Human immunodeficiency virus type 1 evolution in vivo tracked by DNA heteroduplex mobility assays

High mutation rates and strong selective pressures imposed on human immunodeficiency viruses in vivo result in the formation of pools of genetic variants known as quasispecies. DNA heteroduplex mobility and tracking analyses were used to monitor the generation of HIV sequence diversity, to estimate quasispecies complexity, and to assess the turnover of genetic variants to approach an understanding of the relationship between viral quasispecies evolution in vivo and disease progression. Proviral DNA pools were nearly homogeneous soon after sexual transmission. The emergence and clearance of individual variants then occurred at different rates in different individuals. High quasispecies complexity was found in long-term-infected, asymptomatic individuals, while rapid CD4+ cell decline and AIDS were often, but not always, associated with lower quasispecies complexity. Proviral genetic variation was often low following in vitro culture, because of the outgrowth of one or a few variants that often became more abundant only later as proviruses in peripheral blood mononuclear cells. These studies provide insight into the dynamics of human immunodeficiency virus sequence changes in vivo and illustrate the utility of heteroduplex analysis for the study of phenomena associated with rapid genetic changes.     

Lower prevalence and incidence of HIV-1 syncytium-inducing phenotype among injecting drug users compared with homosexual men

OBJECTIVE: To study the prevalence, incidence and predictive value for progression to AIDS of the HIV-1 syncytium-inducing (SI) phenotype in HIV-infected injecting drug users (IDU) compared with HIV-infected homosexual men. DESIGN: Two prospective cohort studies on HIV-1 infection among IDU and homosexual men. METHODS: HIV-infected IDU (n = 225) and homosexual men (n = 366) without AIDS were studied from March 1989 through December 1993. Data on laboratory markers, including the presence of SI variants, demographics, behavioural characteristics and clinical events were collected at every visit. RESULTS: At baseline, SI variants were detected in 4% of IDU and 17% of homosexual men. During the study period 18 IDU and 68 homosexual men switched from non-SI to SI phenotype (4-year cumulative incidence, 14.6 and 28.4%, respectively) before AIDS diagnosis. Among participants with a documented date of HIV infection the cumulative incidence of SI was lower among IDU than homosexual men (4-year cumulative incidence, 6.2 and 20.7%, respectively). At AIDS diagnosis, 21% of all AIDS cases among IDU had the SI phenotype compared with 54% among homosexual men. In both risk groups an accelerated CD4 decline was found after the non-SI-to-SI switch. The SI phenotype appeared to be a predictor of AIDS (multivariate relative hazard, 5.33), independent of CD4 cell count and p24 antigen at baseline. In the multivariate time-dependent analysis, the relative hazard of SI phenotype decreased considerably, which is consistent with the hypothesis that the effect of SI phenotype on progression to AIDS is mediated by CD4 cell count. CONCLUSION: The SI phenotype is associated with accelerated CD4 decline and progression to AIDS in both risk groups. The remarkable lower prevalence and incidence of the SI phenotype among IDU may implicate a difference in pathogenesis and natural history of HIV infection linked to transmission group.     

Adaptive evolution of human immunodeficiency virus-type 1 during the natural course of infection

The rate of progression to disease varies considerably among individuals infected with human immunodeficiency virus-type 1 (HIV-1). Analyses of semiannual blood samples obtained from six infected men showed that a rapid rate of CD4 T cell loss was associated with relative evolutionary stasis of the HIV-1 quasispecies virus population. More moderate rates of CD4 T cell loss correlated with genetic evolution within three of four subjects. Consistent with selection by the immune constraints of these subjects, amino acid changes were apparent within the appropriate epitopes of human leukocyte antigen class I-restricted cytotoxic T lymphocytes. Thus, the evolutionary dynamics exhibited by the HIV-1 quasispecies virus populations under natural selection are compatible with adaptive evolution.     

Low T-cell responses to CD3 plus CD28 monoclonal antibodies are predictive of development of AIDS

OBJECTIVE: Decreased T-cell reactivity in vitro is strongly associated with progression to AIDS and low CD4+ T-cell numbers. Low T-cell responses in vitro induced by CD3 monoclonal antibody (mAb) are predictive for progression to AIDS independent of low CD4+ T-cell counts and high HIV-1 RNA levels. We developed a whole-blood lymphocyte culture system in which T cells were stimulated by a combination of CD3 and CD28 mAb. Combined stimulation of CD28, a costimulatory molecule, and CD3 considerably enhances T-cell responses in vitro and reduces variation coefficients, which may increase the prognostic power of T-cell responses. DESIGN: A prospective study of HIV-1-infected homosexual men followed for 35 months. METHODS: The predictive value of low T-cell responses to CD3 plus CD28 mAb relative to low CD4+ T-cell counts, high HIV-1 RNA levels and the presence of syncytium-inducing (SI) HIV-1 variants was evaluated longitudinally in 202 HIV-1-infected homosexual men followed for 35 months. RESULTS: In multivariate analysis, decreased T-cell responses at baseline were predictive of development of AIDS, independent of low CD4+ T-cell numbers and high HIV-1 RNA levels. In a time-dependent model, HIV-1 RNA levels lost their predictive value, whereas low T-cell responses, low CD4+ T-cell numbers and the presence of SI HIV-1 variants independently predicted AIDS. CONCLUSIONS: These data demonstrate that combined use of virological and immunological markers may be useful in monitoring disease progression and response to antiretroviral therapy.     

Development of AIDS in a chimpanzee infected with human immunodeficiency virus type 1

The condition of a chimpanzee (C499) infected with three different isolates of human immunodeficiency virus type 1 (HIV-1) for over 10 years progressed to AIDS. Disease development in this animal was characterized by (i) a decline in CD4+ cells over the last 3 years; (ii) an increase in viral loads in plasma; (iii) the presence of a virus, termed HIV-1JC, which is cytopathic for chimpanzee peripheral blood mononuclear cells; and (iv) the presence of an opportunistic infection and blood dyscrasias. Genetic analysis of the V1-V2 region of the envelope gene of HIV-1JC showed that the virus present in C499 was significantly divergent from all inoculating viruses (> or = 16% divergent at the amino acid level) and was suggestive of a large quasispecies. Blood from C499 transfused into an uninfected chimpanzee (C455) induced a rapid and sustained CD4+-cell decline in the latter animal, concomitant with high plasma viral loads. These results show that HIV-1 can induce AIDS in chimpanzees and suggest that long-term passage of HIV-1 in chimpanzees can result in the development of a more pathogenic virus.     

Transendothelial migration of lymphocytes from HIV-1-infected donors: a mechanism for extravascular dissemination of HIV-1

To identify factors that cause HIV-1 to establish perivascular foci of infected cells, we studied the transendothelial migration of blood mononuclear leukocytes (MNL) from 76 HIV+ patients and 41 controls. The fraction of patients' lymphocytes that migrated across endothelial cell monolayers in vitro was significantly increased (p < or = 0.03) compared with that of control donors. Migration of patients' CD4+ T cells was particularly enhanced, whereas the migration of monocytes did not differ between patients and controls. Lymphocyte migration correlated with expression of CD11a/CD18 and CD49d/CD29 and with the quantity of TNF-alpha produced as MNLs migrated through the endothelium. Measurement of HIV-1 proviral DNA copies in the patients' MNLs (n = 26) suggested that in half the cases virus-infected cells accumulated preferentially amidst the migratory leukocytes. We observed the same behavior with normal donor MNLs infected, in vitro, with each of 4 strains of HIV-1. The number of HIV-1 proviral DNA copies per million MNLs was 40 to 178 times higher in the migratory population than in the original population added to the endothelium. To test whether only certain strains of HIV-1 stimulate transendothelial migration of infected cells, we used single strand conformation polymorphism analysis to identify quasispecies of HIV-1 in the MNLs. If all strains of HIV-1 were equal in their ability to stimulate transendothelial migration, we expected to find no differences in the quasispecies present in the original and migratory cell populations. In fact the quasispecies differed in 14 of 19 paired samples, suggesting that only certain HIV-1 quasispecies promote transendothelial migration of infected cells.     

Immunologic action of [Met5]enkephalin fragments

To determine the clinical and virological correlates of neuropsychological test performance decline in HIV infection, we measured viral burden in blood in 272 HIV-seropositive men without dementia in the Baltimore arm of the Multicenter AIDS Cohort Study (MACS). These measures were then related to neuropsychological (NP) decline, defined as a decline relative to prior best performance of 2.0 standard deviations or more on one or more neuropsychological tests. A short battery of NP tests (Mini-Screen Battery) was administered to all 272 men. NP test performance decline was identified in 53/272 (19.5%) of participants on the Mini-Screen Battery. Follow-up NP data were available for 204 participants who had undergone the Mini-Screen. The frequency of sustained NP test performance decline was 7.8% for the Mini-Screen Battery. A lower CD4+ cell count was weakly associated with sustained NP test performance decline. After adjustment for CD4+ cell count, hemoglobin, body mass index, and presence of AIDS, none of the viral burden measures (p24 antigenemia, plasma viremia, quantitative culture) correlated with sustained NP test performance decline. We conclude that these measures of blood HIV viral burden are not markers for NP decline, but that a lower CD4+ cell count is.     

Rate of decline of percentage CD4+ cells is faster in HIV-1 than in HIV-2 infection

Increasing evidence suggests that the pathogenesis of HIV-1 is different from that of HIV-2. Thus, we have measured, longitudinally at various times over a median follow-up of 2.1 years, the percentage CD4+ cells of 94 patients infected with HIV-1 and 164 patients infected with HIV-2. The pattern of decline of CD4% over time was linear for patients with either infection. Multilevel statistical modeling techniques showed that after stratifying for HIV status, the rate of decline of CD4% was faster among patients who died than among those who survived (difference in rate of decline = 2.34% CD4+ cells/year; p = 0.0002). After stratifying for survival status, the rate of decline was faster and less variable among patients infected with HIV-1 than among patients infected with HIV-2 (difference in rate of decline = 1.12% CD4+ cells/year; p = 0.05). The proportion of patients who showed no fall in CD4+ cells was higher in HIV-2 than in HIV-1 infection (p = 0.026). These data suggest fundamental differences between the two infections, with HIV-1 being more pathogenic resulting in a faster and more homogeneous rate of decline than HIV-2. In HIV-2 infection, disease in many patients progresses slowly, but in some the advance is just as fast as that in HIV-1 infection. The reasons for this marked heterogeneity need elucidation to understand the disease and to target therapeutic interventions against HIV-2 in those most at risk.     

Quantitative molecular monitoring of HIV-1 RNA during antiretroviral therapy

Plasma HIV-1 RNA testing was used to monitor 43 HIV-1 infected patients newly placed on antiretroviral therapy or whose therapy had been recently changed. A polymerase chain reaction kit was used to measure HIV-1 RNA in clinical samples or frozen plasma. The cutoff of this test was 200 RNA copies/ml. The first group (11 patients) was stable on long-term zidovudine monotherapy when switched to stavudine. The HIV-1 RNA of three patients who had a regular decline in CD4+ T cell count did not change despite this switch, with a mean follow-up of 630 days. The HIV-1 RNA copy numbers of eight patients whose CD4+ T cell counts were stable declined an average of 0.53 log10 between days 90 and 650. The second group (14 patients) was on long-term zidovudine monotherapy and had declining CD4+ T cell counts over the past 6 months. Lamivudine was added to this regimen on day 0. HIV-1 RNA copy number decreased rapidly within 30 d, reaching -0.86 log10 on day 90, and this effect was maintained thereafter, with a mean follow-up of 161 days. There was a concomitant mean gain of +33 CD4+ T cells on day 90. The third group (nine patients) had never received anti-retroviral therapy and was given zidovudine+didanosine. HIV-1 RNA copy number decreased in all cases but one, reaching -1.31 log10 on day 150. This decrease was transient in three cases. The last group (nine patients) had also not had previous anti-retroviral therapy and was given zidovudine + didanosine + lamivudine in combination. HIV-1 RNA copy numbers declined rapidly in all cases, to below the cutoff in eight cases within a mean period of 50.5 days. The CD4+ cell counts increased by 164 cells/microliter on day 14 and by 201 cells/microliter on day 180. The response to therapy of the total population of 43 patients varied according to cases. The relative changes in p24 antigen compared to HIV-1 RNA also differed between patients. Measurement of HIV-1 viremia appears to be a valuable tool in current practice for individualizing therapy.     

Disturbed CD4+ T cell homeostasis and in vitro HIV-1 susceptibility in transgenic mice expressing T cell line-tropic HIV-1 receptors

T cell line-tropic (T-tropic) HIV type 1 strains enter cells by interacting with the cell-surface molecules CD4 and CXCR4. We have generated transgenic mice predominantly expressing human CD4 and CXCR4 on their CD4-positive T lymphocytes (CD4+ T cells). Their primary thymocytes are susceptible to T-tropic but not to macrophage-tropic HIV-1 infection in vitro, albeit with a viral antigen production less efficient than human peripheral blood mononuclear cells. Interestingly, even without HIV infection, transgenic mice display a CD4+ T cell depletion profile of peripheral blood reminiscent of that seen in AIDS patients. We demonstrate that CD4+ T cell trafficking in transgenic mice is biased toward bone marrow essentially due to CXCR4 overexpression, resulting in the severe loss of CD4+ T cells from circulating blood. Our data suggest that CXCR4 plays an important role in lymphocyte trafficking through tissues, especially between peripheral blood and bone marrow, participating in the regulation of lymphocyte homeostasis in these compartments. Based on these findings, we propose a hypothetical model in which the dual function of CXCR4 in HIV-1 infection and in lymphocyte trafficking may cooperatively induce progressive HIV-1 infection and CD4+ T cell decline in patients.     

Amprenavir

Amprenavir is a viral protease inhibitor with specificity for the HIV protease enzyme. The resistance profile of amprenavir appears to differ from that of other protease inhibitors such as saquinavir and indinavir. Twelve hours after single-dose administration of amprenavir 1200mg to HIV-infected individuals, the mean plasma concentration of the drug was more than 10-fold greater than the 50% inhibitory concentration for HIV-1IIIB in peripheral blood lymphocytes. In a small nonblind study, amprenavir monotherapy increased CD4+ cell count and decreased viral load in 37 patients with HIV infection and no previous exposure to protease inhibitor therapy. Combination therapy comprising amprenavir and other antiretroviral agents (abacavir, zidovudine, lamivudine, indinavir, saquinavir or nelfinavir) decreased viral load and increased CD4+ cell counts in patients with HIV infection. Antiviral efficacy was maintained during up to 24 weeks' follow-up. Available data suggest that rash, headache and diarrhoea or loose stools are the most frequent adverse events associated with amprenavir therapy.     

Value of HIV-1 viral load and CD4 lymphocyte count as determinants of progression to AIDS and survival

BACKGROUND: HIV-1 viral load is regarded as a better surrogate marker for progression and death than CD4+ cell counts. Both markers are analysed in a cohort of patients with unknown seroconversion date and advanced HIV infection. PATIENTS AND METHODS: Retrospective cohort analysis of 421 patients, most on antiretroviral therapy, with a median initial CD4+ cell count of 209 x 10(6)/l and a median initial viral load of 4.7 log copies/ml. One thousand two hundred and eighty-six samples were analysed. Univariate and bivariate analysis were performed with initial and sequential CD4+ cell counts and viral load values to estimate the risk of progression and death by Cox regression models. RESULTS: After a median follow up of 763 days, 124 patients developed AIDS and 117 died. Relative risks of progression related to the group that maintained viral load values always < 35,000 copies/ml were: 5-fold (95% CI: 1.4-17.0; p < 0.05) for patients with any viral load value > 35,000 copies/ml but always < 200,000 copies/ml; and 13.6 fold (95% CI: 5.4-34.2; p < 0.0001) for patients who could not maintain viral load < 200.000 copies/ml. CD4+ counts = 100 x 10(6)/l and viral load = 220,000 copies/ml were the threshold values that best fitted to estimate the probability of survival by a bivariate analysis. CONCLUSIONS: The maintenance of sequential viral load values < 35.000 copies/ml is associated with a lower risk of progression. The maintenance of sequential viral load values < 150,000 copies/ml is associated with higher short-term survival rates.     

Presence of the variant mannose-binding lectin alleles associated with slower progression to AIDS. Amsterdam Cohort Study

OBJECTIVE: To examine the association between mannose-binding lectin (MBL) polymorphism and progression to AIDS and death in HIV-1 infection. DESIGN AND METHODS: In 131 HIV-1-infected homosexual seroconverters, survival analyses were performed to determine both the association between MBL genotype and time from HIV-1 seroconversion to AIDS and death, and time from AIDS to death. RESULTS: Of the 131 seroconverters, of whom 61 developed AIDS, 76 were typed as homozygous wild-type and 55 as carriers of variant alleles (52 heterozygous and three homozygous variant alleles). A Survival analyses suggested that HIV-1-infected men with the variant alleles progressed somewhat slower to AIDS [relative hazard (RH), 0.62; 95% confidence interval (CI), 0.36-1.10] and death (RH, 0.73; 95% CI, 0.42-1.25). Interestingly, CD4+ T-cell count determined at the moment of AIDS was found to be significantly lower among persons with the mutation (97 x 10(6)/l versus 204 x 10(6)/l; P=0.03). Furthermore, when AIDS-free times before the diagnosis of an opportunistic infection were compared with those preceding a diagnosis of Kaposi's sarcoma, Kaposi's sarcoma diagnosis was more postponed than that of an opportunistic infection (RH, 0.21; 95% CI, 0.05-0.95; versus RH, 0.67; 95% CI, 0.35-1.27). CONCLUSION: Indications for a weak pre-AIDS protective effect of variant MBL alleles were demonstrated.     

T-cell homeostasis, competition, and drift: AIDS as HIV-accelerated senescence of the immune repertoire

The observation that the density of CD8+ T-lymphocytes increases as the density of CD4+ T-cells declines in adult HIV-1/AIDS patients, together with evidence that the total density of T-cells is regulated (homeostasis) has led to the suggestion that competition between lineages, and classes of T-cells contributes to the pathology of HIV. We use a mathematical model of the interactions between populations of T-cells, HIV, and other parasites to explore the effects of T-cell homeostasis and competition on the progression to AIDS. We demonstrate that as a consequence of parasite-mediated T-cell replication, of competition within and between different T-cell clones, and random processes (T-cell drift), some CD4+ lineages will be represented by relatively few cells, dearths, and some lineages may be lost, leaving holes in the immune repertoire. By killing CD4+ T-lymphocytes, HIV accelerates the rate at which these dearths and holes accumulate and leads to an early breakdown of the immune control of HIV and other parasites, AIDS. When this model allows for intense, but not complete, competition between the CD4+ and CD8+ T-cell populations, it can account for most of the features of an HIV-1 infection in adults, including the gradual decline in CD4+ T-cell densities and concomitant increase in HIV density, as well as the variability in time from infection to AIDS and the decline in the time from infection to AIDS in older patients.     

Abnormal expression of a 170-kilodalton P-glycoprotein encoded by MDR1 gene, a metabolically active efflux pump, in CD4+ and CD8+ T cells from patients with human immunodeficiency virus type 1 infection

Peripheral blood CD4+ and CD8+ T cells from 16 patients with HIV-1 infection, 8 each with CD4+ T cell counts of > 200/mm3 (group I) and with CD4+ T cell counts of < 200/mm3 (group II), and 8 age- and sex-matched controls, were examined for the expression of P-glycoprotein (P-gp), a 170-kDa phosphoglycoprotein encoded by the MDR1 gene, using dual-color flow cytometric analysis. The function of P-glycoprotein was assessed by the accumulation of rhodamine-123 (Rh123) dye in the presence or absence of cyclosporin A (which inhibits Rh123 efflux). A significantly increased proportion of CD4+ T cells from patients with HIV-1 infection expressed P-glycoprotein as compared to controls, resulting in a significantly increased ratio of the proportions of CD4+P-gp+/CD8+P-gp+ cells. The ratio of CD4+P-gp+/CD8+P-gp+ in group II patients was significantly higher (p = 0.02) than in group I patients, suggesting a progressive increase in P-gp expression with the advancement of HIV-1 infection. The proportions of CD4+P-gp+ and CD8+P-gp+ T cells did not differ significantly between those who received AZT and those who were not treated with AZT. Contrary to expectation, both CD4+ and CD8+ T cells from patients accumulated significantly more Rh123 as compared to controls. Furthermore, cyclosporin A failed to increase intracellular accumulation of Rh123 in CD4+ and CD8+ T cells from patients. These data suggest a functionally defective P-gp expression in HIV-1 infection that appears to increase with the progression of HIV-1 infection. A study of a large number of patients with HIV-1 infection is needed to determine the effects of opportunistic infection and antiretroviral therapy on the expression of P-gp and to determine whether the expression of P-gp could serve as another surrogate marker for the progression of HIV-1 infection.     

Inhibition of apoptosis in chlamydia-infected cells: blockade of mitochondrial cytochrome c release and caspase activation

Current clinical gene therapy protocols for the treatment of human immunodeficiency virus type 1 (HIV-1) infection often involve the ex vivo transduction and expansion of CD4+ T cells derived from HIV-positive patients at a late stage in their disease (CD4 count <400). These protocols involve the transduction of T cells by murine leukemia virus (MLV)-based vectors encoding antiviral constructs such as the rev m10 dominant negative mutant or a ribozyme directed against the CAP site of HIV-1 RNA. We examined the efficiency and stability of transduction of CD4+ T cells derived from HIV-infected patients at different stages in the progression of their disease, from seroconversion to AIDS. CD4+ T cells from HIV-positive patients and uninfected donors were transduced with MLV-based vectors encoding beta-galactosidase and an intracellular antibody directed against gp120 (sFv 105) or Tat. (sFvtat1-Ckappa). The expression of marker genes and the effects of the antiviral constructs were monitored in vitro in unselected transduced CD4+ T cells. Efficiency and stability of transduction varied during the course of HIV infection; CD4+ T cells derived from asymptomatic patients were transducible at higher efficiencies and stabilities than CD4+ T cells from patients with acquired immunodeficiency syndrome (AIDS). Expression of the anti-tat intracellular antibody was more effective at stably inhibiting HIV-1 replication in transduced cells from HIV-infected individuals than was sFv 105. The results of this study have important implications for the development of a clinically relevant gene therapy for the treatment of HIV-1 infection.     

Allopurinol hypersensitivity syndrome: hypersensitivity to oxypurinol but not allopurinol

OBJECTIVE: We assessed the clinical, histological, and virological features of anogenital human papillomavirus (HPV) infection, according to their immune status in HIV-1 infected men, referred for an anogenital examination or treatment, in comparison with immunocompetent patients. METHODS: The study population comprised 33 HIV-1 infected heterosexual or homosexual men and 38 HIV negative men seen in a screening and treatment centre for anogenital HPV infections. All patients were examined with a colposcope. Biopsies were carried out on all subjects with anogenital lesions for histological studies and HPV detection by Southern blot. RESULTS: The HIV infected patients had a balanopreputial HPV infection in 70%, anal in 30%, and urethral in 37%, while HIV negative patients had balanopreputial lesion in 72%, anal in 26%, and urethral in 16%. Diffuse anogenital lesions were present in 33% of the HIV infected cases and in 10.5% of HIV negative cases (p < 0.02). Among the HIV infected patients, the genital HPV lesions were condylomatous in 67.5% of the cases and dysplastic in 57%. HIV negative patients had condylomatous lesions in 86% of the cases and dysplasic in 14%. The condylomatous lesions of HIV infected patients had a low grade malignant histological aspect in 36% of the cases and high grade histological criteria were found in 22% of the dysplasias. Oncogenic HPVs were detected more frequently in HIV infected patients (35% v 12%) and more than one HPV type was found in 21.5% of cases. Neither the anogenital diffusion of the HPV lesions nor their morphological, histological, and virological features differed significantly in patient with CD4 cell counts > or < 200 x 10(6)/l. In contrast, patients with CD4 cell counts < 50 x 10(6)/l had a higher risk of several types of HPVs and of developing a diffuse anogenital infection. CONCLUSION: HIV-1 infected patients had an increased frequency of high grade anogenital dysplastic lesions and a higher frequency of HPV infection with multiple and diffuse sites of involvement. These characteristics of HPV infection were independent of the patients' immune status up to CD4 cell counts > 50 x 10(6)/l but showed an increased risk when the CD4 cell count was < 50 x 10(6)/l. The higher frequency of diffuse anogenital infections among HIV infected men calls for rapid treatment, laser or surgery, given the association of histological features of intraepithelial neoplasia and the presence of multiple HPV infection sites which may be the consequence of immune disturbances, most of which are transmissible potentially oncogenic HPVs.     

Detection of infection with human immunodeficiency virus type 1 before seroconversion: correlation with clinical symptoms and outcome

Early (pre-seroconversion) infection with human immunodeficiency virus type 1 (HIV-1) was identified in 50 of 267 participants in the Multicenter AIDS Cohort Study. These 50 men had a positive EIA result, which detected IgM antibody (n = 35), p24 antigen, or serum HIV RNA (n = 15) at their last "seronegative" visit. At that visit, the mean CD4 lymphocyte number (890/mm3 vs. 1038/mm3) was significantly lower than in men who subsequently seroconverted but had no evidence of early infection. The decline in CD4 cells was slower and the duration of AIDS-free time longer in the 19 men who were symptomatic in comparison to the 31 asymptomatic men with early infection, but differences were not significant.