Larval killifish were produced from an in house culture of wild caught specimens.
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 fitted with 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 deionized water and were placed on moist (FW) papers towels in plastic petridishes.
The covered petridishes 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 from FW to SW).
The water was vigorously aerated until eggs hatched (typically with in 10 min 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.