Gut Microbiota and Endothelial Dysfunction Markers in Obese Mexican Children and Adolescents.

Obesity is a metabolic disease characterized by low-grade inflammation and accompanied by dyslipidemia and up-regulation of other bioactive molecules, creating a predisposition to endothelial dysfunction and metabolic syndrome. We studied the association between gut microbiota diversity and endothelial dysfunction (EDF) markers in obese Mexican children and adolescents. We examined clinical data including metabolic factors and EDF markers in blood samples. Gut bacterial diversity was characterized by high-throughput sequencing of V3-16S rDNA libraries. Triglycerides, insulin, homeostasis model assessment-insulin resistant (HOMA-IR), leptin, C-reactive protein (CRP), and EDF marker intercellular adhesion molecule 1 (ICAM-1) were significantly higher in obese children and adolescents. Multivariate analysis showed statistically significant positive associations between vascular cell adhesion molecule 1 (VCAM-1) and Veillonellaceae, and between ICAM-1 and Ruminococcus in obese children. In obese adolescents, there was a statistically significant positive association between total cholesterol and Ruminococcus, and between ICAM-1 and Bacteroides. LEfSe analysis showed that the genus Lactobacillus and family Coriobacteriaceae were enriched in children, and genera Collinsella and Prevotella were enriched in obese adolescents. Obese children and adolescents had higher levels of insulin resistance and metabolic syndrome. These results suggest that obese Mexican children and adolescents had increased levels of CRP and a reduction of adiponectin, which causes higher expression of EDF markers, affecting endothelial function and associating with changes in the gut microbiota.

Detection and quantification of the K103N mutation in HIV reverse transcriptase by pyrosequencing.

Prolonged treatment of human immunodeficiency virus (HIV)-infected patients with nonnucleoside reverse transcriptase inhibitors (NNRTIs) might result in the selection of resistant mutants, the most frequent being the K103N mutation in reverse transcriptase. Resistance mutations are routinely detected by Sanger sequencing of the whole viral population, which does not detect sequence variants with frequencies below 20%. We have developed a pyrosequencing approach for the analysis of codon 103 of the HIV reverse transcriptase gene in the circulating viral population that detects variants below the limit of conventional sequencing. The method was tested with samples from 5 controls (not exposed to NNRTIs), 6 from patients exposed to NNRTIs and having a K103N mutant virus population detected by conventional sequencing, and 9 from patients previously exposed to NNRTIs that had a wild-type virus population by conventional sequencing. In 7 of 9, samples the mutation could not be detected by either the standard assay or pyrosequencing, while in 2 samples persistence of the mutation could be detected by pyrosequencing. The method might be of practical use in detecting minority variants of HIV in the clinical setting, in epidemiological studies with large numbers of samples, or as a complement to more complex approaches.