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PubMed2004

Analysing myocardial tissue from explanted hearts of heart transplant recipients and multi-organ donors for the presence of parvovirus B19 DNA.

BACKGROUND: Parvovirus B19 (PVB19) is an erythrovirus causing diverse clinical manifestations ranging from asymptomatic or mild to more severe outcomes, dependent on the haematological and/or immunological status of the host. Reports of PVB19 infection as a causative agent of paediatric or adult inflammatory cardiac diseases, or of cardiac transplant rejection are rare. OBJECTIVES: To identify PVB19 and other cardiotropic viruses in the myocardium of heart transplant (HTx) recipients and multi-organ donors (MOD). Furthermore, to assess the prevalence of cardiotropic viral infection in inflammatory heart disease. STUDY DESIGN: Heart tissue samples from 110 explants were analysed for PVB19 using primers and a 5'-nuclease probe designed to amplify a 160-basepair PCR product from the VP1/NS1 gene region. Samples tested included those obtained from patients undergoing HTx or from MODs. The findings were correlated with clinical course, histologic analysis and serologic testing. Confirmation of the positive PCR-results was done by sequencing and in situ hybridisation. RESULTS: The new assay described here allows precise quantitation of viral load over 7 orders of magnitude (10(6) to 10(0)IU/assay). Measurable amounts of parvoviral genomes were detected in 4/56 (7%) explanted HTx-hearts and in 5/54 (9%) explanted MOD-hearts. CONCLUSIONS: The newly developed real-time PCR is a rapid, sensitive and specific method to detect PVB19 infection in heart tissue. It will be a useful tool to address important questions regarding viral infections transmitted by transplantation, acute infections, relapses and complications involving late or chronic rejection.

Adolescent↗
PubMed2004

Long-term parvovirus B19 viraemia associated with pure red cell aplasia after allogeneic bone marrow transplantation.

BACKGROUND: Parvovirus B19 infection is associated with a variety of symptoms like erythema infectiosum, anaemia and arthritis. In immunocompetent persons, viraemia is usually cleared a few weeks after infection. OBJECTIVE: An immunocompromised adult female patient was persistently infected with B19 after allogenic bone marrow transplantation (BMT) and developed chronic anaemia. STUDY DESIGN: B19-specific antibodies were determined by ELISA and viral load was assessed using a quantitative real time B19 PCR. The patient was evaluated clinically. RESULTS: Two years after successful BMT, the patient received intensified immunosuppressive treatment, erythropoetin and erythrocyte concentrates due to chronic graft-versus-host disease with renal failure. Despite of this treatment, the aplastic anaemia worsened. PCR revealed B19 viraemia with 10(12) geq/ml serum. After 7 months of repeated applications of immunoglobulins and reduction of immunosuppressive treatment, reticulocyte counts and haemoglobin levels normalized and the viral load finally dropped to 10(3) geq/ml serum. One of the back-up samples of the erythrocyte concentrates tested positive, the respective transfusion had been applied 2 months after the beginning of viraemia. CONCLUSIONS: The source of the primary infection remained unclear, but at least re-infection by blood transfusion is likely. Treatment did not result in virus elimination from peripheral blood but in resolvement of symptoms.

Anemia, Aplastic↗
PubMed2005

Human parvovirus B19 infection during pregnancy--value of modern molecular and serological diagnostics.

BACKGROUND: Over 95% of fetal complications (fetal hydrops and death) occur within 12 weeks following acute parvovirus B19 (B19) infection in pregnancy. Therefore, weekly fetal ultrasound monitoring is generally recommended for this time period. However, in the majority of women, typical symptoms of acute infection (rash or arthropathy) are absent, and during epidemics, B19 infection may be diagnosed incidentally by antibody screening of women at risk. OBJECTIVE: To assess the diagnostic value of currently available molecular and serological methods for reliable diagnosis of primary B19 infection in pregnancy. STUDY DESIGN: Large panels of well-characterized acute-phase or convalescent sera were used to investigate the ability of a VP2 IgM EIA, a Light-Cycler-based B19-DNA PCR, a VP1-IgG avidity EIA and two VP2-IgG epitope-type specificity [ETS] EIAs to pinpoint the time of primary B19 infection in pregnancy. RESULTS: The duration of low-level IgM positivity varied greatly (range 4-26 weeks). Samples collected within the first 2 weeks of infection showed high-level viremia (mean 1.75 x 10(8) geq/ml). During follow-up, low-level DNAemia (mean 9.7 x 10(4)geq/ml) persisted for at least 18 weeks in 91% (20/22) of patients. Considering the first 12 weeks after onset of disease the window of greatest risk for fetal complications, the "acute" phase was extended to cover this full period. In this case, performing the avidity and ETS-EIA sequentially, the positive predictive value was 100% in patients showing concordant avidity and ETS-EIA results. CONCLUSIONS: In the presence of low IgM titres and/or low-level DNAemia the use of supplementary serological assays such as VP1-IgG avidity EIA and VP2-ETS-EIA is advisable for restriction or avoidance of unnecessary fetal ultrasound examinations or invasive diagnostics; and in general for strengthening the reliability of B19 serodiagnosis of pregnant women.

Antibodies, Viral↗
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↗
PubMed2006

Co-infection of human parvovirus B19 in Vietnamese patients with hepatitis B virus infection.

BACKGROUND/AIMS: Human parvovirus B19 (B19) has been identified in the serum of hepatitis B virus (HBV) infected patients. However, the effect of B19-infection on the course of HBV-associated liver disease has not previously been investigated. We examined the prevalence of B19-DNA in HBV-infected Vietnamese patients and analysed the association between co-infection and the clinical outcome of HBV-infection. METHODS: Serum samples from 399 HBV-infected patients and 64 healthy individuals were analysed for the presence of B19-DNA by PCR and DNA-sequencing. RESULTS: B19-DNA was detected in 99/463 (21.4%) individuals. The proportion of HBV-infected patients who were also co-infected with B19 was higher than the healthy controls (P<0.001). B19-DNA was detected more frequently in patients with HBV-associated hepatocellular carcinoma compared to patients with acute and chronic HBV, HBV-associated liver cirrhosis and healthy subjects (P<0.006). A positive correlation was also found between B19-DNA loads and both serum HBV-DNA loads and alanine aminotransferase (rho>0.250 and P<0.05). CONCLUSIONS: Our findings demonstrate that B19-infection is frequent in HBV-infected Vietnamese patients. Also, a significant correlation exists between HBV/B19 co-infection and a greater likelihood of progression to more severe hepatitis B-associated liver disease. Further studies are required to determine the role of B19-infection on HBV-associated pathogenesis.

Adult↗
PubMed2005

Immune reconstitution syndrome associated with parvovirus B19-induced pure red cell aplasia during highly active antiretroviral therapy.

Persistent parvovirus B19 (B19) manifested as chronic pure red cell aplasia (PRCA) has been described in HIV patients. Immune reconstitution syndrome (IRS) is an immune response to an occult opportunistic pathogen or tumor antigen, usually occurs after antiretroviral treatment. We describe the first case of IRS associated with B19-induced PRCA during highly active antiretroviral therapy.

Anti-HIV Agents↗
PubMed2005

Canine parvovirus infection: which diagnostic test for virus?

Five laboratory tests for diagnosis of canine parvovirus type 2 (CPV-2) infection were employed to test 89 faecal samples collected from dogs with diarrhoea. The tests analysed were immunochromatography (IC), haemagglutination (HA), virus isolation (VI), conventional and real-time PCR. IC, HA, VI and conventional or real-time PCR were able, respectively, to detect CPV-2 antigen or nucleic acid in 41, 50, 54, 68 and 73 of the samples. The best correlation was found between conventional and real-time PCR, with an overall agreement of 94.38%. Sixty-eight samples that tested positive by HA, VI or conventional PCR were subjected to antigenic and/or genetic analyses of the CPV-2 strains by monoclonal antibody (MAb), restriction fragment-length polymorphism (RFLP) and/or sequence analyses. In sum, out of the 68 strains analysed, 26 were characterised as CPV-2a, 18 as CPV-2b and 24 as a CPV-2 Glu-426 mutant recently identified in Italy.

Animals↗
PubMed2005

Characterisation of the canine parvovirus type 2 variants using minor groove binder probe technology.

Two minor groove binder (MGB) probe assays were developed for rapid identification and quantitation of the canine parvovirus type 2 (CPV-2) variants in the faecal samples of dogs. The MGB probe assays using type-specific probes labeled with different fluorophores (FAM and VIC) were able to detect the single nucleotide polymorphisms existing between types 2a/2b and 2b/2c. Both the MGB probe assays were found to be highly specific, sensitive and reproducible, ensuring a precise quantitation of the CPV-2 DNA, as confirmed by comparing the obtained viral DNA loads to those calculated using a real-time TaqMan-based PCR assay established previously. A total of 414 faecal specimens collected from diarrhoeic dogs and tested positive to CPV-2 were analysed by the MGB probe assays and by conventional methods (haemagglutination inhibition with MAbs, PCR-RFLP and sequence analysis) for prediction of the CPV-2 antigen specificity, with an overall agreement of 100%. These results make the established MGB probe assays an attractive tool for direct and rapid characterisation of type 2a, 2b and 2c CPVs.

Amino Acid Substitution↗
PubMed2006

A minor groove binder probe real-time PCR assay for discrimination between type 2-based vaccines and field strains of canine parvovirus.

A minor groove binder (MGB) probe assay was developed to discriminate between type 2-based vaccines and field strains of canine parvovirus (CPV). Considering that most of the CPV vaccines contain the old type 2, no longer circulating in canine population, two MGB probes specific for CPV-2 and the antigenic variants (types 2a, 2b and 2c), respectively, were labeled with different fluorophores. The MGB probe assay was able to discriminate correctly between the old type and the variants, with a detection limit of 10(1) DNA copies and a good reproducibility. Quantitation of the viral DNA loads was accurate, as demonstrated by comparing the CPV DNA titres to those calculated by means of the TaqMan assay recognising all CPV types. This assay will ensure resolution of most diagnostic problems in dogs showing CPV disease shortly after CPV vaccination, although it does not discriminate between field strains and type 2b-based vaccines, recently licensed to market in some countries.

Animals↗
PubMed2006

Diagnostic tools based on minor groove binder probe technology for rapid identification of vaccinal and field strains of canine parvovirus type 2b.

TaqMan-based diagnostic tests have been developed for the identification of canine parvovirus type 2 (CPV-2) strains in the faeces of dogs with diarrhoea, including a minor groove binder (MGB) probe assay for identification of type 2-based vaccines and field strains (types 2a, 2b and 2c). Since type 2b vaccines have been licensed recently in Europe, two novel MGB assays were developed for discrimination between type 2b vaccines and field strains of CPV. Such assays have been found to be highly sensitive, specific and reproducible, allowing for simultaneous detection of type 2b vaccinal and field strains present in the same specimens. These new assays will help resolution of the diagnostic problems related to the detection of a type 2b strain in the faeces of dogs shortly after the administration of a type 2b vaccine.

Analysis of Variance↗
PubMed2006

Selective regimen shift and demographic growth increase associated with the emergence of high-fitness variants of canine parvovirus.

The natural evolution of Canine parvovirus (CPV) is characterized by a variety of mutations, mainly in the VP1/VP2 gene. Although positive selection has been previously reported in CPV, little is known about its overall contribution to viral adaptation in the canine population. Herein, the influences of the evolutive constraints on CPV during a period of viral adaptation into a previously uninfected population are more clearly investigated. To do this, 31 sequences of VP1/VP2 gene obtained from symptomatic domestic dogs in Brazil were used, sampled from 1980 to 2000. Marked evolutionary changes in CPV associated with a process of fine-tuning adaptation were observed. Specifically, sequences from the 1980s revealed two distinct antigenic types (i.e. 2a and 2b) cocirculating in Brazil. Moreover, analysis of the selective regimen showed that 90% of the VP2 sites were conserved (d(N)/d(S)=0). In contrast, sequences from the 1990s were composed solely of CPV-2a with 96% of VP2 sites under purifying selection (d(N)/d(S)<1) and site 297 under strong positive selection (omega=4.9). Important features regarding the demographic history of CPV in Brazil were also observed. The viral population size passed through a short period of explosive growth that declined and then stabilized into a constant rate of spread. Remarkably, the explosive growth coincided with the appearance of CPV variants that presented a unique repertoire of mutations never before seen in other worldwide strains. The analysis also showed that the estimated nucleotide substitution was similar to those commonly observed in fast evolving RNA viruses. The present results demonstrated the adaptive potential of CPV to acquire, in short interval of 10 years, key mutations in the VP1/VP 2 gene that increased viral fitness and enabled the virus to disseminate even in vaccinated dogs.

Animals↗
PubMed2004

Molecular method to test if Parvovirus B19 is a new virus to the Old World.

Parvovirus B19 (PVB19) is a non-trivial cause of morbidity and mortality in young individuals from diseases such as aplastic anemia and hydrops fetalis. Curiously, given this significant disease burden, the one genetic phenotype known to protect against PVB19 infection--the p red-cell phenotype characterized by the lack on any P group antigens on erythrocytes--is extremely rare with a prevalence of only approximately 1/200,000. One explanation is that PVB19 is a new virus to the Old World. It is pointed out in this paper that recently developed powerful genetic methods can be used to test this proposal. Confirmation that PVB19 is a new virus could shed light on the etiology of rheumatoid arthritis.

Africa↗
PubMed2006

Parvovirus B19 infection of brain: possible role of gender in determining mental illness and autoimmune thyroid disorders.

Major theories about the etiologies of chronic mental illnesses such as bipolar disorder and schizophrenia include genetic and environmental factors such as famine and infection. It is likely that multiple genes play a role in the pathogenesis of these disorders, but no single gene has been identified as causative. Several viruses have been investigated as potential candidates, but conflicting reports exist. Although a relationship between bipolar disorder and schizophrenia with autoimmune disorders has also been documented for many years, reports are often conflicting. We hypothesize that parvovirus B19 (B19), a common human pathogen, due to its ability to infect the brain and induce autoimmunity, is a strong candidate that may unite prevailing theories. In particular, our preliminary data suggest that B19 may be most likely involved in co-morbid bipolar and autoimmune thyroid disorders in females. In schizophrenics, the gender trend may be reversed. We propose that there is a complex interaction between immuno-genetics, autoimmunity, gender, and B19 infection that leads to at least some forms of bipolar disorder and schizophrenia. Future studies that investigate this hypothesis are warranted and outlined.

Adult↗
PubMed2006

Laboratory assessment and diagnosis of congenital viral infections: Rubella, cytomegalovirus (CMV), varicella-zoster virus (VZV), herpes simplex virus (HSV), parvovirus B19 and human immunodeficiency virus (HIV).

Viral infections during pregnancy may cause fetal or neonatal damage. Clinical intervention, which is required for certain viral infections, relies on laboratory tests performed during pregnancy and at the neonatal stage. This review describes traditional and advanced laboratory approaches and testing methods used for assessment of the six most significant viral infections during pregnancy: rubella virus (RV), cytomegalovirus (CMV), varicella-zoster virus (VZV), herpes simplex virus (HSV), parvovirus B19 and human immunodeficiency virus (HIV). Interpretation of the laboratory tests results according to studies published in recent years is discussed.

Clinical Laboratory Techniques↗
PubMed2004

[Systemic manifestations of Parvovirus B19 infections].

PURPOSE: Parvovirus B19 (B19) causes many clinical disorders, of which the most common are erythema infectiosum, aplastic crisis complicating chronic hemolytic anemia, and hydrops fetalis. In young adults, the skin eruption caused by B19 is accompanied by polyarthritis and polyarthralgia in 60% of the cases. Rheumatoid factors and other antibodies including antinuclear antibodies, anti-ADN, and antiphospholipids can be produced in the wake of B19 infection. CURRENT KNOWLEDGE AND KEY POINTS: These features may simulate systemic diseases as rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) (lupus-like eruption over the cheeks, cytopenia, etc.) or vasculitis (purpura, renal involvement). In addition, there have been a few reports of SLE, vasculitis and other connective tissue diseases developing shortly after a B19 infection associated with virus clearance suggesting that B19 can act as a trigger of systemic disease. However, studies in large series indicate that in fact B19 is probably an extremely rare cause of RA, SLE or vasculitis. FUTURE PROSPECTS AND PROJECTS: In fundamental studies B19 interacts with inflammatory cells by regulation of cytokines. More recently, two studies suggest that viral infection due to B19 may affect the course of SLE, leading to specific biological subsets. These preliminary findings require confirmation to elucidate the significance of the presence of B19 in systemic disease.

Adolescent↗
PubMed2006

[Safety of blood products and B19 parvovirus].

More than 25 years after the discovery of the parvovirus B19 (B19), the issue of the safety of blood components and the screening of this virus in blood donations is still debated. Although more often transmitted by respiratory route, B19 may also be transmitted by transfusion of blood components. This risk of exposure has been estimated to a frequency ranging from 1/625 to 1/50,000, according to the sensitivity of the detection methods and to seasonal epidemiologic circumstances. Usually, B19 is responsible for benign pathologies. However, such an infection can have a serious clinical outcome in three categories of susceptible recipients: (i) patients with shortened red cell survival (thalassemia major, sickle cell disease, other hemolytic diseases); (ii) immunocompromised patients (previously exposed to B19 or not) (iii) and pregnant women (not previously exposed the B19), with a risk of hydrops fetalis or of intrauterine death. Selected blood components, not collected during the short but highly viremic pre-seroconversion phase, could be reserved for these three groups of at-risk recipients. The screening of such viremic donations could be performed with nucleic acid testing (NAT), but an alternate strategy could be the selection of B19 immunised donors far from the primo-infection (positive for B19 IgG and negative for B19 IgM, or only positive for IgG at two controls distant of several months). However, the existence of persistently B19-infected individuals carrying B19 DNA despite the presence of specific IgG (estimated at 1% of blood donors) could constitute a potential threat for transfused immunocompromised recipients. The screening of such donors, which could be performed through a very highly sensitive NAT, would be justified only if the infectivity of such blood donations is demonstrated. If not, a screening of blood donors positive for B19 IgG would be a sufficient preventive measure.

Blood Donors↗
PubMed2005

Prevalence of Parvovirus B19 and Hepatitis A virus in Portuguese blood donors.

INTRODUCTION: In recent years, concern about the safety of blood in regard to the transmission of blood-borne viruses has been decreased. Safety has been achieved with a combination of different strategies, such as careful selection of donors, screening for relevant virological markers and viral inactivation/removal methods. More recently, the implementation of the nucleic acid amplification technologies for the detection of HIV-1, HCV and HBV, has increased safety by reducing the "window period" of the infections. Other viruses, such as Parvovirus B19 (PB19) and Hepatitis A virus (HAV), can cause problems for blood safety. These infections could provoke serious complications in some risk groups, such as pregnant women, patients with hematological problems, children and patients with immunodeficiencies. MATERIALS AND METHODS: An observational study was performed to determine the prevalence of PB19 and HAV in Portuguese blood donors. We gathered, during four months, 5025 plasma donations and made them into 505 pools with no more than 10 donations each. The nucleic acids were isolated using MagNA Pure LC (Roche, Mannheim, Germany). A "Real Time PCR" (LightCycler, Roche) was used to perform the nucleic acid amplification and detection, using kits from the manufacturer. RESULTS: We found a prevalence of 0.12% for PB19 and 0% for HAV. Viraemia levels found in the positive donations range from 7.1x10(4) to 2.1x10(12)IU/ml. DISCUSSION: This study demonstrates the possibility of performing these tests in routine blood banks. We found a similar prevalence of PB19 when compared with other European and USA countries. In the case of HAV, we predict a maximum risk of 0.06% for a donor to be infected. It is necessary to perform other studies, including cost/benefit analysis to evaluate the risks and profits of implementing these methodologies in Transfusion Medicine.

Blood Banks↗
PubMed2006

Recurrent anemia in kidney transplant recipients with parvovirus B19 infection.

Parvovirus B19 (PV B19) infection is known to cause acute anemia in solid organ transplant recipients. Intravenous immunoglobulin combined with reduction of immunosuppression may be of benefit to clear the infection. However, PV B19-associated anemia can be recurrent. We describe three renal transplant recipients with a PV B19 infection. These patients showed recurrent anemia with episodes separated by as much as several months.

Adult↗