[Objective] The spatial variation pattern of ecosystem service importance in karst area of the Yunnan-Guizhou Plateau was examined to provide statistical support for effective management of the ecosystem and maintenance of an ecologically safe pattern in the area. [Methods] The values of three ecosystem services (water conservation, soil and water conservation, and biodiversity conservation) within Qianxinan Prefecture in 2019 were calculated using geographic information technology, and the spatial variation pattern was identified. Spatial heterogeneity of the ecosystem service importance in the region was pinpointed by spatial autocorrelation analysis. [Results] ① The overall ecological health of Qianxinan Prefecture was good. There was a large proportion of land with high ecosystem service importance, providing good ecological services for the region. The proportion of land providing water conservation function and biodiversity conservation function were larger than the proportion of land providing soil and water conservation function. ② Taking the village as a unit of measurement, the Moran’s I index was significantly high (0.65) in valuing the three ecosystem services both individually and aggregately. The study revealed that the research region featured strong spatial clustering with either “strip-shaped” or “block-shaped” ecosystem service clusters, and the functions and values of each cluster differred. ③ Spatial autocorrelation and clustering analyses of biodiversity conservation, water conservation, and soil and water conservation showed similar results. ④ Ecosystem service importance was coupled with soil and water erosion and rocky desertification. Hydrothermal conditions, geological processes, human impacts, and other factors contributed to such spatial variation. [Conclusion] The researched region possesses strong ecosystem service supply capacity with strong spatial clustering and heterogeneity. Such features should be considered in adopting ecosystem management and governance policies. The region should be subcategorized based on the spatial variation patterns of its ecosystem service values, and governed with tailored policies.