Elucidating Mechanisms by which Invertebrate Larval Settlement is Affected by Biofilm Ciliates

Despite extensive studies of benthic invertebrate larvae responding to settlement cues from bacteria and microalgae in biofilms, the roles of protozoa have been largely ignored. We recently showed that an assemblage of biofilm ciliates affected larval settlement and survival rates among two polychaetes, a mussel, and a bryozoan, being inhibitory to some and facilitative to others. Here we investigated settlement inhibition further for the serpulid worm, Galeolaria caespitosa, and the mussel, Mytilus galloprovincialis. Single species of ciliates were capable of inhibiting settlement by up to 68%. The effects were density dependent, with the strength of inhibition being directly related to ciliate abundance. The strength of inhibition also differed significantly among ciliate species, suggesting that both the abundance and makeup of ciliate assemblages could be an important variable determining settlement rates in the field. We studied the mechanisms of inhibition further with G. caespitosa and the ciliate, Euplotes minuta. Filtrate from ciliate cultures failed to inhibit settlement, indicating that dissolved chemicals were not the inhibiting factor. Physical presence of ciliates was inhibitory, as demonstrated by video analysis of larval search behavior. Following contact with a ciliate, larval swimming was disrupted, including retreat from the substratum and significant changes in swimming angles. Ciliates may also have influenced settlement indirectly by altering cues from biofilm bacteria. Although bacterial densities were unaffected by ciliate grazing during the assays, bacterial distributions were significantly more clumped in the presence of ciliates, which could perhaps affect the suitability of the biofilm for larvae. These organism-scale interactions at the biofilm boundary could produce significant constraints on larval recruitment patterns and suggest that further studies are needed on the roles of protozoans in boundary layer processes.