children or adults. In contrast, lymphocytes from convalescent adults produced a typical type 1 response associated with high levels of IL-2 and gamma interferon (IFN-γ). However, we observed a significant (P < 0.001) deficit in the production of IFN-γ in response to VP1 or VP2 from lymphocytes isolated from children. Taken together, these results imply that future parvovirus B19 vaccines designed for children will require the use of conformationally preserved capsid proteins incorporating Th1 driving adjuvants. Furthermore, these data suggest novel mechanisms whereby parvovirus B19 infection may contribute to rheumatoid arthritis and unsuccessful pregnancy.

High-Sensitivity PCR Detection of Parvovirus B19 in Plasma

ABSTRACT Parvovirus B19 (B19) is a human pathogen transmitted to susceptible individuals via respiratory secretions and contaminated blood or blood products. B19 levels in pooled plasma of less than 104 genome equivalents/ml may not be infectious, while those greater than 107/ml are capable of transmitting infection. A World Health Organization (WHO) B19 DNA international standard has been recently introduced. The purpose of the present work was to develop a PCR-enzyme-linked immunosorbent assay (PCR-ELISA) calibrated against the WHO B19 DNA international standard which could easily and reliably detect B19 DNA levels in plasma above 104 IU/ml (6.5 × 103 genome equivalents/ml). A B19 PCR-ELISA system was developed which uses a dinitrophenylated oligonucleotide probe to detect immobilized biotinylated amplicons following single-round PCR amplification. The level of B19 DNA (in international units per milliliter) in individual and pooled plasma specimens was evaluated. Proteinase K treatment of plasma was found to be sufficient to quantitatively release B19 DNA. The B19 PCR-ELISA had a sensitivity of detection of 1.6 × 103 IU/ml B19 DNA and a dynamic range extending from 8 to 1,000 IU of B19 DNA (equivalent to 1.6 × 103 to 2 × 105 IU of B19 DNA/ml). Furthermore, the antibody profile of pooled plasma products was determined in terms of B19 immunoglobulin G (IgG) (in international units per milliliter). The B19 IgG level was found to be 64.7 ± 17.5 IU/ml (mean ± standard deviation). The B19 PCR-ELISA, which is calibrated against the B19 DNA international standard, may have an application for the rapid screening of plasma minipools for B19 DNA, thereby leading to an improvement in blood product safety.

B Cell Memory Is Directed toward Conformational Epitopes of Parvovirus B19 Capsid Proteins and the Unique Region of VP1

BACKGROUND Loss of antibody reactivity against linear epitopes of parvovirus B19 (B19) capsid proteins VP1 and VP2 occurs after infection; however, it is unclear whether B cell memory is established against linear epitopes. METHODS B cell enzyme-linked immunospot assay was used to evaluate B19-specific B cell memory in volunteer donors (n=22). RESULTS B cell memory is maintained against conformational epitopes of VP2 and is absent against linear epitopes of VP2. Individuals seronegative for IgG against the unique region of VP1 have detectable B cell memory, with the potential to mount a humoral response on reexposure to B19. Conversely, in mice immunized with VP2, long-lasting IgG against linear epitopes of VP2 and a strong B cell-memory response are observed. CONCLUSIONS B cell memory is established and maintained against conformational epitopes of VP2 and against linear epitopes of VP1 but not against linear epitopes of VP2. These findings further our understanding of the immune response to B19 and suggest that analysis of B19-specific B cell memory merits consideration for future B19-vaccine studies.

The Immune Response to Parvovirus B19 Exposure in previously Seronegative and Seropositive Individuals

Little information is available on the immune response to parvovirus B19 after the administration of contaminated blood products. In the present study, we found that levels of B19 IgG in B19-seropositive recipients protect against reinfection and, after transfusion with pooled plasma containing B19 DNA (1.6 x 10(8) IU/mL), increase from 19-39 IU/mL to 50-100 IU/mL. We found that, in the presence of 1.6-2.2 x 10(8) IU of B19 DNA/mL in B19-seronegative recipients, a pooled-plasma B19 IgG level of 59.5 IU/mL is insufficient to prevent B19 transmission and subsequent seroconversion. These data should lead to improvements in the assessment of blood-product safety.

Baculovirus expression of parvovirus B19 (B19V) NS1: utility in confirming recent infection

BACKGROUND The presence of anti-parvovirus B19 (B19V) IgM against viral capsid proteins (VP1 and VP2) has long been used to detect recent infection. The utility of antibodies directed against B19V NS1 protein has received less attention as a serological indicator of recent infection, although anti-B19V NS1 IgG has been associated with persistent infection. OBJECTIVES To elucidate the role of anti-B19V NS1 antibody detection in recent infection, full-length B19V NS1 was expressed and purified. The resultant antigen was used to develop both Western blot assays and microplate ELISA for the detection of NS1 antibodies. STUDY DESIGN Serum specimens were obtained from individuals recently infected with B19V (children (n=16), adults (n=40)) and from 17 individuals with no evidence of recent B19V infection. All specimens were screened for anti-B19V NS1 IgG and IgM. RESULTS It was observed that 68.8% (11/16) of children recently infected with B19V were anti-B19V NS1 IgG seropositive. Furthermore, 27.5% (11/40) anti-B19V VP2 IgM positive specimens also contained anti-B19V NS1 IgM when tested by ELISA, while no reactivity was observed following Western blot analysis, possibly due to the absence of conformational epitopes. CONCLUSIONS Anti-B19V NS1 IgM detection may have utility in the confirmation of recent infection with B19V.

Evidence of Serological Cross-Reactivity between Genotype 1 and Genotype 3 Erythrovirus Infections

Candotti and colleagues (1) reported the prevalence of a third strain of human erythrovirus, genotype 3 (V9), in the Ghanaian population and, in part, concluded that a genotype 1 (B19)-based assay failed to detect genotype 3 immunoglobulin G (IgG) in 38.5% of Ghanaian samples containing genotype 3 antibodies. We disagree with this conclusion for the following reasons.

Improved detection of acute parvovirus B19 infection by immunoglobulin M EIA in combination with a novel antigen EIA

Background and Objectives  Although parvovirus B19 is a significant blood product contaminant, few methods other than polymerase chain reaction (PCR) have been developed to detect the presence of the virus.

Human Parvovirus B19: Molecular Virology, Clinical Features, Prevalence, Diagnosis and Control

Publisher Summary Human parvovirus B19 (B19) was first identified in 1975 by Yvonne Cossart. The virus was first associated with disease in 1981, when it was linked to an aplastic crisis in a patient with sickle-cell disease. Subsequently, B19 has since been shown to be the causative agent of erythema infectiosum (EI), spontaneous abortion, and some forms of acute arthritis. Parvovirus B19 (B19) is an erythrovirus, and recent studies have classified B19 as genotype 1 erythrovirus with genotype 2 (erythrovirus K71 or A6) and 3 (erythrovirus V9) also present in the human population. B19 is a significant human pathogen that can cause fetal hydrops and foetal death, if maternal infection followed by transplacental fetal infection occurs during pregnancy. The virus is also transmitted by inter-personal contact and potentially via blood product administration. Symptoms of B19 infection include malaise, rash, and anthralgia. Significantly, maternal B19 infection during pregnancy can be asymptomatic and so careful monitoring of at-risk pregnancies is recommended. Both antibody and cell-mediated immunity play an important role in the anti-viral response and effective diagnostic test systems, for both B19 antibody and DNA detection, are now available. B19-induced fetal hydrops can be effectively treated by intrauterine blood transfusion; however, no vaccine is available to prevent infection at present.