To understand the effect of different farming systems on the dynamics of diversity of sorghum (Sorghum bicolor (L.) Moench) crop, genetic structure of widely used landraces and modern varieties collected from two contrasting agroecosystems, in eastern Sudan and western Kenya, were analysed with 16 polymorphic microsatellite markers. A total of 1104 accessions, grouped into 46 samples from individual farmers, were genotyped. Cluster analysis of the samples from the two countries displayed contrasting patterns. Most strikingly, differently named landraces from western Kenya formed widely overlapping clusters, indicating weak genetic differentiation, while those from eastern Sudan formed clearly distinguishable groups. Similarly, samples of the modern variety from Sudan displayed high homogeneity, whereas the most common modern variety from western Kenya was very heterogeneous. The high degree of fragmentation of farmlands of western Kenya, coupled with planting of different sorghum varieties in the same fields, increases the likelihood of inter-variety gene flow. This may explain the low genetic differentiation between the differently named landraces and heterogeneity of the modern variety from western Kenya. This study highlights the important role of farmers in shaping the genetic variation of their crops and provides population parameter estimates allowing forecasting of the fate of ‘modern’ germplasm (conventional or genetically modified) when introduced into subsistence farming systems.