Manipulation of the Symbiodiniaceae microbiome confers multigenerational impacts on symbioses and reproductive ecology of its <i>Exaiptasia diaphana</i> host

Symbiodiniaceae-associated microbiota strongly influence cnidarian symbioses. We systematically reduced the bacterial and fungal communities associated with Symbiodiniaceae to study potential effects on the cnidarian holobiont Exaiptasia diaphana (Aiptasia). Clonal anemones were inoculated with xenic Breviolum minutum (SSB01) and microbiome-manipulated cultures after antibacterial or antifungal treatment. The asexual reproduction of pedal laceration allowed for three generations of clonal aposymbiotic Aiptasia to be utilised in this study, from the initial adult generation (G0), to the first (G1), and second (G2) generation. We inoculated small and large G1 Aiptasia with SSB01 algae and monitored onset of symbiosis, rate of algal proliferation, and holobiont characteristics. Sequencing the 16S and 18S rRNA gene regions identified significant differences in the bacterial and fungal communities of the G0 and G1 generations, alongside differences between the size classes of small and large G1 anemones. The microbiome of larger G1 individuals was distinct to the smaller G1 anemones, suggesting a microbiome maturation process. Control Breviolum minutum cultures exhibited a significantly greater proliferation rate in large G1 anemones when compared to antibacterial or antifungal treated cultures, whereas the opposite trend was documented in the small G1 anemones. Although no differences were observed between algal photochemical parameters, or the growth and polyp activity of G1 juveniles, we observed a significant influence in the production of G2 clones between treatments. Overall, we provide strong ecological implications of manipulating Symbiodiniaceae microbiome, not for the algae themselves, but for the maturation of the host Aiptasia, as well as for the cnidarian holobiont over multiple generations.