Assessing the applicability of sequential eye lens stable isotope analysis for reconstructing dietary histories in amphibians

Retrospective isotope analysis using metabolically inert tissues is a powerful tool for reconstructing historical environmental conditions experienced by animals. Although this technique has been successfully applied to fish and squids using eye lenses, its applicability to metamorphosing organisms, such as amphibians, across multiple life-history stages has not been thoroughly assessed. In this study, we examined whether stable isotope ratios in frog eye lenses reflect dietary data from both the larval and post-metamorphic stages through feeding trials of two species, Rana ornativentris and Babina subaspera, using diets with distinct δ¹³C values. Additionally, we analyzed samples collected from the wild and assessed the potential future use of this method. Our feeding trials demonstrated that δ¹³C values in the central and outer lens sections were closely aligned with the larval and adult food, respectively, confirming the applicability of the method. The transition point of stable isotope ratios was closely aligned with the actual metamorphosis point, indicating that this method can effectively identify metamorphosis. However, the two wild specimens exhibited no discernible patterns in isotope ratios across their life stages. This indicates that applying this method in the field requires careful selection of the environmental conditions and a comprehensive understanding of the stable isotope ratios of potential food sources.