Native fish populations in Australia’s Murray-Darling Basin (MDB) are in a highly degraded state, with an estimated decline in native fish abundance of >90% since European settlement. Much of this decline has been attributed to historical and contemporary human disturbance, including changes to land use, impacts of river flow regulation and consumptive use of water, over-fishing, invasive species, drought, large-scale bushfires and blackwater-related hypoxia. To date, most estimates of native fish population decline have relied heavily upon expert knowledge due to a lack of long-term data on fish population trends. While incorporation of expert knowledge is a critical input to fish management and conservation, there is also a need for quantitative estimates of fish population status to facilitate formal analysis of temporal trends. The aim of this study was to analyse an extensive fish monitoring database to examine multi-decadal trends (1997 – 2022) in the relative abundance of key large-bodied fish in the NSW portion of the MDB. Using mixed models, in a two-step modelling process, we analysed both overall and region-specific relative abundance trends across the NSW MDB for six key large-bodied species. We found evidence of regional specific trends including, for example, evidence of increasing relative abundance of Murray cod in the Namoi and Murrumbidgee regions and possible declines in other catchments. These region-specific trends in species abundance highlight the importance of using long-term to contextualise the current condition of fish populations and emphasise the need to manage species at appropriate spatial and temporal scales.