Multi-use marine protected areas (MUMPAs) are a commonly applied tool for marine conservation in developing countries, particularly where large no-take reserves are not socially or politically feasible. Although MUMPAs have produced benefits around the world, the persistence of moderate fishing pressure reduces the likelihood of achieving the primary objective of these areas, which is the conservation of ecosystems. In this study we used traditional and functional metrics to evaluate how fish assemblages changed through time in a MUMPA, including shifts in species responses and in ecological processes. We conducted visual censuses of fishes at Espíritu Santo Island, México (MUMPA; N = 320; 24°N, 110°W) from 2005 to 2017 to assess fish richness, size-distribution and density. Three functional indices were calculated using six traits (size, mobility, period of activity, aggregation, position in water column and diet): functional richness (volume occupied by species), dispersion (complementarity between species) and originality (inverse of functional redundancy). We compared fish diversity among three management zone types (sustainable fishing, traditional fishing and no-take zones), through a 13-year period, assessing which species increased or decreased in occurrence, density, and biomass, and how indices respond over time. Despite a general increase in biomass and stability in density and originality, we detected a reduction in fish biodiversity in the form of declines in species and functional richness, which could imply the risk of local extinction and decrease in certain ecosystem processes. In addition, changes in functional dispersion showed that some functions are losing representation through time. Although no single cause is apparent, such factors as competitive interactions, habitat loss and persistence of fishing pressure potentially explain these decreases. The rise in biomass was associated with a general increase in the average size, rather than increased biomass of commercial species, as the latter remained stable during the study period. Expansion of no-take areas, enforcement of fishing regulations, and surveillance in core zones, should be implemented to reverse the decline in particular species and to promote conservation of fish functional diversity in this MUMPA.