Objectives: We assessed the contamination with Legionella pneumophila (Lp) of the hot water network (HWN) of a hospital, mapped the risk of contamination, and evaluated the relatedness of isolates. We further validated phenotypically the biological features that could account for the contamination of the network., Methods: We collected 360 water samples from October 2017 to September 2018 in 36 sampling points of a HWN of a building from a hospital in France. Lp were quantified and identified with culture-based methods and serotyping. Lp concentrations were correlated with water temperature, date and location of isolation. Lp isolates were genotyped by pulsed-field gel electrophoresis and compared to a collection of isolates retrieved in the same HWN two years later, or in other HWN from the same hospital., Results: 207/360 (57.5%) samples were positive with Lp. In the hot water production system, Lp concentration was negatively associated with water temperature. In the distribution system, the risk of recovering Lp decreased when temperature was >55 °C (p < 10 -3 ), the proportion of samples with Lp increased with distance from the production network (p < 10 -3 ), and the risk of finding high loads of Lp increased 7.96 times in summer (p = 0.001). All Lp isolates (n = 135) were of serotype 3, and 134 (99.3%) shared the same pulsotype which is found two years later (Lp G). In vitro competition experiments showed that a 3-day culture of Lp G on agar inhibited the growth of a different pulsotype of Lp (Lp O) contaminating another HWN of the same hospital (p = 0.050). We also found that only Lp G survived to a 24h-incubation in water at 55 °C (p = 0.014)., Conclusion: We report here a persistent contamination with Lp of a hospital HWN. Lp concentrations were correlated with water temperature, season, and distance from the production system. Such persistent contamination could be due to biotic parameters such as intra-Legionella inhibition and tolerance to high temperature, but also to the non-optimal configuration of the HWN that prevented the maintenance of high temperature and optimal water circulation., Competing Interests: Declaration of competing interest None of the authors have anything to disclose., (Copyright © 2023 Elsevier GmbH. All rights reserved.)