DNA methylation-based ageing in a deuterostome invertebrate: an epigenetic clock for the crown-of-thorns seastar (Acanthaster cf. solaris).

Accurate and reliable ageing tools are essential for wildlife conservation and management. While DNA methylation has emerged as a promising tool for age estimation in vertebrates, its application to invertebrates remains contested and has been limited to arthropods. Here, we develop an epigenetic clock(definition) for the Pacific crown-of-thorns seastar (CoTS; Acanthaster cf. solaris), a destructive coral predator contributing to habitat degradation across Indo-Pacific reefs. Using Oxford Nanopore Technologies, we generated whole-genome DNA methylation profiles across five age groups and identified 1910 CpG sites with methylation patterns significantly associated with age. We then fitted age prediction models using elastic net regression and evaluated predictive performance with leave-one-out cross-validation (LOOCV), achieving a mean absolute error of 0.31 ± 0.22 years, corresponding to 4-6% of the CoTS lifespan (5-8 years). This accuracy suggests the potential to differentiate annual cohorts, supporting future management-relevant inference. To facilitate practical implementation, we constructed an optimized epigenetic clock from 14 CpG sites consistently selected across LOOCV iterations. Our results demonstrate that DNA methylation-based age estimation is feasible in a deuterostome invertebrate, extending epigenetic ageing approaches beyond arthropods and establishing their potential to advance age determination and management in invertebrates that lack reliable ageing methods.