Complete Genome Sequence of Germline Chromosomally Integrated Human Herpesvirus 6A and Analyses Integration Sites Define a New Human Endogenous Virus with Potential to Reactivate as an Emerging Infection.

Human herpesvirus-6A and B (HHV-6A, HHV-6B) have recently defined endogenous genomes, resulting from integration into the germline: chromosomally-integrated "CiHHV-6A/B". These affect approximately 1.0% of human populations, giving potential for virus gene expression in every cell. We previously showed that CiHHV-6A was more divergent than CiHHV-6B by examining four genes in 44 European CiHHV-6A/B cardiac/haematology patients. There was evidence for gene expression/reactivation, implying functional non-defective genomes. To further define the relationship between HHV-6A and CiHHV-6A we used next-generation sequencing to characterize genomes from three CiHHV-6A cardiac patients. Comparisons to known exogenous HHV-6A showed CiHHV-6A genomes formed a separate clade; including all 85 non-interrupted genes and necessary cis-acting signals for reactivation as infectious virus. Greater single nucleotide polymorphism (SNP) density was defined in 16 genes and the direct repeats (DR) terminal regions. Using these SNPs, deep sequencing analyses demonstrated superinfection with exogenous HHV-6A in two of the CiHHV-6A patients with recurrent cardiac disease. Characterisation of the integration sites in twelve patients identified the human chromosome 17p subtelomere as a prevalent site, which had specific repeat structures and phylogenetically related CiHHV-6A coding sequences indicating common ancestral origins. Overall CiHHV-6A genomes were similar, but distinct from known exogenous HHV-6A virus, and have the capacity to reactivate as emerging virus infections

Allelic variations in coding regions of the vitamin D receptor gene in dairy cows and potential susceptibility to periparturient hypocalcaemia

Periparturient hypocalcaemia (milk fever) is a disorder of Ca metabolism in dairy cattle primarily affecting multiparous cows. The major reasons for the rapid decrease of blood Ca concentration after calving are the prompt increase of Ca secretion into the colostrum and the delayed activation of Ca regulation mechanisms including calcitriol, a metabolite of vitamin D. In man, vitamin D receptor (VDR) gene polymorphisms are reported to be associated with disturbances of Ca metabolism, whereas data confirming the same in dairy cows are still missing. Moreover, polymorphisms that only affect non-coding regions are sometimes difficult to ascribe to a specific disorder as pathways and unequivocal links remain elusive. Therefore, the idea of the present study was to investigate in a small group of dairy cows with documented clinical records whether polymorphisms in the coding regions of the VDR gene existed and whether these potentially found variations were correlated with the incidence of periparturient hypocalcaemia. For this purpose, blood DNA was isolated from 26 dairy cows in their 4th to 6th lactation, out of which 17 had experienced hypocalcaemia at least once, whereas 9 cows had never undergone periparturient hypocalcaemia in their lifetime. The 10 VDR exons and small parts of adjacent introns were sequenced and compared with the Bos taurus VDR sequence published on NCBI based on the DNA of one Hereford cow. In total, 8 sequence alterations were detected in the fragments, which were primarily heterozygous. However, only 4 of them were really located on exons thereby potentially causing changes of the encoded amino acid of the VDR protein, but were not correlated with the incidence of periparturient hypocalcaemia. Certainly, this lack of statistical correlation could be due to the small number of animals included; anyhow, it was not encouraging enough to initiate a larger study with hundreds of cows and document blood Ca levels post partum for at least four lactations

Evolutionary History of Endogenous Human Herpesvirus 6 Reflects Human Migration out of Africa

Human herpesvirus 6A and 6B (HHV-6) can integrate into the germline, and as a result, similar to 70 million people harbor the genome of one of these viruses in every cell of their body. Until now, it has been largely unknown if 1) these integrations are ancient, 2) if they still occur, and 3) whether circulating virus strains differ from integrated ones. Here, we used next-generation sequencing and mining of public human genome data sets to generate the largest and most diverse collection of circulating and integrated HHV-6 genomes studied to date. In genomes of geographically dispersed, only distantly related people, we identified clades of integrated viruses that originated from a single ancestral event, confirming this with fluorescent in situ hybridization to directly observe the integration locus. In contrast to HHV-6B, circulating and integrated HHV-6A sequences form distinct clades, arguing against ongoing integration of circulating HHV-6A or "reactivation" of integrated HHV-6A. Taken together, our study provides the first comprehensive picture of the evolution of HHV-6, and reveals that integration of heritable HHV-6 has occurred since the time of, if not before, human migrations out of Africa

NS1 Specific CD8(+) T-Cells with Effector Function and TRBV11 Dominance in a Patient with Parvovirus B19 Associated Inflammatory Cardiomyopathy

Background: Parvovirus B19 (B19V) is the most commonly detected virus in endomyocardial biopsies (EMBs) from patients with inflammatory cardiomyopathy (DCMi). Despite the importance of T-cells in antiviral defense, little is known about the role of B19V specific T-cells in this entity. Methodology and Principal Findings: An exceptionally high B19V viral load in EMBs (115,091 viral copies/mg nucleic acids), peripheral blood mononuclear cells (PBMCs) and serum was measured in a DCMi patient at initial presentation, suggesting B19V viremia. The B19V viral load in EMBs had decreased substantially 6 and 12 months afterwards, and was not traceable in PBMCs and the serum at these times. Using pools of overlapping peptides spanning the whole B19V proteome, strong CD8(+) T-cell responses were elicited to the 10-amico-acid peptides SALKLAIYKA (19.7% of all CD8(+) cells) and QSALKLAIYK (10%) and additional weaker responses to GLCPHCINVG (0.71%) and LLHTDFEQVM (0.06%). Real-time RT-PCR of IFN gamma secretion-assay-enriched T-cells responding to the peptides, SALKLAIYKA and GLCPHCINVG, revealed a disproportionately high T-cell receptor Vbeta (TRBV) 11 expression in this population. Furthermore, dominant expression of type-1 (IFN gamma, IL2, IL27 and Tbet) and of cytotoxic T-cell markers (Perforin and Granzyme B) was found, whereas gene expression indicating type-2 (IL4, GATA3) and regulatory T-cells (FoxP3) was low. Conclusions: Our results indicate that B19V Ag-specific CD8(+) T-cells with effector function are involved in B19V associated DCMi. In particular, a dominant role of TRBV11 and type-1/CTL effector cells in the T-cell mediated antiviral immune response is suggested. The persistence of B19V in the endomyocardium is a likely antigen source for the maintenance of CD8(+) T-cell responses to the identified epitopes

Evolutionary History of Endogenous Human Herpesvirus 6 Reflects Human Migration out of Africa.

Human herpesvirus 6A and 6B (HHV-6) can integrate into the germline, and as a result, ∼70 million people harbor the genome of one of these viruses in every cell of their body. Until now, it has been largely unknown if 1) these integrations are ancient, 2) if they still occur, and 3) whether circulating virus strains differ from integrated ones. Here, we used next-generation sequencing and mining of public human genome data sets to generate the largest and most diverse collection of circulating and integrated HHV-6 genomes studied to date. In genomes of geographically dispersed, only distantly related people, we identified clades of integrated viruses that originated from a single ancestral event, confirming this with fluorescent in situ hybridization to directly observe the integration locus. In contrast to HHV-6B, circulating and integrated HHV-6A sequences form distinct clades, arguing against ongoing integration of circulating HHV-6A or "reactivation" of integrated HHV-6A. Taken together, our study provides the first comprehensive picture of the evolution of HHV-6, and reveals that integration of heritable HHV-6 has occurred since the time of, if not before, human migrations out of Africa

Cardiomyopathies - The special entity of myocarditis and inflammatory cardiomyopathy

Cardiomyopathy is a heart muscle disease with structural and functional myocardial abnormalities in the absence of coronary artery disease, hypertension, valvular disease, and congenital heart disease. However, it has become clear that diverse etiologies and clinical manifestations (e.g. arrhythmogenic right-ventricular cardiomyopathy/dysplasia (ARVC/D), ARVD/C, left-ventricular non-compaction cardiomyopathy (LVNC)) are responsible for the clinical picture of dilated cardiomyopathy (DCM). The American Heart Association (AHA) classification grouped cardiomyopathies into genetic, mixed and acquired forms, while the European Society of Cardiology (ESC) classification proposed the subgrouping of each major type of cardiomyopathy into familial or genetic, and nonfamilial or nongenetic, forms [1-4]. Cardiomyopathies are clinically heterogeneous diseases, and there are differences in sex, age of onset, rate of progression, risk of development of overt heart failure and likelihood of sudden death within each cardiomyopathy subtype [5]. Because of the complex etiology and clinical presentation, the diagnostic spectrum in cardiomyopathies spans the entire range of non-invasive and invasive cardiological examination techniques including genetic analysis. The exact verification of certain cardiomyopathies necessitates additional investigations. So, histological, immunohistological and molecular biological/virological investigations of endomyocardial biopsies are the gold standard to confirm the diagnosis of an inflammatory cardiomyopathy (DCMi) [6-10]. This review focuses on myocarditis and inflammatory cardiomyopathies underlying an immune-mediated process or persistent viral infection

Telbivudine Reduces Parvovirus B19-Induced Apoptosis in Circulating Angiogenic Cells

Aims: Human parvovirus B19 (B19V) infection directly induces apoptosis and modulates CXCR4 expression of infected marrow-derived circulating angiogenic cells (CACs). This leads to dysfunctional endogenous vascular repair. Treatment for B19V-associated disease is restricted to symptomatic treatment. Telbivudine, a thymidine analogue, established in antiviral treatment for chronic hepatitis B, modulates pathways that might influence induction of apoptosis. Therefore, we tested the hypothesis of whether telbivudine influences B19V-induced apoptosis of CAC. Methods and Results: Pretreatment of two CAC-lines, early outgrowth endothelial progenitor cells (eo-EPC) and endothelial colony-forming cells (ECFC) with telbivudine before in vitro infection with B19V significantly reduced active caspase-3 protein expression (−39% and −40%, both p < 0.005). Expression of Baculoviral Inhibitor of apoptosis Repeat-Containing protein 3 (BIRC3) was significantly downregulated by in vitro B19V infection in ECFC measured by qRT-PCR. BIRC3 downregulation was abrogated with telbivudine pretreatment (p < 0.001). This was confirmed by single gene PCR (p = 0.017) and Western blot analysis. In contrast, the missing effect of B19V on angiogenic gene expression postulates a post-transcriptional modulation of CXCR4. Conclusions: We for the first time show a treatment approach to reduce B19V-induced apoptosis. Telbivudine reverses B19V-induced dysregulation of BIRC3, thus, intervening in the apoptosis pathway and protecting susceptible cells from cell death. This approach could lead to an effective B19V treatment to reduce B19V-related disease.Peer Reviewe