Viral safety of a new highly purified factor VIII (OCTATE).

The inactivation of both transfusion-relevant and model viruses by modified pasteurisation (10 hours at 63 degrees C in solution) has been evaluated following the established guidelines of the EU CPMP Ad Hoc Working Party on Biotechnology/Pharmacy. This heat treatment was introduced into the manufacturing process of OCTAVI, a very high purity factor VIII concentrate stabilized only by von Willebrand factor, in the presence of a proprietary mixture of low molecular weight stabilizers. Both enveloped (human immunodeficiency virus, Sindbis virus, herpes simplex virus, pseudorabies virus) and nonenveloped viruses (poliovirus, Coxsackievirus, hepatitis A virus) were inactivated by this heating step by more than 4.7 log10. The combination of the solvent/detergent step used in the manufacture of OCTAVI with this modified pasteurization leads to a double virus-inactivated factor VIII concentrate (OCTATE) with a viral safety distinctly superior to monoinactivated products.

Influenza vaccines.

This is a report about influenza vaccines. Recommendations for the 1995/1996 influenza season vaccine components are cited. There are many different types of influenza vaccine, but only inactivated vaccines are licensed for parenteral administration in humans. Experimental live vaccines have been developed and evaluated for their usefulness in the immunoprophylaxis of influenza in humans, especially for those at high risk of death from influenza virus infection. The current inactivated vaccines do not provide complete protection.

Influenza surveillance: experiences from establishing a sentinel surveillance system in Germany.

OBJECTIVES: Before and during peak influenza periods there is increased morbidity from other respiratory tract disorders. Sentinel networks of primary care physicians can be very effective in the early detection of influenza epidemics and the German network, the Arbeitsgemeinschaft Influenza (AGI), began its work in this area in 1992. METHODS: Data are transmitted weekly from the doctor's computer via Btx to a central computer. The numerator is the weekly number of acute respiratory infections (ARI) in five age groups and the denominator is the weekly number of patient consultations. Data on hospitalisation, mortality, and days of sick leave from work or school are also collected. Swabs for influenza specimens are collected in 30 physicians' offices each Monday and sent to three reference centres. FINDINGS: During the last recording period, from week 46 1993 to week 15 1994, 411 physicians' offices participated in the network. For 16 to 22 weeks, more than 60% of the participants transmitted data. During both the 1992-93 and 1993-94 influenza seasons, peaks were observed in the rate of ARI. There was a corresponding increase in sick leave from work and school. Rates for hospitalisation and deaths due to influenza showed no peaks during either season. CONCLUSIONS: Although the German sentinel network for influenza experienced some technical problems in the first year, it was possible to solve these. Reporting rates were very satisfactory in the second year. The network will now be expanded to include 750 physicians in order to receive 600 weekly reports and obtain a solid baseline for an early warning system.

High proportion of HIV-2 and HIV-1/2 double-reactive sera in two Indian states, Maharashtra and Goa: first appearance of an HIV-2 epidemic along with an HIV-1 epidemic outside of Africa.

The seroprevalence of HIV-1 and HIV-2 infections was determined in selected groups of patients from an STD clinic and in female prostitutes from two different states of India. A total of 809 sera from Maharashtra and of 61 sera from Goa were analyzed. Of the unscreened sera from Bombay, 39% were positive as were 15% of the sera from Goa. Among the HIV-positive sera from Bombay, 79% were positive for HIV-1, 4% were positive for HIV-2 and 17% were positive for HIV-1 and 2. In the Goa group, 67% of the infected sera were HIV-1-positive and 33% were HIV-2 positive. Due to the nearly even distribution of infections between men and women, a mainly heterosexual mode of transmission is evident. HIV-2 infections were first detected in India in 1990 (Rübsamen et al., Lancet, 1991). Before that time, Asia was believed to be free HIV-2. The present study represents the first documentation of a large proportion of these infections, alone and in combination with HIV-1, in the Asian continent. An HIV-2 epidemic running parallel to an HIV-1 epidemic so far has never been observed outside of Africa. This would indicate that a further spread of HIV-2 worldwide is to be expected and that HIV-2 should be included in programs of vaccine development for humans.

HIV-1 and HIV-2 infections in a high-risk population in Bombay, India: evidence for the spread of HIV-2 and presence of a divergent HIV-1 subtype.

A high-risk population (patients of a sexually transmitted disease clinic and the GT hospital in Bombay) was tested for antibodies against HIV-1 and HIV-2. Among 405 serum samples, 226 had previously been classified HIV-positive in India using different locally available enzyme-linked immunosorbent assay (ELISA) tests. The serology of 179 samples was unknown. All 405 samples were tested at the Georg-Speyer-Haus (GSH) with the Pasteur HIV-1/2-Combi-ELISA. Positive samples were further analyzed with HIV-1 and HIV-2 Western blot kits from Dupont and Pasteur, respectively. A very high seroprevalence of HIV was found in this population. Among the 179 unscreened samples, 69 (38.5%) were positive in the ELISAs as well as the Western blots for HIV-1 or HIV-2. Among the prescreened samples, only 174 (77%) were confirmed HIV-positive. Altogether, 243 of 405 sera were HIV-positive. Of these, 184 (76%) were reactive with HIV-1, 10 (4%) were reactive with HIV-2, and 49 (20%) had dual reactivity to HIV-1 and HIV-2. Previous data from the Indian Council of Medical Research had already suggested a possible high prevalence of HIV-1 in India. Our results confirm this view. The finding of a substantial spread of HIV-2 infection was, however, totally unexpected in India, but confirms our previous study which had already demonstrated the existence of HIV-2 in this country. Asia can thus no longer be considered free of HIV-2, and testing for HIV-2 appears mandatory, at least in India.(ABSTRACT TRUNCATED AT 250 WORDS)

Markers for HIV-disease progression in untreated patients and patients receiving AZT: evaluation of viral activity, AZT resistance, serum cholesterol, beta 2-microglobulin, CD4+ cell counts, and HIV antigen.

In order to find parameters which allow the assessment of the clinical state of HIV patients with or without antiviral therapy, viral cultures on lymphocytes and monocytes/macrophages, CD4-cell counts, HIV antigen, beta 2-microglobulin and serum cholesterol were evaluated for their predictive value. As had been shown previously for lymphocytes, the efficiency of viral isolation on macrophages also depends on the disease stage (CDC) of the patients and thus has a high predictive value. A multivariant discriminant analysis showed that the combination of beta 2-microglobulin, viral antigen, CD4+ cell count and HDL cholesterol predicted the outcome of viral cultures with 80% accuracy. While viral antigen, CD4+ cell counts and beta 2-microglobulin had been known, HDL cholesterol deserves further evaluation as prognostic parameter. The analysis of HIV derived from patients with AZT showed a 20-200-fold in vitro drug resistance after seven to 24 months of therapy. DNA sequence determination of such strains isolated from AZT patients over time showed only two of the amino acid exchanges described in the literature for resistant strains and an additional Val60-Ile transition after 32 months of therapy.

Isolation frequency and growth properties of HIV-variants: multiple simultaneous variants in a patient demonstrated by molecular cloning.

The biological properties and efficiency of isolation of different HIV (LAV/HTLV III, ARV, and AAV) subtypes were evaluated by recovering and growing HIV on fresh peripheral human lymphocytes. Cultures for virus isolation were performed from more than 180 German AIDS, ARC, LAS, and virus-exposed asymptomatic patients. The virus isolation rate depended on the state of health of the patients being close to 80% in AIDS patients, 30-40% in ARC/LAS patients, and lower in asymptomatic HIV seropositive patients. The cytopathic effects of the HIV isolates obtained on lymphocyte-cell cultures ranged from no effect to marked syncytia formation and cytopathogenicity. Marked differences were also observed in the replication rate of the various isolates. These properties were stable in all in vitro passages of the viruses performed so far and allowed to tentatively define four subtypes of HIV. In the majority of AIDS cases with neurological symptoms well-growing strains were obtained from peripheral blood, while all but two isolates from the cerebrospinal fluid of the same patients grew remarkably slowly and to only low titres on lymphocytes, suggesting that selection of variants for growth at specific sites of the body occurs. For one of the most cytopathogenic strains the influence of several variables of culture conditions (cell type, corticosteroids, IL-2, and polybrene) on virus replication was studied. Apart from polybrene, all parameters strongly influenced replication.

Humoral immune response after hepatitis-B-vaccination: kinetics of anti-HBs antibodies and demonstration of HLA antigens.

Three injections monthly of 50 micrograms/ml hepatitis-B-vaccine (Hevac-B-Pasteur) and one booster injection after 12 months from the beginning of vaccination were given to 676 adult recipients. Anti-HBs was detected in 45.9 percent of persons one month after the first dose of vaccine, in 79.3 percent after the third dose and in 98.4 percent after the booster injection. A follow-up of 142 persons 3 years after the first dose of vaccine indicated that protective levels of anti-HBs were maintained in 95.1 per cent of all participants. HLA antigens were typed in 13 vaccine recipients with low anti-HBs titers (less than or equal to 10 ImU/ml) and in 26 recipients with high anti-HBs titers (greater than or equal to 20,000 ImU/ml). Significant differences of antigen frequency (p less than 0.05) were found for HLA A3 (decrease in high responders), and HLA DR7 (increase of in low, decrease in high responders). The results indicate genetic differences between high and low responding recipients of HBV vaccine.

Isolation of variants of lymphocytopathic retroviruses from the peripheral blood and cerebrospinal fluid of patients with ARC or AIDS.

LAV/HTLV-III/AAV viruses were isolated from 20 German patients with ARC/AIDS in order to investigate strain variation. Virus was isolated from the peripheral blood and/or cerebrospinal fluid (CSF) in umbilical cord peripheral blood lymphocyte (PBL) cultures. Isolates were identified by their cytopathic effect (CPE), by reverse transcriptase assays on cell-free infected culture supernatant fluid (SNF), and one or more of the following: immunofluorescence assays on infected cells for viral antigen using HTLV-III reference sera, Western blot analysis of cell-free infected culture SNF, electron microscopy of infected cells, and Southern blot restriction analysis and specific HTLV-III probing of DNA extracted from infected cultured PBL. The isolates could be classified into three groups according to differences in growth rate and cytopathic effect: Most showed what was regarded as the typical CPE, while some either grew rapidly and induced a striking CPE and others grew slowly with minimal CPE. In one patient, virus producing typical CPE was isolated from the peripheral blood while the isolate from his filtered cell-free CSF produced atypical slow CPE, suggesting that antigenic variation may occur with persistent infection or that superinfection may occur. Southern blot DNA restriction analysis of the DNA of three selected isolates showed that two of the isolates were similar but that the restriction pattern of all three differed from patterns previously published. Our results supplement the accumulating evidence of genetic variation among LAV/HTLV-III strains. The extent of this variation needs to be evaluated for any effect on the sensitivity of diagnostic tests, on the strategy of vaccine development, on tissue tropism by altering the viral surface receptor-binding sites, and possibly on the development of specific chemotherapy.

Detection of antibodies to HTLV-III in commercially available immunoglobulin preparations.

Samples of 96 polyvalent and virus-specific immunoglobulin batches commercially available in West-Germany were tested by enzyme-linked immunoassay and immunoblot for the presence of anti-HTLV-III antibodies. 37% of the polyvalent and 87% of the virus-specific batches were positive. It was concluded that these preparations are still safe and because of their reportedly low titer neutralizing antibodies possibly beneficial in certain cases, such as newborns of HTLV-III positive mothers or after accidental exposure to infectious material in clinics or laboratories.

Seroepidemiology of HTLV-III (LAV) in the Federal Republic of Germany.

In 1984 10,281 sera were collected in the FRG and examined for antibodies to HTLV-III (LAV) with an enzyme-linked immunosorbent assay and confirmative tests. Of the German AIDS patients 81% have antibodies. Individuals belonging to AIDS risk groups, homosexuals, haemophiliacs and i.v. drug abusers, have antibody frequencies between 25%-72%. The detection of HTLV-III antibodies in blood donors indicates that the virus is being transmitted by blood transfusions.

Serum antibodies to human T-cell leukaemia virus type I in different ethnic groups and in non-human primates in South Africa.

The prevalence of humoral antibodies to human T-cell leukaemia virus type I (HTLV-I) was investigated in different ethnic groups and in non-human primates in South Africa. Serum antibody levels were determined by enzyme-linked immunosorbent assay (ELISA) using either disrupted whole HTLV-I or purified p24 core protein (p24 HTLV-I) as antigens. ELISA was complemented by direct radio-immunoprecipitation assays using either purified iodinated p24 HTLV-I or radiolabelled lysates of an HTLV-producing cell line as antigen followed by sodium dodecyl sulphate polyacrylamide gel electrophoresis of the immunoprecipitates, and by immunofluorescence using the HTLV-I-producing cell line HUT-102 as antigen. Antibodies were demonstrated in 3,5% of Asians, 3,5% of blacks and 4,1% of coloureds, but not in whites, and also in 29% of vervet monkeys and 33% of baboons. We conclude that HTLV-I or closely related viruses cause widespread infection in non-human primates in South Africa and in a lower percentage of humans, including apparently healthy blood donors. We are currently isolating retroviruses from seropositive reactors and investigating the possible relevance to disease in South Africa.