BACKGROUND: Data on the safety and pharmacokinetics of dolutegravir in children with HIV and tuberculosis are scarce. We aimed to determine the pharmacokinetics and safety of dolutegravir 50 mg twice daily in children receiving rifampicin, and to predict exposures for once-daily dolutegravir with rifampicin. METHODS: ORCHID is an open-label, sequential, prospective cohort study in children (<
18 years) weighing 20-35 kg initiated on a rifampicin-based tuberculosis regimen and dolutegravir in Durban, South Africa. We collected seven plasma samples over one dosing interval from each patient while on dolutegravir 50 mg twice daily during tuberculosis treatment and while on dolutegravir 50 mg once daily after tuberculosis treatment discontinuation. Pharmacokinetic data were analysed using population modelling in NONMEM version 7.5. The final model was used to perform Monte Carlo simulations in silico of once-daily dolutegravir dosing and time below target concentration (0·064 mg/L). Participants underwent regular clinical and safety visits. HIV viral load was measured at weeks 8, 12, 24, and 48. Primary outcomes were trough concentration (C FINDINGS: Between Aug 19, 2021, and Aug 17, 2023, we enrolled and followed up 13 children, with a median weight of 23·8 kg (IQR 21·7-24·8) and median age 10 years (range 5·9-13·0). Seven were male, six female, and 13 Black. Typical dolutegravir clearance was 0·584 L/h (95% CI 0·492-0·724), with an increase in clearance of 99·1% (73·2-120) with rifampicin. Median C INTERPRETATION: Twice-daily dolutegravir with rifampicin in children weighing 20-35 kg achieved therapeutic concentrations and was well tolerated with high rates of viral suppression. Simulations suggest that once-daily dolutegravir during rifampicin co-administration in children weighing 20-35 kg should be investigated in clinical studies. FUNDING: National Institutes of Health and South African Medical Research Council.