The present study aimed to delve into the local ecological knowledge of fisheries in the Meghna River Basin (MRB) of Bangladesh by exploring the insights and perspectives of local communities. A survey was administered among six fishing communities from five districts along the MRB between August 2015 and January 2016 to accumulate data for this study. The study sites were selected meticulously based on three crucial criteria: upstream river, coastal area, and fish sanctuaries, which covered three major rivers, namely the Meghna, Andharmanik, and Payra. The study employed participatory rural appraisal (PRA) tools, including 120 individual interviews using purposive sampling, 25 focus group discussions, and 36 key informant interviews. The study identified the ten most frequently caught fish species along with their temporal and spatial variation as reported by the respondents. Nine of these species fell into the least concern category, which indicate their stable population status. Meanwhile, six out of ten species cited as highly caught in the previous one to two decades belong to the threatened or near-threatened category. Findings also reveal that fishers are able to recognize important microhabitats of the study area and their significance for fish species. In addition, fishers identified the negative drivers of ecosystem degradation as well as suggested several management measures to address these challenges. The results of this study underscore the critical role of engaging with local communities and integrating their ecological knowledge into initiatives for the sustainable exploitation and conservation of aquatic resources in the MRB.