Standardized comparison of the relative impacts of HIV-1 reverse transcriptase (RT) mutations on nucleoside RT inhibitor susceptibility.

Determining the phenotypic impacts of reverse transcriptase (RT) mutations on individual nucleoside RT inhibitors (NRTIs) has remained a statistical challenge because clinical NRTI-resistant HIV-1 isolates usually contain multiple mutations, often in complex patterns, complicating the task of determining the relative contribution of each mutation to HIV drug resistance. Furthermore, the NRTIs have highly variable dynamic susceptibility ranges, making it difficult to determine the relative effect of an RT mutation on susceptibility to different NRTIs. In this study, we analyzed 1,273 genotyped HIV-1 isolates for which phenotypic results were obtained using the PhenoSense assay (Monogram, South San Francisco, CA). We used a parsimonious feature selection algorithm, LASSO, to assess the possible contributions of 177 mutations that occurred in 10 or more isolates in our data set. We then used least-squares regression to quantify the impact of each LASSO-selected mutation on each NRTI. Our study provides a comprehensive view of the most common NRTI resistance mutations. Because our results were standardized, the study provides the first analysis that quantifies the relative phenotypic effects of NRTI resistance mutations on each of the NRTIs. In addition, the study contains new findings on the relative impacts of thymidine analog mutations (TAMs) on susceptibility to abacavir and tenofovir; the impacts of several known but incompletely characterized mutations, including E40F, V75T, Y115F, and K219R; and a tentative role in reduced NRTI susceptibility for K64H, a novel NRTI resistance mutation.

Molecular characterization of stool microbiota in HIV-infected subjects by panbacterial and order-level 16S ribosomal DNA (rDNA) quantification and correlations with immune activation.

BACKGROUND: The relationship between gut microbial community composition at the higher-taxonomic order level and local and systemic immunologic abnormalities in HIV disease may provide insight into how bacterial translocation impacts HIV disease. METHODS: Antiretroviral-naive patients with HIV underwent upper endoscopy before and 9 months after starting antiretroviral treatment. Duodenal tissue was paraffin-embedded for immunohistochemical analysis and digested for fluorescence activated cell sorting for T-cell subsets and immune activation (CD38+/HLA-DR+) enumeration. Stool samples were provided from patients and control subjects for comparison. Metagenomic microbial DNA was extracted from feces for optimized 16S ribosomal RNA gene (rDNA) real-time quantitative polymerase chain reaction assays designed to quantify panbacterial loads and the relative abundances of proinflammatory Enterobacteriales order and the dominant Bacteroidales and Clostridiales orders. RESULTS: Samples from 10 HIV subjects before initiating and from six subjects receiving antiretroviral treatment were available for analysis. There was a trend for a greater proportion of Enterobacteriales in HIV-positive subjects compared with control subjects (P = 0.099). There were significant negative correlations between total bacterial load and duodenal CD4 and CD8 T-cell activation levels (r = -0.74, P = 0.004 and r = -0.67, P = 0.013, respectively). The proportions of Enterobacteriales and Bacteroidales were significantly correlated with duodenal CD4 T-cell depletion and peripheral CD8 T-cell activation, respectively. CONCLUSIONS: These data represent the first report of quantitative molecular and cellular correlations between total/universal and order-level gut bacterial populations and gastrointestinal-associated lymphoid tissue levels of immune activation in HIV-infected subjects. The correlations between lower overall 16S rDNA levels and tissue immune activation suggest that the gut microbiome may contribute to immune activation and influence HIV progression.