Objective:To identify relationships between insulin resistance (IR) and mitochondrial respiration in perinatally HIV-infected youth.Design:Case-control study.Methods:Mitochondrial respiration was assessed in perinatally HIV-infected youth in Tanner stages 2-5, 25 youth with IR (IR+) and 50 without IR (IR-) who were enrolled in the Pediatric HIV/AIDS Cohort Study. IR was defined as a homeostatic model of assessment for IR value at least 4.0. A novel, high-throughput oximetry method was used to evaluate cellular respiration in peripheral blood mononuclear cells. Unadjusted and adjusted differences in mitochondrial respiration markers between IR+ and IR- were evaluated, as were correlations between mitochondrial respiration markers and biochemical measurements.Results:IR+ and IR- youth were similar on age, sex, and race/ethnicity. Mean age was 16.5 and 15.6 years in IR+ and IR-, respectively. The IR+ group had significantly higher mean BMI and metabolic analytes (fasting glucose, insulin, cholesterol, triglycerides, and venous lactate and pyruvate) compared with the IR-. Mitochondrial respiration markers were, on average, lower in the IR+ compared with IR-, including basal respiration (417.5 vs. 597.5pmol, P=0.074), ATP production (11513 vs. 15202pmol, P=0.078), proton leak (584.6 vs. 790.0pmol, P=0.033), maximal respiration (1815 vs. 2399pmol, P=0.025), and spare respiration capacity (1162 vs. 2017pmol, P=0.032). Nonmitochondrial respiration did not differ by IR status. The results did not change when adjusted for age.Conclusion:HIV-infected youth with IR have lower mitochondrial respiration markers when compared to youth without IR. Disordered mitochondrial respiration may be a potential mechanism for IR in this population.