Kings Bay in northwest Florida, USA, is an important winter home of the largest aggregation of Florida manatee (Trichechus manatus latirostris), and the only location in the United States where visitors legally swim and interact with manatees. In addition to ambient conditions, visitors to the area and management actions have the potential to influence manatee behaviors. We tracked 32 manatees with satellite-linked global position system (GPS) telemetry tags in Kings Bay from 2006 to 2018. Also, personnel at Crystal River National Wildlife Refuge collected manatee counts at Three Sisters Springs from 2014 to 2017. Our objectives were to document the use of springs and other habitat components in Kings Bay relative to ambient water temperature, time of day, tide stage, and management actions within Three Sisters Springs and other manatee sanctuaries in the bay. Manatees that we tagged in Kings Bay spent a median 87% of the winter within the local study area, compared to 47% for animals that we tagged in the Florida Panhandle. Manatees showed preferences for King and Magnolia springs basins at low tide, indicating that they function as tidal refuges, when other locations may be less accessible; we also recorded within spring basin movements. Magnolia Spring Basin showed a significant diel pattern overall, and within basins, King Spring, Tarpon Hole, and Three Sisters Springs were used more during the night than during the day. All areas outside Kings Bay were used more at warmer ambient gulf water temperatures, while all of the springs basins were used more when gulf temperature was colder, especially Three Sisters Springs Basin. Management of Three Sisters Springs had an influence on manatee use during times of cold ambient gulf water temperatures; manatees used the springs more when the spring was closed to visitors, versus when the springs were completely open or when the spring lobes alone were closed. Manatee decisions were consistent with avoiding human interactions.