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Increasing global concern has been raised about the expansion of hypoxia in coastal waters and its potential to impact benthic ecosystems. Upwelling areas offer opportunities to study the effects of hypoxia on benthic communities under natural conditions. We used a biological trait-based approach and estimated functional diversity indices to assess macrobenthic community functioning along a depth gradient associated with naturally increasing hypoxia and concentrations of organic matter in the upwelling zone of northern Chile (South-East Pacific) over two years. Our results highlighted the increasing dominance of opportunistic biological traits associated with hypoxia and high organic matter content. Habitat filtering was the main process affecting the studied communities. Functional diversity patterns were persistent overtime despite the occurrence of a pulse of oxygenation. This study contributes to our understanding of how natural hypoxia impacts macrobenthic communities, providing useful information in the context of increasing eutrophication due to human influence on coastal areas. • Three types of functional communities were found along a natural gradient of hypoxia. • Habitat filtering was the main process affecting benthic community diversity. • Hypoxia threshold of 2 mL/L dissolved oxygen for taxonomic and functional richness. • Opportunistic traits related to organic matter and hypoxia tolerance were identified. • Functional diversity in hypoxic habitats was not impacted by an oxygenation pulse. [ABSTRACT FROM AUTHOR]