2. Materials and methods
2.1. Experimental animals
Larval killifish (F. heteroclitus) were produced from an inhouse culture of wild caught specimens (St. Augustine, FL).
Adult killifish were maintained in flow-through SW at ambient temperature and were fed ad libitum commercially available
dry pellet food. Groups consisting of one male and three to four females were placed in separate tanks. For spawning, tanks were fittedwith egg laying substrates overnight. Egg laying substrates consisted of short (2 cm) lengths of PVC tubing covered with fine plastic mesh in the bottom and a wider plastic mesh on the top. Thewider plasticmesh on the top of the PVCtubing allowed for eggs to fall through and collect on the fine mesh in the bottom while preventing the adults from having access to the eggs. Each morning eggs were removed from the spawning tanks, rinsed briefly in de-ionizedwater andwere placed onmoist freshwater papers towels in plastic petri-dishes. The covered petri-dishes were placed in an environmental chamber at 25 ◦C and fungus infected eggs and unfertilized eggs were removed daily. At day
14 post-fertilization, eggs were placed in 1 l Tripour® plastic beakers containing 0.8 l of water at the planned test salinity (a total of seven salinities ranging fromFWto SW). The water was vigorously aerated until eggs hatched (typicallywithin 10min of immersion) afterwhich aerationwas reduced and newly hatched Artemia nauplii were added. Hatching success of eggs that had developed through 14 days of incubation was >90% regardless of salinity and larvae were fed Artemia nauplii ad libitum daily. All toxicity tests were initiated with 7-day-old fish.