Differential gene expression of cytokines, receptors, and miRNAs in individuals living with HIV-1 and vaccinated against yellow fever.

Between 2016 and 2018, Brazil faced a yellow fever (YF) outbreak, which led to an expansion of vaccination coverage. The coexistence of the YF outbreak and the HIV-1 epidemic in Brazil raised concerns regarding the immune response and vaccine effectiveness in individuals living with HIV (PLWH). The aim of this study was to investigate the immune response to YF vaccination in PLWH and HIV-uninfected individuals as controls. Transcript levels of immunomodulatory molecules, including IL-6, IL-10, IL-21, TGF-ß, CD19, CD163, miR-21, miR-146, and miR-155, were measured using RTqPCR. TCD4+ cells were evaluated by cytometry, and neutralizing antibody (Nab) titers were detected by a micro plaque-reduction neutralization test. The findings of our study revealed several noteworthy observations. First, there was a notable reduction in the circulation of TCD4+ cells postvaccination. Among people living with HIV (PLWH), we observed an increase in the expression of IL-10 following vaccination, while IL-6 expression was diminished in PLWH with lower TCD4+ counts. Furthermore, we identified the downregulation of CD19 and TGF-ß, along with the upregulation of IL-21 and CD163. Notably, we observed positive correlations between the levels of IL-10/IL-21, IL-10/CD163, and IL-6/CD19. Additionally, there was a positive correlation between miRNAs 146 and 155. It is important to emphasize that all participants exhibited robust neutralizing antibody responses after receiving 17DD YF vaccination. In this context, the gene expression data presented can be useful for biomarker studies of protective antibodies induced by YF vaccination. This study sheds light on immune mechanisms in individuals living with HIV and YF vaccination.

Increased biomarkers of cardiovascular risk in HIV-1 viremic controllers and low persistent inflammation in elite controllers and art-suppressed individuals.

HIV controllers (HICs) are models of HIV functional cure, although some studies have shown persistent inflammation and increased rates of atherosclerosis in HICs. Since immune activation/inflammation contributes to the pathogenesis of cardiovascular diseases (CVD), we evaluated clinical data and inflammation markers in HIV-1 viremic controllers (VC), elite controllers (EC), and control groups (HIV positive individuals with virological suppression by antiretroviral therapy-cART; HIV negative individuals-HIVneg) to assess whether they presented elevated levels of inflammation markers also associated with CVD. We observed the highest frequencies of activated CD8+ T cells in VCs, while EC and cART groups presented similar but slightly altered frequencies of this marker when compared to the HIVneg group. Regarding platelet activation, both HICs groups presented higher expression of P-selectin in platelets when compared to control groups. Monocyte subset analyses revealed lower frequencies of classical monocytes and increased frequencies of non-classical and intermediate monocytes among cART individuals and in EC when compared to HIV negative individuals, but none of the differences were significant. For VC, however, significant decreases in frequencies of classical monocytes and increases in the frequency of intermediate monocytes were observed in comparison to HIV negative individuals. The frequency of monocytes expressing tissue factor was similar among the groups on all subsets. In terms of plasma markers, VC had higher levels of many inflammatory markers, while EC had higher levels of VCAM-1 and ICAM-1 compared to control groups. Our data showed that VCs display increased levels of inflammation markers that have been associated with CVD risk. Meanwhile, ECs show signals of lower but persistent inflammation, comparable to the cART group, indicating the potential benefits of alternative therapies to decrease inflammation in this group.

A case report of HIV-1 superinfection in an HIV controller leading to loss of viremia control: a retrospective of 10 years of follow-up.

BACKGROUND: HIV controllers (HICs) are a rare group of HIV-1-infected individuals able to naturally control viral replication. Several studies have identified the occurrence of HIV dual infections in seropositive individuals leading to disease progression. In HICs, however, dual infections with divergent outcomes in pathogenesis have been described. CASE PRESENTATION: Here, we present a case report of a HIC diagnosed in late 1999 who displayed stable CD4+ T cell levels and low plasmatic viral load across 12 years of follow-up. In early 2013, the patient started to present an increase in viral load, reaching a peak of 10,000 copies/ml in early 2014, followed by an oscillation of viremia at moderate levels in the following years. The genetic diversity of env proviral quasispecies from peripheral blood mononuclear cells (PBMCs) was studied by single genome amplification (SGA) at six timepoints across 2009-2017. Phylogenetic analyses of env sequences from 2009 and 2010 samples showed the presence of a single subtype B variant (called B1). Analyses of sequences from 2011 and after revealed an additional subtype B variant (called B2) and a subsequent dominance shift in the proviral quasispecies frequencies, with the B2 variant becoming the most frequent from 2014 onwards. Latent syphilis related to unprotected sexual intercourse was diagnosed a year before the first detection of B2, evidencing risk behavior and supporting the superinfection hypothesis. Immunologic analyses revealed an increase in CD8+ and CD4+ T cell immune activation following viremia increase and minor T cell subset alterations during follow-up. HIV-specific T cell responses remained low throughout the follow-up period. CONCLUSIONS: Altogether, these results show that loss of viremia control in the HIC was associated with superinfection. These data alert to the negative consequences of reinfection on HIV pathogenesis, even in patients with a long history of viremia control and an absence of disease progression, reinforcing the need for continued use of adequate prevention strategies.

Reduction of inflammation and T cell activation after 6 months of cART initiation during acute, but not in early chronic HIV-1 infection.

OBJECTIVES: To investigate the impact of early combined antiretroviral therapy (cART) on inflammation biomarkers and immune activation during acute and early chronic HIV-1 infection. METHODS: We included 12 acute (AHI), 11 early chronic (EcHI), and 18 late chronic HIV-1-infected (LcHI) individuals who were treated with cART and 18 HIV-1-uninfected (HIV-neg) individuals. Plasmatic levels of inflammation biomarkers, CD8+CD38+HLA-DR+ T cell frequencies, CD4 T cell counts, CD4/CD8 ratio, total HIV-1 DNA and plasmatic viral load were evaluated. Mann-Whitney test, Pearson and Spearman correlation, and linear regression models were used for statistical analyses. RESULTS: IP-10, IL-18, and sCD163 were significantly elevated at pre-ART in the AHI and EcHI groups, showing a significant reduction after 6 months of cART in the AHI group, achieving similar levels to the HIV-neg group. For the EcHI group, the IP-10 and sCD163 levels were also significantly reduced on M6-ART; however, IP-10 levels remained higher than in the HIV-neg group, and no significant reduction of IL-18 levels was observed. The CD8+ T cell activation levels were elevated in the AHI and EcHI groups at pre-ART and showed a significant reduction on M6-ART, but they were similar to levels seen for HIV-neg only after 12 months of cART. At pre-ART, IP-10 levels but not IL-18 levels were positively correlated with HIV-1 viral load in the AHI group. CONCLUSIONS: Early initiation of cART in HIV infection can reduce systemic inflammation, but the earlier normalization of the inflammation markers was only observed when cART was initiated in the acute phase of infection. A slower dynamic of reduction was observed for CD8+ T cell activation.

Next-generation sequencing analyses of the emergence and maintenance of mutations in CTL epitopes in HIV controllers with differential viremia control.

BACKGROUND: Despite the low level of viral replication in HIV controllers (HICs), studies have reported viral mutations related to escape from cytotoxic T-lymphocyte (CTL) response in HIV-1 plasma sequences. Thus, evaluating the dynamics of the emergence of CTL-escape mutants in HICs reservoirs is important for understanding viremia control. To analyze the HIV-1 mutational profile and dynamics of CTL-escape mutants in HICs, we selected 11 long-term non-progressor individuals and divided them into the following groups: (1) viremic controllers (VCs; n = 5) and (2) elite controllers (ECs; n = 6). For each individual, we used HIV-1 proviral DNA from PBMCs related to earliest (VE) and latest (VL) visits to obtain gag and nef sequences using the Illumina HiSeq system. The consensus of each mapped gene was used to assess viral divergence, and next-generation sequencing data were employed to identify SNPs and variations within and flanking CTL epitopes. RESULTS: Divergence analysis showed higher values for nef compared to gag among the HICs. EC and VC groups showed similar divergence rates for both genes. Analysis of the number of SNPs showed that VCs present more variability in both genes. Synonymous/non-synonymous mutation ratios were < 1 for gag among ECs and for nef among ECs and VCs, exhibiting a predominance of non-synonymous mutations. Such mutations were observed in regions encoding CTL-restricted epitopes in all individuals. All ECs presented non-synonymous mutations in CTL epitopes but generally at low frequency (< 1%); all VCs showed a high number of mutations, with significant frequency changes between VE and VL visits. A higher frequency of internal mutations was observed for gag epitopes, with significant changes across visits compared to Nef epitopes, indicating a pattern associated with differential genetic pressure. CONCLUSIONS: The high genetic conservation of HIV-1 gag and nef among ECs indicates that the higher level of viremia control restricts the evolution of both genes. Although viral replication levels in HICs are low or undetectable, all individuals exhibited CTL epitope mutations in proviral gag and nef variants, indicating that potential CTL escape mutants are present in HIC reservoirs and that situations leading to a disequilibrium of the host-virus relationship can result in the spread of CTL-escape variants.

Highly divergent patterns of genetic diversity and evolution in proviral quasispecies from HIV controllers.

BACKGROUND: Ongoing intra-host HIV-1 evolution has been shown in individuals that naturally suppress the viremia to low levels (HIV controllers) by the analysis of the RNA in plasma compartment. Detection of evolution at the DNA proviral compartment in HIV controllers, however, has been more challenging and the precise correlation between the systemic viral suppression level and rate of reservoir's reseeding in those individuals is not fully understood. In this sense, we examined the proviral DNA quasispecies by single genome amplification of the env gene in a cohort of 23 HIV controllers from Brazil, divided in three groups, according to the level of systemic viral suppression: (1) elite controllers with persistent undetectable viral load (PEC, n = 6); (2) elite controllers with occasional episodes of transient (51-400 copies/mL) viremia (EEC, n = 7); and (3) viremic controllers with persistent low-level (80-2000 copies/mL) viremia (VC, n = 10). RESULTS: The HIV-1 diversity of the PBMC-associated proviral quasispecies in EC was significantly (P < 0.01) lower than in VC, but not significantly different between PEC and EEC groups. We detected a considerable variation in the average pairwise nucleotide distance and proportion of unique sequences in the HIV-1 proviral quasispecies of PEC and EEC. Some PEC and EEC displayed highly homogenous proviral populations with large clusters of identical sequences, while others exhibited relatively diverse proviral populations with a high proportion of unique sequences comparable to VC subjects. The long-term (10-15 years) follow-up of the HIV-1 proviral populations revealed a complete evolutionary stasis in one PEC and measurable divergence rates in one EEC [3.1 (1.2-5.6) × 10-3 substitutions/site/year and one VC [2.9 (0.7-5.1) × 10-3 substitutions/site/year]. CONCLUSIONS: There is no simple relationship between systemic viral suppression and intra-host proviral diversity or rate of reservoir's reseeding in chronically infected HIV controllers. Our results demonstrate that very divergent patterns of intra-host viral diversity and divergence could be detected in the setting of natural suppression of HIV-1 replication and that ongoing evolution and reseeding of the PBMC proviral reservoir occurs in some elite controllers.

Association of single nucleotide polymorphisms in the lens epithelium-derived growth factor (LEDGF/p75) with HIV-1 infection outcomes in Brazilian HIV-1+ individuals.

The lens epithelium-derived growth factor p75 (LEDGF/p75), coded by the PSIP1 gene, is an important host co-factor that interacts with HIV-1 integrase to target integration of viral cDNA into active genes. The aim of this study was to investigate the association of SNPs in the PSIP1 gene with disease outcome in HIV-1 infected patients. We performed a genetic association study in a cohort of 171 HIV-1 seropositive Brazilian individuals classified as rapid progressors (RP, n = 69), typical progressors (TP, n = 79) and long-term nonprogressors (LTNP, n = 23). The exonic SNP rs61744944 and 9 tag SNPs were genotyped. A group of 192 healthy subjects was analyzed to determine the frequency of SNPs and haplotypes in the general population. Linkage disequilibrium (LD) analyses indicated that the SNPs analyzed were not in high LD (r2<0.8). Logistic regression models suggested that patients carrying the T allele rs61744944 (472L) were more likely to develop a LTNP phenotype (OR = 4.98; p = 0.05) as compared to TP group. The same trend was observed when LTNPs were compared to the RP group (OR = 3.26). Results of haplotype analyses reinforced this association, since the OR values obtained for the haplotype carrying allele T at rs61744944 also reflected an association with LTNP status (OR = 6.05; p = 0.08 and OR = 3.44; p = 0.12 for comparisons to TP and RP, respectively). The rare missense variations Ile436Ser and Thr473Ile were not identified in the patients enrolled in this study. Gene expression analyses showed lower LEDGF/p75 mRNA levels in peripheral blood mononuclear cells obtained from HIV-1 infected individuals. However, these levels were not influenced by any of the SNPs investigated. In spite of the limited number of LTNPs, these data suggest that the PSIP1 gene could be associated with the outcome of HIV-1 infection. Further analyses of this gene may guide the identification of causative variants to help predict disease course.