David J. Blackbourn

Antibodies to human herpesvirus type 8 in the general population and in Kaposi's sarcoma patients

BACKGROUND Much of the evidence that human herpesvirus type 8 (HHV-8) is associated with Kaposis sarcoma (KS) has come from molecular studies of HHV-8 DNA. Seroepidemiological studies have been hampered by the lack of a reliable assay. METHODS The serological data reported here were obtained by means of a mouse monoclonal antibody-enhanced immunofluorescence assay for antibodies to lytic and latent HHV-8 antigens. 1435 single samples of serum (or plasma) from many different disease groups and parts of the world were assayed. FINDINGS All patients with African endemic KS and 96% of American patients with AIDS-associated KS were seropositive for lytic antigen, as were 90% of American HIV-infected homosexual men; by contrast only 23% of HIV-seropositive drug users and 21% of HIV-seropositive women were positive for HHV-8 antibody. Factor VIII treatment before 1983 did not increase the risk of HHV-8 infection in patients with haemophilia. In the American general population, about 25% of adults (including volunteer blood donors) and 2-8% of children had antibodies to HHV-8. INTERPRETATION Our data are consistent with HHV-8 being primarily associated with sexual transmission, but the HHV-8 seropositivity rate in American children suggests that there is a non-sexual route of HHV-8 infection also. On the evidence available so far, the risk of parenteral transmission is low.

Infectious human herpesvirus 8 in a healthy North American blood donor

BACKGROUND Molecular studies have provided strong evidence for the association of human herpesvirus 8 (HHV-8) with Kaposis sarcoma. These data have been supported by serological studies, which have also suggested that HHV-8 can be found in the healthy population. We report the presence of infectious HHV-8 in a healthy donor to a North American blood bank. METHODS We examined the peripheral blood mononuclear cells or CD19 cells of blood donors by PCR for evidence of HHV-8 infection. The CD19 cells were separated from peripheral blood mononuclear cells by immunomagnetic-bead selection. To enhance detection of HHV-8, the CD19 cells from eleven unsystematically selected blood donors were activated with phorbol ester and recombinant interleukin-6; the culture fluid was filtered and inoculated onto HHV-8-negative target CD19 cells that had been prepared from phytohaemagglutinin-stimulated peripheral blood mononuclear cells. These inoculated target cells were cultured for 3 days and then analysed for HHV-8 sequences by PCR. Serum samples were tested for antibodies to HHV-8 by an indirect immunofluorescence assay. FINDINGS One blood donor was consistently found to be infected with HHV-8 by PCR after the cell-culture activation procedure. He was seropositive for the virus. The HHV-8 recovered was infectious, as shown by a reverse-transcription-PCR technique that detected HHV-8 RNA in the inoculated target cells. INTERPRETATION These data provide the first indication that HHV-8 can be recovered from the blood of a healthy individual, a blood donor, and that the virus is infectious. This observation suggests that HHV-8 could be transmitted by blood transfusion, a possibility that merits further study.

Human herpesvirus 8 detection in nasal secretions and saliva

The presence of human herpesvirus 8 (HHV-8) was determined by polymerase chain reaction (PCR) in nasal secretions and saliva from 14 HHV-8-seropositive persons, including 8 Kaposis sarcoma patients: 7 were human immunodeficiency virus type 1-infected, 6 of whom were asymptomatic. HHV-8 was detected in one or both body fluids in 8 (57%) of 14 subjects. Parallel PCR testing revealed the concomitant presence of cytomegalovirus, Epstein-Barr virus, and HHV-6 in various combinations in these body fluids. These data indicate frequent shedding of multiple herpesviruses in nasal secretions and saliva, particularly in Kaposis sarcoma patients. Both body fluids are therefore potential sources HHV-8 by nonsexual transmission.

The restricted cellular host range of human herpesvirus 8

DesignA selection of primary and transformed cell types were evaluated for their susceptibility to infection with human herpesvirus 8 (HHV-8)/Kaposis sarcoma- associated herpesvirus. MethodsSources of HHV-8 included Kaposis sarcoma lesion punch biopsies that were either cocultured directly with target cells or that were first cocultured with human lymphocytes to derive HHV-8-containing fluids that were inoculated onto target cells. HHV-8 was also obtained from primary effusion lymphoma-derived cell lines. Techniques to detect infection included the PCR, immunofluorescence assays and in situ hybridization. ResultsSusceptible cells included human umbilical cord blood mononuclear cells (UCMC), adult CD19 B cells, macrophages and certain endothelial cells of human and animal origin, including some that are transformed with human papilloma virus type 16 E6 and E7 genes. The infection of lymphocytes did not yield established lymphoblastoid cell lines (LCL) and virus infection persisted for only 4–7 days. However, long-term HHV-8 infection of UCMC could be achieved by coinfection with Epstein–Barr virus. HHV-8 could also infect UCMC LCL recently derived by Epstein–Barr virus transformation, but long-established LCL could not be infected with HHV-8. ConclusionsThese data provide further biological evidence in cell culture for the limited cellular host range of HHV-8 to CD19 B cells, macrophages, and certain endothelial cells.

CD8 Cell Noncytotoxic Antiviral Activity in Human Immunodeficiency Virus-Infected and -Uninfected Children

CD8 cells from human immunodeficiency virus (HIV)-infected adults and children can show cytotoxic as well as noncytotoxic activity against viral replication. The noncytotoxic anti-HIV response, measured by suppression of acute viral infection of CD4 cells, has also been observed in uninfected adults who have a history of exposure to HIV. This CD8 cell antiviral activity was found to be detectable as well in approximately 50% of uninfected children born of infected mothers. The findings could reflect a protective response of the children to HIV after being exposed to the virus.

Suppression of human immunodeficiency virus type 1 replication by a soluble factor produced by CD8+ lymphocytes from HIV-2-infected baboons.

Human immunodeficiency virus type 2 (HIV-2)-infected baboons (Papio cynocephalus) provide a valuable animal model for the study of acquired immunodefidency syndrome (AIDS) pathogenesis since many features of disease progression resemble HIV-1-infection of humans. In some HIV-2-infected baboons that are clinically healthy, a CD8+ cell antiviral response, that is partly mediated by a soluble factor, controls viral replication in vitro. In the present study, we demonstrate that CD8+ cells derived from HIV-2-infected baboon peripheral blood, lymph nodes, adenoids and tonsils had antiviral activity in co-cultures of CD8+ and CD4+ cells that inversely correlates with viral load. A soluble factor was found to be active against the chemokine-resistant, syncytium-inducing HIV-1SF2 and HIV-1SF33 isolates and was relatively heat stable at 100 degrees C for 10 min. Moreover, inhibition of the transcription from the long terminal repeat of HIV-1 was observed in 1G5 cells after activation with phorbol 12-myristate 13-acetate. Therefore, the soluble suppressing activity of CD8+ cells in HIV-2-infected baboons may be analogous to the CD8+ cell antiviral factor described in human HIV-infected asymptomatic people.

Immune responses in baboons vaccinated with HIV‐2 genetic expression libraries

Abstract: Immunization using genetic expression libraries may be an improvement over conventional DNA immunization using a single gene because more epitopes are simultaneously presented to the immune system. In this study, we evaluated the effectiveness of an HIV‐2 vaccine made from a genomic expression library in baboons. We found that HIV‐2 expression library immunization induced HIV‐2‐specific memory responses but low levels of CD8+ cell anti‐viral responses and neutralizing antibodies. After intravenous virus challenge using a homologous pathogenic variant, HIV‐2UC2/9429, viral loads were similar in the HIV‐2‐immunized and control baboons. We conclude that although immunization using HIV‐2 expression libraries induces immune responses, this approach does not provide protection in baboons against intravenous challenge with HIV‐2.