Gorai River is one of the important rivers in Bangladesh for rich aquatic biodiversity. The river is originated from the Ganges-Padma River system, a trans-boundary river between India and Bangladesh. Once the river was rich in fish biodiversity, but due to man-made and natural causes the availability of fish reduced drastically. A comprehensive analysis of fish diversity indices, gear efficiency, catch composition and decline causes of fish diversity in Gorai River, Bangladesh was accomplished. The data were collected on monthly basis from January to December 2018 from three major fishing sites of the river. A total of 62 fish and 2 prawn species under 12 orders and 24 families were recorded. Cypriniformes was the leading order consisting 27% of the total catch. The mean values of Shannon-Weaver diversity (H\'), Simpson\'s index (1-D), Margalef\'s richness (d) and Pielou\'s evenness (J\') indices were recorded as, 1.478 ± 0.495, 0.57 ± 0.197, 15.115 ± 4.435 and 0.481 ± 0.152, respectively. At the similarity of 58.7%, two groups were attained in the cluster analysis and the Non-metric Multidimensional Scaling (nMDS) showed 40% similarity among the three sites in twelve months based on the Bray-Curtis similarity matrix. The highest and lowest CPUE were recorded from seine net (5.2 ± 1.72 kg gear-1 haul-1) and hook & long lines (0.0135 ± 0.0015 kg gear-1haul-1), respectively whereas, highest and lowest gear efficiency were recorded from lift net (0.321 ± 0.036 kg gear-1person-1hour-1) and fish trap (0.0005 ± 0.0002 kg gear-1person-1hour-1), respectively. Alternatively, the highest fish catch was recorded on April (21228 ± 464.38 kg) and lowest on August (3855 ± 138.21 kg). According to the fishermen fish biodiversity of the Gorai River declined day by day due to overexploitation, destructive fishing practice, pollution, construction of obstacles for fish movement, and natural causes like siltation. Proper implementation of fish acts and regulations, use of authorized fishing gear, community-based fisheries management, sanctuary establishment and management, stocking of fish fingerling, and raising public awareness can play a great role in enhancing and conserving fish biodiversity in the Gorai River of Bangladesh.