One hundred and nine Malus sieversii accessions from four geographical populations located at Kuerdening in Mohe town, Gongliu County, Jiaowutuohai, in Xinyuan County, Daxigou in Houcheng County of Ily State, and Baerluke Mountain in Yumin County of Tacheng State, Xinjiang Uygur Autonomous Region of China were studied by SSR markers. The purpose of the study was to determine the genetic structure and diversity in these eco-geographical populations with eight pair SSR primers of apple. The results indicated that: an average of 16 bands was detected in the four populations. The percentage of polymorphic bands in Gongliu population (89.06%) was the highest in the four populations. The average Nei's gene diversity index was 0.257 for all the loci. Totally, 128 polymorphic loci were detected and the percentage of polymorphic loci (P) was 100%, 88.28%, 84.83%, 87.50%, and 87.12%, respectively, at the species level and Gongliu, Xinyuan, Huocheng, and Yumin population levels. The Nei's gene diversity index (H = 0.2619) and Shannon's information index (I = 0.4082) in the species level were higher than in the population level. The Nei's gene diversity index and Shannon's information index in the four populations were Gongliu > Huocheng > Xinyuan > Yumin. Gongliu population and Xinyuan population were the highest in genetic identity and the closest in genetic distance. Gene flow between the populations was 7.265 based on genetic differentiation coefficient (G(ST) = 0.064). The UPGMA cluster analysis indicated that the genetic relationships between the Gongliu and Xinyuan population were the closest, and the Yumin population were the farthest with the other three populations. The UPGMA cluster analysis indicated that the four geographical populations located in Gongliu, Xinyuan, Huocheng, and Yumin were relatively independent populations. Concurrently, there was also mild gene exchange between the populations. On the basis of the study of population genetic structure and the highest genetic diversity, Gongliu population should be given a high priority consideration in Malus sieversii population's in situ germplasm conservation.