Dharam V. Ablashi

Genomic polymorphism, growth properties, and immunologic variations in human herpesvirus-6 isolates

Fifteen human herpesvirus-6 (HHV-6) isolates from normal donors and patients with AIDS, systemic lupus erythematosis, chronic fatigue syndrome, collagen-vascular disease, leukopenia, bone marrow transplants, Exanthem subitum (roseola), and atypical polyclonal lymphoproliferation were studied for their tropism to fresh human cord blood mononuclear cells, growth in continuous T cell lines, reactivity to monoclonal antibodies, and by restriction enzyme banding patterns. All isolates replicated efficiently in human cord blood mononuclear cells, but mitogen stimulation of the cells prior to infection was required. The ability to infect continuous T-cell lines varied with the isolates. Isolates similar to GS prototype infected HSB2 and Sup T1 cells and did not infect Molt-3 cells, whereas isolates similar to Z-29 infected Molt-3 cells but not HSB2 and Sup T1 cells. Some of the monoclonal antibodies directed against the HHV-6 (GS) isolate showed reactivity with all isolates tested, but others only reacted with HHV-6 isolates similar to the GS isolate and not with those similar to Z-29 isolate. Restriction enzyme analysis using EcoRI, BamHI, and HindIII revealed that HHV-6 isolates from roseola, bone marrow transplant, leukopenia, and an HIV-1-positive AIDS patient from Zaire (Z-29) were closely related but distinct from GS type HHV-6 isolates. Based on the above findings, we propose that, like herpes simplex virus types 1 and 2, the 15 HHV-6 isolates analyzed can be divided into group A (GS type) and group B (Z-29 type).

The latent human herpesvirus-6A genome specifically integrates in telomeres of human chromosomes in vivo and in vitro

Previous research has suggested that human herpesvirus-6 (HHV-6) may integrate into host cell chromosomes and be vertically transmitted in the germ line, but the evidence—primarily fluorescence in situ hybridization (FISH)—is indirect. We sought, first, to definitively test these two hypotheses. Peripheral blood mononuclear cells (PBMCs) were isolated from families in which several members, including at least one parent and child, had unusually high copy numbers of HHV-6 DNA per milliliter of blood. FISH confirmed that HHV-6 DNA colocalized with telomeric regions of one allele on chromosomes 17p13.3, 18q23, and 22q13.3, and that the integration site was identical among members of the same family. Integration of the HHV-6 genome into TTAGGG telomere repeats was confirmed by additional methods and sequencing of the integration site. Partial sequencing of the viral genome identified the same integrated HHV-6A strain within members of families, confirming vertical transmission of the viral genome. We next asked whether HHV-6A infection of naïve cell lines could lead to integration. Following infection of naïve Jjhan and HEK-293 cell lines by HHV-6, the virus integrated into telomeres. Reactivation of integrated HHV-6A virus from individuals’ PBMCs as well as cell lines was successfully accomplished by compounds known to induce latent herpesvirus replication. Finally, no circular episomal forms were detected even by PCR. Taken together, the data suggest that HHV-6 is unique among human herpesviruses: it specifically and efficiently integrates into telomeres of chromosomes during latency rather than forming episomes, and the integrated viral genome is capable of producing virions.

Chromosomally integrated human herpesvirus 6: questions and answers

Chromosomally integrated human herpesvirus 6 (ciHHV‐6) is a condition in which the complete HHV‐6 genome is integrated into the host germ line genome and is vertically transmitted in a Mendelian manner. The condition is found in less than 1% of controls in the USA and UK, but has been found at a somewhat higher prevalence in transplant recipients and other patient populations in several small studies. HHV‐6 levels in whole blood that exceed 5.5 log10 copies/ml are strongly suggestive of ciHHV‐6. Monitoring DNA load in plasma and serum is unreliable, both for identifying and for monitoring subjects with ciHHV‐6 due to cell lysis and release of cellular DNA. High HHV‐6 DNA loads associated with ciHHV‐6 can lead to erroneous diagnosis of active infection. Transplant recipients with ciHHV‐6 may be at increased risk for bacterial infection and graft rejection. ciHHV‐6 can be induced to a state of active viral replication in vitro. It is not known whether ciHHV‐6 individuals are put at clinical risk by the use of drugs that have been associated with HHV‐6 reactivation in vivo or in vitro. Nonetheless, we urge careful observation when use of such drugs is indicated in individuals known to have ciHHV‐6. Little is known about whether individuals with ciHHV‐6 develop immune tolerance for viral proteins. Further research is needed to determine the role of ciHHV‐6 in disease. Copyright

Elevated antibody titers to Epstein-Barr virus in Hodgkin's disease.

Sera from 63 patients with Hodgkins disease and 42 patients of comparable age with other lymphomas were tested for antibody to Epstein‐Barr virus (EBV) by indirect immunofluorescence. The geometric mean titer (GMT) of EBV antibody in the patients with Hodgkins disease was significantly higher (1:367) than the GMT of the other lymphoma patients (1:132) and 85 normal controls (1:90). Higher EBV titers in untreated patients with Hodgkins disease were associated with a longer duration of symptoms, more advanced disease, shorter survival, and a histologic picture of lymphocyte depletion. Treated patients had significantly higher titers than untreated patients. When the same sera were tested for antibody to 4 other herpes viruses (herpes simplex type I and type II, cytomegalovirus, and varicella), no differences in titer between patient and control groups were found. Although this study associates elevated EBV titers with those factors relating to a poor prognosis in patients with Hodgkins disease, the data do not distinguish between an etiologic role for EBV and that of a passenger virus which produces high titers as a result of the disease process.

Seroprevalence of human herpesvirus-8 (HHV-8) in countries of Southeast Asia compared to the USA, the Caribbean and Africa

Seroprevalence of HHV-8 has been studied in Malaysia, India, Sri Lanka, Thailand, Trinidad, Jamaica and the USA, in both healthy individuals and those infected with HIV. Seroprevalence was found to be low in these countries in both the healthy and the HIV-infected populations. This correlates with the fact that hardly any AIDS-related Kaposi’s sarcoma has been reported in these countries. In contrast, the African countries of Ghana, Uganda and Zambia showed high seroprevalences in both healthy and HIV-infected populations. This suggests that human herpes virus-8 (HHV-8) may be either a recently introduced virus or one that has extremely low infectivity. Nasopharyngeal and oral carcinoma patients from Malaysia, Hong Kong and Sri Lanka who have very high EBV titres show that only 3/82 (3.7%) have antibody to HHV-8, demonstrating that there is little, if any, cross-reactivity between antibodies to these two gamma viruses.

Human B-lymphotropic virus (human herpesvirus-6).

Human B-lymphotropic virus (HBLV), also known as human herpesvirus-6 (HHV-6) was first isolated in 1986 from AIDS patients and patients with other lymphoproliferative disorders. HBLV is distinct from known human herpesviruses, biologically, immunologically and by molecular analysis. HBLV can infect and replicate in fresh and established lines of hemopoietic cells and cells of neural origin, suggesting wide tropism. The prevalence of HBLV antibody in the normal population was 26% though clear differences between different populations were observed. The prevalence of HBLV antibody an elevated antibody titer was higher in sera from certain malignancies, Sjögrens syndrome and sarcoidosis. Antibody to HBLV was also elevated in AIDS patients and patients with chronic fatigue syndrome. HBLV-DNA was detected in some B-cell lymphomas. The broad in vitro tropism, combined with immunological and molecular evidence of HBLV infection in individuals raise the question of the pathogenicity of this virus in some diseases. Because in vitro co-infection of CD4 cells by HBLV and HIV leads to enhanced degeneration, this raises the possibility that infection in AIDS patients by both viruses can aggravate the HIV-induced immunodeficiency. Specific reagents and immunological and molecular assays are currently being investigated, which will aid in virus detection in cells from patients, and in elucidating the possible pathogenesis of HBLV.

Multicenter comparison of serologic assays and estimation of human herpesvirus 8 seroprevalence among US blood donors

BACKGROUND:  As part of assessing the possibility of transfusion transmission of human herpesvirus 8 (HHV‐8 or Kaposis sarcoma‐associated herpesvirus), HHV‐8 seroprevalence was estimated among US blood donors, the performance of HHV‐8 serologic tests was compared, and the presence of HHV‐8 DNA was tested for in donated blood.

Multicenter Comparison of PCR Assays for Detection of Human Herpesvirus 8 DNA in Semen

ABSTRACT Human herpesvirus 6 (HHV-6) is a ubiquitous virus with which infections have been associated with pathologies ranging from delayed bone marrow engraftment to a variety of neurological diseases. The lack of a standardized assay that can be used to detect and estimate HHV-6 DNA contents in various clinical specimens can lead and has led to discordant results among investigators and on the potential association of HHV-6 to diseases. To identify the most reliable and sensitive assays, an identical set of 11 coded serum samples spiked with various quantities of the HHV-6A variant (range, 4 to 400,000 genome copies/ml) was sent to eight independent laboratories around the world. Each laboratory was asked to estimate the HHV-6 DNA content by use of its own protocols and assays. Among the various assays, three TaqMan-based real-time PCR assays yielded quantities that were closest to the quantity of HHV-6 that had been spiked. To provide better homogeneity between the results from the different laboratories working on HHV-6, we propose that investigators interested in quantifying HHV-6 in clinical samples adopt one of these assays.

Classification of HHV-6A and HHV-6B as distinct viruses

Shortly after the discovery of human herpesvirus 6 (HHV-6), two distinct variants, HHV-6A and HHV-6B, were identified. In 2012, the International Committee on Taxonomy of Viruses (ICTV) classified HHV-6A and HHV-6B as separate viruses. This review outlines several of the documented epidemiological, biological, and immunological distinctions between HHV-6A and HHV-6B, which support the ICTV classification. The utilization of virus-specific clinical and laboratory assays for distinguishing HHV-6A and HHV-6B is now required for further classification. For clarity in biological and clinical distinctions between HHV-6A and HHV-6B, scientists and physicians are herein urged, where possible, to differentiate carefully between HHV-6A and HHV-6B in all future publications.

Isolation of the Human T-Cell Leukemia/Lymphotropic Virus Type III From the Cornea

Corneoscleral donor tissue from a donor with a positive serum antibody to HTLV-III but without the overt clinical signs of the acquired immune deficiency syndrome (AIDS) was cultured for the presence of the human T-cell leukemia/lymphotropic virus type III (HTLV-III). The virus was isolated from the two corneal specimens in this patient after the tissue had been stored for four days in McCarey-Kaufman medium. The presence of HTLV-III was confirmed by the detection of viral core proteins (approximately 24,000 protein, termed P24 gag), by immunofluorescence of a touch preparation of the corneal epithelium as well as in cells cultured in vitro. The percentage of immunofluorescent cells detected by HTLV-III anti-P24 antibody ranged between 2% and 3%. These findings emphasize the possibility of transmission of this virus via corneal transplantation surgery. Although no documented cases of AIDS have occurred in corneal transplant recipients, serologic screening of donors before the use of the tissue for transplantation is advisable.