Epidemiologic Analysis

Table 1. Table 1. Baseline Characteristics of the Participants, According to Study Group.

In total, 223 participants were infected with HIV-1 after enrollment; 191 were infected with HIV-1 only (161 men and 30 women), and 32 were infected with both HIV-1 and HIV-2 (26 men and 6 women) (Table 1). Among the participants with dual infection, 12 had concomitant dual seroreactivity and 20 had HIV-2 seroreactivity before dual seroreactivity occurred. Dual infection and concomitant dual seroreactivity could have occurred by means of three potential infection scenarios: HIV-1 infection preceding dual infection, HIV-2 infection preceding dual infection, or simultaneous HIV-1 and HIV-2 infection. In the cohort and apart from the 32 participants with dual infection described above, 1 participant was infected with HIV-1 after enrollment but before conversion to dual seropositivity. This participant was excluded from the study because no conclusions could be drawn about this group from 1 person and because of the aim of the study: to investigate whether HIV-2 infection could affect the rate of HIV-1 disease progression during the complete course of HIV-1 infection (including the important acute phase, during which the level of infection is established).

The median time to AIDS was 104 months (95% confidence interval [CI], 75 to 133) in participants with both HIV-1 and HIV-2 infection and 68 months (95% CI, 60 to 76) in participants with HIV-1 infection only (P=0.003 by the log-rank test) (Fig. S1 in the Supplementary Appendix). Participants with HIV-1 infection only and those with dual infection did not differ significantly in age (P=0.10 by a two-tailed Student's t-test) or sex (P=0.61 by a two-tailed Fisher's exact test), and a similar proportion of participants in each of the two groups had data right-censored during the study period (Supplementary Appendix). Stratification according to sex showed a slightly more pronounced difference in the time to AIDS between the groups (P=0.001 by the log-rank test). The adjusted hazard ratio for progression to AIDS among HIV-1–infected participants versus participants with dual infection was 2.81 in a Cox proportional-hazards model controlling for age and sex (95% CI, 1.55 to 5.09; P<0.001 by the Wald test).

Figure 1. Figure 1. Kaplan–Meier Analysis of Time to the Development of AIDS, According to Study Group. Tick marks indicate participants with censored data. Asterisks indicate the time point in each group when five participants are still at risk for the development of AIDS.

To evaluate the effect of the order of infection (with HIV-2 preceding or being simultaneously recorded with HIV-1), we stratified the dual-infection group into the 20 participants with HIV-2 seroreactivity preceding dual seroreactivity and the 12 with simultaneous HIV-1 and HIV-2 seroreactivity recorded first. Kaplan–Meier analysis showed that the 20 participants who had HIV-2 seroreactivity first had a longer time to AIDS (median, 129 months [95% CI, 93 to 165]) than participants with HIV-1 infection only (P=0.007 by the log-rank test) (Figure 1). Even though the time to AIDS in the group with dual infection and simultaneous HIV-1 and HIV-2 seroreactivity recorded first was intermediate (88 months; 95% CI, 69 to 106), this time did not differ significantly from that in either the group with dual infection and HIV-2 seroreactivity first (P=0.50 by the log-rank test) or the group with HIV-1 infection only (P=0.13 by the log-rank test). The hazard ratio for progression to AIDS (after controlling for age and sex) was 3.12 (95% CI, 1.52 to 6.41; P=0.002 by the Wald test) for HIV-1 infection only as compared with dual infection with HIV-2 seroreactivity first and was 2.27 (95% CI, 0.90 to 5.68; P=0.08 by the Wald test) for HIV-1 infection only as compared with dual infection with simultaneous HIV-1 and HIV-2 seroreactivity recorded first. The hazard ratio for dual infection with simultaneous HIV-1 and HIV-2 seroreactivity recorded first as compared with dual infection with HIV-2 seroreactivity first was 1.37 (95% CI, 0.45 to 4.17; P=0.58 by the Wald test).

CD4+ and CD8+ T Cells

Table 2. Table 2. CD4+ or CD8+ T-Cell Percentage of the Total Lymphocyte Count, According to Study Group.

We used a full-factorial mixed model with time since seroconversion as the covariate to compare the extrapolated level at seroconversion and rate of decline in CD4+ T-cell percentage in participants infected with HIV-1 only and those infected with both HIV-1 and HIV-2. The rate of decline in CD4+ T-cell percentage was similar with HIV-1 infection only and dual infection, with an average decline of 1.2% per year (P=0.36, with the use of a mixed model with interaction term removed). However, the CD4+ T-cell percentage was significantly higher in participants with dual infection (31.3%) than in those with HIV-1 infection only (23.3%) (P<0.001) (Table 2). A stratified analysis showed similar decreases in CD4+ T-cell percentages in the group with dual infection and simultaneous HIV-1 and HIV-2 seroreactivity recorded first and the group with dual infection and HIV-2 seroreactivity first. The latter group had, however, a significantly higher CD4+ T-cell percentage (32.3%) than those with HIV-1 infection only (P<0.001). The group with dual infection and simultaneous HIV-1 and HIV-2 seroreactivity recorded first showed an intermediate CD4+ T-cell percentage (28.1%), with no significant difference from either the group infected with HIV-1 only or the group with dual infection and HIV-2 seroreactivity first (Table 2). Next, we examined differences in CD8+ T-cell percentages over time, finding a slower increase in CD8+ T-cell percentages among participants with dual infection (1.5% per year) than among those with HIV-1 infection only (3.0% per year, P=0.03) (Table 2). A significant difference was also found between HIV-1 infection only and dual infection with HIV-2 seroreactivity first (P=0.002) but not between HIV-1 infection only and dual infection with simultaneous HIV-1 and HIV-2 seroreactivity recorded first (P=0.52).

Since the percentages and rates of change in T-cell populations may differ among disease stages, we also analyzed T-cell counts during the asymptomatic stage of infection (defined as CD4+ T-cell counts >200 or >14% of the total). As found in the analysis of the complete data set, the rate of decline in CD4+ T-cell percentages was similar among participants with HIV-1 infection only and those with dual infection, with an average decline of 0.8% per year (P=0.20, with the use of a mixed model with interaction term removed). The CD4+ T-cell percentage at the extrapolated level at seroconversion was significantly higher with dual infection (30.1%) than with HIV-1 infection only (24.3%) (P=0.005) (Table S3 in the Supplementary Appendix). The stratified analysis showed rates of decreasing CD4+ T-cell percentages that were similar with dual infection with simultaneous HIV-1 and HIV-2 seroreactivity recorded first and dual infection with HIV-2 seroreactivity first. Again, the latter group had a significantly higher CD4+ T-cell percentage (31.0%) than the group infected with HIV-1 only (P=0.003), whereas the group with dual infection and simultaneous HIV-1 and HIV-2 seroreactivity recorded first had an intermediate percentage, which did not differ significantly from that in either the HIV-1–only group or the group with dual infection and HIV-2 seroreactivity first (Table S3 in the Supplementary Appendix).

Whereas CD8+ T-cell percentages differed significantly over time (i.e., throughout the stages of infection) between the group with HIV-1 infection only and the group with dual infection, there was no significant between-group difference in CD8+ percentages during the asymptomatic stage of infection (P=0.09) (Table S3 in the Supplementary Appendix). However, the rate of increase in CD8+ T-cell percentages was greater with HIV-1 infection only (2.7% per year) than with dual infection with HIV-2 seroreactivity first (0.9% per year, P=0.02) (Table S3 in the Supplementary Appendix). The differences in the changes in CD8+ T-cell percentages prompted us to analyze differences in the immune activation markers β 2 -microglobulin and neopterin; however, we did not find any significant differences between the participants infected with HIV-1 only and those infected with both HIV-1 and HIV-2 (see the Supplementary Appendix).

Molecular Evolution of HIV-1

HIV-1 evolution is characterized by high mutation rates, rapid viral turnover, and high recombination rates. The evolution can be quantified by diversity (the genetic variation at a given time point) and divergence (the genetic distance to a reference point, e.g., a founder strain). Several studies have presented evidence of a positive correlation between diversity and time from seroconversion during the asymptomatic stage of infection.21-23 HIV-1 diversity has also been positively correlated with viral load and viral fitness.24,25 The rate of divergence of HIV-1 has been shown to be relatively constant (reflected by a linear increase in divergence) during the asymptomatic stage of infection.22,26 Studies comparing divergence rates with disease progression rates show conflicting results.26,27

Figure 2. Figure 2. Disease Progression and Evolution of HIV-1 Diversity. Panel A shows the epidemiologic estimates of the time to the development of AIDS (time to AIDS) in participants with HIV-1 infection only (68 months) and those with HIV-1 and HIV-2 infection (dual infection) (104 months). The difference between the study groups was 36 months. Panel B shows the data from models on the rate of evolution of HIV-1 genetic diversity (the genetic variation at a given time point, as measured in substitutions per site) for HIV-1 infection only and dual infection. The indicated diversity threshold (as measured in substitutions per site) for development of AIDS is based on the 68-month time-to-AIDS estimate for participants with HIV-1 infection only (dashed horizontal line). We estimated the time to reach that threshold in participants with dual infection by using the global mean diversity rate (1.75×10–3 substitutions per site per year) to extrapolate from the mean diversity among participants with dual infection to the diversity threshold. For the time to AIDS in participants with dual infection, both the maximum-likelihood estimate (104.93 months) and the Bayesian estimates (104.95 months) were close to the epidemiologic estimate (104 months) (see Table S6 in the Supplementary Appendix).

Since maximum-likelihood and Bayesian analyses showed high concordance (Tables S5 and S6 in the Supplementary Appendix), we present only maximum-likelihood estimates here. The average increase in HIV-1 sequence diversity over time was similar in participants with HIV-1 infection only and those with dual infection, with an average of 1.75×10−3 substitutions per site per year (P=0.81 by the t-test). At similar time points after seroconversion, the diversity was significantly lower among participants with dual infection (5.67×10−3±1.61×10−3 substitutions per site) than among those with HIV-1 infection only (11.04×10−3±1.28×10−3 substitutions per site; P=0.01 by the t-test) (Figure 2).

Despite detailed evolutionary analyses, we found no significant differences between the HIV-1–infection group and the dual-infection groups in the rates of HIV-1 divergence or synonymous or nonsynonymous substitutions (Table S5 in the Supplementary Appendix).