Understanding biodiversity patterns and how they are driven at different spatial scales is a crucial issue in ecological studies. This is particularly evident for the deep sea, the largest biome of the biosphere, where information on the scales of spatial variation is very scant. Here, we investigated deep-sea nematodes species richness, turnover and functional diversity, and life strategies at different spatial scales (from local to macro-regional) to identify the factors that shape regional (<i>γ</i>) and macro-regional (<i>ϵ</i>) deep-sea diversity. This study was conducted in several deep-sea habitats (canyons, open slopes, deep-water corals, and bathyal plains) over > 2000 km across the whole Mediterranean basin, at a fixed depth of ca. 1000 m. Our results indicate that the patterns of local (<i>α</i>) diversity across the deep Mediterranean follow the gradients of the trophic conditions, which decrease from the Western to the Eastern basins. For all of the sites and habitats, the <i>α</i>-diversity is generally low. Conversely, the turnover diversity changes significantly among habitats (<i>β</i>-diversity) and between regions (<i>δ</i>-diversity), showing values of dissimilarity (based on species presence/absence) between 59% and 90% for <i>β</i>-diversity and between 81% and 89% for <i>δ</i>-diversity. This suggests that patterns and values of regional (<i>γ</i>) and macro-regional (<i>ϵ</i>) diversity in the deep Mediterranean Sea are related to turnover diversity among habitats and between regions (<i>β</i>- and <i>δ</i>-diversity), rather than to the local biodiversity (<i>α</i>-diversity). These results indicate that the differences in <i>β</i>-diversity and <i>δ</i>-diversity are even more important than those for the <i>α</i>-diversity for the understanding of the drivers of biodiversity in the deep Mediterranean Sea. These data also allow us to conclude that habitat heterogeneity (and type) and gradients in environmental conditions, by promoting a high turnover diversity across the deep Mediterranean Sea, are crucial players for the nematode diversity levels