Africa hosts the greatest human genetic diversity globally, but legacies of ancient population interactions and dispersals across the continent remain understudied. Here, we report genome-wide data from 20 ancient sub-Saharan African individuals, including the first reported ancient DNA from the DRC, Uganda, and Botswana. These data demonstrate the contraction of diverse, once contiguous hunter-gatherer populations, and suggest the resistance to interaction with incoming pastoralists of delayed-return foragers in aquatic environments. We refine models for the spread of food producers into eastern and southern Africa, demonstrating more complex trajectories of admixture than previously suggested. In Botswana, we show that Bantu ancestry post-dates admixture between pastoralists and foragers, suggesting an earlier spread of pastoralism than farming to southern Africa. Our findings demonstrate how processes of migration and admixture have markedly reshaped the genetic map of sub-Saharan Africa in the past few millennia and highlight the utility of combined archaeological and archaeogenetic approaches.