Baseline studies of South Atlantic oceanic islands and seamounts are scarce compared with those from other regions. Indeed, mesophotic depths are rarely assessed and their study contributes to the global knowledge gaps about reef ecosystems. Here, we present the first detailed quantitative assessment of benthic communities across an abrupt depth gradient (euphotic to mesophotic) in the smallest and most isolated tropical rocky reefs in the world, the St. Peter and St. Paul Archipelago (SPSPA). The influence of biotic (abundance of fish trophic groups) and abiotic (benthic complexity, depth, and light) variables on benthic community structure was evaluated with scuba and a remotely operated vehicle. We recorded 77 benthic taxa in six groups (macroalgae, Porifera, Cnidaria, Annelida, Tunicata, and Bryozoa). Macroalgae was the richest group (41 infrageneric taxa), with 17 new records for the SPSPA. Turf algae was the most abundant group in both the euphotic and mesophotic samples (26 species), but only eight turf-forming species occurred in the latter. Two distinct benthic assemblages were detected: (1) a euphotic assemblage (0–30 m) composed primarily of the zoanthid Palythoa caribaeorum (Duchassaing and Michelotti, 1861), the fleshy alga Caulerpa racemosa (Forsskål) J. Agardh, crustose coralline algae (CCA), and Bryopsis spp.; and (2) a mesophotic assemblage (30–60 m) dominated by CCA, bryozoans, and scleractinian corals. Depth and light were the most important predictors of benthic community structure.