Fishes of Pseudogobius, distributed widely in the Indian and West Pacific Oceans, are represented by 3 species (P. javanicus, P. masago, Pseudogobius sp.) in Japanese waters. Although P. javanicus is a common species at Okinawa-jima Island, P. masago is scarce and listed as “Vulnerable” in the Red List of the Japan Ministry of Environment. In the present study, micro–habitat use (water temperature, salinity, water depth, flow velocity, median grain size and substratum hardness, percentage of mud, and percentages of litter cover and stone cover) by P. javanicus and P. masago were examined in 72 quadrats on Sashiki tidal flat, Okinawa–jima Island, from May to September 2013. In total, 36 P. javanicus and 97 P. masago were collected, the species occurring in 8 and 16 quadrats, respectively. A significant difference in quadrat occupation was apparent between the species. The relationships between number of individuals of each species and environmental factors were analyzed using generalized linear models. Water depth, percentage of litter cover and median grain size (for P. javanicus), and substratum hardness, salinity, and percentages of stone cover and mud (for P. masago) were significant factors in the ten top models. In particular, the presence of both species was affected by substratum condition (percentages of litter and stone), which may be related to predation risk. The results demonstrated that microhabitat uses of the two species were different. However, the Sashiki tidal flat includes suitable environments for both P. javanicus and P. masago at present, the accumulated soft substratum being preferred by both species. It is important, however, for continued conservation of both species, that landfills and riverbank revetment works around Nakagusuku Bay be kept to a minimum, in order to provide and prevent an outflow of soft substratum to the tidal flat.