We isolated and characterized seven microsatellite markers including di-, tri-, and tetra-nucleotide repeats for Cynops ensicauda, which is a near-threatened species and endemic to the Central Ryukyus, Japan, using next-generation sequencing. Six out of the seven loci that showed polymorphism in Cynops ensicauda popei on Okinawajima Island were also amplified in C. e. ensicauda on Amamioshima Island. Expected heterozygosities of these markers were 0.00–0.738 (mean, 0.429) and 0.00–0.791 (mean, 0.459) in C. e. popei and C. e. ensicauda, respectively. These markers will be useful for genetic studies, such as evaluating the genetic diversity within and between populations of this species.