In this experiment, larvae were placed into feeding treatments with different types of phytoplankton to determine larval nutritional strategies.

Larvae hatched on Sept 24, 2009 and were distributed into tubs on 500mL filtered sea water (FSW). Three larval feeding treatments with different species of phytoplankton, all at 10 x 104 cells mL-1: Isochrysis galbana (‘‘Iso’’ treatment), Dunaliella tertiolecta (‘‘Dun’’ treatment), a 1:1 ratio of I. galbana and D. tertiolecta (‘‘Mixed’’ treatment), plus an Unfed treatment in which larvae were raised in FSW. Investigators used a hemocytometer to count algal cells and calculate densities of phytoplankton stocks and amount of stock to add to containers for each treatment. At 10- post-hatch, larvae were placed in settlement challenges. These were done in plastic containers with 200 mL FSW and two small fragments (*2–3 cm diam.) of coral rubble. Before use in settlement challenges, the coral rubble fragments were rinsed in freshwater and lightly scrubbed to remove any associated macrofauna. Each piece of rubble had 25–50% cover of live coralline algae, but no apparent other live or boring organisms associated with them. Coral rubble was chosed because a pilot experiment had shown that after 24 h of exposure to coral rubble, fed, swimming larvae were capable of completely losing or reabsorbing the velum, and subsequently actively crawling on the substratum. The settlement challenge on day 10 consisted of pooling three larvae from each replicate container within each food treatment and redistributing them into the settlement challenge containers. Thus, nine larvae were added to each of three replicate containers for the settlement challenge (N = 27 larvae total from each food treatment). Larvae were examined every day for 3 days for loss of velum or complete metamorphosis. Complete metamorphosis was evident by several obvious morphological changes: the previously long and active foot was lost, the margin of the protoconch aperture became light pink, and early growth of the juvenile tube. In most cases at metamorphosis, the protoconch became cemented to the substratum (either the coral rubble or the bottom of the container), but in others the shell remained unattached. Investigators left swimming larvae in the containers and removed dead larvae. The water in the settlement challenge containers was changed each day.

Phillips_2011_Expt1_SettlementChallenge10.csv