This research aimed to assess the fish biodiversity and present status of the Dholai river in Companigonj Upazila under Sylhet district. A semi-structured questionnaire survey was used to gather the relevant information from 64 fishers from November 2019 to April 2020. This research identified 56 fish species under 21 families and ten orders. The most dominant order was Cypriniformes 32.14%, following Siluriformes 28.57%, Perciformes 14.28%, Channiformes 7.14%, Synbranchiformes 5.36%. The species availability status was observed in four categories and achieved as 17 species of commonly available (30.36%), 14 species of moderately available (25%), 17 species of less available (30.36%), and eight species of fishes were rarely found available (14.28%). A total number of five nets, one trap, one hook, and a line were recorded. The peak amount of mean Catch per Unit effort (CPUE) in the current jal was 3.67±1.50 kg/day, and the lowest was in borshi at 0.09±0.01kg/day. The highest diversity index value was 2.55 in January, and the lowest was 1.21 in April, with a mean value of 1.98±0.05. The peak richness index value was 2.76 in January, and the lowest was 1.79 in April, with a mean value of 2.36±0.1. The peak evenness value was 0.73 in January, and the lowest value was 0.41 in April, with the mean evenness value 0.61±0.05. The peck amount of fish production was in January as 478±70 kg/day, and less value was in March at 109±12 kg/day. The local fishermen identified several threats to fish biodiversity, fish habitat, and fish production of the Dholai river. The establishment of the fish sanctuary, control of river pollution, minimizing exploitation of sand and rock, conservation of angling gears ever, and maintaining fisheries rules and regulations will be effective for the conservation of fish biodiversity of the Dholai River.