Isolated waterholes and fragmented habitats are common in low- and no-flow rivers of the arid Murray-Darling Basin (MDB). During these periods, remaining water must provide sufficient oxygen and cool refugia for native fish. Water quality is influenced by an intricate combination of in-stream, morphological, riparian, and broader environmental factors, making habitat conservation challenging. This study tested relationships between water temperature and dissolved oxygen (DO) and environmental components under low- and no-flow conditions. We examined factors such as how near-stream vegetation alters climate or how inflows and flood pulses affect water quality. Using coupled three-dimensional ecohydrology models, we studied two sites in the MDB: a turbid waterhole on the Culgoa River and the Darling River at Menindee Lakes. These models incorporated synthesized microclimate, riparian vegetation structure, and biogeochemical processes, validated by continuous temperature and DO measurements. Common drivers of water quality at both sites included air temperature and solar radiation, while vegetation had a weak correlation with solar variability. Wind had a positive impact on water quality in the waterhole but not in the flowing river. Watercourse morphology influenced water quality differently under flowing and non-flowing conditions, with larger watercourses linked to lower water temperatures and higher DO in the isolated waterhole, and the opposite occurring at the flowing site. Understanding these drivers is crucial for managing native fish communities, and the study's methods offer a framework for identifying key environmental controls on water quality. This study will inform the next phase of research, an Australian Research Council Linkage project, Between a Hot Place & Hypoxia.