The composition of the demersal nekton assemblage inhabiting the Mediterranean shelf-break of west-central Italy was examined to investigate the diel variation in species composition and abundance. Fish were obtained during four 24-h sampling periods that represented the four annual seasons. Thirty-two hauls (eight in each season) were conducted in a shelf-break area at 140-160 m depth. The density of fish, crustaceans, and decapods exhibited differences on short- and long-term time scales: Multidimensional scaling suggested that season and light intensity may be important factors influencing fish assemblage composition. At the diel scale two main assemblages separating nocturnal and diurnal hauls were identified. The typical assemblage structure of shelf-break bottoms was represented during daylight by species such as Merluccius merluccius, Capros aper, Serranus hepatus, Macroramphosus scolopax and Trisopterus minutus capelanus. At night, the assemblage showed a drastic change due to the increase of small nektobenthic species, Argentina sphyraena and Glossanodon leioglossus, together with cave-dwelling fish (Gnathophis mystax, Chlopsis bicolor, Nettastoma melanurum) and nektobenthic crustaceans (i.e., Parapenaeus longirostris and Solenocera membranacea). The above-mentioned differences in species composition may be related to changes in vertical distribution for trophic needs (M. Merluccius) and/or in the shoal structure (A. sphyraena and G. leioglossus). The lack of data on the pattern of trawl catchability of Mediterranean species during the day may lead to an inaccurate estimation of species abundance when only diurnal trawl survey data are used.