Confirmation of the local establishment of alien invasive turtle, Pseudemys peninsularis, in South Korea, using eggshell DNA.

Alien invasive species are posing conservation challenges worldwide. Pet trade, one of the many ways, is worsening the situation. Especially, pet turtles have been released into nature due to their longer life span and peoples' religious and traditional beliefs. In addition, unwanted and undesired pets are also released. While information on the successful local establishment and subsequent dispersal into new habitats is required to designate an invasive and ecosystem-disturbing species, alien freshwater turtle nests have always been hard to find and identify in nature. Because one should identify nests by the eggs, which do not always guide properly, as adults abandon the sites quickly. We thought the recent advancement in DNA technology may help improve the situation. We studied Pseudemys peninsularis, one of the most traded freshwater turtle pet species, which has already been reported from a wide range of wild areas in South Korea. Yet, it is not designated as ecosystem-disturbing species due to a lack of adequate information on their local reproduction and establishment. We conducted surveys and found two nests in Jeonpyeongje Neighborhood Park, Maewol-dong, Seo-gu, Gwangju. We developed the methodology for extracting DNA from the eggshells and successfully identified the nests by phylogenetic analysis and verified through egg characteristics and morphological features of artificially hatched juveniles. This was the first successful initiative to extract DNA from freshwater turtle eggshells. We believe it will help future researchers identify the alien invasive turtle nests and develop their control and management policies. In addition, our study also included comparative descriptions and schematic diagrams of the eggs of eight freshwater turtles, including a native and three ecosystem-disturbing species, from South Korea. We urged an immediate designation of P. peninsularis as an ecosystem-disturbing species considering its local establishment, distribution range, and potential negative impact on native ecosystems