Seroepidemiology of Kaposi's sarcoma-associated herpesvirus (KSHV).

Since the Kaposi's sarcoma-associated herpesvirus (KSHV also referred to as HHV-8, human herpesvirus-8) was discovered it has been shown that the virus is associated with all cases of Kaposi's sarcoma (KS) classical, endemic, or AIDS associated. In the numerous countries where the seroprevalence of this virus has been studied, data demonstrate that the virus is not ubiquitous in general healthy human populations as is the case with other human herpesviruses. Many seroprevalence studies to detect antibodies to HHV-8 have now been conducted using a variety of immunologic techniques. While these assays are not in total agreement and may overstate or understate the positivity of sera in the general population, they all show similar general antibody trends. For general populations the seroprevalence in sub-Saharan Africa is the highest, approximately 40% positive; in Mediterranean countries the seroprevalence is approximately 10%; whereas northern European, southeast Asian, and Caribbean countries have seroprevalence rates in the 2-4% range. In the United States, a 'mixing bowl' country the seroprevalence is in the range of 5-20%. In people with KS whether AIDS associated, classical, or endemic and other HHV-8 associated diseases such as multicentric Castleman's disease and certain body cavity lymphomas (BCL), also called primary effusion lymphoma (PEL) the seroprevalency rates are >90%. In populations with HIV-1 infection but no diagnosis of KS, the seroprevalency rates are elevated (20-50%) above those in the general population except in southeast Asia and the Caribbean where no AIDS associated KS has been reported. No correlation has been found between the presence of KSHV antibodies and other malignancies.

Propagation and characterization of human herpesvirus-7 (HHV-7) isolates in a continuous T-lymphoblastoid cell line (SupT1).

After initial culture of HHV-7 in PHA-stimulated human cord blood mononuclear cells (HCBMC), six HHV-7 isolates were propagated successfully in an immature continuous T-lymphoblastoid cell line SupT1. All six isolates infected efficiently the SupT1 cells, and the infected cells became grossly enlarged and multinucleated 7-21 days post-infection. Various stages of HHV-7 morphogenesis were detected. Cell-free supernatants from HHV-7-infected SupT1 cells were infectious to HCBMC as well as to SupT1 cells. The HHV-7-infected SupT1 and HCBMC cell lysates contained more infectious virus than the centrifuged cell culture fluid supernates from the same culture. The HHV-7 isolates H7-2, H7-3, JHC, and JB, concentrated 500 times, had average infectivity titers of 10(3.0) TCID50/ml while strains H7-4 and KHR titered approximately 1-2 logs higher. When all six HHV-7 isolates were propagated in SupT1 and culture fluid supernatants were examined 14-21 days post-infection by negative stain electron microscopy they contained an average of 1.9 x 10(9) virus particles/liter. IFA and ELISA, using HHV-7/SupT1 cell lysate as an antigen, seem to correlate well in detecting high and low HHV-7 antibody in sera from chronic fatigue patients and healthy donors as controls. HHV-7 from SupT1 cell culture was free of HHV-6 and other human herpesviruses as tested by PCR, and the HHV-7 PCR signal was still strong when the viral preparation was diluted to 4.82 x 10(2) genome copies. Since HCBMC are expensive to obtain and available in only small amounts, it is difficult to obtain large quantities of HHV-7 antigen. On the other hand, the SupT1 cell is an excellent source to produce consistently sufficient quantities of HHV-7 for purification studies, development of immunodiagnostics, in vivo infectivity studies, evaluation of antiviral drugs, and molecular biological studies.

Detection and titration of human herpesvirus-8-specific antibodies in sera from blood donors, acquired immunodeficiency syndrome patients, and Kaposi's sarcoma patients using a whole virus enzyme-linked immunosorbent assay.

A human herpesvirus-8 (HHV-8) enzyme-linked immunosorbent assay (ELISA) with a whole virus lysate as antigen was developed and used to measure the seroprevalence rate and levels of IgG antibodies to HHV-8 in sera/plasma of various patient groups and blood donors. The virus antigen was prepared from the KS-1 cell line, which produces lytic virus, and therefore contains a broad array of viral proteins. Seroprevalence studies using this ELISA showed the following: 10 of 91 blood donors (11%) had an average HHV-8 antibody titer of 118; 67 of 72 (93%) classic Kaposi's sarcoma (KS) patients were positive with an average titer of 14,111; and 57 of 62 (92%) KS/human immunodeficiency virus (HIV) patients were positive with an average titer of 4,000. A study on a very limited number of serial serum samples from patients before and after diagnosis with KS showed highly elevated antibody titers to HHV-8 virus after KS lesions developed. Preliminary data show that 50% of the sera from HIV-1(+) homosexual patients contain IgG antibodies to HHV-8 suggesting that this population is at high risk for developing KS. Antibody results correlated well with the confirmatory immunofluorescent assays (IFA) using KS-1 cells as the substrate. This HHV-8 IgG antibody detection ELISA is sensitive and specific and does not cross-react with Epstein-Barr virus (EBV) or other human herpesviruses. The results of this HHV-8 antibody survey suggest that this rapid ELISA assay can be used to screen large numbers of sera to find those at risk for developing KS.

Longitudinal follow-up of two AIDS patients for active human herpesvirus-6 (HHV-6) infection.

Because of the demonstrated presence of Human Herpesvirus-6 (HHV-6) in various tissues and organs of HIV-1 infected AIDS patients, and its implication in the pathogenesis of AIDS, we followed two AIDS patients for active HHV-6 infection for two years. Evidence of active HHV-6 infection was demonstrated by immunovirologic assays in both patients; however, the profile of infection in the two patients varied. One patient showed appearance and disappearance of HHV-6, indicating virus reactivation, whereas the other patient showed chronic or persistent HHV-6 infection. Both patients' peripheral blood mononuclear cells (PBMC) contained HIV-1 when their CD4 cell count was approximately 200 mm3. No active HHV-6 was demonstrated in a healthy donor, who was used as a control for this study.

Inhibition of the human immunodeficiency virus-1 protease and human immunodeficiency virus-1 replication by bathocuproine disulfonic acid Cu1+.

The protease encoded by the human immunodeficiency virus-1 (HIV-1) is essential for processing viral polyproteins which contain the enzymes and structural proteins required for the infectious virus. It was previously found that cupric chloride, in the presence of dithiothreitol or ascorbic acid, could inhibit the HIV-1 protease. It was suggested that a Cu1+ chelate was the moiety responsible for inhibition of the protease. This hypothesis has now been investigated directly by utilizing the stable Cu1+ chelate, bathocuproine disulfonic acid Cu1+ (BCDS-Cu1+). BCDS-Cu1+ inhibited the HIV-1 wild type protease as well as a mutant HIV-1 protease lacking cysteines. BCDS-Cu1+ was a competitive inhibitor of the mutant HIV-1 protease with an apparent Ki of 1 microM. Replication of HIV-1 in human lymphocytes and the cytotoxic effect of HIV-1 in CEM cells was inhibited by micromolar BCDS-Cu1+. Inhibition of the protease and of HIV replication by BCDS-Cu1+ was dependent on the presence of Cu1+ as BCDS alone was ineffective. EDTA blocked the inhibition of the protease by Cu1+ but was unable to block inhibition of the protease by BCDS-Cu1+, indicating that the Cu1+ complex was the inhibitory agent. The apparent IC50 for BCDS-Cu1+ on the inhibition of replication by primary isolates of HIV-1 was 5 microM. However, BCDS-Cu1+ did not affect polyprotein processing in an H9 cell line chronically infected with HIV-1, indicating that BCDS-Cu1+ acts by yet another mechanism to block HIV infection. Other possible targets for BCDS-Cu1+ include inhibition of viral adsorption and/or inhibition of the HIV-1 integrase.