Feeding live prey organisms in a commercial marine fish hatchery requires significant labor and resources. The effects of feeding frequency of live rotifers and Artemia on growth, survival, hyposalinity stress resistance, and Na+/K+ ATPase activity in black sea bass larvae Centropristis striata were evaluated. Newly hatched larvae (day 0 post-hatching=d0ph) were stocked (53larvae/L) into twenty-four 30-L tanks supported by a recirculating seawater (35g/L) system in a controlled-environment laboratory. Temperature (22°C), aeration (225mL/min), light intensity (~1000lx), and photoperiod (18L: 6 D) were constant. Beginning on d3ph, larvae were fed a prescribed daily ration of live prey (enriched s-type rotifers and/or Artemia nauplii), but distributed at different frequencies: (1) 1×/d at 0800h, (2) 2×/d at 0800 and 1600h, (3) 3×/d at 0800, 1200 and 1600h, and (4) 4×/d at 0800, 1200, 1600 and 2000h, with six replicate tanks per treatment. The daily ration of live prey (maximum of 750,000 rotifers and 90,000 Artemia) was divided equally among feedings; therefore, prey density at each feeding (i.e., meal size) decreased at higher frequencies. By d31ph, larvae fed 4×/d showed significantly (P<0.05) greater notochord length and SGR (mean±s.e.=7.05±0.20mm and 20.1±0.467%/d) compared to larvae fed 1×/d (6.02±0.11mm, and 18.1±0.204%/d) or 2×/d (6.08±0.12mm and 18.4±0.399%/d). Feeding frequency had no effect on larval survival (23.8–38.2%) or whole body fatty acid composition. Ability to withstand an acute hyposalinity (10g/L) challenge on d32ph was significantly poorer in larvae fed 1×/d than in those fed 2×/d, 3×/d or 4×/d. Following transfer to a sub-lethal salinity (15g/L) on d32ph, Na+/K+ ATPase activity (μmol ADP/mg protein/h) was higher in larvae fed 3×/d (5.61) than in those fed 4×/d (5.22) or 1×/d (5.20). The largest larvae (4×/d treatment) showed reduced Na+/K+ ATPase activity (vs 3×/d), likely due to accelerated development and a shifting of the Na+/K+ ATPase from a global distribution to specialized osmoregulatory sites. The results demonstrated that, for a prescribed total daily ration, feeding lower prey densities (smaller meals) at higher frequencies (3×/d or 4×/d) improved larval growth and resistance to hyposalinity stress and may increase osmoregulatory ability, without affecting survival. These findings delineate more judicious use of limited live prey resources to maximize larval growth and vitality during hatchery production of pre-metamorphic stage black sea bass. Statement of relevanceLive prey (rotifers and Artemia) are expensive to produce and can limit production of juvenile fish in a commercial marine fish hatchery. The results show that given a prescribed (albeit moderate) total daily ration of prey, feeding lower prey concentrations at higher frequencies optimized larval growth performance and vitality in black sea bass raised from first-feeding through pre-metamorphic stages.