Coffea stenophylla is a rare Coffea species boasting a flavor profile comparable to Arabica coffee (Coffea arabica) and has a good adaptability to lowland tropical climates. This species faces increasing threats from climate change, deforestation, and habitat fragmentation in its West African homeland. Using 1037 novel SNP markers derived from Genotyping-by-Sequencing (GBS), we revealed the presence of three distinct natural populations (mean Fst = 0.176) in Sierra Leone. Evidence of recent bottlenecks and small effective population size (118–140) was found across all three populations, reflecting the impact of recent anthropogenic disturbances on this species. Using a model-flexible inference approach, we unveiled a strong ancient bottleneck approximately 23,000 years ago, coinciding with the last glacial maximum (LGM), followed by post-glacial expansion and divergence into distinct genetic clusters. A comparative analysis between ex situ genebanks and natural populations detected a significant gap in genetic diversity, with two out of three natural populations missing from the ex situ genebank collection. These findings highlight the urgent need to improve conservation practices for C. stenophylla in Sierra Leone. The novel SNP markers developed in this study provided valuable tools to support future efforts in conservation and utilization of C. stenophylla genetic resources in West Africa.