Antifouling Potential of Compounds

Antifouling activity was assessed via inhibition of settlement and metamorphosis of larvae of the Pacific transparent sea squirt (Ciona savignyi) and the blue mussel (Mytilus galloprovincialis). Methods followed those published by Grant et al. [81 (link)]. Adults of both species were collected from coastal populations in the Nelson region of New Zealand, held in a recirculating seawater system (18 ± 1 °C, 33 ± 1 PSU) and fed bulk-cultured Isochorysis galbana until ready to spawn. Larval spawning and rearing procedures followed previously described methods for C. savignyi [82 (link)] and M. gallorprovincialis [83 ]. Competent larvae were diluted in artificial seawater to yield 3 ± 1 larvae/mL. Aliquots of these larval suspensions were added to 12-well tissue culture plates (Corning Co-Star) containing serial dilutions of 1 ranging from 0.1–100 µg/mL. Controls were included, and three replicates were performed in all cases. After 5 days of incubation at 18 ± 1 °C, the number of successfully settled and metamorphosed individuals were counted in each well. Sigmoidal dose-response relationships were explored using R Statistical Software [84 ] to determine whether inhibition had occurred relative to the controls.