Recently, three distinct genotypes (1, 2 and 3) of human parvovirus B19 (B19) have been identified. However, the characteristics and distribution of B19 genotypes in Vietnam have not been investigated. Phylogenetic analysis using 49 subgenomic NS1/VP1u regions and two coding NS1-VP1/VP2 regions has been applied to investigate the prevalence of B19 genotypes in Vietnamese patients co-infected with Hepatitis B virus. Genetic analysis of the subgenomic NS1/ VP1u region of B19 revealed that two genotypes of B19 were identified in these populations, with predominance of genotype 1 (47/49, 96 %) followed by genotype 2 (2/49, 4 %), but not genotype 3. Further, phylogenetic analysis of subgenomic B19 genomes revealed two major subgroups within genotype 1 (B19-1A and B19-1B) with an estimated nucleotide difference of >5 % between each subgroup, forming different branches. The mean percentage of amino acid variation between subgroup B19-1A and B19-1B was >2 % of the NS1, VP1 and VP2 proteins. Our results indicated that two of the three known genotypes of B19 were present in Vietnamese patients, with genotype 1 predominating, and that this genotype can be classified into at least two subgroups, B19-1A and B19-1B. INTRODUCTIONHuman parvovirus B19 (B19) is a member of the genus Erythrovirus within the family Parvoviridae . Parvoviruses are non-enveloped and are amongst the smallest DNA-containing viruses that are capable of infecting mammalian cells . B19 has a diverse spectrum of clinical manifestations, including erythema infectiosum, hydrops fetalis, aplastic anaemia (Anderson et al., 1983;Young & Brown, 2004), arthritis (Takahashi et al., 1998;Moore, 2000), myocarditis (Bültmann et al., 2003;Bock et al., 2005; Tschöpe et al., 2005;Bock, 2006), vasculitic syndromes (Finkel et al., 1994;Dingli et al., 2000), neurological disorders (Barah et al., 2001) and hepatic inflammation (Naides et al., 1996;Yoto et al., 1996;Hillingsø et al., 1998;Karetnyi et al., 1999; He et al., 2003).The genome of B19 consists of a linear, single-stranded DNA molecule of about 5600 nt (Morinet, 1992;Zhi et al., 2004), which contains three open reading frames encoding the non-structural protein NS1 (77 kDa) and the two structural proteins VP1 (84 kDa) and VP2 (58 kDa) (Cotmore et al., 1986). Genetic diversity among B19 strains has been shown to be very low, with <1-2 % nucleotide divergence of the full-length sequences. Partial sequence data from different coding regions of the viral genome have confirmed a slightly higher degree of variability with a larger number of strains from distinct epidemiological settings and geographical areas (Erdman et al., 1996). However, some B19 strainsThe GenBank/EMBL/DDBJ accession numbers for the nucleotide sequences of B19 samples determined in this study are DQ357064 (V147) and DQ357065 (V115). obtained from patients with persistent infection show a higher degree of variability, particularly in the VP1-unique (VP1u) region, demonstrating 4 and 8 % divergence at the DNA and protein levels, respectively (Hemaue...

The pathogenic mechanism by which parvovirus B19 may induce inflammatory cardiomyopathy (iCMP) is complex but is known to involve inflammatory processes, possibly including activation of JAK/STAT signaling. The nonstructural B19 protein NS1 acts as a transactivator triggering signaling cascades that eventually lead to activation of interleukin 6 (IL-6). We examined the impact of NS1 on modulation of STAT signaling in human endothelial cells (HMEC-1). The NS1 sequences were identified from B19 DNA isolated from the myocardia of patients with fatal iCMP. B19 infection as well as NS1 overexpression in HMEC-1 cells produced a significant upregulation in the phosphorylation of both tyrosine 705 and serine 727 STAT3 (P < 0.05). The increased STAT3 phosphorylation was accompanied by dimerization, nuclear translocation, and DNA binding of pSTAT3. In contrast, NS1 expression did not result in increased STAT1 activation. Notably, the expression levels of the negative regulators of STAT activation, SOCS1 and SOCS3, were not altered by NS1. However, the level of PIAS3 was upregulated in NS1-expressing HMEC-1 cells. Analysis of the transcriptional activation of target genes revealed that NS1-induced STAT3 signaling was associated with upregulation of genes involved in immune response (e.g., the IFNAR1 and IL-2 genes) and downregulation of genes associated with viral defense (e.g., the OAS1 and TYK2 genes). Our results demonstrate that B19 NS1 modulates the STAT/PIAS pathway. The NS1-induced upregulation of STAT3/PIAS3 in the absence of STAT1 phosphorylation and the lack of SOCS1/SOCS3 activation may contribute to the mechanisms by which B19 evades the immune response and establishes persistent infection in human endothelial cells. Thus, NS1 may play a critical role in the mechanism of viral pathogenesis in B19-associated iCMP.Human parvovirus B19 is emerging as an important pathogenic agent in the etiology of inflammatory cardiomyopathy (iCMP). Recent studies have indicated an association between infection with B19 and acute myocarditis in both children and adults (5, 44). However, the role of B19 infection in the development of chronic iCMP patients is still unclear. We have recently demonstrated that endothelial cells but not cardiac myocytes are B19-specific target cells in patients with B19-associated myocarditis (21). Furthermore, B19 could be detected frequently in patients with unexplained isolated diastolic dysfunction (2, 49).The B19 single-stranded DNA genome of 5,600 base pairs contains two open reading frames encoding the nonstructural protein NS1 and two structural capsid proteins, VP1 and VP2, by a combination of alternative splicing (15). A functional phospholipase A2-like activity has been demonstrated recently in the VP1 region which is involved in intracellular Ca 2ϩ regulation (28). In addition, three small viral proteins of unknown function have been described previously (39,55).The main function of NS1 includes transactivation of the viral P6 promoter, which is important for viral replication in a proces...

Background: Immune checkpoint blockade such as ipilimumab and anti-PD1 monoclonal antibodies have significantly improved survival in advanced melanoma. Biomarkers are urgently needed as a majority of patients do not respond, despite treatment-related toxicities. We analysed pre-treatment 18 F-fluorodeoxyglucose positron emission tomography/computerised tomography (FDG PET/CT) parameters to assess its correlation with patient outcome. Methods: This retrospective study evaluated pre-treatment FDG PET/CT scans in a discovery cohort of patients with advanced melanoma treated with ipilimumab or anti-PD1. Pre-treatment scans were assessed for maximum tumoral standardised uptake value (SUVmax), metabolic tumour volume (MTV) and spleen to liver ratio (SLR). Progression-free survival (PFS) and overall survival (OS) were characterised and modelled using univariable and multivariable analyses. Correlation of SLR and OS was validated in an independent cohort. Blood parameters and stored sera of patients from the discovery cohort was analysed to investigate biological correlates with SLR. Results: Of the 90 evaluable patients in the discovery cohort: 50 received ipilimumab monotherapy, 20 received anti-PD1 monotherapy, and 20 patients received ipilimumab followed by anti-PD1 upon disease progression. High SLR > 1.1 was associated with poor PFS (median 1 vs 3 months; HR 3.14, p = 0.008) for patients treated with ipilimumab. High SLR was associated with poor OS after ipilimumab (median 1 vs 21 months; HR 5.83, p = 0.0001); as well as poor OS after first line immunotherapy of either ipilimumab or anti-PD1 (median 1 vs 14 months; HR 3.92, p = 0.003). The association of high SLR and poor OS after ipilimumab was validated in an independent cohort of 110 patients (median 2.3 months versus 11.9 months, HR 3.74). SLR was associated with poor OS in a multi-variable model independent of stage, LDH, absolute lymphocyte count and MTV. Conclusions: Pre-treatment Spleen to liver ratio (SLR) > 1.1 was associated with poor outcome after ipilimumab in advanced melanoma. This parameter warrants prospective evaluation.

Recent reports demonstrated an association of human parvovirus B19 with inflammatory cardiomyopathy (iCMP), which is accompanied by endothelial dysfunction. As intracellular Ca 2؉ activity is a key regulator of cell function and participates in mechanisms leading to endothelial dysfunction, the present experiments explored the effects of the B19 capsid proteins VP1 and VP2. A secreted phospholipase A2 (PLA2)-like activity has been located in the VP1 unique region of the B19 minor capsid protein. As PLA2 has recently been shown to activate the store-operated or capacitative Ca 2؉ channel I CRAC , we analyzed the impact of the viral PLA2 motif on Ca 2؉ entry. We cloned the VP1 and VP2 genes isolated from a patient suffering from fatal B19 iCMP into eukaryotic expression vectors. We also generated a B19 replication-competent plasmid to demonstrate PLA2 activity under the control of the complete B19 genome. After the transfection of human endothelial cells (HMEC-1), cytosolic Ca Human parvovirus B19, a nonenveloped virus of about 22 to 24 nm in diameter, is a member of the genus Erythrovirus within the family of Parvoviridae (35). B19 infection occurs frequently in humans, and this is documented by the high prevalence of specific immunoglobulin G (IgG) antibodies in young children (5% to 15%), adults (60%), and seniors older than 69 years (85%) (15). B19 is the causative agent of erythema infectiosum (fifth disease), hydrops fetalis, and transient aplastic anemia (1,86). Several studies disclosed an association between B19 and a variety of diseases (43,48,63,80), such as arthritis (56, 77), vasculitic syndromes (19, 24, 31, 80), hepatitis (27,36,38,75,85), and neurological disorders (2, 85). Specifically, B19 infections have been observed to be associated with acute and chronic myocarditis (13,16,30,44,53,(58)(59)(60)71). Moreover, the development of endothelial and isolated left ventricle diastolic dysfunction has been associated with B19 infection (81). During pregnancy, parvovirus B19 infection may cause maternal and fetal myocarditis, congenital abnormalities, stillbirth, and abortion (10, 21, 39, 61). The particularly severe course of the antenatal disease may relate to the preference of B19 for proliferating tissues (79).The cellular receptor for B19 infection has been regarded as a blood group P antigen based on the failure of B19 infection in a patient with a hereditary P antigen defect (12). The P antigen is necessary for B19 binding but not sufficient for virus entry into cells. In this regard, the ␣5␤1 integrin and the recently identified Ku80 autoantigen act as cellular coreceptors for human parvovirus B19 infection (57, 82). Therefore, target cells of B19 are mainly erythroid progenitor cells expressing high levels of P antigen as well as the coreceptors ␣5␤1 integrin and Ku80 autoantigen. However, nonerythroid cell lineages, such as fetal myocytes, follicular dendritic cells, and endothelial cells can be infected by B19 (12,28,29,57,82). We have recently localized B19 genomes in endothelial cells of my...

The human parvovirus B19 (PVB19), an erythrovirus causing diverse clinical manifestations ranging from asymptomatic or mild to more severe outcomes such as hydrops fetalis, is the only known human pathogenic parvovirus so far. Although enteroviruses have long been considered the most common cause of inflammatory cardiomyopathy, PVB19 is emerging as a important candidate. Recent studies have indicated an association of PVB19 with paediatric and adult inflammatory cardiac disease. However, whether or not PVB19 has an impact on inflammatory cardiomyopathy in adult patients is still unclear. The first hints for a possible aetiopathogenetic role of the PVB19-infection and the development of cardiac dysfunction were demonstrated by molecular biology utilizing in situ hybridization (ISH) and polymerase chain reaction (PCR). According to available evidence, PVB19-associated inflammatory cardiomyopathy is characterized by infection of endothelial cells of small intracardiac arterioles and venules, which may be associated with endothelial dysfunction, impairment of myocardial microcirculation, and penetration of inflammatory cells into the myocardium.