Species diversity is generally considered one of the key factors of ecosystem resilience in response to anthropogenic pressures, including fishing. In this context, the spatial and temporal changes in demersal fish assemblages and species diversity were investigated in the northern Gulf of St. Lawrence (Canada), over a 20-year period (1990–2010). Data were obtained from the summer research survey conducted by the Department of Fisheries and Oceans, and include commercial and non-commercial species. The study covers the period of groundfish fishery collapse, the moratorium period, and the post-moratorium period, and reflects various modifications in management. Multivariate statistical methods revealed two communities. A coastal community corresponds to strata located above 200 m depth and a deeper community located in the deep channels. Interannual differences in the composition of fish assemblages were observed and are mainly due to the changes in the relative biomass of some dominant species. Three diversity indices (Shannon–Wiener, Simpson's Index of Diversity and Motomura's constant) indicate a slight but significant increase of the diversity over time. This trend is due to the increase of the relative biomass of low-rank species, which may have been favoured by the prohibition of groundfish trawling after 1997 in that region. The geographical distribution of the Shannon–Wiener index also shows temporal dynamics reflecting the biomass distribution of dominant demersal species.