HIV-1C env and gag Variation in the Cerebrospinal Fluid and Plasma of Patients with HIV-Associated Cryptococcal Meningitis in Botswana.

HIV-1 compartmentalization in reservoir sites remains a barrier to complete HIV eradication. It is unclear whether there is variation in HIV-1 env and gag between cerebrospinal fluid (CSF) and plasma of individuals with HIV-associated cryptococcal meningitis (CM). We compared HIV-1 env characteristics and the gag cytotoxic T-lymphocyte (CTL) escape mutations from CSF and plasma samples. Employing population-based Sanger sequencing, we sequenced HIV-1 env from CSF of 25 patients and plasma of 26 patients. For gag, 15 CSF and 21 plasma samples were successfully sequenced. Of these, 18 and 9 were paired env and gag CSF/plasma samples, respectively. There was no statistically significant difference in the proportion of CCR5-using strains in the CSF and plasma, (p = 0.50). Discordant CSF/plasma virus co-receptor use was found in 2/18 pairs (11.1%). The polymorphisms in the HIV-1 V3 loop were concordant between the two compartments. From the HIV-1 gag sequences, three pairs had discordant CTL escape mutations in three different epitopes of the nine analyzed. These findings suggest little variation in the HIV-1 env between plasma and CSF and that the CCR5-using strains predominate in both compartments. HIV-1 gag CTL escape mutations also displayed little variation in CSF and plasma suggesting similar CTL selective pressure.

Quantification of entry phenotypes of macrophage-tropic HIV-1 across a wide range of CD4 densities.

Defining a macrophage-tropic phenotype for HIV-1 to assess a role in pathogenesis is complicated by the fact that HIV-1 isolates vary continuously in their ability to enter monocyte-derived macrophages (MDMs) in vitro, and MDMs vary in their ability to support HIV-1 entry. To overcome these limitations, we identified consistent differences in entry phenotypes between five paired blood-derived, T cell-tropic HIV-1 env genes, four of which are CCR5-using (R5) and one of which is CXCR4-using (X4), and cerebrospinal fluid (CSF)-derived, R5 macrophage-tropic env genes. We performed entry assays using the CD4- and CCR5-inducible Affinofile cell line, expressing a range of CD4 levels that approximates the range from MDMs to CD4(+) T cells. The macrophage-tropic viruses were significantly better at infecting cells expressing low levels of CD4 than the T cell-tropic viruses from the same subjects, with the titration of CD4 providing a distinctive and quantitative phenotype. This difference in CD4 utilization was not due to macrophage-tropic viruses being CD4 independent. Furthermore, macrophage-tropic viruses did not differ from paired T cell-tropic viruses in their ability to use low levels of CCR5 (tpaired = -1.39; P = 0.24) or their use of an alternative conformation of CCR5. We also infected MDMs with a panel of viruses and observed that infectivity of each virus differed across four donors and between three preparations from a single donor. We concluded that the evolutionary transition from replication in T cells to that in macrophages involves a phenotypic transition to acquire the ability to infect cells expressing low levels of CD4 and that this phenotype is more reliably measured in Affinofile cells than in macrophages. IMPORTANCE HIV-1 typically infects memory T cells by using CD4 and CCR5 to enter cells. The virus evolves to infect new cell types by changing the coreceptor from CCR5 to CXCR4 to infect naive T cells or adapting to the use of low levels of CD4 to infect macrophages. However, defining the phenotype of macrophage tropism has been difficult due to inherent variability in the use of macrophages generated in culture to support entry of HIV-1. We describe the use of Affinofile cells with inducible and variable levels of CD4 to identify a signature phenotype for macrophage-tropic HIV-1. The ability to define HIV-1 variants that have evolved an entry phenotype that allows more efficient entry into cells with low levels of CD4 sets the stage for a clearer placement of these variants in HIV-associated pathogenesis.

Discordant genotypic resistance and HIV-1 genetic diversity from paired plasma and cerebrospinal fluid samples in Chinese settings.

In the highly active antiretroviral therapy era, the incidence of human immunodeficiency virus (HIV)-associated neurocognitive disorder remains high with the improved survival. The prevalence of resistance differs across geographical areas and HIV subtypes. Currently, little information on the resistance patterns in the central nervous system (CNS) is available in Chinese settings. In this study, we sequenced and analyzed the pol gene from paired cerebrospinal fluid and plasma samples of 34 Chinese HIV-infected patients. We found that eight subjects harbored mutations that confer drug resistance, and of these, six subjects exhibited discordant resistance patterns between the CNS and the blood. The levels of viral diversity in the CNS were significantly higher than those in the blood (p < 0.0001). Our results contribute to improving our understanding of HIV neuropathogenesis and help to optimize neuro-acquired immunodeficiency syndrome treatment.

Genetic analysis of HIV type 1 env gene in cerebrospinal fluid and plasma of infected Chinese paid blood donors.

HIV infection in the central nervous system (CNS) can progress to AIDS dementia complex (ADC). Currently, the HIV-1 env gene in the CNS of infected Chinese paid blood donors (PBDs) has not been well characterized. In the study, the C2-V5 regions of the HIV env gene were cloned and sequenced from both cerebrospinal fluid (CSF) and plasma samples of six HIV-infected Chinese PBDs. Sequence analysis revealed that the sequences from Henan province clustered closely with subtypes B' and B, and the levels of diversity from the CNS were significantly lower than those from blood (p<0.0001). In addition, all viral quasispecies from CNS use CCR5 as the coreceptor. These data provide valuable information on HIV pathogenesis in the CSF and plasma of infected Chinese PBDs, and our findings could enhance insights into HIV-associated neurological disease.