The immediate-early protein 1 of human herpesvirus 6B interacts with NBS1 and inhibits ATM signaling.

Viral infection often trigger an ATM serine/threonine kinase (ATM)-dependent DNA damage response in host cells that suppresses viral replication. Viruses evolved different strategies to counteract this antiviral surveillance system. Here, we report that human herpesvirus 6B (HHV-6B) infection causes genomic instability by suppressing ATM signaling in host cells. Expression of immediate-early protein 1 (IE1) phenocopies this phenotype and blocks homology-directed double-strand break repair. Mechanistically, IE1 interacts with NBS1, and inhibits ATM signaling through two distinct domains. HHV-6B seems to efficiently inhibit ATM signaling as further depletion of either NBS1 or ATM do not significantly boost viral replication in infected cells. Interestingly, viral integration of HHV-6B into the host's telomeres is not strictly dependent on NBS1, challenging current models where integration occurs through homology-directed repair. Given that spontaneous IE1 expression has been detected in cells of subjects with inherited chromosomally-integrated form of HHV-6B (iciHHV-6B), a condition associated with several health conditions, our results raise the possibility of a link between genomic instability and the development of iciHHV-6-associated diseases.

HHV-6B detection and host gene expression implicate HHV-6B as pulmonary pathogen after hematopoietic cell transplant.

Limited understanding of the immunopathogenesis of human herpesvirus 6B (HHV-6B) has prevented its acceptance as a pulmonary pathogen after hematopoietic cell transplant (HCT). In this prospective multicenter study of patients undergoing bronchoalveolar lavage (BAL) for pneumonia after allogeneic HCT, we test blood and BAL fluid (BALF) for HHV-6B DNA and mRNA transcripts associated with lytic infection and perform RNA-seq on paired blood. Among 116 participants, HHV-6B DNA is detected in 37% of BALs, 49% of which also have HHV-6B mRNA detection. We establish HHV-6B DNA viral load thresholds in BALF that are highly predictive of HHV-6B mRNA detection and associated with increased risk for overall mortality and death from respiratory failure. Participants with HHV-6B DNA in BALF exhibit distinct host gene expression signatures, notable for enriched interferon signaling pathways in participants clinically diagnosed with idiopathic pneumonia. These data implicate HHV-6B as a pulmonary pathogen after allogeneic HCT.

Detection of HHV-6 Virus in specimen of a ductal pancreatic adenocarcinoma with comparison in tumor and normal tissue.

AIMS: The association of human herpesvirus 6 (HHV-6) species with pancreatic cancer is controversially discussed. The aim of this study was to further investigate the postulated association and to identify the basis of HHV-6 DNA positivity reported for pancreatic cancer tissue. METHODS: All samples of patients with pancreatic cancer (cancer and surrounding tissue) were analyzed for presence of HHV-6 DNA by PCR and then selected cases by immunohistochemistry. RESULTS: Sixty eight per cent (68% = 52/77) of all patients were HHV-6 DNA positive in any of the samples, 49% (38/77) were positive in tumor tissue. Specimens of just one patient were HHV-6A DNA positive, all other patients were positive for HHV-6B. Immunohistochemical analysis of HHV-6 DNA positive samples did not reveal any specific HHV-6B protein positive tumor cell. In contrast, supposed immune cells presented intra- and peritumorally expressed HHV-6B-protein. The cause of presence of these cells in the tumor stroma is unknown, as of yet. CONCLUSIONS: HHV-6 DNA-positivity of pancreatic cancer tissue described by us and others is probably not due to the infection of pancreatic cells by HHV-6, but rather due to the migration of HHV-6 positive immune cells into the pancreas. Based on our data, we suppose that there is no direct evidence for HHV-6 as a causative agent of pancreatic cancer, but further in-depth studies (including investigation of immune status of patients) are necessary to make definitive conclusions.

Inherited IRAK-4 Deficiency in Acute Human Herpesvirus-6 Encephalitis.

Human herpesvirus-6 (HHV-6) infection can rarely cause life-threatening conditions, such as encephalitis, in otherwise healthy children, with unclear pathogenesis. We studied a child who presented with acute HHV-6 encephalitis at the age of 10 months and who was homozygous for a novel missense mutation in IRAK4, encoding interleukin-1 receptor-associated kinase 4, identified by whole-exome sequencing. We tested the damaging impact of this mutation in silico by molecular dynamics simulations and in vitro by biochemical and functional experiments utilizing cell lines and patient's cells. We found that the mutation is severely hypomorphic, impairing both the expression and function of IRAK-4. Patient's leukocytes had barely detectable levels of IRAK-4 and diminished anti-viral immune responses to various stimuli inducing different Toll-like receptors and cytosolic nucleic acid sensors. Overall, these findings suggest that acute HHV-6 encephalitis can result from inborn errors of immunity to virus. This study represents the first report of isolated acute HHV-6 infection causing encephalitis in an inherited primary immunodeficiency, notably autosomal recessive (AR) partial IRAK-4 deficiency, and the first report of AR IRAK-4 deficiency presenting with a severe viral disease, notably HHV-6 encephalitis upon an acute infection, thereby expanding the clinical spectrum of IRAK-4 deficiency.

Impact of Host Telomere Length on HHV-6 Integration.

Human herpesvirus 6A and 6B are two closely related viruses that infect almost all humans. In contrast to most herpesviruses, HHV-6A/B can integrate their genomes into the telomeres during the infection process. Both viruses can also integrate in germ cells and subsequently be inherited in children. How HHV-6A/B integrate into host telomeres and the consequences of this remain a subject of active research. Here, we developed a method to measure telomere length by quantitative fluorescence in situ hybridization, confocal microscopy, and computational processing. This method was validated using a panel of HeLa cells having short or long telomeres. These cell lines were infected with HHV-6A, revealing that the virus could efficiently integrate into telomeres independent of their length. Furthermore, we assessed the telomere lengths after HHV-6A integration and found that the virus-containing telomeres display a variety of lengths, suggesting that either telomere length is restored after integration or telomeres are not shortened by integration. Our results highlight new aspects of HHV-6A/B biology and the role of telomere length on virus integration.

MicroRNA expression profiling of cerebrospinal fluid/serum exosomes in children with human herpesvirus 6-associated encephalitis/encephalopathy by high-throughput sequencing.

Primary human herpesvirus 6 (HHV-6) infection is sometimes accompanied by acute encephalopathy with reduced subcortical diffusion (AED) in immunocompetent children. We investigated exosomal microRNA (miRNA) expression profiles in cerebrospinal fluid (CSF) and sera of patients with HHV-6-associated AED (n = 5) and febrile seizure (FS) (n = 5) using high-throughput sequencing. A total of 176 and 663 miRNAs were identified in CSF and serum exosomes, respectively. Comparative analysis determined that some miRNAs (miR-381-3p, miR-155) were exclusively expressed in the CSF exosomes of AED but not of FS patients, suggesting their potential application as novel diagnostic biomarkers for AED.

[The importance of telomeres in human herpesvirus-6A/B infections]. / L'importance des télomères dans les infections par les Herpèsvirus humains-6A/B.

Herpesviruses are undisputed masters of disguise. The ability to become invisible to the immune system effectors is a complex process resting on a variety of stealth approaches. Among these, human herpesviruses-6A and -6B (HHV-6A/B) have developed the unique ability to integrate their genome within the ends of chromosomes allowing viral persistence in the absence of viral protein expression. This aptitude, unique to HHV-6A/B among human herpesviruses, requires close interactions between the telomeric regions of chromosomes and the viral genome. In this review article, the biology of telomeres and the mechanisms responsible for viral integration are discussed. In closing, the possible biological consequences of HHV-6A/B integration into chromosomal DNA are discussed.

TITLE: L'importance des télomères dans les infections par les Herpèsvirus humains-6A/B. ABSTRACT: Les Herpèsvirus sont des maîtres incontestés du camouflage. En effet, ces virus utilisent divers moyens pour assurer leur persistance chez l'hôte infecté. Les Herpèsvirus humains-6A et -6B (HHV-6A/B) ont ainsi développé une approche unique, en intégrant l'ensemble de leur génome au sein des extrémités des chromosomes des cellules qu'ils infectent. Cette capacité, propre aux HHV-6A/B parmi les Herpèsvirus humains, requiert des interactions étroites entre les régions télomériques des chromosomes de l'hôte et le génome viral. Dans cette revue, la biologie des télomères et les mécanismes responsables de l'intégration virale seront abordés et les conséquences biologiques de l'intégration des HHV-6A/B au sein de l'ADN chromosomique seront discutées.

Modulation of microRNome by Human Cytomegalovirus and Human Herpesvirus 6 Infection in Human Dermal Fibroblasts: Possible Significance in the Induction of Fibrosis in Systemic Sclerosis.

Human cytomegalovirus (HCMV) and Human herpesvirus 6 (HHV-6) have been reportedly suggested as triggers of the onset and/or progression of systemic sclerosis (SSc), a severe autoimmune disorder characterized by multi-organ fibrosis. The etiology and pathogenesis of SSc are still largely unknown but virological and immunological observations support a role for these beta-herpesviruses, and we recently observed a direct impact of HCMV and HHV-6 infection on the expression of cell factors associated with fibrosis at the cell level. Since miRNA expression has been found profoundly deregulated at the tissue level, here we aimed to investigate the impact on cell microRNome (miRNome) of HCMV and HHV-6 infection in in vitro infected primary human dermal fibroblasts, which represent one of the main SSc target cells. The analysis, performed by Taqman arrays detecting and quantifying 754 microRNAs (miRNAs), showed that both herpesviruses significantly modulated miRNA expression in infected cells, with evident early and late effects and deep modulation (>10 fold) of >40 miRNAs at each time post infection, including those previously recognized for their key function in fibrosis. The correlation between these in vitro results with in vivo observations is strongly suggestive of a role of HCMV and/or HHV-6 in the multistep pathogenesis of fibrosis in SSc and in the induction of fibrosis-signaling pathways finally leading to tissue fibrosis. The identification of specific miRNAs may open the way to their use as biomarkers for SSc diagnosis, assessment of disease progression and possible antifibrotic therapies.

Severe herpes virus 6 interstitial pneumonia in an infant with three variants in genes predisposing to lung disease.

Infections due to human herpesvirus 6 (HHV-6) are frequent during early childhood. Usually, they have a favorable clinical course. Conversely, HHV-6 congenital infections occur in about 1% of neonates and may present with more severe clinical pictures. HHV-6 can be found in lung tissues and bronchoalveolar lavage (BAL) samples from patients with pneumonia and in immunocompromised patients can cause mild to severe pneumonia. In neonates, the role of HHV-6 in the genesis of severe pneumonia is poorly defined still now. We describe a healthy infant with a late-onset (15 days of life) severe interstitial pneumonia and heavy HHV-6 genome load, persistently detected in its BAL fluid. The baby underwent high-frequency oscillatory ventilation, hydroxychloroquine, steroids, and ganciclovir for 6 weeks and at 9 months she died. Next-generation sequencing of genes known to cause neonatal respiratory insufficiency revealed the presence of a "probably pathogenetic" heterozygous variant in the autosomal recessive DRC1 gene, a heterozygous variant of unknown significance (VUS) in the autosomal recessive RSPH9 gene, and a heterozygous VUS in the autosomal recessive MUC5B gene. HHV-6 infection should be considered in the differential diagnosis of late-onset severe respiratory distress in neonates and the co-occurrence of genetic predisposing factors or modifiers should be tested by specific molecular techniques. The intensity of HHV-6 genome load in BAL fluid could be an indicator of the response to antiviral therapy.

The cross-talk between STAT1/STAT3 and ROS up-regulates PD-L1 and promotes the release of pro-inflammatory/immune suppressive cytokines in primary monocytes infected by HHV-6B.

Programmed death ligand 1 (PD-L1) up-regulation on antigen presenting cells induces T cell dysfunction, strongly impairing immune response. Human Herpesviruses (HHV) 6B is a ß-herpesvirus that, although displays a higher tropism for T cells, can infect other immune cells including monocytes and dendritic cells (DCs) and neuronal cells. We have previously shown that HHV-6B infection of primary monocytes reduced autophagy and induced Endoplasmic Reticulum (ER) stress/ Unfolded Protein Response (UPR), impairing their survival and differentiation into DCs. In this study, we found that PD-L1 expression was up-regulated by HHV-6B on the surface of infected monocytes and that its extracellular release also increased, effects known to lead to an impairment of anti-viral immune response. At molecular level, PD-L1 up-regulation correlated with the activation of a positive regulatory circuit between the increase of intracellular ROS and the activation of STAT1 and STAT3 induced by HHV-6B, accompanied by a high release of pro-inflammatory/immune suppressive cytokines. In conclusion, this study unveils new strategies put in place by HHV-6B to induce immune dysfunction and the underlying molecular pathways that could be targeted to counteract such immune suppressive effects.

The Promyelocytic Leukemia Protein facilitates human herpesvirus 6B chromosomal integration, immediate-early 1 protein multiSUMOylation and its localization at telomeres.

Human herpesvirus 6B (HHV-6B) is a betaherpesvirus capable of integrating its genome into the telomeres of host chromosomes. Until now, the cellular and/or viral proteins facilitating HHV-6B integration have remained elusive. Here we show that a cellular protein, the promyelocytic leukemia protein (PML) that forms nuclear bodies (PML-NBs), associates with the HHV-6B immediate early 1 (IE1) protein at telomeres. We report enhanced levels of SUMOylated IE1 in the presence of PML and have identified a putative SUMO Interacting Motif (SIM) within IE1, essential for its nuclear distribution, overall SUMOylation and association with PML to nuclear bodies. Furthermore, using PML knockout cell lines we made the original observation that PML is required for efficient HHV-6B integration into host chromosomes. Taken together, we could demonstrate that PML-NBs are important for IE1 multiSUMOylation and that PML plays an important role in HHV-6B integration into chromosomes, a strategy developed by this virus to maintain its genome in its host over long periods of time.

Role for the shelterin protein TRF2 in human herpesvirus 6A/B chromosomal integration.

Human herpesviruses 6A and 6B (HHV-6A/B) are unique among human herpesviruses in their ability to integrate their genome into host chromosomes. Viral integration occurs at the ends of chromosomes within the host telomeres. The ends of the HHV-6A/B genomes contain telomeric repeats that facilitate the integration process. Here, we report that productive infections are associated with a massive increase in telomeric sequences of viral origin. The majority of the viral telomeric signals can be detected within viral replication compartments (VRC) that contain the viral DNA processivity factor p41 and the viral immediate-early 2 (IE2) protein. Components of the shelterin protein complex present at telomeres, including TRF1 and TRF2 are also recruited to VRC during infection. Biochemical, immunofluorescence coupled with in situ hybridization and chromatin immunoprecipitation demonstrated the binding of TRF2 to the HHV-6A/B telomeric repeats. In addition, approximately 60% of the viral IE2 protein localize at cellular telomeres during infection. Transient knockdown of TRF2 resulted in greatly reduced (13%) localization of IE2 at cellular telomeres (p<0.0001). Lastly, TRF2 knockdown reduced HHV-6A/B integration frequency (p<0.05), while no effect was observed on the infection efficiency. Overall, our study identified that HHV-6A/B IE2 localizes to telomeres during infection and highlight the role of TRF2 in HHV-6A/B infection and chromosomal integration.

Inherited chromosomally integrated HHV-6 possibly modulates human gene expression.

Approximately, 1% of human population possesses a copy of human herpesvirus 6A and 6B (HHV-6A/B) in the genome. This viral element is referred to as inherited chromosomally integrated HHV-6A/B (iciHHV-6A/B) and is encoded in all of their cells. A recent study revealed that iciHHV-6A/B potentially increases the immune responses against HHV-6. However, it remains unclear whether iciHHV-6A/B affects human gene expression. Here, we perform global transcriptome analysis using the datasets obtained from various human tissues. We detected two and four individuals positive for iciHHV-6A and iciHHV-6B, respectively, and revealed that the transcriptional expression of iciHHV-6A/B was sporadic in the human body. Transcriptome analysis identified the human genes differentially expressed between iciHHV-6A/B-positive and -negative individuals. Particularly, the expression of some genes encoding immunoglobulins decreased in sigmoid colon of iciHHV-6A/B-positive individuals. Our findings suggest that iciHHV-6A/B may be associated with human health maintenance.

Detection of human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7) DNA in endocervical samples from a positive and negative HPV woman of Córdoba, Argentina.

AIMS: The purpose of the present study was to elucidate the presence of human herpesvirus 6A (HHV-6A), HHV-6B and HHV-7 in samples of the uterine cervix through detection of viral DNA. We analysed normal tissues, samples with low-grade squamous intraepithelial lesions (LSILs) and high-grade squamous intraepithelial lesions (HSILs). We correlated the presence of HHV-6 and HHV-7 with the finding of human papillomavirus (HPV) in mucosal samples. METHODS: Cervical samples were examined and grouped as follows: group 1 (n=29), normal cytology; group 2 (n=61), samples with LSIL; group 3 (n=35), samples with HSIL. Molecular biology examinations were performed in all samples to detect HHV-6, HHV-7 and HPV DNA and to typify HHV-6 species. RESULTS: Group 1: normal cytology and HPV (-): HHV-6: 6.8% (2/29), HHV-7: 79.3% (23/29); group 2: LSIL and HPV (-): HHV-6: 93.1% (27/29), HHV-7: 96.5% (28/29); LSIL and HPV (+): HHV-6: 0% (0/32), HHV-7: 90.6% (29/32); group 3: HSIL and HPV (-): HHV-6: 20% (2/10), HHV-7: 70% (7/10); HSIL HPV (+): HHV-6: 12% (3/25), HHV-7: 68% (17/25). HHV-6A DNA was not detected in any samples. CONCLUSIONS: (1) Both HHV-6 and HHV-7 infect the mucosal cells of the cervix with higher prevalence of HHV-7. (2) The higher prevalence of HHV-6 in LSIL HPV (-) samples compared with those with normal cytology indicates that it constitutes a possible risk factor for atypia production. (3) The presence of HHV-7 in all samples questions its role in the production of atypia. (4) The finding of HHV-6 and HHV-7 suggests that the cervical mucosa is a possible transmission pathway for these viruses.

Inherited Chromosomally Integrated Human Herpesvirus 6 Demonstrates Tissue-Specific RNA Expression In Vivo That Correlates with an Increased Antibody Immune Response.

Human herpesviruses 6A and 6B (HHV-6A and HHV-6B) are human viruses capable of chromosomal integration. Approximately 1% of the human population carries one copy of HHV-6A/B integrated into every cell in their body, referred to as inherited chromosomally integrated human herpesvirus 6A/B (iciHHV-6A/B). Whether iciHHV-6A/B is transcriptionally active in vivo and how it shapes the immunological response are still unclear. In this study, we screened DNA sequencing (DNA-seq) and transcriptome sequencing (RNA-seq) data for 650 individuals available through the Genotype-Tissue Expression (GTEx) project and identified 2 iciHHV-6A- and 4 iciHHV-6B-positive candidates. When corresponding tissue-specific gene expression signatures were analyzed, low levels HHV-6A/B gene expression was found across multiple tissues, with the highest levels of gene expression in the brain (specifically for HHV-6A), testis, esophagus, and adrenal gland. U90 and U100 were the most highly expressed HHV-6 genes in both iciHHV-6A- and iciHHV-6B-positive individuals. To assess whether tissue-specific gene expression from iciHHV-6A/B influences the immune response, a cohort of 15,498 subjects was screened and 85 iciHHV-6A/B+ subjects were identified. Plasma samples from iciHHV-6A/B+ and age- and sex-matched controls were analyzed for antibodies to control antigens (cytomegalovirus [CMV], Epstein-Barr virus [EBV], and influenza virus [FLU]) or HHV-6A/B antigens. Our results indicate that iciHHV-6A/B+ subjects have significantly more antibodies against the U90 gene product (IE1) than do non-iciHHV-6-positive individuals. Antibody responses against EBV and FLU antigens or HHV-6A/B gene products either not expressed or expressed at low levels, such as U47, U57, and U72, were identical between controls and iciHHV-6A/B+ subjects. CMV-seropositive individuals with iciHHV-6A/B+ have more antibodies against CMV pp150 than do CMV-seropositive controls. These results argue that spontaneous gene expression from integrated HHV-6A/B leads to an increase in antigenic burden that translates into a more robust HHV-6A/B-specific antibody response.IMPORTANCE HHV-6A and -6B are human herpesviruses that have the unique property of being able to integrate into the telomeric regions of human chromosomes. Approximately 1% of the world's population carries integrated HHV-6A/B genome in every cell of their body. Whether viral genes are transcriptionally active in these individuals is unclear. By taking advantage of a unique tissue-specific gene expression data set, we showed that the majority of tissues from iciHHV-6 individuals do not show HHV-6 gene expression. Brain and testes showed the highest tissue-specific expression of HHV-6 genes in two separate data sets. Two HHV-6 genes, U90 (immediate early 1 protein) and U100 (glycoproteins Q1 and Q2), were found to be selectively and consistently expressed across several human tissues. Expression of U90 translates into an increase in antigen-specific antibody response in iciHHV-6A/B+ subjects relative to controls. Future studies will be needed to determine the mechanism of gene expression, the effects of these genes on human gene transcription networks, and the pathophysiological impact of having increased viral protein expression in tissue in conjunction with increased antigen-specific antibody production.

Genome-Wide Approach to the CD4 T-Cell Response to Human Herpesvirus 6B.

Human herpesvirus 6 (HHV-6) and cytomegalovirus (CMV) are population-prevalent betaherpesviruses with intermittent lytic replication that can be pathogenic in immunocompromised hosts. Elucidation of the adaptive immune response is valuable for understanding pathogenesis and designing novel treatments. Knowledge of T-cell antigens has reached the genome-wide level for CMV and other human herpesviruses, but study of HHV-6 is at an earlier stage. Using rare-cell enrichment combined with an HLA-agnostic, proteome-wide approach, we queried HHV-6B-specific CD4 T cells from 18 healthy donors with each known HHV-6B protein. We detected a low abundance of HHV-6-specific CD4 T cells in blood; however, the within-person CD4 T-cell response is quite broad: the median number of open reading frame (ORF) products recognized was nine per person. Overall, the data expand the number of documented HHV-6B CD4 T-cell antigens from approximately 11 to 60. Epitopes in the proteins encoded by U14, U90, and U95 were mapped with synthetic peptides, and HLA restriction was defined for some responses. Intriguingly, CD4 T-cell antigens newly described in this report are among the most population prevalent, including U73, U72, U95, and U30. Our results indicate that selection of HHV-6B ORFs for immunotherapy should consider this expanded panel of HHV-6B antigens.IMPORTANCE Human herpesvirus 6 is highly prevalent and maintains chronic infection in immunocompetent individuals, with the potential to replicate widely in settings of immunosuppression, leading to clinical disease. Antiviral compounds may be ineffective and/or pose dose-limiting toxicity, and therefore, immune-based therapies have garnered increased interest in recent years. Attempts at addressing this unmet medical need begin with understanding the cellular response to HHV-6 at the individual and population levels. The present study provides a comprehensive assessment of HHV-6-specific T-cell responses that may inform the development of cell-based therapies directed at this virus.

RNA Sequencing of the In Vivo Human Herpesvirus 6B Transcriptome To Identify Targets for Clinical Assays Distinguishing between Latent and Active Infections.

Human herpesvirus 6B (HHV-6B) DNA is frequently detected in human samples. Diagnostic assays distinguishing HHV-6B reactivation from latency are limited. This has impaired strategies to diagnose and treat HHV-6B-associated diseases. We used RNA sequencing to characterize and compare the HHV-6B transcriptome in multiple sample types, including (i) whole blood from hematopoietic cell transplant (HCT) recipients with and without HHV-6B plasma viremia, (ii) tumor tissue samples from subjects with large B cell lymphoma infected with HHV-6B, (iii) lymphoblastoid cell lines (LCLs) from subjects with inherited chromosomally integrated HHV-6B or latent infection with HHV-6B, and (iv) HHV-6B Z29 infected SupT1 CD4+ T cells. We demonstrated substantial overlap in the HHV-6B transcriptome observed in in vivo and in vitro samples, although there was variability in the breadth and quantity of gene expression across samples. The HHV-6B viral polymerase gene U38 was the only HHV-6B transcript detected in all next-generation RNA sequencing (RNA-seq) data sets and was one of the most highly expressed genes. We developed a novel reverse transcription-PCR assay targeting HHV-6B U38, which identified U38 mRNA in all tested whole-blood samples from patients with concurrent HHV-6B viremia. No HHV-6B U38 transcripts were detected by RNA-seq or reverse transcription-real-time quantitative PCR (RT-qPCR) in whole-blood samples from subjects without HHV-6B plasma detection or from latently infected LCLs. A RT-qPCR assay for HHV-6B U38 may be useful to identify lytic HHV-6B infection in nonplasma samples and samples from individuals with inherited chromosomally integrated HHV-6B. This study also demonstrates the feasibility of transcriptomic analyses for HCT recipients.IMPORTANCE Human herpesvirus 6B (HHV-6B) is a DNA virus that infects most children within the first few years of life. After primary infection, HHV-6B persists as a chronic, latent infection in many cell types. Additionally, HHV-6B can integrate into germ line chromosomes, resulting in individuals with viral DNA in every nucleated cell. Given that PCR to detect viral DNA is the mainstay for diagnosing HHV-6B infection, the characteristics of HHV-6B infection complicate efforts to distinguish between latent and active viral infection, particularly in immunocompromised patients who have frequent HHV-6B reactivation. In this study, we used RNA sequencing to characterize the HHV-6B gene expression profile in multiple sample types, and our findings identified evidence-based targets for diagnostic tests that distinguish between latent and active viral infection.

Induction of Proinflammatory Multiple Sclerosis-Associated Retrovirus Envelope Protein by Human Herpesvirus-6A and CD46 Receptor Engagement.

The aberrant expression of human endogenous retrovirus (HERV) elements of the HERV-W family has been associated with different diseases, including multiple sclerosis (MS). In particular, the expression of the envelope protein (Env) from the multiple sclerosis-associated retrovirus (MSRV), a member of HERV-W family and known for its potent proinflammatory activity, is repeatedly detected in the brain lesions and blood of MS patients. Furthermore, human herpesvirus 6 (HHV-6) infection has long been suspected to play a role in the pathogenesis of MS and neuroinflammation. We show here that both HHV-6A and stimulation of its receptor, transmembrane glycoprotein CD46, induce the expression of MSRV-Env. The engagement of extracellular domains SCR3 and SCR4 of CD46-Cyt1 isoform was required for MSRV-env transactivation, limiting thus the MSRV-Env induction to the CD46 ligands binding these domains, including C3b component of complement, specific monoclonal antibodies, and both infectious and UV-inactivated HHV-6A, but neither HHV-6B nor measles virus vaccine strain. Induction of MSRV-Env required CD46 Cyt-1 singling and was abolished by the inhibitors of protein kinase C. Finally, both membrane-expressed and secreted MSRV-Env trigger TLR4 signaling, displaying thus a proinflammatory potential, characteristic for this viral protein. These data expand the specter of HHV-6A effects in the modulation of the immune response and support the hypothesis that cross-talks between exogenous and endogenous viruses may contribute to inflammatory diseases and participate in neuroinflammation. Furthermore, they reveal a new function of CD46, known as an inhibitor of complement activation and receptor for several pathogens, in transactivation of HERV env genes, which may play an important role in the pathogenesis of inflammatory diseases.

Comparison of HHV-6 DNA detection in plasma and whole blood in allogeneic hematopoietic stem cell transplant recipients: frequent false-positive results for active HHV-6 infection using whole blood samples.

In this prospective observational study, we compared the human herpesvirus-6 (HHV-6) DNA load in serially collected paired plasma and whole blood (WB) samples from allogeneic hematopoietic stem cell transplantation (HSCT) recipients. A total of 721 paired samples were collected from 68 recipients. The positive rate for HHV-6 DNA was 9.7 and 35.0% in plasma and WB samples, respectively (P < 0.001). The correlation of HHV-6 DNA load between plasma and WB was poor (R2 = 0.250). After reaching peak levels, HHV-6 DNA showed a delayed decrease in WB in comparison with plasma (median, 28 versus 7 days, P < 0.001). We additionally tested HHV-6 mRNA status in 95 samples from eight patients. To identify positive HHV-6 mRNA, plasma HHV-6 DNA showed 55.0% sensitivity and 100% specificity, whereas WB HHV-6 DNA showed 90.0% sensitivity and 68.0% specificity. The false-positive rate for identifying positive HHV-6 mRNA was 0% for plasma HHV-6 DNA and 32.0% for WB HHV-6 DNA. Although WB was more sensitive than plasma for detecting HHV-6 reactivation, the rates of false positivity for active HHV-6 infection were higher for WB than for plasma.