Mt. Hakodate, Hokkaido, is a small mountain geographically isolated by the sea and urban area, and some red foxes (Vulpes vulpes) live there. In consideration of the specific geographical environment, we made a hypothesis that the fox population on Mt. Hakodate is small in size and isolated, and consequently its genetic variation is low, and then tested it. To investigate the genetic variation, we genotyped microsatellite loci using fecal samples collected noninvasively from 2009 to 2011. As the result, we successfully identified 35 foxes for the three years (12 in 2009, 11 in 2010, and 22 in 2011), and presumed eight of them were adults, indicating that the population size is extremely small. The genetic structures and degree of differentiations showed that the Mt. Hakodate population was genetically differentiated from those of the other populations. The urban area of Hakodate City could have played a role of a dispersal barrier for foxes. Additionally, the genetic diversity of the Mt. Hakodate population was lower than those of the other populations in Hokkaido. Our results suggested that the Mt. Hakodate population have low genetic diversity due to restricted gene flow and the small population size, supporting our hypothesis.