Spotted seatrout and Southern flounder are highly important fishery species, and both have declined in Alabama in recent decades. Stock assessments identified factors regulating juvenile survival as critical for stock sustainability. Alabama’s coastal waters include valuable nursery habitats of intermixed SAV and emergent marsh, with extensive areas in Mississippi Sound at high salinities, and the Mobile Delta at low salinities. Access to suitable nursery habitat may be regulated by salinity tolerances. We evaluated the salinity tolerance of juveniles of both species through a combination of the analysis of historic state fishery independent monitoring data and a series of laboratory salinity tolerance experiments. The State data suggests strongly contrasting salinity preferences, with the highest catch per unit effort of flounder from 0 ppt and few individuals above 10 ppt, while trout were most commonly captured above 25 ppt with few individuals below 5 ppt. Mesocosm experiments with hatchery-reared juveniles showed that both species could tolerate acute exposure to freshwater, and that juvenile flounder tolerated instantaneous transitions of up to 5 ppt without impacting growth. In 5-9 day chronic experiments, flounder acclimated to near 0 ppt continued to grow strongly with no mortality, while juvenile trout rapidly became lethargic with declines in feeding and growth, and some loss of equilibrium, suggesting that juvenile trout in the wild would not survive exposure to near 0 ppt for more than 2-3 days. Multi-stressor experiments with juvenile trout showed that salinity, temperature, and DO interact with trout acclimated to near 0 ppt, having lower tolerance to high temperatures and low DO. These findings suggest that interannual fluctuations in river flows and future changes in salinity regimes in the region could shift the suitability of extensive areas of nursery habitat in the Delta to favor one species or the other, with subsequent impacts to the stocks.