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AMP-activated protein kinase (AMPK) is a cellular energy sensor regulating metabolic homeostasis. In this study, we investigated the role of AMPK in response to human herpesvirus 6A (HHV-6A) infection. We show that HHV-6A infection significantly downregulates the active phosphorylated state of AMPK in infected T cells. Pharmacological activation of AMPK highly attenuated HHV-6A propagation. Mechanistically, we found that the activation of AMPK by AICAR blocked HHV-6-induced glycolysis by inhibiting glucose metabolism and lactate secretion, as well as decreasing expressions of key glucose transporters and glycolytic enzymes. In addition, mTOR signaling has been inactivated in HHV-6A infected T cells by AICAR treatment. We also showed that HHV-6A infection of human umbilical cord blood mononuclear cells (CBMCs) reduced AMPK activity whereas the activation of AMPK by metformin drastically reduced HHV-6A DNA replication and virions production. Taken together, this study demonstrates that AMPK is a promising antiviral therapeutic target against HHV-6A infection., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background and Aims: An increase in the number of cases of acute hepatitis of unknown origin (HUO) in children was observed in 2021. Adenovirus and adeno-associated virus 2 (AAV2) infections have been suggested as possible triggers. However, the potential etiology is still unclear. We aimed to characterize a cohort of children with HUO in Israel in view of the COVID-19 pandemic., Method: Demographics, clinical data, and laboratory results on the children compatible with the CDC criteria for HUO were collected by the established registry of the Ministry of Health. Available specimens were sent to the Central Virology Laboratory., Results: A total of 39 children were included in the registry. A total of 20 were enrolled prospectively, in which human herpes virus 6 (HHV6) infection or reactivation was identified in 11/19, adenovirus was found in 4/19 of the cases, and AAV2 was detected in 2/16. Past COVID-19 exposure was recorded for 24/39 of the children. A total of 10 children underwent liver biopsy, and 8 were successfully treated with steroids and 2 underwent liver transplantation., Conclusions: The COVID-19 pandemic and the related containment measures combined with reactivation or active infection with other viruses could have been a trigger for the HUO outbreak. In our cohort, HHV6 was the most abundant finding.

HHV-6B reactivation affects approximately half of all allogeneic hematopoietic cell transplant (HCT) recipients. HHV-6B is the most frequent infectious cause of encephalitis following HCT and is associated with pleiotropic manifestations in this setting, including graft-versus-host disease, myelosuppression, pneumonitis, and CMV reactivation, although the causal link is not always clear. When the virus inserts its genome in chromosomes of germ cells, the chromosomally integrated form (ciHHV6) is inherited by offspring. The condition of ciHHV6 is characterized by the persistent detection of HHV-6 DNA, often confounding diagnosis of reactivation and disease-this has also been associated with adverse outcomes. Recent changes in clinical practice in the field of cellular therapies, including a wider use of post-HCT cyclophosphamide, the advent of letermovir for CMV prophylaxis, and the rapid expansion of novel cellular therapies require contemporary epidemiological studies to determine the pathogenic role and spectrum of disease of HHV-6B in the current era. Research into the epidemiology and clinical significance of HHV-6B in chimeric antigen receptor T cell (CAR-T cell) therapy recipients is in its infancy. No controlled trials have determined the optimal treatment for HHV-6B. Treatment is reserved for end-organ disease, and the choice of antiviral agent is influenced by expected toxicities. Virus-specific T cells may provide a novel, less toxic therapeutic modality but is more logistically challenging. Preventive strategies are hindered by the high toxicity of current antivirals. Ongoing study is needed to keep up with the evolving epidemiology and impact of HHV-6 in diverse and expanding immunocompromised patient populations.

Background: Drug Reaction with Eosinophilia and Systemic Symptoms (DRESS) is a severe adverse event (mortality of 10%). Its pathophysiology involves herpesviruses, particularly HHV-6, but the exact mechanisms are still poorly understood., Objective: To describe severe cases of DRESS and especially their association with herpesvirus reactivation., Methods: This study was a multicentre case series conducted between 2007 and 2021 at five University Hospital Centres in France. The study included patients who had severe DRESS, which was defined as death, transfer to the intensive care unit (ICU), or severe damage to internal organs. We excluded patients without blood PCR sample, without a drug formally attributed or with RegiSCAR score < 6. We collected data on severity, causative drug, associated visceral damage and results of viral blood PCRs. HHV-6 reactivation was studied in skin biopsies by detection of small non-coding transcripts (HHV-6 miR-aU14) and a late viral protein (GP82/105)., Results: Fifty-two patients were included (29 female, median age 62, interquartile range (IQR) [37;72]). Eight patients (15%) died, 13 (27%) were admitted to ICU. Most patients (n = 34; 65%) had multisystem involvement: most frequent was liver (n = 46; 88%), then renal failure (n = 24; 46%). Forty patients (77%) had at least one blood viral reactivation among HHV-6, EBV or CMV, of which 21 (53%) had at least two. Median time of blood HHV-6 reactivation was 24 days (IQR [20;35]). HHV-6 reactivation was demonstrated in 15 out of 20 skin biopsies, with a median time of 11 days [9;17]., Conclusions: We confirmed the high rate of HHV-6 reactivation in severe DRESS and demonstrated cutaneous HHV-6 reactivation using small non-coding transcripts (HHV-6 miR-aU14), which preceded viral PCR positivity in blood. These results suggest that HHV-6 reactivation during DRESS may start in skin. Furthermore, search for miR-aU14 in skin biopsy could become a useful diagnostic tool for early detection of HHV-6 reactivation., (© 2023 European Academy of Dermatology and Venereology.)

Cell therapies have yielded durable clinical benefits for patients with cancer, but the risks associated with the development of therapies from manipulated human cells are understudied. For example, we lack a comprehensive understanding of the mechanisms of toxicities observed in patients receiving T cell therapies, including recent reports of encephalitis caused by reactivation of human herpesvirus 6 (HHV-6) 1 . Here, through petabase-scale viral genomics mining, we examine the landscape of human latent viral reactivation and demonstrate that HHV-6B can become reactivated in cultures of human CD4 + T cells. Using single-cell sequencing, we identify a rare population of HHV-6 'super-expressors' (about 1 in 300-10,000 cells) that possess high viral transcriptional activity, among research-grade allogeneic chimeric antigen receptor (CAR) T cells. By analysing single-cell sequencing data from patients receiving cell therapy products that are approved by the US Food and Drug Administration 2 or are in clinical studies 3-5 , we identify the presence of HHV-6-super-expressor CAR T cells in patients in vivo. Together, the findings of our study demonstrate the utility of comprehensive genomics analyses in implicating cell therapy products as a potential source contributing to the lytic HHV-6 infection that has been reported in clinical trials 1,6-8 and may influence the design and production of autologous and allogeneic cell therapies., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Human herpesvirus 6 (HHV-6) can reactivate after allogeneic hematopoietic stem cell transplant (allo-HSCT) and may lead to severe symptoms. HHV-6-specific immune responses after HSCT are largely unexplored. We conducted a prospective observational study on 208 consecutive adult patients who received allo-HSCT to investigate HHV-6 reactivations and specific immune responses. Interferon gamma-producing HHV-6-specific T cells were quantified using enzyme-linked immunospot assay (ELISpot). HHV-6 reactivation occurred in 63% of patients, at a median of 25 days from allo-HSCT. Only 40% of these presented a clinically relevant infection, defined by the presence of classical HHV-6 end-organ diseases (EODs), based on European Conference on Infections in Leukaemia (ECIL) guidelines, and other possible HHV6-related EODs. Using multivariate analysis, we identified risk factors for HHV-6 reactivation: previous allo-HSCT, posttransplant cyclophosphamide (PT-Cy), and time-dependent steroids introduction. The use of PT-Cy and steroids were associated with clinically relevant infections, whereas higher CD3+ cell counts seemed to be protective. Interestingly, circulating HHV-6-specific T cells were significantly higher in patients with reactivated virus. Moreover, HHV-6-specific T-cell responses, quantified at >4 days after the first viremia detection, predicted clinically relevant infections (P < .0001), with higher specificity (93%) and sensitivity (79%) than polyclonal CD3+ cells per μL. Overall survival and transplant-related mortality were not affected by time-dependent HHV-6 reactivation, whereas a significant association was observed between clinically relevant infections and acute graft-versus-host disease. These results shed light on the role of HHV-6 in allo-HSCT and may affect HHV-6 monitoring and treatment., (© 2023 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Introduction: Human herpesvirus-6 (HHV-6) is a ubiquitous lymphotropic betaherpesvirus that can reactivate in drug rash with eosinophilia and systemic symptoms (DRESS). Despite recent publications advancing our understanding of HHV-6 in DRESS, the exact role of HHV-6 in disease pathogenesis remains unclear., Methods: A scoping review with the PubMed query "(HHV 6 AND (drug OR DRESS OR DIHS)) OR (HHV6 AND (drug OR DRESS OR DIHS))" was conducted in accordance with PRISMA guidelines. Articles containing original data on at least one DRESS patient with HHV-6 testing were included., Results: Our search returned a total of 373 publications, of which 89 met eligibility criteria. HHV-6 reactivation occurred in 63% of DRESS patients (n = 748), which was significantly more often than other herpesviruses. HHV-6 reactivation was associated with worse outcomes and greater severity in controlled studies. Case reports have demonstrated sometimes fatal HHV-6-related multi-organ involvement. Temporally, HHV-6 reactivation typically occurs 2 to 4 weeks after DRESS onset and has been linked to markers of immunologic signaling, such as OX40 (CD134), an HHV-6 entry receptor. Efficacy of antiviral or immunoglobulin treatment has only been demonstrated anecdotally, and steroid use may affect HHV-6 reactivation., Conclusion: HHV-6 is implicated in DRESS more than in any other dermatologic condition. It is still unclear whether HHV-6 reactivation is cause or consequence of DRESS dysregulation. Similar pathogenic mechanisms precipitated by HHV-6 in other contexts may be relevant in DRESS. Future randomized controlled studies to assess effects of viral suppression on clinical outcomes is needed.

Human herpesvirus 6 (HHV-6) reactivation is common after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and is associated with higher mortality and increased transplantation-related complications. We hypothesized that preemptive treatment with a short course of foscarnet at a lower cutpoint of plasma HHV-6 viral load would be effective in treating early HHV-6 reactivation, preventing complications and precluding hospitalization of these patients. We reviewed outcomes of adult patients (age ≥18 years) who received preemptive treatment with once-daily foscarnet 60 to 90 mg/kg for 7 days for HHV-6 reactivation after allo-HSCT at our institution between May 2020 and November 2022. Plasma HHV-6 viral load was monitored by quantitative PCR twice monthly in the first 100 days post-transplantation and twice weekly after reactivation until resolution. Eleven patients with a median age of 46 years (range, 23 to 73 years) were included in the analysis. HSCT was performed with a haploidentical donor in 10 patients and with an HLA-matched related donor in 1 patient. The most common diagnosis was acute leukemia (9 patients). Myeloablative and reduced-intensity conditioning regimens were used in 4 and 7 patients, respectively. Ten of the 11 patients received post-transplantation cyclophosphamide-based graft-versus-host disease prophylaxis. The median follow-up was 440 days (range, 174 to 831 days), and the median time to HHV-6 reactivation was 22 days post-transplantation (range, 15 to 89 days). The median viral load at first reactivation was 3,100 copies/mL (range, 210 to 118,000 copies/mL), and the median peak viral load was 11,300 copies/mL (range, 600 to 983,000 copies/mL). All patients received a short course of foscarnet at either 90 mg/kg/day (n = 7) or 60 mg/kg/day (n = 4). In all patients, plasma HHV-6 DNA was undetectable at completion of 1 week of treatment. No HHV-6 encephalitis or pneumonitis occurred. All patients achieved neutrophil and platelet engraftment after a median of 16 days (range, 8 to 22 days) and 26 days (range, 14 to 168 days), respectively, with no secondary graft failure. No complications related to foscarnet administration were noted. One patient with very high HHV-6 viremia had recurrent reactivation and received a second course of foscarnet as an outpatient. A short course of once-daily foscarnet is effective in treating early HHV-6 reactivation post-transplantation and may reduce the incidence of HHV-6-related and treatment-related complications and preclude hospitalization in these patients., (Copyright © 2023 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

Background: Immunocompromised patients face reactivation of latent viruses that increase the risk of morbidity., Aim: The study aimed to detect human herpes virus 6 (HHV-6) reactivation among allogeneic (allo) and autologous (auto) hematopoietic stem cell transplant (HSCT) recipients and to correlate potentially attributed clinical manifestations to HHV-6 DNA plasma level., Methods: A prospective study included all (forty) patients undergoing allo and auto-HSCT from Jan 2020 till June 2022. Plasma samples were collected for HHV-6 serology, and for HHV-6 quantitative PCR at post-transplantation weeks 2, 4, 6. Demographic and clinical data were recorded., Results: Out of 40 peripheral blood stem cell transplant (PBSCT) recipients, 34 (85%) were HHV-6 IgG positive pre-HSCT. Of which, fourteen patients (14/34, 41.2%) showed positive HHV-6 DNaemia. HHV-6 DNAemia (15/40, 37.5%) was significantly higher among allo (8/12, 66.7%) versus auto (7/28, 25%) HSCT recipients (p = 0.030). Patients with HHV-6 DNAemia developed fever, delayed engraftment and bone marrow suppression in 6/15, 40%, thrombocytopenia (5/15, 33.3%), rash and pneumonitis (2/15, 13.3%), acute GVHD (aGVHD) (1/15, 6.7%). HHV-6 DNAemia ranged from 101 to 102,000 copies/mL. Univariate analysis identified conditioning with busulfan-cyclophosphamide as a significant risk (p = 0.043), while receiving BEAM protocol was a protective factor (p = 0.045). In multivariate analysis, receiving BEAM protocol retained significance (p = 0.040)., Conclusion: Frequent HHV-6 reactivation was detected after HSCT, especially in allo-HSCT recipients with clinical manifestations which could not be otherwise explained. To our best knowledge this is the first study of HHV6 reactivation in HSCT recipients from Egypt. Raising awareness for HHV-6 reactivation manifestations and screening in HSCT recipients could be lifesaving., (© 2023. The Author(s).)

A number of studies have suggested that human herpesvirus 6A (HHV-6A) may play a role in multiple sclerosis (MS). Three possible hypotheses have been investigated: (1) U24 from HHV-6A (U24-6A) mimics myelin basic protein (MBP) through analogous phosphorylation and interaction with Fyn-SH3; (2) U24-6A affects endocytic recycling by binding human neural precursor cell (NPC) expressed developmentally down-regulated protein 4-like WW3* domain (hNedd4L-WW3*); and (3) MS patients who express Killer Cell Immunoglobulin Like Receptor 2DL2 (KIR2DL2) on natural killer (NK) cells are more susceptible to HHV-6 infection. In this contribution, we examined the validity of these propositions by investigating the interactions of U24 from HHV-6B (U24-6B), a variant less commonly linked to MS, with Fyn-SH3 and hNedd4L-WW3* using heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) titrations and isothermal titration calorimetry (ITC). In addition, the importance of phosphorylation and the specific role of U24 in NK cell activation in MS patients were examined. Overall, the findings allowed us to shed light into the models linking HHV-6 to MS and the involvement of U24.

The stimulus-induced cAMP response element (CRE)-binding protein (CREB) family of transcription factors bind to CREs to regulate diverse cellular responses, including proliferation, survival, and differentiation. Human herpesvirus 6A (HHV-6A), which belongs to the Betaherpesvirinae subfamily, is a lymphotropic herpesvirus frequently found in patients with neuroinflammatory diseases. Previous reports implicated the importance of CREs in the HHV-6A life cycle, although the effects of the binding of transcription factors to CREs in viral replication have not been fully elucidated. In this study, we analyzed the role of the CREB family of transcription factors during HHV-6A replication. We found that HHV-6A infection enhanced phosphorylation of the CREB family members CREB1 and activating transcription factor 1 (ATF1). Knockout (KO) of CREB1 or ATF1 enhanced viral gene expression and viral replication. The increase in viral yields in supernatants from ATF1-KO cells was greater than that in supernatants from CREB1-KO cells. Transcriptome sequencing (RNA-seq) analysis showed that sensors of the innate immune system were downregulated in ATF1-KO cells, and mRNAs of beta interferon (IFN-β) and IFN-regulated genes were reduced in these cells infected with HHV-6A. IFN-β treatment of ATF1-KO cells reduced progeny viral yields significantly, suggesting that the enhancement of viral replication was caused by a reduction of IFN-β. Taken together, our results suggest that ATF1 is activated during HHV-6A infection and restricts viral replication via IFN-β induction. IMPORTANCE Human herpesvirus 6A (HHV-6A) is a ubiquitous herpesvirus implicated in Alzheimer's disease, although its role in its pathogenesis has not been confirmed. Here, we showed that the transcription factor ATF1 restricts HHV-6A replication, mediated by IFN-β induction. Our study provides new insights into the role of ATF1 in innate viral immunity and reveals the importance of IFN-β for regulation of HHV-6A replication, which possibly impairs HHV-6A pathogenesis.

Beyond their pulmonary disease, many COVID-19 patients experience a complex constellation of characteristics, including hyperinflammatory responses, autoimmune disorders, and coagulopathies. However, the pathogenesis of these aspects of COVID-19 is obscure. More than 90% of people are latently infected with the lymphotropic herpesviruses Epstein-Barr Virus (EBV) and/or Human Herpesvirus-6 (HHV-6). Some of the inflammatory features of COVID-19 resemble clinical syndromes seen during EBV and HHV-6 infection, and these latent viruses can be reactivated by inflammatory mediators. We hypothesized that EBV and HHV-6 reactivation might be a common feature of early COVID-19, particularly in patients with more inflammation. We tested for EBV and HHV-6 reactivation in 67 patients acutely hospitalized with COVID-19 using previously validated quantitative PCR assays on the plasma. In our cohort, we found that 15/67 (22.4%) patients had detectable EBV and 3/67 (4.5%) had detectable HHV-6. This frequency of activation is somewhat more than the frequency reported for some healthy cohorts, such as blood donors and other healthy control cohorts. There was no association between EBV or HHV-6 and markers indicative of more inflammatory disease. We conclude that EBV and HHV-6 activation at about day 7 of hospitalization occurred in a modest fraction of our cohort of COVID-19 patients and was not associated with high levels of inflammation. In the modest fraction of patients, EBV and HHV-6 reactivation could contribute to some features of acute disease and pre-disposition to post-acute sequelae in a subset of patients.

Within the family Herpesviridae , sub-family β-herpesvirinae , and genus Roseolovirus , there are only three human herpesviruses that have been described: HHV-6A, HHV-6B, and HHV-7. Initially, HHV-6A and HHV-6B were considered as two variants of the same virus (i.e., HHV6). Despite high overall genetic sequence identity (~90%), HHV-6A and HHV-6B are now recognized as two distinct viruses. Sequence divergence (e.g., >30%) in key coding regions and significant differences in physiological and biochemical profiles (e.g., use of different receptors for viral entry) underscore the conclusion that HHV-6A and HHV-6B are distinct viruses of the β-herpesvirinae . Despite these viruses being implicated as causative agents in several nervous system disorders (e.g., multiple sclerosis, epilepsy, and chronic fatigue syndrome), the mechanisms of action and relative contributions of each virus to neurological dysfunction are unclear. Unresolved questions regarding differences in cell tropism, receptor use and binding affinity (i.e., CD46 versus CD134), host neuro-immunological responses, and relative virulence between HHV-6A versus HHV-6B prevent a complete characterization. Although it has been shown that both HHV-6A and HHV-6B can infect glia (and, recently, cerebellar Purkinje cells), cell tropism of HHV-6A versus HHV-6B for different nerve cell types remains vague. In this study, we show that both viruses can infect different nerve cell types (i.e., glia versus neurons) and different neurotransmitter phenotypes derived from differentiated human neural stem cells. As demonstrated by immunofluorescence, HHV-6A and HHV-6B productively infect VGluT1-containing cells (i.e., glutamatergic neurons) and dopamine-containing cells (i.e., dopaminergic neurons). However, neither virus appears to infect GAD67-containing cells (i.e., GABAergic neurons). As determined by qPCR, expression of immunological factors (e.g., cytokines) in cells infected with HHV-6A versus HHV6-B also differs. These data along with morphometric and image analyses of infected differentiated neural stem cell cultures indicate that while HHV-6B may have greater opportunity for transmission, HHV-6A induces more severe cytopathic effects (e.g., syncytia) at the same post-infection end points. Cumulatively, results suggest that HHV-6A is more virulent than HHV-6B in susceptible cells, while neither virus productively infects GABAergic cells. Consistency between these in vitro data and in vivo experiments would provide new insights into potential mechanisms for HHV6-induced epileptogenesis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bahramian, Furr, Wu and Ceballos.)

CD46 is a receptor for human herpesvirus 6A (HHV-6A) and is in some cells also important for infection with HHV-6B. CD46 has several isoforms of which the most commonly expressed can be distinguished by expression of a BC domain or a C domain in a serine-threonine-proline-rich (STP) extracellular region. Using a SupT1 CD46 CRISPR-Cas9 knockout model system reconstituted with specific CD46 isoforms, we demonstrated that HHV-6A infection was more efficient when BC isoforms were expressed as opposed to C isoforms, measured by higher levels of intracellular viral transcripts and recovery of more progeny virus. Although the B domain contains several O -glycosylations, mutations of Ser and Thr residues did not prevent infection with HHV-6A. The HHV-6A infection was blocked by inhibitors of clathrin-mediated endocytosis. In contrast, infection with HHV-6B was preferentially promoted by C isoforms mediating fusion-from-without, and this infection was less affected by inhibitors of clathrin-mediated endocytosis. Taken together, HHV-6A preferred BC isoforms, mediating endocytosis, whereas HHV-6B preferred C isoforms, mediating fusion-from-without. This demonstrates that the STP region of CD46 is important for regulating the mode of infection in SupT1 cells and suggests an epigenetic regulation of the host susceptibility to HHV-6A and HHV-6B infection. IMPORTANCE CD46 is the receptor used by human herpesvirus 6A (HHV-6A) during infection of T cells, but it is also involved in infection of certain T cells by HHV-6B. The gene for CD46 allows expression of several variants of CD46, known as isoforms, but whether the isoforms matter for infection of T cells is unknown. We used a genetic approach to delete CD46 from T cells and reconstituted them with separate isoforms to study them individually. We expressed the isoforms known as BC and C, which are distinguished by the potential inclusion of a B domain in the CD46 molecule. We demonstrate that HHV-6A prefers the BC isoform to infect T cells, and this occurs predominantly by clathrin-mediated endocytosis. In contrast, HHV-6B prefers the C isoform and infects predominantly by fusion-from-without. Thus, CD46 isoforms may affect susceptibility of T cells to infection with HHV-6A and HHV-6B.

Background: Infections with Epstein-Barr virus (EBV) and human herpesvirus-6 (HHV-6) have been implicated in multiple sclerosis (MS) onset but little work has studied their relationships in early disease., Objective: Evaluate associations between markers of EBV and HHV-6 infection/reactivation and MS conversion, relapse and EDSS/MSSS amongst 205 CIS participants with EBV/HHV-6 data followed over 5 years., Method: Baseline serological and viral load measures of EBV and HHV-6 exposure/reactivation were measured and infectious mononucleosis (IM) history recorded. Conversion to MS and relapses were assessed annually, and EDSS/MSSS measured at 5-year review. Determinants of MS conversion and relapse assessed by Cox regression, and disability progression by linear regression., Results: IM history showed a strong positive trend with higher relapse risk (aHR=1.45,95%CI=0.97-2.16) but was not associated with MS conversion (aHR=0.92,95%CI=0.57-1.48). Anti-HHV-6 IgG titre>40 also showed strong positive trends with higher relapse (aHR=1.61,95%CI=0.99-2.63) and MS conversion risks (aHR=1.48,95%CI=0.89-2.46). Anti-HHV-6 IgG titre≥640 was significantly associated with higher MSSS (0.15(95%CI=0.00,0.30) and also showed a strong positive trend with higher EDSS 0.10(95%CI=-0.02,0.21). HHV-6 DNA detection showed strong positive trends with 83%(95%CI=-6-357) and 77%(95%CI=-4-328) higher MS conversion and relapse risk. Anti-EBV-EA-D IgG titre was associated with a lower annualised disability progression by EDSS (p trend =0.037) and also showed strong positive trend with higher MSSS (p trend =0.053). No associations were seen for other serological or viral load markers., Conclusion: Overall, our data provides evidence that higher HHV-6 IgG was associated with increased risk of MS conversion and relapse but of borderline significance, and greater annualised disability progression, while that for EBV was more limited., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Introduction: This study investigated the spontaneous clinical course of patients with endomyocardial biopsy (EMB)-proven lymphocytic myocarditis and cardiac human herpesvirus 6 (HHV6) DNA presence, and the effectiveness of steroid-based intervention in HHV6-positive patients., Results: 756 heart failure (HF) patients underwent an EMB procedure to determine the underlying cause of unexplained HF. Low levels of HHV6 DNA, detectable by nested PCR only, were found in 10.4% of the cases ( n = 79) of which 62% ( n = 49) showed myocardial inflammation. The spontaneous course of patients with EMB-proven HHV6 DNA-associated lymphocytic myocarditis ( n = 26) showed significant improvements in the left ventricular ejection fraction (LVEF) and clinical symptoms, respectively, in 15/26 (60%) patients, 3-12 months after disease onset. EMB mRNA expression of components of the NLRP3 inflammasome pathway and protein analysis of cardiac remodeling markers, analyzed by real-time PCR and MALDI mass spectrometry, respectively, did not differ between HHV6-positive and -negative patients. In another cohort of patients with ongoing symptoms related to lymphocytic myocarditis associated with cardiac levels of HHV6-DNA copy numbers <500 copies/µg cardiac DNA, quantified by real-time PCR, the efficacy and safety of steroid-based immunosuppression for six months was investigated. Steroid-based immunosuppression improved the LVEF (≥5%) in 8/10 patients and reduced cardiac inflammation in 7/10 patients, without an increase in cardiac HHV6 DNA levels in follow-up EMBs., Conclusion: Low HHV6 DNA levels are frequently detected in the myocardium, independent of inflammation. In patients with lymphocytic myocarditis with low levels of HHV6 DNA, the spontaneous clinical improvement is nearby 60%. In selected symptomatic patients with cardiac HHV6 DNA copy numbers less than 500 copies/µg cardiac DNA and without signs of an active systemic HHV6 infection, steroid-based therapy was found to be effective and safe. This finding needs to be further confirmed in large, randomized trials.

A patient with an ultimate diagnosis of human herpesvirus-6 (HHV-6) encephalitis developed central nervous system (CNS) symptoms 13 days after undergoing myeloablative haploidentical allogeneic hematopoietic stem cell transplant (HSCT). Due to the patient's body habitus, magnetic resonance (MR) imaging was not obtained until the onset of retrograde amnesia on day +24. MR imaging and other clinical findings eliminated all skepticism of HHV-6 encephalitis and HHV-6 antivirals were initiated on day +28, leading to gradual recovery. This case demonstrates some of the factors that may complicate the diagnosis of post-alloHSCT HHV-6 encephalitis. Because HHV-6 encephalitis and viremia can occur without warning, a single negative study should not exclude future development, especially if CNS symptoms are present. Acute graft-versus-host disease and cord blood transplantation are both significant risk factors for HHV-6 encephalitis. Human leukocyte antigen (HLA) mismatch, engraftment complications, or certain HLA alleles have also been associated with HHV-6 encephalitis. Chromosomally integrated HHV-6 must also be ruled out to prevent inappropriate and potentially harmful administration of antivirals. Due to the severe short- and long-term sequelae of HHV-6 encephalitis, appropriate treatment should be administered as soon as possible.

Human endogenous retrovirus W family envelope proteins (pHERV-W ENV/syncytin-1) have been repeatedly associated with multiple sclerosis (MS). Here, we have focused on the study of pHERV-W ENV/syncytin-1 expression levels in MS patients (relapsing and progressive forms) and in healthy donors (HD) and on exploring their possible relationship with Epstein-Barr virus (EBV) and human herpesvirus-6A/B (HHV-6A/B). We included blood samples from 101 MS patients and 37 HD to analyze antiviral antibody titers by ELISA and pHERV-W ENV/syncytin-1 expression levels by flow cytometry as well as by qPCR. Patients with relapsing MS forms showed significantly higher pHERV-W ENV/syncytin-1 protein and gene expression levels than HD. Progressive MS patients also showed significantly higher protein and gene expression levels than both HD and relapsing MS patients. Regarding antiviral antibodies titers, anti-HHV-6A/B IgM levels were positively correlated with pHERV-W ENV/syncytin-1 protein expression levels in patients with relapsing MS, while in the progressive forms patients this correlation was found with anti-HHVA/B IgG levels. Therefore, pHERV-W ENV could be involved in MS pathogenesis, playing a role in relapsing and progressive forms. Besides, anti-HHV-6A/B antibodies positively correlated with pHERV-W ENV expression. Further studies are needed to better understand this possible relationship., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Pérez-Pérez, Domínguez-Mozo, García-Martínez, García-Frontini, Villarrubia, Costa-Frossard, Villar, Arroyo and Álvarez-Lafuente.)

The coevolution of the human immune system and herpesviruses led to the emergence and diversification of both cellular danger molecules recognized by immune cells on the one hand and viral countermeasures that prevent the expression of these proteins on infected cells on the other. There are eight ligands for the activating receptor NKG2D in humans - MICA, MICB, ULBP1-6. Several of them are induced and surface-expressed on herpesvirus-infected cells to serve as danger signals to activate the immune system. Therefore, these ligands are frequently targeted for suppression by viral immune evasion mechanisms. Mechanisms to downregulate NKG2D ligands and thereby escape immune recognition have been identified in all other human herpesviruses (HHV), except for HHV-6A. In this study, we identify two HHV-6A encoded immunoevasins, U20 and U21, which suppress the expression of the NKG2D ligands ULBP1 and ULBP3, respectively, during infection. Additionally, MICB is targeted by a so far unexplored viral protein. Due to the diminished NKG2D ligand surface expression on infected cells, recognition of HHV-6A infected cells by innate immune cells is impaired. Importantly, our study indicates that immune escape mechanisms between the related herpesviruses HHV-6A and HHV-6B are evolutionary conserved as the same NKG2D ligands are targeted. Our data contribute an additional piece of evidence for the importance of the NKG2D receptor - NKG2D ligand axis during human herpesvirus infections and sheds light on immune evasion mechanisms of HHV-6A., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Chaouat, Seliger, Mandelboim and Schmiedel.)

Background: Systemic reactivation of herpesviruses may occur in intensive care unit (ICU) patients and is associated with morbidity and mortality. Data on severe Coronavirus disease-19 (COVID-19) and concomitant reactivation of herpesviruses are lacking., Methods: We selected patients admitted to ICU for confirmed COVID-19 who underwent systematic testing for Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human-herpes virus-6 (HHV-6) DNAemia while in the ICU. We retrospectively analysed frequency, timing, duration and co-occurrence of viral DNAemia., Results: Thirty-four patients were included. Viremia with EBV, CMV, and HHV-6 was detected in 28 (82%), 5 (15%), and 7 (22%) patients, respectively. EBV reactivation occurred early after ICU admission and was associated with longer ICU length-of-stay., Conclusions: While in the ICU, critically ill patients with COVID-19 are prone to develop reactivations due to various types of herpesviruses., (© 2021 Elsevier Masson SAS. All rights reserved.)

Human herpesviruses 6A (HHV-6A) and human herpesvirus 6B (HHV-6B)-collectively, HHV-6A/B-are recently-discovered but ancient human viruses. The vast majority of people acquire one or both viruses, typically very early in life, producing an ineradicable lifelong infection. The viruses have been linked to several neurological, pulmonary and hematological diseases. In early human history, the viruses on multiple occasions infected a germ cell, and integrated their DNA into a human chromosome. As a result, about 1% of humans are born with the full viral genome present in every cell, with uncertain consequences for health. HHV-6A may play a role in 43% of cases of primary unexplained infertility. Both the inherited and acquired viruses may occasionally trigger several of the factors that are important in the pathogenesis of preeclampsia. Transplacental infection occurs in 1-2% of pregnancies, with some evidence suggesting adverse health consequences for the child. While emerging knowledge about these viruses in reproductive diseases is not sufficient to suggest any changes in current practice, we write this review to indicate the need for further research that could prove practice-changing., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Komaroff, Rizzo and Ecker.)

IRAK4 deficiency is an inborn error of immunity predisposing patients to invasive pyogenic infections. Currently, there is no established simple assay that enables precise characterization of IRAK4 mutant alleles in isolation. Anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis is an autoimmune condition that is characterized by psychiatric symptoms, involuntary movement, seizures, autonomic dysfunction, and central hypoventilation. It typically occurs in adult females associated with tumors. Only a few infantile cases with anti-NMDAR encephalitis have been so far reported. We identified a 10-month-old boy with IRAK4 deficiency presenting with anti-NMDAR encephalitis and human herpes virus 6 (HHV6) reactivation. The diagnosis of IRAK4 deficiency was confirmed by the identification of compound heterozygous mutations c.29&#95;30delAT (p.Y10Cfs*9) and c.35G>C (p.R12P) in the IRAK4 gene, low levels of IRAK4 protein expression in peripheral blood, and defective fibroblastic cell responses to TLR and IL-1 (TIR) agonist. We established a novel NF-κB reporter assay using IRAK4-null HEK293T, which enabled the precise evaluation of IRAK4 mutations. Using this system, we confirmed that both novel mutations identified in the patient are deleterious. Our study provides a new simple and reliable method to analyze IRAK4 mutant alleles. It also suggests the possible link between inborn errors of immunity and early onset anti-NMDAR encephalitis.

Next-generation sequencing technologies allowed sequencing of thousands of genomes. However, there are genomic regions that remain difficult to characterize, including telomeres, centromeres, and other low-complexity regions, as well as transposable elements and endogenous viruses. Human herpesvirus 6A and 6B (HHV-6A and HHV-6B) are closely related viruses that infect most humans and can integrate their genomes into the telomeres of infected cells. Integration also occurs in germ cells, meaning that the virus can be inherited and result in individuals harboring the virus in every cell of their body. The integrated virus can reactivate and cause disease in humans. While it is well established that the virus resides in the telomere region, the integration locus is poorly defined due to the low sequence complexity (TTAGGG)n of telomeres that cannot be easily resolved through sequencing. We therefore employed genome imaging of the integrated HHV-6A and HHV-6B genomes using whole-genome optical site mapping technology. Using this technology, we identified which chromosome arm harbors the virus genome and obtained a high-resolution map of the integration loci of multiple patients. Surprisingly, this revealed long telomere sequences at the virus-subtelomere junction that were previously missed using PCR-based approaches. Contrary to what was previously thought, our technique revealed that the telomere lengths of chromosomes harboring the integrated virus genome were comparable to the other chromosomes. Taken together, our data shed light on the genetic structure of the HHV-6A and HHV-6B integration locus, demonstrating the utility of optical mapping for the analysis of genomic regions that are difficult to sequence., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Human herpesvirus 6A (HHV-6A) and 6B (HHV-6B) are two closely related viruses that can infect cells of the central nervous system (CNS). The similarities between these viruses have made it difficult to separate them on serological level. The broad term HHV-6 remains when referring to studies where the two species were not distinguished, and as such, the seroprevalence is over 90% in the adult population. HHV-6B has been detected in up to 100% of infants with the primary infection roseola infantum, but less is known about the primary infection of HHV-6A. Both viruses are neurotropic and have capacity to establish lifelong latency in cells of the central nervous system, with potential to reactivate and cause complications later in life. HHV-6A infection has been associated with an increased risk of multiple sclerosis (MS), whereas HHV-6B is indicated to be involved in pathogenesis of epilepsy. These two associations show how neurological diseases might be caused by viral infections, but as suggested here, through completely different molecular mechanisms, in an autoimmune disease, such as MS, by triggering an overreaction of the immune system and in epilepsy by hampering internal cellular functions when the immune system fails to eliminate the virus. Understanding the viral mechanisms of primary infection and reactivation and their spectrum of associated symptoms will aid our ability to diagnose, treat and prevent these severe and chronic diseases. This review explores the role of HHV-6A and HHV-6B specifically in MS and epilepsy, the evidence to date and the future directions of this field., (© 2020 The Authors. Scandinavian Journal of Immunology published by John Wiley & Sons Ltd on behalf of The Scandinavian Foundation for Immunology.)

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.

Viral reactivation occurs frequently in the context of immunodeficiency and immunosuppression after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and can cause severe complications. The aim of this single-center retrospective analysis was to characterize viral infections in the first year after HSCT, to investigate risk factors and to study the impact of viral infections on transplantation outcome. This will facilitate the identification of at-risk patients and the development of new preventive strategies. 107 pediatric allo-HSCT from January 2005 through December 2015 were analyzed for infections with Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpesvirus 6 (HHV-6), adenovirus (ADV), herpes simplex virus (HSV) and varicella zoster virus (VZV). Viral infections were detected after 68.2% of transplantations. The viruses most commonly encountered were HHV-6 (36/107) and EBV (30/107). Severe viral disease was rare (7/107) and none of the patients died as result of viral reactivation. Important risk factors for viral infections were higher age at HSCT, donor type and occurrence of acute graft-versus-host disease (aGvHD). Especially for EBV, transplant from an unrelated donor and in-vivo T-cell depletion (TCD) had a significant effect on infection rates, whereas for CMV the strongest effect was seen by donor and recipient serostatus with recipient seropositivity most predictive for reactivation. The occurrence of severe aGvHD was associated with EBV and ADV infections. For HSV, the recipient serostatus was identified as prognostic factor for HSV infections, while we found higher age at time of HSCT as risk factor for VZV infections. The overall survival of patients with or without viral infections did not differ significantly. Interestingly, when looking at the 85 patients in our cohort who had received an HSCT for a malignant disease, a tendency towards lower relapse rates was seen in patients affected by viral infections (HR 0.51, 95% CI 0.25 - 1.06, p = 0.072). Viral reactivations are common after pediatric allo-HSCT, though severe complications were rare in our collective. Determining risk factors for viral reactivations may help to identify patients in need of intensified monitoring and to individualize preventive strategies., Competing Interests: The authors have declared that no competing interests exist.

Background: Human herpesvirus-6B (HHV-6B) infection after allogenic hematopoietic stem cell transplantation (allo-HSCT) is known to be associated with post-transplant limbic encephalitis in adults. Meanwhile, the association between HHV-6B infection and central nervous system complications remains unclear in pediatric allo-HSCT patients., Methods: In this study, HHV-6B infection was monitored for more than 50 days after HSCT using virus isolation and real-time PCR. Clinical information such as patient background and encephalitis status was collected retrospectively from medical records. Risk factors for HHV-6B infection were determined by the Cox proportional hazards model, and the clinical features of HHV-6B encephalitis in pediatric allo-HSCT patients were elucidated., Results: Human herpesvirus-6B infection was observed in 74 (33.8%) of 219 patients at 3-47 days (median 18, interquartile range 13-20). Risk factors identified in multivariable analysis were hematological malignancy (hazards ratio [HR], 5.0; 95% confidence interval [CI], 2.3/12.5; P < .0001), solid tumor (HR, 4.8; CI, 1.5/16.3; P = .0104), unrelated donor (HR, 2.1; CI, 1.0/4.6; P = .0378), and sex-mismatched donor (HR 1.8; CI, 1.1/3.0; P = .0257). HHV-6B encephalitis occurred in only one of the 219 patients (0.46%); this patient demonstrated the typical clinical course of posterior reversible encephalopathy syndrome., Conclusion: Hematological malignancy, solid tumor, unrelated donor, and sex-mismatched donor were significant risk factors for HHV-6B infection after pediatric allo-HSCT. In pediatric allo-HSCT patients, the incidence of HHV-6B encephalitis was low and the clinical features differed from those in adult patients., (© 2019 Wiley Periodicals, Inc.)

The human betaherpesviruses, human cytomegalovirus (HCMV; species Human betaherpesvirus 5) and human herpesviruses 6A, 6B, and 7 (HHV-6A, -6B, and -7; species Human betaherpesviruses 6A, 6B, and 7) are highly prevalent and can cause severe disease in immune-compromised and immune-naive populations in well- and under-developed communities. Herpesvirus virion assembly is an intricate process that requires viral orchestration of host systems. In this review, we describe recent advances in some of the many cellular events relevant to assembly and egress of betaherpesvirus virions. These include modifications of host metabolic, immune, and autophagic/recycling systems. In addition, we discuss unique aspects of betaherpesvirus virion structure, virion assembly, and the cellular pathways employed during virion egress., (© 2020 Elsevier Inc. All rights reserved.)

Human herpesvirus (HHV)-6A or HHV-6B involvement in multiple sclerosis (MS) etiology has remained controversial mainly due to the lack of serological methods that can distinguish the two viruses. A novel multiplex serological assay measuring IgG reactivity against the immediate-early protein 1 from HHV-6A (IE1A) and HHV-6B (IE1B) was used in a MS cohort (8,742 persons with MS and 7,215 matched controls), and a pre-MS cohort (478 individuals and 476 matched controls) to investigate this further. The IgG response against IE1A was positively associated with MS (OR = 1.55, p = 9 × 10 -22 ), and increased risk of future MS (OR = 2.22, p = 2 × 10 -5 ). An interaction was observed between IE1A and Epstein-Barr virus (EBV) antibody responses for MS risk (attributable proportion = 0.24, p = 6 × 10 -6 ). In contrast, the IgG response against IE1B was negatively associated with MS (OR = 0.74, p = 6 × 10 -11 ). The association did not differ between MS subtypes or vary with severity of disease. The genetic control of HHV-6A/B antibody responses were located to the Human Leukocyte Antigen (HLA) region and the strongest association for IE1A was the DRB1 * 13:01-DQA1 * 01:03-DQB1 * 06:03 haplotype while the main association for IE1B was DRB1 * 13:02-DQA1 * 01:02-DQB1 * 06:04. In conclusion a role for HHV-6A in MS etiology is supported by an increased serological response against HHV-6A IE1 protein, an interaction with EBV, and an association to HLA genes., (Copyright © 2019 Engdahl, Gustafsson, Huang, Biström, Lima Bomfim, Stridh, Khademi, Brenner, Butt, Michel, Jons, Hortlund, Alonso-Magdalena, Hedström, Flamand, Ihira, Yoshikawa, Andersen, Hillert, Alfredsson, Waterboer, Sundström, Olsson, Kockum and Fogdell-Hahn.)

HHV-6A and HHV-6B are ubiquitous human betaherpesviruses sharing more than 80% homology. HHV-6B is the most common cause of encephalitis in transplant patients and its primary infection may cause the exanthema subitum and febrile seizures in infants. HHV-6A and HHV-6B are able to infect several immune cell types such as T cells, monocytes and dendritic cells (DCs). In this study we found that HHV-6 B derived from patients affected by exanthema subitum impaired monocyte differentiation into DCs, as the infected cells acquired less CD1a DC marker and retained more CD14 monocyte marker. In agreement with the previous finding that HHV-6B dysregulated autophagy and induced endoplasmic reticulum (ER) stress in cells in which it replicated, here we found that these effects occurred also in differentiating monocytes and that ER stress relief, by using the chemical chaperone sodium 4-phenylbutirate (PBA), partially restored DC formation. This suggests that the induction of ER stress, likely exacerbated by autophagy inhibition, could contribute to the immune suppression induced by HHV-6B derived from exanthema subitem patients., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Myalgic encephalomyelitis, also referred to as chronic fatigue syndrome (ME/CFS) is a debilitating disease characterized by myalgia and a sometimes severe limitation of physical activity and cognition. It is exacerbated by physical and mental activity. Its cause is unknown, but frequently starts with an infection. The eliciting infection (commonly infectious mononucleosis or an upper respiratory infection) can be more or less well diagnosed. Among the human herpesviruses (HHV-1-8), HHV-4 (Epstein-Barr virus; EBV), HHV-6 (including HHV-6A and HHV-6B), and HHV-7, have been implicated in the pathogenesis of ME/CFS. It was therefore logical to search for serological evidence of past herpesvirus infection/reactivation in several cohorts of ME/CFS patients (all diagnosed using the Canada criteria). Control samples were from Swedish blood donors. We used whole purified virus, recombinant proteins, and synthetic peptides as antigens in a suspension multiplex immunoassay (SMIA) for immunoglobulin G (IgG). The study on herpesviral peptides based on antigenicity with human sera yielded novel epitope information. Overall, IgG anti-herpes-viral reactivities of ME/CFS patients and controls did not show significant differences. However, the high precision and internally controlled format allowed us to observe minor relative differences between antibody reactivities of some herpesviral antigens in ME/CFS versus controls. ME/CFS samples reacted somewhat differently from controls with whole virus HHV-1 antigens and recombinant EBV EBNA6 and EA antigens. We conclude that ME/CFS samples had similar levels of IgG reactivity as blood donor samples with HHV-1-7 antigens. The subtle serological differences should not be over-interpreted, but they may indicate that the immune system of some ME/CFS patients interact with the ubiquitous herpesviruses in a way different from that of healthy controls.

Human herpes virus 6B (HHV-6B) is a widespread virus that infects most people early in infancy and establishes a chronic life-long infection with periodic reactivation. CD4 T cells have been implicated in control of HHV-6B, but antigenic targets and functional characteristics of the CD4 T-cell response are poorly understood. We identified 25 naturally processed MHC-II peptides, derived from six different HHV-6B proteins, and showed that they were recognized by CD4 T-cell responses in HLA-matched donors. The peptides were identified by mass spectrometry after elution from HLA-DR molecules isolated from HHV-6B-infected T cells. The peptides showed strong binding to matched HLA alleles and elicited recall T-cell responses in vitro. T-cell lines expanded in vitro were used for functional characterization of the response. Responding cells were mainly CD3 + CD4 + , produced IFN-γ, TNF-α, and low levels of IL-2, alone or in combination, highlighting the presence of polyfunctional T cells in the overall response. Many of the responding cells mobilized CD107a, stored granzyme B, and mediated specific killing of peptide-pulsed target cells. These results highlight a potential role for polyfunctional cytotoxic CD4 T cells in the long-term control of HHV-6B infection., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Objective: There has been little research on human herpesvirus 6B (HHV-6B) in healthy adults and prevalences in different age groups have been unclear. Therefore, the major objective of this study was to evaluate seroprevalence to HHV-6 antibodies in ordinary working people and examine the effect of aging on seroprevalence. Also, as HHV-6B is reactivated in saliva, another objective was to investigate an association between age and HHV-6B reactivation based on measured salivary HHV-6 DNA levels., Methods: Our subjects were 77 ordinary office workers who underwent a health checkup. In this population, we measured anti-HHV-6 antibody titers using enzyme-linked immunosorbent assay and salivary HHV-6 DNA levels. In addition to examining an association with age, we examined associations with body mass index, smoking habit, and alcohol consumption as confounding factors., Results: There was a significant decrease in the seropositivity of HHV-6 antibodies in subjects of 50 years and older, and age was significantly negatively correlated with anti-HHV-6 antibody titers. Age and salivary HHV-6 DNA levels were also significantly negatively correlated but there were no significant correlations with other factors., Conclusions: Our results suggest that HHV-6B reactivation is attenuated by aging. Thus, HHV-6 antibodies steadily decrease in the body with aging., (© 2019 Wiley Periodicals, Inc.)

Objectives: Kawasaki disease (KD) is one of the most common childhood vasculitides. Some serological studies have suggested an etiological relationship between KD and human herpesvirus (HHV)-6 or HHV-7. However, primary or reactivated HHV-6 and -7 has not been fully investigated in patients with KD. Methods: Twenty-three patients with KD were prospectively enrolled in this study. Peripheral blood was collected in the acute and convalescence phases, and HHV-6 and -7 viral loads were measured by real-time PCR. Results: In the acute phase, HHV-6 and -7 DNA was detected in 7 (30%) patients each, compared to 13 (57%) and 9 (39%) patients in the convalescence phase, respectively. HHV-6 and -7 DNA loads were significantly higher in the convalescence phase than in the acute phase. Significant increases in HHV-6 and -7 DNA loads were not observed in disease control patients. Taking into account HHV-6 and -7 serostatus, reactivation of HHV-6 and -7 was observed in 7 and 9 patients, respectively. KD patients with HHV-6 reactivation showed higher C-reactive protein levels and more frequently required steroid therapies than patients without reactivation. Conclusion: HHV-6 and -7 reactivation is frequent in KD patients. HHV-6 reactivation might exacerbate the severity of KD.

Human herpesvirus 6 (HHV-6) infects >90% of the population and establishes a latent infection with asymptomatic episodes of reactivation. However, HHV-6 reactivation is associated with morbidity and sometimes mortality in immunocompromised patients. To date, control of the virus in healthy virus carriers and the failure to control it in patients with disease remain poorly understood. In particular, knowledge of HHV-6-specific T-cell responses is limited. Here, we characterized HHV-6A- and HHV-6B-specific CD4 + and CD8 + T-cell responses from peripheral blood mononuclear cells (PBMCs) of healthy donors. We studied the phenotype of effector HHV-6-specific T cells ex vivo , as well as of induced specific suppressive regulatory CD4 + T cells in vitro poststimulation, in comparison to human cytomegalovirus (HCMV) responses. Compared to that for HCMV, we show that ex vivo T-cell reactivity in peripheral blood is detectable but at very low frequency, both for HHV-6A and -6B viruses. Interestingly, the phenotype of the specific T cells also differs between the viruses. HHV-6A- and HHV-6B-specific CD4 + T lymphocytes are less differentiated than HCMV-specific T cells. Furthermore, we show a higher frequency of HHV-6-specific suppressive regulatory T cells (eTregs) than HCMV-specific eTregs in coinfected individuals. Despite the strong similarity of HHV-6 and HCMV from a virologic point of view, we observed immunological differences, particularly in relation to the frequency and phenotype of effector/memory and regulatory virus-specific T cells. This suggests that different immune factors are solicited in the control of HHV-6 infection than in that of HCMV infection. IMPORTANCE T cells are central to an effective defense against persistent viral infections that can be related to human cytomegalovirus (HCMV) or human herpesvirus 6 (HHV-6). However, knowledge of HHV-6-specific T-cell responses is limited. In order to deepen our knowledge of T-cell responses to HHV-6, we characterized HHV-6A- and HHV-6B-specific CD4 + and CD8 + T-cell responses directly ex vivo from healthy coinfected blood donors. Despite the strong similarity of HHV-6 and HCMV from a virologic point of view, we observed immunological differences, particularly in relation to the frequency and phenotype of effector/memory and regulatory virus-specific T cells. This suggests that different immune factors are solicited in the control of HHV-6 infection than in that of HCMV infection. Our findings may encourage immunomonitoring of patients with viral replication episodes to follow the emergence of effector versus regulatory T cells., (Copyright © 2019 American Society for Microbiology.)

Background: Human leukocyte antigen (HLA) mismatch and the administration of immunosuppressive agents are considered risks for human herpesvirus 6 (HHV-6) reactivation after stem cell transplantation (SCT). However, the incidence of HHV-6 reactivation in HLA-mismatched related SCT remains unknown., Methods: We monitored plasma HHV-6 DNA loads weekly using real-time quantitative polymerase chain reaction for 5 weeks after SCT and compared serum IL-6 levels in HLA-mismatched SCT groups., Results: Compared with detection in all 11 umbilical cord blood transplantation (CBT) patients (100%), plasma HHV-6 DNA was detected in only 3 of 42 haplo-SCT patients (7.1%) despite the use of methylprednisolone and antithymocyte globulin as graft-vs-host disease prophylaxis and a reduced-intensity conditioning regimen, respectively. Correspondingly, serum IL-6 levels in haplo-SCT patients were significantly lower than those in CBT patients. No HHV-6-associated encephalitis developed in either groups., Conclusions: Neither HLA disparity nor the use of methylprednisolone and antithymocyte globulin were risk factors for HHV-6 reactivation in our haplo-SCT patients. Rather than increasing risk, the administration of immunosuppressive agents potentially prevented HHV-6 reactivation after haplo-SCT by suppressing IL-6 production., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Viral infection is a serious complication that can greatly affect patient mortality and morbidity after allogenic hematopoietic stem cell transplantation (allo-HSCT). For the early identification of patients at high risk for viral infection, we evaluated the impact of lymphocyte area under the curve (AUC) value as a new predictive factor for early immune reconstitution after allo-HSCT against viral infection. This study included 286 patients who underwent their first allo-HSCT at Kyoto University Hospital between 2005 and 2017. Lymphocyte AUC from day 0 to day +15 was calculated in the analysis of human herpesvirus 6 (HHV-6), and lymphocyte AUC from day 0 to day +30 was calculated in the analysis of other viruses (cytomegalovirus [CMV], adenovirus, BK virus, JC virus, and varicella zoster virus). The risk factors for each viral reactivation/infection were assessed by multivariate analysis. The median age at transplantation was 51years (range, 17 to 68 years). The median lymphocyte AUC was 63/μL (range, 0 to 5620/μL) at day +15 and 3880 (range, 0 to 118,260/μL) at day +30. An increase in lymphocyte AUC was significantly associated with a high frequency of HHV-6 reactivation (P = .033) and a low frequency of CMV antigenemia (P = .014). No apparent association was found between lymphocyte AUC and reactivation/infection of other viruses. Aplastic anemia as a primary disease (hazard ratio [HR], 5.34; P < .001) and cord blood as a donor source (HR, 3.05; P = .006) were other risk factors for HHV-6 reactivation. Other risk factors for CMV antigenemia included the occurrence of acute graft-versus-host disease (HR 2.21; P < .001) and recipient age (HR 1.55; P = .017). Higher lymphocyte AUC at day +30 was significantly associated with low treatment-related mortality (HR, .47; P = .045). Lymphocyte AUC may be a good predictive factor for immune reconstitution against CMV reactivation. It also provides valuable information for predicting HHV-6 reactivation and treatment-related mortality., (Copyright © 2018 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.)

In this chapter, we present an overview on betaherpesvirus entry, with a focus on human cytomegalovirus, human herpesvirus 6A and human herpesvirus 6B. Human cytomegalovirus (HCMV) is a complex human pathogen with a genome of 235kb encoding more than 200 genes. It infects a broad range of cell types by switching its viral ligand on the virion, using the trimer gH/gL/gO for infection of fibroblasts and the pentamer gH/gL/UL128/UL130/UL131 for infection of other cells such as epithelial and endothelial cells, leading to membrane fusion mediated by the fusion protein gB. Adding to this scenario, however, accumulating data reveal the actual complexity in the viral entry process of HCMV with an intricate interplay among viral and host factors. Key novel findings include the identification of entry receptors platelet-derived growth factor-α receptor (PDGFRα) and Netropilin-2 (Nrp2) for trimer and pentamer, respectively, the determination of atomic structures of the fusion protein gB and the pentamer, and the in situ visualization of the state and arrangement of functional glycoproteins on virion. This is covered in the first part of this review. The second part focusses on HHV-6 which is a T lymphotropic virus categorized as two distinct virus species, HHV-6A and HHV-6B based on differences in epidemiological, biological, and immunological aspects, although homology of their entire genome sequences is nearly 90%. HHV-6B is a causative agent of exanthema subitum (ES), but the role of HHV-6A is unknown. HHV-6B reactivation occasionally causes encephalitis in patients with hematopoietic stem cell transplant. The HHV-6 specific envelope glycoprotein complex, gH/gL/gQ1/gQ2 is a viral ligand for the entry receptor. Recently, each virus has been found to recognize a different cellular receptor, CD46 for HHV 6A amd CD134 for HHV 6B. These findings show that distinct receptor recognition differing between both viruses could explain their different pathogenesis., (© 2019 Elsevier Inc. All rights reserved.)

Human herpesvirus 6B (HHV-6B) is a ubiquitous pathogen that infects most individuals before the age of three years. HHV-6B reactivates in approximately 40% of transplant recipients where it has been associated with a number of important outcomes, especially in allogeneic transplant recipients. This article will review the epidemiology, clinical manifestations, diagnosis, and treatment of HHV-6B infection.

Human herpesvirus-6A and -6B (HHV-6A and -6B) are two closely related betaherpesviruses that infect humans. Upon primary infection they establish a life-long infection termed latency, where the virus genome is integrated into the telomeres of latently infected cells. Intriguingly, HHV-6A/B can integrate into germ cells, leading to individuals with inherited chromosomally-integrated HHV-6 (iciHHV-6), who have the HHV-6 genome in every cell. It is known that telomeric repeats flanking the virus genome are essential for integration; however, the protein factors mediating integration remain enigmatic. We have previously shown that the putative viral integrase U94 is not essential for telomere integration; thus, we set out to assess the contribution of potential viral recombination proteins U41 and U70 towards integration. We could show that U70 enhances dsDNA break repair via a homology-directed mechanism using a reporter cell line. We then engineered cells to produce shRNAs targeting both U41 and U70 to inhibit their expression during infection. Using these cells in our HHV-6A in vitro integration assay, we could show that U41/U70 were dispensable for telomere integration. Furthermore, additional inhibition of the cellular recombinase Rad51 suggested that it was also not essential, indicating that other cellular and/or viral factors must mediate telomere integration.

Background: Human herpesvirus 6B (HHV-6B) is a neurotropic virus that has been repeatedly associated with mesial temporal lobe epilepsy (MTLE). However, the mechanism behind this suggested association is not known. Therefore, the aim of this study was to investigate what genes were affected by HHV-6B, possibly revealing HHV-6B induced disease causing mechanisms., Material and Method: First, gene expression in MTLE tissue positive for HHV-6B DNA (n = 10) and negative for HHV-6B DNA (n = 14) was compared using the Affymetrix® Human Gene 2.1 ST Array. Secondly, in vitro experiments were conducted where Molt-3 T cells were infected with HHV-6B and gene expression of MAP2K4 (MKK4) and 89 other genes in the MAPK signaling pathway was investigated using qPCR. In addition, phosphorylated MKK4 was assessed using IFA and the DNA methylation investigated with Illumina Infinium HumanMethylation450 BeadChip array., Results: MAP2K4 was one of the most differently expressed genes in the Affymetrix array, suggesting an upregulation by HHV-6B infection in MTLE tissue. No gene reached statistical significance but MAP2K4 was selected for further investigation in vitro, where it was clearly upregulated by HHV-6B infection both on gene expression and protein expression level. Further investigating expression of genes in the MAPK pathways in vitro revealed that several genes were affected by HHV-6B infection, but none of these genes displayed viral induced changes in DNA methylation., Conclusions: As the MAPK pathways are involved in transforming different stimuli (like stress) into a cellular responses (like apoptosis or inflammation), it may not be surprising that genes in these pathways are affected by virus infection. This is the first report of HHV-6B's effect on these signaling cascades and given that both dysregulation of the MAPK pathways and an association with HHV-6B have been previously observed in epilepsy, a possible link of infection induced dysregulation of MAPK in epilepsy warrant further investigation., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: We sought to determine whether late-phase human herpesvirus 6B (HHV-6B) infection in hematopoietic stem cell transplant (HSCT) recipients was associated with serious outcomes and mortality., Methods: The occurrence and course of HHV-6B infection was monitored for at least 60 days after transplant using virus isolation and real-time polymerase chain reaction. Risk factors for late-phase HHV-6B infection were examined, and the propensity score was calculated with significant risk factors. The inverse probability-weighted multivariable logistic regression analysis was performed to estimate odds ratios (ORs) and the 95% confidence intervals (95% CI) for mortality., Results: Late-phase HHV-6B infection was observed in 12/89 (13.5%) of the HSCT recipients. Older age (OR: 10.3, 95% CI: 2.1/72.9, P = .0027), hematologic malignancy (OR: 10.3, 95% CI: 1.8/97.1, P = .0063), unrelated donor transplantation (OR: 5.3, 95% CI: 1.1/36.0, P = .0345), and sex-mismatched donor transplantation (OR: 6.3, 95% CI: 1.4/39.5, P = .0149) were identified as risk factors for late-phase HHV-6B infection. Fifteen subjects died (17%). Inverse probability-weighted multivariable logistic model analysis revealed that late-phase HHV-6B infection was an independent risk factor for mortality (OR: 4.2, 95% CI: 1.7/11.0, P = .0012). Among 5 of the fatal cases of late-phase HHV-6B infection, viral infection might be associated with severe clinical manifestations., Conclusion: Late-phase HHV-6B infection in HSCT recipients was associated with worse outcomes. The full spectrum of clinical features of the infection has not been fully elucidated, and therefore, recipients with high-risk factors for late-phase HHV-6B infection should be carefully monitored., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Human herpesvirus 6A (HHV-6A) replicates in peripheral blood mononuclear cells (PBMCs) and various T-cell lines in vitro. Intriguingly, the virus can also establish latency in these cells, but it remains unknown what influences the decision between lytic replication and the latency of the virus. Incoming virus genomes are confronted with the nuclear domain 10 (ND10) complex as part of an intrinsic antiviral response. Most herpesviruses can efficiently subvert ND10, but its role in HHV-6A infection remains poorly understood. In this study, we investigated if the ND10 complex affects HHV-6A replication and contributes to the silencing of the virus genome during latency. We could demonstrate that ND10 complex was not dissociated upon infection, while the number of ND10 bodies was reduced in lytically infected cells. Virus replication was significantly enhanced upon knock down of the ND10 complex using shRNAs against its major constituents promyelocytic leukemia protein (PML), hDaxx, and Sp100. In addition, we could demonstrate that viral genes are more efficiently silenced in the presence of a functional ND10 complex. Our data thereby provides the first evidence that the cellular ND10 complex plays an important role in suppressing HHV-6A lytic replication and the silencing of the virus genome in latently infected cells.

HHV-6A and HHV-6B are found as inherited and chromosomally integrated forms (iciHHV-6A and -6B) into all germinal and somatic cells and vertically transmitted in a Mendelian manner in about 1% of the population. They were occasionally shown to be horizontally transmitted through hematopoietic stem cell transplantation. Here, we present a clinical case of horizontal transmission of iciHHV-6A from donor to recipient through liver transplantation. Molecular analysis performed on three viral genes (7.2 kb) in the recipient and donor samples supports transmission of iciHHV-6A from the graft. Transmission was followed by reactivation, with high viral loads in several compartments. The infection was uncontrollable, leading to severe disease and death, despite antiviral treatments and the absence of resistance mutations. This case highlights the fact that physicians should be aware of the possible horizontal transmission of iciHHV-6 and its consequences in case of reactivation in immunocompromised patients., (© 2018 The American Society of Transplantation and the American Society of Transplant Surgeons.)