Dataset encompasses identification and quantitative information on megabenthic fauna and substrate characterization resulted from the processing of ROV video transects acquired during the research expedition MEDWAVES (MEDiterranean out flow WAter and Vulnerable EcosystemS), as part of the H2020 European project A Trans-Atlantic assessment and Deep-Water Ecosystem-Based Spatial Management Plan for Europe (ATLAS, https://www.eu-atlas.org/). This data were collected at four geomorphological features that represent different types of seafloor elevations, covering a gradient of influence of the Mediterranean Outflow Water (MOW) from the western Mediterranean to the central North Atlantic: The Seco de los Olivos bank (SdO) in the Alboran Sea (Western Mediterranean), the Gazul mud volcano (GMV) in the Gulf of Cádiz, the Ormonde seamount (OS) in the Portuguese margin and the Formigas seamount (FS), SE Azores. To obtain quantitative data, a series of still images were captured from the video footage of ROV video transects at 5 m intervals. Each record at the datasets (fauna and substrate) represent a georreferenced still image. Images with poor quality (i.e., sediment resuspension, lack of focus, ROV too far from the seafloor, close-ups for species identifications and not visible lasers, sampling events) were disregarded for the analyses. At each still image, the abundance (total count) of species or operational taxonomic units (OTUs) (fauna dataset) and the percentage (from 0 to 20) of each substrate type (substrate dataset) were recorded. Technical details related to methodology of data collection is available in the Cruise Summary Report - MEDWAVES survey (MEDiterranean out flow WAter and Vulnerable EcosystemS): https://zenodo.org/record/556516#.YnP8jy0rxUODetails of the methodology used for fauna and substrate identification and process ROV video transects to obtain quantitative data were published in Puerta et al 2022 Deep-Sea Research I (DOI: 10.1016/j.dsr.2022.103791)