Sampling details.

Ocean acidification (OA), driven by rising atmospheric CO2, presents a serious threat to marine biodiversity, especially within coral reef ecosystems. Natural analogue sites, such as the high-pCO2 seep at Iōtorishima Island in Japan, offer insights into future conditions. This study investigated the holobiont communities of Symbiodiniaceae and bacteria in the zoantharian Palythoa tuberculosa at Iōtorishima and compared them to specimens from control sites in Okinawa and Hawaiʻi. Using amplicon sequencing of the dinoflagellate internal transcribed spacer 2 (ITS2) region of ribosomal DNA and microbial 16S rRNA gene, we detected significant shifts in both Symbiodiniaceae and bacterial communities under high-pCO2 conditions at Iōtorishima. Specifically, P. tuberculosa at the seep site had reduced Symbiodiniaceae diversity, predominatly featuring Cladocopium C1 and C3 types. Additionally, its bacterial communities showed lower richness with distinct taxonomic profiles, including increased levels of Mollicutes and Vibrio spp. These results highlight the potentially adverse effects of OA on hexacoral holobionts and emphasize the need for detailed, high-resolution studies across various holobiont species and geographic locations. The shifts observed specifically in Symbiodiniaceae and bacterial communities at the Iōtorishima seep suggest that holobionts may exhibit plasticity in response to environmental stress, which has implications for resilience and adaptation of zoantharians and other reef organisms amid climate change. This research provides crucial baseline data for predicting future coral reef compositions in an OA-affected world.