Human neutralizing monoclonal antibodies of the IgG1 subtype protect against mucosal simian-human immunodeficiency virus infection.

Although maternal human immunodeficiency virus type 1 (HIV-1) transmission occurs during gestation, intrapartum and postpartum (by breast-feeding), 50-70% of all infected children seem to acquire HIV-1 shortly before or during delivery. Epidemiological evidence indicates that mucosal exposure is an important aspect of intrapartum HIV transmission. A simian immunodeficiency virus (SIV) macaque model has been developed that mimics the mucosal exposure that can occur during intrapartum HIV-1 transmission. To develop immunoprophylaxis against intrapartum HIV-1 transmission, we used SHIV-vpu+ (refs. 5,6), a chimeric simian-human virus that encodes the env gene of HIV-IIIB. Several combinations of human monoclonal antibodies against HIV-1 have been identified that neutralize SHIV-vpu+ completely in vitro through synergistic interaction. Here, we treated four pregnant macaques with a triple combination of the human IgG1 monoclonal antibodies F105, 2G12 and 2F5. All four macaques were protected against intravenous SHIV-vpu+ challenge after delivery. The infants received monoclonal antibodies after birth and were challenged orally with SHIV-vpu+ shortly thereafter. We found no evidence of infection in any infant during 6 months of follow-up. This demonstrates that IgG1 monoclonal antibodies protect against mucosal lentivirus challenge in neonates. We conclude that epitopes recognized by the three monoclonal antibodies are important determinants for achieving substantial protection, thus providing a rational basis for AIDS vaccine development.

Interaction of dendritic cells with skin endothelium: A new perspective on immunosurveillance.

The goal of this study was to determine the mechanisms by which dendritic cells (DCs) in blood could interact with endothelium, a prerequisite to extravasation into tissues. Our results indicate that DCs express both HECA-452-reactive and nonreactive isoforms of P-selectin glycoprotein ligand 1 (PSGL-1) and can tether and roll efficiently on E- and P-selectin under flow conditions in vitro. Freshly isolated blood DCs were further observed to roll continuously along noninflamed murine dermal endothelium in vivo. This interaction is strictly dependent on endothelial selectins, as shown by experiments with blocking antibodies and with E- and P-selectin-deficient mice. We hypothesize that DCs in blood are constitutively poised at the interface of blood and skin, ready to extravasate upon induction of inflammation, and we showed that cutaneous inflammation results in a rapid recruitment of DCs from the blood to tissues. We propose that this is an important and previously unappreciated element of immunosurveillance.

Inhibition of HIV-1 replication by an aqueous extract of Spirulina platensis (Arthrospira platensis).

An aqueous extract of the blue-green filamentous algae Arthrospira platensis (previously called Spirulina platensis) inhibited HIV-1 replication in human T-cell lines, peripheral blood mononuclear cells (PBMC), and Langerhans cells (LC). Extract concentrations ranging between 0.3 and 1.2 microg/ml reduced viral production by approximately 50% (50% effective concentration [EC50]) in PBMCs. The 50% inhibitory concentration (IC50) of extract for PBMC growth ranged between 0.8 and 3.1 mg/ml. Depending on the cell type used, therapeutic indices ranged between 200 and 6000. The extract inactivated HIV-1 infectivity directly when preincubated with virus before addition to human T-cell lines. Fractionation of the extract revealed antiviral activity in the polysaccharide fraction and also in a fraction depleted of polysaccharides and tannins. We conclude that aqueous A platensis extracts contain antiretroviral activity that may be of potential clinical interest.

Oral SIV, SHIV, and HIV type 1 infection.

Several strains of simian immunodeficiency virus (SIV), including uncloned and molecularly cloned SIV strains, can cross intact mucosal surfaces after oral exposure in both adult and neonatal rhesus macaques, resulting in viremia and disease. Cell-free SIV strains as well as infected whole blood have resulted in systemic infection after oral inoculation. Neonatal macaques, exposed orally to the chimeric SHIV-vpu+, a derivative of SIVmac239 that encodes the env gene of the T cell-tropic HIV-IIIB, have also become persistently infected. These data indicate that oral exposure to various virus strains, including T cell-tropic variants, leads to infection. After nontraumatic inoculation, the oral route was more efficient than the rectal route in permitting SIV entry in adult macaques. Infection and AIDS resulting from oral exposure of adult macaques have implications for the transmission of the human immunodeficiency virus type 1 (HIV-1) during oral-genital contact.

Human immunodeficiency virus-1 entry into purified blood dendritic cells through CC and CXC chemokine coreceptors.

Blood dendritic cells (DC) are susceptible to both macrophage (M) and T-cell line (T) tropic human immunodeficiency virus type 1. The CC chemokines RANTES, macrophage inflammatory protein-1alpha (MIP-1alpha), MIP-1beta, eotaxin, and, to a lesser extent, monocyte chemoattractant protein-1 (MCP-1) and MCP-4 blocked entry of M-tropic virus into blood DC. The CXC chemokine, SDF-1, a fusin (CXCR4 chemokine receptor) ligand, and an antifusin antibody inhibited DC entry by T-tropic virus. Purified blood DC contained CCR1, CCR2, CCR3, and CCR5 as well as the CXCR4 chemokine receptor RNA transcripts and high levels of fusin on the cell surface. The coexpression of multiple chemokine receptors offers a molecular mechanism to explain the permissiveness of DC for both M- and T-tropic viruses.

Full-length sequence of an ethiopian human immunodeficiency virus type 1 (HIV-1) isolate of genetic subtype C.

Genetic subtype C of the human immunodeficiency virus type-1 (HIV-1) has established foci of infection in India and in at least eight African countries, and is expected to contribute significantly to the global pandemic. Here we report the first almost full-length sequence of a subtype C HIV-1 from Ethiopia. Clone C2220, 9031 nt in length, was derived by long PCR amplification of proviral DNA from virus cultured on primary peripheral blood mononuclear cells, and contains all but 74 nt of the unique sequence information of the HIV-1 genome. This clone resembles HIV-1 isolates of subtypes A, B, and D in its genome organization with one notable exception: the core promoter contains not two, but three potential binding sites for the transcription factor NF-kB. The extra NF-kB site was found in all other Ethiopian strains analyzed, as well as in subtype C viruses from Zambia, suggesting it is typical for the C-subtype of HIV-1. The phylogenetic relationship of C2220 to other HIV-1 isolates is also presented. Subtype C viruses circulating in Ethiopia exhibit the low interisolate diversity typical of other, newly established HIV-1 epidemics, and C2220 is both representative of Ethiopian subtype C viruses and a suitable prototype for the development of vaccines against HIV-1 subtype C.

PIP: Foci of HIV-1 subtype C infection exist in India and at least eight African countries. HIV-1 subtype C will likely contribute significant numbers of cases to the global AIDS pandemic. The first almost full-length sequence of a subtype C HIV-1 from Ethiopia is presented. Clone C2220, 9031 nucleotides long, was derived by long polymerase chain reaction amplification of proviral DNA from virus cultured on primary peripheral blood mononuclear cells and contains all but 74 nucleotides of the unique sequence information of the HIV-1 genome. The clone's genome organization resembles HIV-1 isolates of subtypes A, B, and D except that its core promoter contains three rather than two potential binding sites for transcription factor NF-kB. The extra NF-kB site was found in all other Ethiopian strains analyzed, as well as in subtype C viruses from Zambia, suggesting that the configuration is typical for subtype C HIV-1. The phylogenetic relationship of C2220 to other HIV-1 isolates is also presented.

HIV-1 proviral DNA sequences of env gp41 PCR amplificates from Tanzania.

The polymerase chain reaction (PCR) and direct DNA sequencing were used to detect and characterize selected regions of the HIV-1 proviral genome in whole blood samples from Tanzania. Specific PCR amplification products were obtained in gag and/or env (gp41) regions from 15 of the 19 HIV-1 seropositive samples investigated. Env regions from 12 different amplificates were further characterized using the dideoxy sequencing method. Preliminary results indicate that, despite scattered nucleotide mismatches, HIV-1 gp41 amino acid sequences from Tanzania conform to the 1990 Los Alamos African consensus sequence and resemble the HIV-1 subtype A or D consensus sequences in the characterized regions.

Circulating human dendritic cells differentially express high levels of a 55-kd actin-bundling protein.

This study was initiated to examine the differential expression of an evolutionary conserved human 55-kd actin-bundling (p55) protein that is induced in B lymphocytes by Epstein-Barr virus infection. Our study demonstrates that p55 is specifically expressed at constitutively high levels in human peripheral blood dendritic cells and lymph node (interdigitating) dendritic cells. Blood dendritic cells constitute a minority (< 2%) of all blood leukocytes but are a distinct population of potent antigen-presenting cells. Immunofluorescence microscopy with a monoclonal antibody specific for p55 showed that 87% of peripheral blood dendritic cells stained brightly in the cytoplasm and in the veiled cytoplasmic extensions. In contrast, monocytes, granulocytes, T cells, and B lymphocytes showed no expression of the p55 protein. Western blot analysis confirmed that only the dendritic cell component of peripheral blood expressed high levels of p55. Staining of human lymph node sections demonstrated selective expression of the p55 antigen by dendritic cells in the T-cell-dependent areas but not in the B cell follicles. p55 is likely to be involved in the organization of a specialized microfilament cytoskeleton in the dendritic cells, and the anti-p55 antibody should be useful for further characterization of this important population of antigen-presenting cells in clinical transplantation, HIV-1 pathogenesis, and autoimmune diseases.

Acutely infected Langerhans cells are more efficient than T cells in disseminating HIV type 1 to activated T cells following a short cell-cell contact.

Most human immunodeficiency virus type 1 (HIV-1) infections involve sexual contact and virus passage across mucosal surfaces. While Langerhans cells (LCs) and dendritic cells (DCs) have been implicated in mucosal infection, their role is undefined. Here we demonstrate that acutely HIV-1-infected LCs and DCs effectively transmit virus to uninfected, activated T cells. Cocultivation of these cells results in massive virus production that requires a short cell-cell contact; as little as 30 min contact time is sufficient for HIV-1-pulsed DCs to infect their target T cells. Furthermore, surface-bound virus inactivation by trypsin does not significantly decrease the efficiency of virus transmission by LC/DCs, suggesting rapid internalization of virus. This effective virus transfer by infected LCs and blood-derived DCs requires prior activation of T cells. Surprisingly, cocultivation of acutely infected T cells with uninfected, activated target T cells results only in low virus production, even with T cell-tropic virus. We conclude that LCs and DCs are not only important targets of HIV-1 infection, but may also play a key role in the early dissemination of virus to T cells they encounter in skin or lymphoid tissue.

Elevated levels of circulating tumor necrosis factor alpha in human immunodeficiency virus type 1-infected Africans living in Sweden.

Elevated levels of tumor necrosis factor alpha in serum were found in human immunodeficiency virus type 1 (HIV-1)-infected Africans to a higher extent than in matched HIV-1-infected Caucasians, both groups living in Sweden. The results suggest that factors not related to the environment contribute to enhanced synthesis of tumor necrosis factor alpha in HIV-1-infected patients.

Sequence analysis of selected regions of the env (V3 loop and gp41) and gag (p7) reading frames of Ethiopian human immunodeficiency virus type 1 strains.

Amplified polymerase chain reaction (PCR) products, corresponding to the V3 loop and gp41 of the env, and p7 of the gag region, from proviral DNA of several Ethiopian and Swedish HIV-1 strains were sequenced. Of the six amino acids (GPGRAF) that constitute the principal neutralizing determinant (PND) within the V3 loop, the Ethiopian isolates all showed two amino acid changes (GPGQTF). Four to five other substitutions were found in the amino acids flanking the PND. Substitution of alanine (A) for threonine (T) should result in a change in the predicted secondary structure, i.e., disappearance of a coil structure. Percentage similarity data on a stretch of 22 amino acids within the V3 loop showed a concordance of the Ethiopian HIV-1 isolates with the sequences of published macrophage-T-cell tropic HIV isolates. Additionally derived protein sequences in two other regions showed two common substitutions in p7 and one to two substitutions in gp41 compared to a recent consensus sequence. These changes are hitherto unique for the Ethiopian strains, and suggest the presence of a clustering of a divergent HIV-1 strain in Addis Ababa, Ethiopia.

Raised levels of tumour necrosis factor-alpha and neopterin, but not interferon-alpha, in serum of HIV-1-infected patients from Ethiopia.

Serum levels of tumour necrosis factor-alpha (TNF-alpha), neopterin and interferon-alpha (IFN-alpha) were determined by immunoradiometric assays in 60 HIV-1+ and 20 HIV-1- subjects from Ethiopia. Swedish samples were used as reference material. The Ethiopian HIV-1+ subjects were found to have significantly increased TNF-alpha and neopterin, but not IFN-alpha levels. Increased levels of TNF-alpha and neopterin were frequently found in Ethiopian asymptomatic subjects (37% and 47%), and the concentration increased in patients with AIDS (83% and 90% respectively). The levels of the two substances and the proportion of patients with higher TNF-alpha values were lower in the corresponding Swedish subjects. The proportion of sera with raised levels of IFN-alpha was very low (asymptomatic 4%, and AIDS 7%) in Ethiopian subjects. These results suggest a very early increase in the TNF-alpha production and activation of the cellular immune response, and a low level of IFN-alpha synthesis in the natural course of HIV infection in Ethiopia. The aberrations may contribute to a rapid progress of immunodeficiency and cachexia often seen in Ethiopian patients.

Response of Ethiopian human immunodeficiency virus type 1 isolates to antiviral compounds.

Human immunodeficiency virus type 1 (HIV-1) isolates of 8 Ethiopian and 8 Swedish untreated AIDS-patients were examined for their sensitivity to 3'-azido-3'-deoxythymidine (AZT), 2',3'-dideoxyinosine (ddI) and leukocyte-derived interferon-alpha (IFN-alpha). No significant difference in drug sensitivity was found between Ethiopian and Swedish isolates, which all were sensitive to AZT, ddI and IFN-alpha except for one Swedish isolate. This isolate exhibited a mutation at amino acid position 215. These results suggest that it should be possible to perform clinical trials in Ethiopia using the same dose regimens as in Sweden.

PIP: Human immunodeficiency virus (HIV-1) isolates from 8 Ethiopian and 8 Swedish AIDS patients, none of them treated with antiviral drugs, were compared for sensitivity to azido-deoxy-thymidine (AZT), dideoxy-inosine (ddI) and interferon-alpha. HIV was isolated from peripheral blood mononuclear class, identified by Western blot and nucleotide sequencing, and passaged 1-3 times. Sensitivity to the 3 drugs, expressed as ED50s relative to positive controls, was determined by culturing HIV in the presence of drugs in a range of concentrations and assaying the supernatant for p24 antigen and the virus pellet for reverse transcriptase (RT). Dose-dependent anti-HIV activity for AZT was seen in the 8 Ethiopian isolates, and ED50s for p24 antigen and RT activity were correlated. 1 Ethiopian HIV isolate was sensitive to ddI, and another, to interferon-alpha. 1 Swedish HIV was resistant to AZT, and on analysis had a mutation from threonine to tyrosine at position 215. There were no significant differences between ED50s for interferon in the Swedish and Ethiopian HIVs. Combined data for each drug showed correlation between the p24 antigen and RT activities of the Ethiopian and Swedish HIVs. Since there was no resistance observed in the Ethiopian HIV to AZT or ddI, low-dose treatment would probably slow progression of HIV infection in Ethiopians, if these drugs could be made available for clinical trials.

Relationship between cell-free viraemia, antigenaemia and antibody levels in HIV-1-infected Ethiopian patients.

OBJECTIVE: To determine the relationship and occurrence of cell-free viraemia, free or immune-complexed p24-antigen and p24-antibody levels in blood from HIV-1-infected patients in Ethiopia. METHODS: Peripheral blood was obtained from 66 Ethiopian and 137 Swedish HIV-1-seropositive patients. Blood samples were analysed for free or immune-complex bound p24 antigen by enzyme-linked immunosorbent assay before and after acid hydrolysis of immune complexes for infectious virus in plasma and peripheral blood mononuclear cells (PBMC), and for p24-antibody levels. We compared the kinetics of viral replication of Ethiopian with Swedish isolates in vitro. RESULTS: Infectious virus was isolated from PBMC in 95% and from plasma in 81% of Ethiopian AIDS patients. In contrast, p24 antigen was detected in only 5% of AIDS patients from Ethiopia, compared with 76% of those from Sweden. p24-antibody levels were much higher and more persistent in Ethiopian than in Swedish subjects. The ratio between reverse transcriptase activity and p24 antigen was significantly higher in Ethiopian isolate culture than in those of the Swedish isolates. CONCLUSIONS: Our results show that relationships between viraemia, p24 antigenaemia and p24-antibody levels in HIV-1-infected Ethiopian patients differ from those found in comparable Swedish patients. This pattern may partly explain the differences seen in the natural course of HIV-1 infection in Ethiopia and Sweden.

PIP: Researchers conducted hematologic tests on 66 HIV-1 positive adults from Ethiopia and compared the results with those of 137 HIV-1 patients in Stockholm, Sweden, to determine the incidence of cell-free viremia, free or complexed p24 antigen, and p24 antibody levels. They isolated HIV-1 from peripheral blood mononuclear cells in 95% and from plasma in 81% of the Ethiopian subjects. The corresponding percentages for the Swedish subjects were 95% and 92%. They found p24 antigen in only 5% of AIDS patients from Ethiopia (none for asymptomatic HIV-1 subjects) compared with 76% of Swedish patients (p .01). The Ethiopian subjects had significantly higher p24 antibody levels than did the Swedish subjects (85% vs. 52%; p = .008). The ratio between reverse transcriptase activity and p24 antigen concentration stood much higher in the Ethiopians than in the Swedes (7.5 vs. 3.6; p = .0019). These results suggested that the HIV-1 strains in the Ethiopian subjects resembled rapid high HIV-1 strains. In addition, the high degrees of cell-free viremia, relative lack of free or immune complexed p24 antigen, and a persistence of p24 antibody during the entire course of infection in Ethiopian HIV-1 infected subjects intimated that the interaction between HIV-1 and the Ethiopians may be different in Africa than it is in Europe and North America. The results of another study conducted by the researchers supported this conclusion. They included high levels of tumor necrosis factor-alpha and neopterin and low levels of interferon-alpha in HIV-1 positive Ethiopians.

HIV-1 in Ethiopia: phylogenetic divergence from other HIV-1 strains.

Phylogenetic tree analysis was performed on selected polymerase chain reaction (PCR)-amplified and sequenced regions of the gag and env reading frames of several Ethiopian and Swedish human immunodeficiency virus type 1 (HIV-1) strains. These regions are considered to be conserved parts of the HIV-1 genome and correspond to the p7 of the core (gag) and part of the carboxy terminal of the gp41 protein of env respectively. Comparisons were made with all available HIV-1 sequences. The tree analysis showed that gag sequences from nine and env sequences from four Ethiopian strains all grouped together in separate branches distinct from all other sequenced European, North American, and African HIV-1 isolates. Thus, the Ethiopian strains seem to represent a highly divergent group of HIV-1, which might have developed during a relatively early stage of HIV-1 evolution.

Differences in PCR reactivity between HIV proviruses from individuals in Ethiopia and Sweden.

The polymerase chain reaction (PCR), using primer pairs in the gag, pol, and env regions, was used in a comparative study of HIV-1 DNA in peripheral mononuclear blood cells from HIV-1-seropositive individuals in Ethiopia and Sweden. Although all Swedish samples were positive by PCR, the reactivity was more pronounced in samples from late stages than in those from early stages of infection. Six of nine Ethiopian samples from HIV-1-seropositive patients were positive by PCR, but the reactions were much weaker than those observed for Swedish samples, and in most cases seen with one primer pair only. These results suggest that the burden of HIV-1 DNA in peripheral mononuclear blood cells increases with advancing disease. PCR using primer pairs designed to detect HIV-1 infection in Europe and North America is not always suitable for the detection of HIV-1 infection in Ethiopia. The differences in PCR reactivity could possibly be a consequence of differences regarding host responses to the virus in the two countries, but more likely due to genomic differences between HIV-1 strains prevalent in Ethiopia and Sweden.

HIV seropositivity in a 40-day-old child in Addis Ababa.

In this communication Human Immunodeficiency Virus (HIV) seropositivity is described in a 40-day-old Ethiopian child born to parents infected with HIV-I.

Acquired immunodeficiency syndrome: seven cases in an Addis Ababa hospital.

PIP: 7 confirmed cases of Acquired Immune Deficiency Syndrome (AIDS), seen in the Department of Medicine, Yekatit 12 Hospital, Addis Ababa, Nigeria, are presented. These are the 1st cases to be reported in Ethiopia. The ELISA and Western Blot tests were performed at the National Research Institute of Health in Addis Ababa. Attempts to trace possible sexual contacts of 3 cases were unsuccessful. Family members of 2 cases were tested for HIV infection. 3 of the 7 patients presented with tuberculosis, 2 with pneumocystis carinii pneumonia, 1 with generalized lymphadenopathy, and 1 with chronic diarrhea. All had AIDS; 6 of the 7 had 2 major clinical criteria, and all had 2 or more minor clinical criteria. 4 of the patients died. The diagnosis of 7 clinically obvious cases within 1 year in 1 general hospital at a time when it was somewhat difficult to have serological tests performed suggests the urgency of education of hospital workers in the safe handling of blood, blood products and body fluids, and through sterilization of needles and other equipment used by and for all patients.^ieng