The expansion of bamboo (Phyllostachys edulis) affects the growth status of trees in colonized forests, but there has been insufficient research on changes in tree water physiology. In this study, we used stable δ2H, δ18O, and 13C isotope ratios to analyze the water sources and water use efficiency (WUE) of bamboo, deciduous broadleaf trees (Alniphyllum fortunei), and evergreen broadleaf trees (Machilus pauhoi and Castanopsis eyrei) in a bamboo-expended broadleaf forest (BEBF), a bamboo-absent broadleaf forest (BABF), and a bamboo forest (BF). We found that the expansion of bamboo had no significant effect on the water sources and WUE of deciduous broadleaf trees, but altered the water sources of evergreen broadleaf trees. During the growing season, evergreen broadleaf trees decrease their uptake fractions of surface soil water by 7.1% to 9.6% and increased their uptake fractions of middle soil water by 5.8%~9.4%. Conversely, during the non-growing season, they increased their uptake fractions of surface soil water by 11.9% and decreased their uptake fractions of deeper soil water by 5.6%~12.9%. Additionally, after expanding into broadleaf forests, bamboo increased its uptake proportion of surface and shallow soil water by 20.0% and 9.4% during the growing season. Its WUE also improved, increasing by 20.0 μmol/mol and 13.0 μmol/mol during the growing and non-growing seasons, respectively. These results indicate that as bamboo expands into broadleaf forests, it enhances its competitiveness for water resources by changing its water use strategy. Compared to deciduous broadleaf trees, evergreen broadleaf trees exhibit more flexible water use strategies under the conditions of bamboo expansion. Our research reveals, for the first time, how broadleaf trees adjust their water use strategies in response to bamboo expansion, and uncovers the mechanisms behind bamboo expansion into evergreen broadleaf forests from the perspective of water use strategies. This will aid future forest management under the conditions of bamboo expansion.