In recent decades, invasive animals have disturbed the native ecosystem, via intraspecific and interspecific genetic hybridization. Previously, maternally-inherited mitochondrial DNA analysis detected two genetically distinct lineages of the sika deer (Cervus nippon) in the Japanese Archipelago and revealed the coexistence of both lineages mainly in the central part of Toyama Prefecture in Japan. One of the lineages was thought to be derived from introduced individuals. In the present study, using 13 microsatellite DNA markers, the level of hybridization between the two lineages was investigated in Toyama Prefecture. The results revealed deviation from the Hardy-Weinberg equilibrium in both the entire population and individuals from maternally-inherited samples. The Bayesian estimation elucidated the contribution of genetically divergent ancestral sources to the studied population due to the introgression of introduced gene pool and subsequent continuous mating across generations.