Viral load and CD4+ T-cell dynamics in primary HIV-1 subtype C infection

Abstract

Background: Most knowledge of primary HIV-1 infection is based on subtype B studies, whereas the evolution of viral parameters in the early phase of HIV-1 subtype C infection is not well characterized.

Methods: The kinetics of viral RNA, proviral DNA, CD4+ T-cell count, and subsets of CD4+ T cells expressing CCR5 or CXCR4 were characterized in 8 acute and 62 recent subtype C infections over the first year postseroconversion.

Results: The viral RNA peak was 6.25 +/- 0.92 log10 copies per milliliter. After seroconversion, heterogeneity among acute cases was evident by patterns of change in viral load and CD4+ T-cell count over time. The patterns were supported by the rate of viral RNA decline from peak (P = 0.022), viral RNA means (P = 0.005), CD4 levels (P < 0.001), and CD4 decline to 350 (P = 0.011) or 200 (P = 0.046). Proviral DNA had no apparent peak and its mean was 2.59 +/- 0.69 log10 per 106 peripheral blood mononuclear cell. In recent infections, viral RNA set point was 4.00 +/- 0.97 log10 and viral RNA correlated inversely with CD4+ T cells (P < 0.001) and directly with proviral DNA (P < 0.001).

Conclusions: Distinct patterns of viral RNA evolution may exist shortly after seroconversion in HIV-1 subtype C infection. The study provides better understanding of the early phase of subtype C infection.

FIGURE 1

Primary HIV-1 infection cohort. Two sources for identification of acute and recent HIV-1 infections are presented: a postnatal cohort and voluntary counseling and testing (VCT)–based referrals. A total of 324 HIV-negative postnatal women were enrolled in the prospective cohort in Botswana and were followed for up to 12 months; 2 cases were enrolled in the primary HIV-1 infection cohort (1 acute and 1 recent infection) from March 2004 to March 2005. The VCT-based referrals have been described elsewhere. A total of 8 acute and 70 recent HIV-1 infections were enrolled by VCT-based referrals from November 2004 to December 2007. One acutely and 9 recently infected enrollees completed fewer than 4 study visits and were excluded from analysis. Therefore, a total of 8 acute and 62 recent HIV-1 infections including 1 acute and 1 recent case enrolled in the postnatal cohort were analyzed in this study.

FIGURE 2

ML phylogenetic tree provides evidence that all subjects genotyped in this study were infected with HIV-1 subtype C. Eight of 8 acutely infected subjects and 41 of 62 recent infections are presented. Three sequences per subject spanning the V1C5 region of env were included in the analysis. The tree was generated by PhyML using the HKY model, and visualized in FigTree v.1.1.2. HIV-1 subtype C reference sequences are highlighted by circles. The HIV-1 M group non–subtype C references are shown at the bottom.

FIGURE 3

Intrapatient evolution of viral RNA, proviral DNA, and CD4⁺ T cells in acute HIV-1 subtype C infection. The timeline shows days from detected seroconversion. Time 0 corresponds to the first seropositive test. Plasma viral RNA load is expressed as log₁₀ copies per milliliter of plasma. Measurements of viral RNA before time 0 are preseroconversion. Cell-associated proviral DNA load is expressed as log₁₀ copies per million of PBMC. T cells expressing CCR5 and CXCR4 coreceptors are shown as percentage of CD4⁺ T cells. The percentage of CCR5 and CXCR4 on the surface of CD4⁺ T cells do not necessarily add up to 100%, reflecting a continuum of fluorescent intensity of CCR5⁺CD4⁺ and CXCR4⁺CD4⁺ T cells. Numbers at top correspond to patient cases. Dotted lines with arrows indicate initiation of ART.

FIGURE 4

Summary of viral RNA evolution in acute HIV-1 subtype C infection. The timeline shows days from detected seroconversion. Time 0 corresponds to the first seropositive test. Plasma viral RNA load is expressed as log₁₀ copies per milliliter of plasma. Post-ART data were excluded. A, viral RNA measurements in subjects with slow decline of viral RNA—1811, 2865, 3312, and 5018—are shown with solid circles. Measurements in subjects with fast decline of viral RNA—3430, 3505, 3603, and 5582—are shown with open circles. B, Summary of viral RNA levels per subject within the first 2monthspostseroconversion. C,Summary of viral RNA levels per subject from 2 to 6 months postseroconversion. D, Kaplan–Meier curve of CD4⁺ T cells decline to 350 between groups with slow (red) and fast (green) RNA decline. E, Kaplan–Meier curve of CD4⁺ T cells decline to 200 between groups with slow (red) and fast (green) RNA decline.

FIGURE 5

Viral RNA (A–D), proviral DNA (E–H), and CD4⁺ T cells (I–L) in recent HIV-1 subtype C infection, n = 62. The timeline shows days from detected seropositivity after adjusting for Fiebig stage. Plasma viral RNA load is expressed as log₁₀ copies per milliliter of plasma. Proviral DNA load is expressed as log₁₀ copies per million of PBMC. In the box plots (C, G, and K), the boundary of the box closest to zero indicates the 25th percentile, a solid line within the box marks the mean value, a dashed line within the box shows the median, and the boundary of the box farthest from zero indicates the 75th percentile. Whiskers above and below the box indicate the 10th and 90th percentiles. Points above and below the whiskers indicate outliers outside the 10th and 90th percentiles. A, Viral RNA. Regression curve represents summarized viral set point. The distribution of dots at the bottom of the graph reflects the threshold for detection of viral RNA by standard reverse transcriptase–polymerase chain reaction (400 copies/mL) and ultrasensitive reverse transcriptase–polymerase chain reaction (50 copies/mL). B, Individual regression curves of viral RNA over follow-up. C, Viral RNA means box plot. D, Distribution of viral RNA means. E, Proviral DNA. Regression curve represents summarized proviral DNA set point. F, Individual regression curves of proviral DNA over follow-up. G, Proviral DNA means box plot. H, Distribution of proviral DNA means. I, CD4⁺ T-cell counts. Regression curve represents summarized CD4⁺ T-cell counts. J, Individual regression curves of CD4⁺ T-cell counts over follow-up. K, CD4⁺ T-cell means box plot. L, Distribution of CD4⁺ T-cell count means.

FIGURE 6

Associations in recent HIV-1 subtype C infection, n = 62 (linear regression analyses). Data is adjusted for Fiebig stage. A, Direct correlation between means of viral RNA and proviral DNA. B, Inverse correlation between means of CD4⁺ T-cell counts and viral RNA. C, Inverse correlation between means of CD4⁺ T-cell counts and proviral DNA. D, Direct correlation between means and range of CD4⁺ T-cell counts. E, Inverse correlation between age and viral RNA means.

FIGURE 7

CD4⁺ T-cell counts and proviral DNA within viral RNA quartiles, n = 62. Data is adjusted for Fiebig stage. Boundaries of viral RNA quartiles are shown at the bottom of each box plot: the first quartile included patients with mean RNA load of more than 4.74 log₁₀ copies per milliliter, the second quartile was >4.10 log₁₀ and ≤4.74 log₁₀, the third quartile was >3.36 log₁₀ and ≤4.10 log₁₀, and the fourth quartile was ≤3.36 log₁₀. In the box plots, the boundary of the box closest to zero indicates the 25th percentile, a line within the box marks the median, and the boundary of the box farthest from zero indicates the 75th percentile. Whiskers above and below the box indicate the 10th and 90th percentiles. Points above and below the whiskers indicate outliers outside the 10th and 90th percentiles. Comparisons between groups are based on t tests or the Mann–Whitney sum rank test if the normality test failed within the group. The P values obtained by Mann–Whitney sum rank test are shown with asterisks. A, CD4⁺ T-cell counts. 95% CIs for difference of means are −474 to −151 between the first and fourth quartiles. B, Proviral DNA. 95% CIs for difference of means are 0.38 to 1.19 between the first and third quartiles, 0.72–1.47 between the first and fourth quartiles, 0.02–0.75 between the second and third quartiles, 0.39–1.01 between the second and fourth quartiles, and 0.01–0.61 between the third and fourth quartiles.