Oysters (Crassostrea virginica) were cultured as spat-on-shell at the Auburn University Shellfish Laboratory (AUSL) on Dauphin Island, AL starting in late May 2019 using standard techniques (Congrove et al. 2009). Oyster larvae were settled onto sun-bleached oyster shells to create spat-on-shell. After 3 days, when oyster spat were approximately 1.0 millimeters, they were exposed to either exudate from predatory blue crabs or empty cage controls in four flow-through holding tanks (length = 2.4 meters, width = 0.9 meters, water depth = 0.4 meters) supplied with unfiltered seawater pumped directly from the Gulf of Mexico. The number of spat per shell varied from approximately 5 – 40 and we elected to not alter the initial density to mimic natural settlement during the induction period. Oysters were suspended above the tank bottom in oyster aquaculture baskets (64 x 23 x 14 centimeters with 140 spat-covered per shell basket) to prevent sediment buildup from suffocating oysters. Seven oyster baskets were present in each tank (28 total).
Spat were exposed to blue crab predator cues by holding four live caged adult blue crabs (Callinectes sapidus) in two of the tanks (8 crabs total), whereas the remaining two tanks contained empty cages (control) to mimic conditions where oysters regularly experience predator cues or are limited in their exposure from cues. Water volumes and crab densities were informed from established procedures (Belgrad et al. 2021). Crabs in each tank were held in four separate cages (32 x 23 x 14 centimeters) to prevent crabs from consuming the experimental oysters or each other. Every crab was fed one adult oyster daily (approximately 5.0 centimeters in length) to maximize predator cue intensity as experimental oysters would be exposed to exudates from predators and damaged conspecifics. This ensured that oysters were exposed to the most natural set of cues indicative of a predation event, which produces a strong response in oysters (Scherer et al. 2016). Crabs were replaced during the experiment as needed due to mortality. Experimental oyster baskets were rotated around the crab cages daily to reduce differences in oyster growth due to proximity to predator cues, and no differences among cages were found. The induction period was 2 months.
Oyster survival experiment
We conducted a field experiment on a healthy intertidal oyster reef to test whether our laboratory findings were consistent with effects in the field. Oyster spat-on-tile were taken from the above culturing work. Oysters were scraped to 10 spat per tile to standardize individual predation risk, and transported in aerated coolers filled with seawater to a large (more than 1 kilometer long x 10 meters wide x 1 meter high) healthy oyster reef at Skidaway Island, GA (31°57'52.2" N; 81°00'49.4" W) on August 11th, 2020. This reef is a contiguous set of approximately 10-100 meter live oyster patches separated by regions of mud and shell hash containing smaller oyster clusters. The site is located along the Wilmington River in upper Wassaw Sound, adjacent to the Skidaway Institute of Oceanography and bordered by a robust Spartina alterniflora zone. Mud crab densities within the oyster reef are roughly 30 square meters and resident mud crabs experience heavy predation by blue crabs, particularly in areas of bare substrate and shell hash (Hill and Weissburg 2013).
Sixty pairs of induced and control spat-covered tiles were zip tied to 60 pieces of 1.5 meter-long rebar. Rebar was set in 15 transects that ran perpendicular to the shoreline with over a 5-meter separation between each transect. Each transect contained 4 rebar poles planted in the following locations: the upper tidal zone of the reef, the lower tidal zone of the reef, the reef edge, and in the mud with at least a 1-meter distance from the reef. Poles typically were separated from each other by 1.5 meters (total transect length = 7 – 8 meters) and were planted so that the spat tiles rested just above the substrate. Individual survival of all oysters was checked 48 hours after planting (n = 1,200 spat total; 150 spat per treatment).