Serological differences between human T-lymphotropic virus type-I (HTLV-I) and HTLV-III/LAV

Sera from each of five preselected groups of patients with acquired immune deficiency syndrome (AIDS), AIDS-related complex (ARC), hemophilia, adult T-cell leukemia (ATL), and healthy controls were examined for antibodies to human T-cell leukemia (T-lymphotropic) virus type-I (HTLV-I) and HTLV-III by indirect immunofluorescence (IF) and radioimmunoprecipitation (RIP) methods. All sera from five patients with AIDS, ARC, and hemophilia reacted at titers from 1 : 512 to 1 : 5,120 with fixed H9/HTLV-III cells by IF but not with fixed MT-1 cells carrying HTLV-I. Similarly, sera from patients with AIDS, ARC, and hemophilia precipitated HTLV-III-specific polypeptides of 120K, 46K, and 24K. In contrast, sera from five patients with ATL did not react with fixed H9/HTLV-III cells, but reacted with fixed MT-1 cells. Moreover, HTLV-I-specific polypeptides of 68K, 28K, and 24K were precipitated with sera from ATL-patients but not with anti-HTLV-III-positive sera. Recently, we infected HTLV-I-carrying MT-4 cells with HTLV-III and provoked strong cytopathic effects. This system enabled testing for neutralizing antibodies to HTLV-III. Neutralizing titers to HTLV-III of five anti-HTLV-III-positive sera ranged from 1 : 720 to 1 : 9,000. In contrast, all five seronegative controls showed no or only low reactivity to HTLV-III envelope (1 : 80 and 100). However, three out of five anti-HTLV-I-positive sera exhibited weak cross-reactivities with HTLV-III. The reactivities were expressed as less than 1 : 160, except for one case (1 : 720). They were considered to be nonspecific since they were negative for HTLV-III antibodies in the radioimmunoprecipitation studies.     

Isolation of a lentivirus from a macaque with lymphoma: comparison with HTLV-III/LAV and other lentiviruses.

A retrovirus has been isolated on the human T-cell line HuT 78 after cocultivation of a lymph node from a pig-tailed macaque (Macaca nemestrina) that had died with malignant lymphoma in 1982 at the University of Washington primate center. This isolate, designated MnIV (WPRC-1) (M. nemestrina immunodeficiency virus, Washington Primate Research Center) shows the characteristic morphology of a lentivirus and replicates to high titers in various lymphocyte lines of human and primate origin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified MnIV revealed multiple bands of structural proteins, including a major viral gag protein of 28 kilodaltons, that did not comigrate with the viral proteins of a human immunodeficiency virus (HIV [FRE-1]) that was also isolated on HuT 78 cells. The relatedness of MnIV to other lentiviruses (HTLV-III/LAV, EIAV, and visna) was examined in radioimmunoassays, by immunoblot techniques, and by N-terminal amino acid sequence analysis of the viral p28 gag protein. The immunoassays revealed cross-reactivity only between MnIV p28 and HTLV-III/LAV p24, and sequence analysis showed that 14 of the 24 N-terminal residues of MnIV p28 and HTLV-III/LAV p24 are identical. These results indicate that MnIV belongs to the same lentivirus family as HTLV-III/LAV but is only partially related to these human acquired immune deficiency syndrome retroviruses.     

Antibodies to the structural and nonstructural gag-coded proteins of type-D retroviruses in patients with lymphadenopathy

Screening of antibodies to structural and nonstructural gag gene-coded proteins in humans with lymphadenopathy and AIDS was performed by means of radioimmunoprecipitation (RIP) and western blotting. Pr78gag precursor of gag-coded proteins of type-D retrovirus from Hep-2 cells served as an antigen in RIP tests. Total number of sera (of humans with lymphoadenopathy) under RIP analysis was 18 and one sera of AIDS patient. Six of them reacted with Pr78gag and one out of one AIDS serum. Over 80 sera samples of humans with lymphadenopathy have been tested by means of western blotting with proteins of Mason-Pfizer monkey virus as antigens. Antibodies to p27 (major internal protein of Mason-Pfizer monkey virus) were detected in 12 sera samples of those with lymphadenopathy (dilution 1:100) and in 9 out of 12 sera of AIDS patients (dilution 1:100-1:400). Results obtained make it possible to predict that type-D retroviruses are associated with acquired immunodeficiency syndrome and generalized lymphadenopathy and could play some role in development of this illness in humans.     

Sera from HTLV-III/LAV antibody-positive individuals mediate antibody-dependent cellular cytotoxicity against HTLV-III/LAV-infected T cells

The causative agent of the acquired immunodeficiency syndrome (AIDS) has been shown to be a human retrovirus called human T lymphotropic virus (HTLV)-III or lymphadenopathy-associated virus (LAV). The nature of the protective immune response against this virus is currently unknown. We report here results using an antibody-dependent cellular cytotoxicity (ADCC) assay which has been developed for measuring a specific immune response against HTLV-III/LAV. Forty-four sera were examined for their ability to mediate ADCC against HTLV-III/LAV-infected T cells. Sera from healthy HTLV-III/LAV seropositive individuals in the presence of mononuclear cells from healthy HTLV-III/LAV seronegative donors exhibited significantly higher levels of ADCC activity compared to sera from patients with AIDS. Western blot analysis of serum samples indicated that antibody reactivity with the p24 protein of HTLV-III/LAV correlated with higher levels of ADCC activity than did reactivity with Gp120/160. The observation that sera from healthy HTLV-III/LAV seropositive individuals mediated higher levels of ADCC activity than did sera obtained from subjects with AIDS suggests that ADCC may represent a protective immune response to infection with HTLV-III/LAV.     

IgG-antibodies to HTLV-III in patients with AIDS, LAS, and persons at risk of AIDS in West Germany

In a first seroepidemiological study on the prevalence of the human T-lymphotropic retrovirus HTLV-III in West Germany, sera of 26 patients with acquired immunodeficiency syndrome (AIDS), 33 patients with lymphadenopathy syndrome (LAS) or AIDS related complex (ARC), and 113 homosexual men at risk of AIDS were screened for IgG antibodies to HTLV-III by an enzyme linked immunosorbent assay (ELISA). 22 out of 26 AIDS-patients (84.6%), 24 out of 33 LAS-patients (72.7%), and 44 out of 113 healthy homosexual men with increased risk of AIDS (38.9%) were found positive for antibodies to HTLV-III. Heterosexual controls including healthy laboratory workers and medical personnel with contact to AIDS patients did not show antibodies to HTLV-III. The HTLV-III antibodies analyzed predominantly recognize a protein of molecular weight 41,000 (p41).     

IgM and IgG antibodies to human T cell lymphotropic retrovirus (HTLV-III) in lymphadenopathy syndrome and subjects at risk for AIDS in Italy.

A study was performed to assess the prevalence of specific antibodies to human T cell lymphotropic retrovirus (HTLV-III) in patients with lymphadenopathy syndrome, patients with the acquired immune deficiency syndrome (AIDS), and those at risk of AIDS. Serum samples were obtained from these groups and from healthy controls in selected cities in Italy, and antibodies to HTLV-III were measured by immunofluorescence assay and, in a few patients, by Western blotting. In addition, IgM antibody values were measured in 82 of those positive for HTLV-III. Altogether, 235 out of 320 patients with lymphadenopathy syndrome had antibodies to HTLV-III, the proportions being highest in haemophiliacs, homosexuals, and drug addicts from Rome; 11 out of 12 patients with AIDS had antibodies; 78 out of 439 subjects at risk for AIDS had antibodies; and six out of 30 patients with lymphadenopathy syndrome and positive for HTLV-III antibodies and nine of 52 patients at risk of AIDS had a detectable titre of IgM. HTLV-III is widespread in groups at risk of AIDS in Italy, and antibodies to HTLV-III are highly prevalent in patients with lymphadenopathy syndrome. A higher proportion of drug abusers were positive for antibodies than in previous studies. HTLV-III "infection" would appear to be spread mainly in compromised hosts, as none of the controls were positive for antibodies.     

Delineation of type-specific regions on the envelope glycoproteins of human T cell leukemia viruses.

Two different approaches were used to map the type-specific regions on human T cell leukemia virus (HTLV) envelope glycoproteins. 1) Antibody reactivities of polymerase chain reaction-confirmed HTLV-I or HTLV-II carriers' sera were analyzed by Western blot assay with seven recombinant proteins containing different regions of HTLV-I or HTLV-II envelope proteins. 2) Rabbit antibodies elicited by nine HTLV-I Env synthetic peptides were used to react with the native HTLV envelope glycoproteins in an antibody-dependent cellular cytotoxicity (ADCC) assay. The results of the Western blot analysis showed that RP-B2, which contains amino acid residues 166 to 213 from HTLV-II exterior glycoprotein, was specifically reactive with 90.6% (48 of 53) of the HTLV-II carriers' sera but not with any of the HTLV-I carriers' serum (0 of 71). In contrast, RP-B, which contains amino acid residues 166 to 229 from HTLV-I exterior glycoprotein, was reactive with 85.1% (114 of 134) of the HTLV-I carriers' sera but not with any HTLV-II carrier serum (0 of 62). Furthermore, anti-HTLV-I Env synthetic peptide antibody-mediated ADCC identified several distinguishing HTLV-I ADCC epitopes in the middle region (amino acid residues 177 to 257) of the HTLV-I exterior glycoprotein. Therefore, HTLV type-specific epitopes reside mainly in a 69-amino acid sequence bounded by two cysteine residues (amino acids 157 and 225 for HTLV-I and 153 and 221 for HTLV-II), in the middle region of the exterior envelope glycoproteins.     

The Rex proteins of human T-cell leukemia virus type II differ by serine phosphorylation.

The Rex proteins of human T-cell leukemia virus types I and II (HTLV-I and HTLV-II) induce cytoplasmic expression of unspliced gag-pol mRNA and singly spliced env mRNA and are critical for virus replication. Two rex gene products, p27rex and p21rex of HTLV-I and p26rex and p24rex of HTLV-II, have been detected in HTLV-infected cells; however, the structural and biological relationship of the proteins has not been clearly elucidated. Endoproteinase digestion and phosphoamino acid analysis of HTLV-II Rex indicated that p24rex has the same amino acid backbone as p26rex and that the larger apparent molecular size of p26rex is attributable to serine phosphorylation.     

Dual Infection of HIV-1 and HTLV-I in South India: A Study on a Patient with AIDS-Related Complex

A dually HIV-and HTLV-infected ARC patient was found by serological studies in South India. These viruses were isolated and molecular study showed that the patient had both HIV-1 and HTLV-I but not HIV-2 and HTLV-II. In addition to this, 9 other dually infected persons which include another full-blown AIDS case have been identified as on July 1993 in South India. Our findings provide an opportunity to clarify geographical distribution of these human retroviruses.     

Isolation of a type D retrovirus from B-cell lymphomas of a patient with AIDS.

An atypical syncytial variant of a high-grade Burkitt's-type B-cell lymphoma from a patient with AIDS who was seropositive for human immunodeficiency virus type 1 was studied. A productive type D retrovirus infection was identified in early-passage cell lines derived from two lymphomas from this patient. Nucleotide and amino acid sequence analysis as well as immunologic reactivity indicated that the isolated virus was highly related to Mason-Pfizer monkey virus (MPMV). MPMV is an immunosuppressive type D retrovirus that causes an AIDS-like syndrome in rhesus macaques. Amplification of DNA from the patient's diagnostic bone marrow biopsy specimen by polymerase chain reaction generated the appropriate MPMV-specific fragments and indicated that the patient was infected with the MPMV-like retrovirus. In addition, the patient's serum contained antibodies which recognized type D viral env proteins (gp70 and gp20) and gag proteins (p27 and p14). Although there have been reports of human cell lines infected with type D retroviruses and of type D-reactive human sera, this is the first evidence of a type D retrovirus infection in a human confirmed by virus isolation, serum reactivity, and viral DNA identification in tumor tissue.     

HIV and HTLV-I antibody studies: pregnant women in the 1960s, patients with AIDS, homosexuals, and individuals with tropical spastic paraparesis

To investigate the possible occurrence of human immunodeficiency virus (HIV) or human T-cell lymphotropic virus, type I (HTLV-I) infections in the United States prior to 1979-1981, when acquired immune deficiency syndrome (AIDS) was first recognized, we tested sera from 310 pregnant women who participated in the Collaborative Perinatal Project during the period 1959-1964 for HIV and HTLV-I antibody. These samples included sera from 53 pregnant women who were intravenous drug users. The remainder were from women who had cervical epithelial abnormalities, who developed cervical carcinomas, who had had children with erythroblastosis fetalis, who had had children that developed malignant neoplasms early in life, or normal pregnant women. None of the 310 women had confirmed HIV or HTLV-I antibody. The rate of false-positive reactions with the HIV enzyme-linked immunosorbent assay (ELISA) antibody test in these long-frozen samples was similar to that observed in fresh sera. HIV antibody was detected in homosexual patients with AIDS; HTLV-I antibody was not detected in any of these sera. HTLV-I antibody was detected in 17 of 20 patients with tropical spastic paraparesis (TSP) and in two of seven patients with other neurological diseases diagnosed as transverse myelopathy and multiple sclerosis, and in none of nine normal controls; HIV antibody was not detected in any of these sera patients. Thus, we conclude that there was no serological evidence of infection with HIV or HTLV-I in the pregnant women studied; however, HIV antibody was present in all AIDS patients tested, and HTLV-I antibody was found in the majority of patients with TSP.     

Immunoglobulin subclasses of antibodies to human T-cell leukemia/lymphoma virus I-associated antigens in acquired immune deficiency syndrome and lymphadenopathy syndrome

Immunoglobulin G (IgG) and IgM antibodies to human T-cell leukemia/lymphoma virus-I (HTLV-I)-associated membrane antigens (HTLV-I-MA) were assayed by indirect cytospin immunofluorescence, and IgG and IgM antibodies to purified HTLV-I were assayed by enzyme-linked immunosorbent assay in sera from 119 immunologically well-characterized promiscuous male homosexuals in The Netherlands, of whom 9 suffered from acquired immune deficiency syndrome (AIDS), 18 suffered from lymphadenopathy syndrome (LAS), and 5 suffered from gay bowel syndrome. Antibodies to HTLV-I-MA were present in four of nine AIDS patients, including one patient with antibodies to purified HTLV-I. Antibodies to HTLV-I-MA were present in 6 of 18 LAS patients, including 3 patients with antibodies to purified HTLV-I. Of five patients with gay bowel syndrome, one had IgG and IgM antibodies to HTLV-I-MA. Of the four HTLV-I seropositive AIDS patients, two had IgG and IgM antibodies to HTLV-I or HTLV-I-MA, one had only IgG antibodies, and one had only IgM antibodies. Of the six HTLV-I seropositive LAS patients, four had IgG and IgM antibodies to HTLV-I or HTLV-I-MA, and two had only IgM antibodies. In the sera from 27 healthy homosexuals with and 60 without T-cell subset imbalances, no antibodies to HTLV-I or HTLV-I-MA were detected.     

HIV-1 drug-resistance and drug-dependence

Background

The Deltaretrovirus genus comprises viruses that infect humans (HTLV), various simian species (STLV) and cattle (BLV). HTLV-I is the main causative agent in adult T-cell leukemia in endemic areas and some of the simian T-cell lymphotropic viruses have been implicated in the induction of malignant lymphomas in their hosts. BLV causes enzootic bovine leukosis in infected cattle or sheep. During the past few years several new Deltaretrovirus isolates have been described in various primate species. Two new HTLV-like viruses in humans have recently been identified and provisionally termed HTLV-III and HTLV-IV. In order to identify a broad spectrum of Deltaretroviruses by a single PCR approach we have established a novel consensus PCR based on nucleotide sequence data obtained from 42 complete virus isolates (HTLV-I/-II, STLV-I/-II/-III, BLV). The primer sequences were based on highly interspecies-conserved virus genome regions. We used this PCR to detect Deltaretroviruses in samples from adult patients with a variety of rare T-cell neoplasms in Germany.

Results

The sensitivity of the consensus PCR was at least between 10^-2 and 10^-3 with 100% specificity as demonstrated by serial dilutions of cell lines infected with either HTLV-I, HTLV-II or BLV. Fifty acute T-cell lymphoblastic leukemia (T-ALL) samples and 33 samples from patients with various rare mature T-cell neoplasms (T-PLL, Sézary syndrome and other T-NHL) were subsequently investigated. There were no cases with HTLV-I, HTLV-II or any other Deltaretroviruses.

Conclusion

The results rule out a significant involvement of HTLV-I or HTLV-II in these disease entities and show that other related Deltaretroviruses are not likely to be involved. The newly established Deltaretrovirus PCR may be a useful tool for identifying new Deltaretroviruses.     

Cytomegalovirus but not human T lymphotropic virus type III/lymphadenopathy associated virus detected by in situ hybridisation in retinal lesions in patients with the acquired immune deficiency syndrome.

Paraffin sections of retinal tissue from five patients who died from the acquired immune deficiency syndrome (AIDS) and retinopathy were examined by in situ hybridisation experiments with deoxyribonucleic acid (DNA) labelled with sulphur-35 of lentivirus, human T lymphotropic virus type III/lymphadenopathy associated virus (HTLV-III/LAV), and cytomegalovirus. HTLV-III/LAV ribonucleic acid (RNA) was not detected in any of the tissue sections. Cytomegalovirus RNA was identified, however, in three of the five patients. Retinopathy induced by cytomegalovirus may thus be one of the many syndromes potentiated by the immunosuppression caused by HTLV-III/LAV.     

Gag protein epitopes recognized by CD4(+) T-helper lymphocytes from equine infectious anemia virus-infected carrier horses

Antigen-specific T-helper (Th) lymphocytes are critical for the development of antiviral humoral responses and the expansion of cytotoxic T lymphocytes (CTL). Identification of relevant Th lymphocyte epitopes remains an important step in the development of an efficacious subunit peptide vaccine against equine infectious anemia virus (EIAV), a naturally occurring lentivirus of horses. This study describes Th lymphocyte reactivity in EIAV carrier horses to two proteins, p26 and p15, encoded by the relatively conserved EIAV gag gene. Using partially overlapping peptides, multideterminant and possibly promiscuous epitopes were identified within p26. One peptide was identified which reacted with peripheral blood mononuclear cells (PBMC) from all five EIAV-infected horses, and three other peptides were identified which reacted with PBMC from four of five EIAV-infected horses. Four additional peptides containing both CTL and Th lymphocyte epitopes were also identified. Multiple epitopes were recognized in a region corresponding to the major homology region of the human immunodeficiency virus, a region with significant sequence similarity to other lentiviruses including simian immunodeficiency virus, puma lentivirus, feline immunodeficiency virus, Jembrana disease virus, visna virus, and caprine arthritis encephalitis virus. PBMC reactivity to p15 peptides from EIAV carrier horses also occurred. Multiple p15 peptides were shown to be reactive, but not all infected horses had Th lymphocytes recognizing p15 epitopes. The identification of peptides reactive with PBMC from outbred horses, some of which encoded both CTL and Th lymphocyte epitopes, should contribute to the design of synthetic peptide or recombinant vector vaccines for EIAV.