Background:There is limited evidence on the optimal timing of antiretroviral therapy (ART) initiation in children 2-5 y of age. We conducted a causal modelling analysis using the International Epidemiologic Databases to Evaluate AIDS-Southern Africa (IeDEA-SA) collaborative dataset to determine the difference in mortality when starting ART in children aged 2-5 y immediately (irrespective of CD4 criteria), as recommended in the World Health Organization (WHO) 2013 guidelines, compared to deferring to lower CD4 thresholds, for example, the WHO 2010 recommended threshold of CD4 count <750 cells/mm3 or CD4 percentage (CD4%) <25%.Methods and Findings:ART-naïve children enrolling in HIV care at IeDEA-SA sites who were between 24 and 59 mo of age at first visit and with ≥1 visit prior to ART initiation and ≥1 follow-up visit were included. We estimated mortality for ART initiation at different CD4 thresholds for up to 3 y using g-computation, adjusting for measured time-dependent confounding of CD4 percent, CD4 count, and weight-for-age z-score. Confidence intervals were constructed using bootstrapping.The median (first; third quartile) age at first visit of 2,934 children (51% male) included in the analysis was 3.3 y (2.6; 4.1), with a median (first; third quartile) CD4 count of 592 cells/mm3 (356; 895) and median (first; third quartile) CD4% of 16% (10%; 23%). The estimated cumulative mortality after 3 y for ART initiation at different CD4 thresholds ranged from 3.4% (95% CI: 2.1-6.5) (no ART) to 2.1% (95% CI: 1.3%-3.5%) (ART irrespective of CD4 value). Estimated mortality was overall higher when initiating ART at lower CD4 values or not at all. There was no mortality difference between starting ART immediately, irrespective of CD4 value, and ART initiation at the WHO 2010 recommended threshold of CD4 count <750 cells/mm3 or CD4% <25%, with mortality estimates of 2.1% (95% CI: 1.3%-3.5%) and 2.2% (95% CI: 1.4%-3.5%) after 3 y, respectively. The analysis was limited by loss to follow-up and the unavailability of WHO staging data.Conclusions:The results indicate no mortality difference for up to 3 y between ART initiation irrespective of CD4 value and ART initiation at a threshold of CD4 count <750 cells/mm3 or CD4% <25%, but there are overall higher point estimates for mortality when ART is initiated at lower CD4 values.Please see later in the article for the Editors' Summary. © 2013 Schomaker et al.