Aspergillus fumigatus is the primary pathogen causing aspergillosis. Recent molecular population genetic studies have demonstrated that A. fumigatus exhibits high local genetic diversity and with evidence for limited differentiation among geographic populations. However, research on the impacts of geomorphological factors on shaping the population diversity patterns of this species remains scarce. In this study, large-scale sampling and in-depth population genetic analysis were performed on soil-derived A. fumigatus from Guizhou Province, a representative karst landscape in southern China. This area is dominated by plateaus and mountains (accounting for 92.5% of the total area) and rep-resents a classic example of conical karst landscapes. A total of 206 A. fumigatus strains were isolated from 9 sampling sites across Guizhou. Genetic diversity, genetic differen-tiation, and population structure of these strains were analyzed based on 9 loci short tandem repeats (STRs). The results revealed that A. fumigatus in the karst region of Guizhou harbors abundant novel alleles and genotypes, with high genetic diversity. Gene flow among geographical populations was infrequent, and significant genetic differentiation was detected between multiple pairs of geographical populations, with the overall regional genetic differentiation reaching PhiPT = 0.061. Furthermore, the Guizhou populations showed significant differences from those reported in other regions world-wide. Surprisingly, only one of the 206 (0.49%) A. fumigatus isolates from this region exhibited resistance to the two medical triazoles commonly used for treating aspergillosis, and this resistance frequency was far lower than those reported in previous studies from other regions