Immuno-modulation of settlement cues in the barnacle, Amphibalanus amphitrite: significance of circulating haemocytes

Gregarious settlement in barnacles has been attributed to chemical cues originating from the associated microorganisms, lectins, and settlement-inducing protein complex from arthropodin, a glycoprotein present in their conspecifics/adults. In this study, we explored the influence of haemolymph haemocytes on the metamorphosis of barnacle, Amphibalanus amphitrite cyprids. The barnacle haemocytes were also characterized for the first time using flow cytometry. Cyprid metamorphosis was significantly higher in the presence of surface-bound haemocytes compared to water-borne cues from haemolymph or adult glycoprotein, a known settlement inducer. The flow cytometry-assisted haemocyte characterization, sorting and their subsequent microscopic evaluation indicated the presence of hyalinocytes and granulocytes. Tagging of these haemocytes with lectins revealed that a significant number of haemocytes had d-glucose/d-mannose and N-acetyl-d-galactosamine, which are important settlement cues. These glycoconjugates were also detected in the cyprid larva pointing out the origin of conspecific cues. The retention of these glycoconjugates in the adult haemocytes and their occurrence in adult leachants indicate pelago-benthic coupling in sessile barnacles wherein these glycoconjugates play a significant role in their gregarious settlement. Characterization of such a coupling process in other sessile benthic invertebrates will pave the way to the understanding of complexities in their population dynamics.