Environment conservation of the Three Gorges Reservoir (TGR) requires reliable assessment of fish resources and variation of their distribution. The goal of this study was to determine the best season and the diurnal period to estimate fish density in the TGR using acoustic surveys as a monitoring tool. To find out diurnal migration pattern of fishes in the TGR, we conducted reduplicated 24-h hydroacoustical surveys in the forequarter of the TGR, which covered different habitats (mainstream, tributary bay), seasons (summer, winter) and years (2012-2015). Measurements were performed using SIMRAD EY60 split beam echosounder with frequency 120 kHz. Additionally, zooplankton and environmental parameters were measured. The results showed more than ten times differences between the day and night fish density estimates, probably resulting either from the horizontal migrations between the littoral and open water or vertical migrations. Differences were higher during winter than during summer, and higher in the tributary bay than in the mainstream. Clear vertical migrations were observed both in the tributary bay (amplitude 15 m) and in the mainstream (amplitude 35 m), but only during winter time. Inter-annual estimates of fish density showed generally decreasing trend with years. especially well pronounced in the tributary bay. We also observed high seasonal and diurnal variability in fish distributions, which can significantly affect the accuracy and precision of biomass estimate, indicating importance of choosing appropriate timing for fish monitoring. According to our results, in order to receive the most reliable estimates of fish density in the TGR, hydroacoustic surveys should be performed during summer at night time. (C) 2017 Elsevier GmbH. All rights reserved.