Water changes are predicted to regulate physiological activities of bryophytes characterized by poikilohydric gametophytes. In montane forest ecosystems, nitrogen(N)-fixing bryophyte-cyanobacteria associations are main N resources. The aim of this study was to assess how bryophyte-associated microbiomes and their nitrogenase activity response to instant or long-term water content changes. We investigated the cyanobacterial colonization and nitrogenase activity of four epiphytic bryophyte species in a subtropical montane cloud forest during the dry and rainy seasons in Ailao Mountains, Yunnan, southwestern China. We also evaluated the nitrogenase activity of bryophyte-cyanobacteria associations in response to different water contents in laboratory experiment. The degree of cyanobacterial colonization was evaluated using ultraviolet fluorescence microscopy, and nitrogenase activity of bryophyte-cyanobacteria associations were measured using the acetylene reduction assay (ARA). Cyanobacteria showed an association with all four bryophyte species, with 1.04-3.37% area colonized of the shoot and 10.16-20.21 nmol C2H4.g(-1).h(-1) average nitrogenase activity. Nitrogenase activity was positively related to cyanobacterial colonization (R = 0.742; P = 0.0349). The relationship between water content and nitrogenase activity was unimodal, and both water surplus and shortage inhibited N fixation. Furthermore, long-term drought conditions reduced the degree of cyanobacteria colonization on bryophyte shoots, resulting in decreased nitrogenase activity in the dry season. These results indicate that different response strategies of N fixation operate in bryophyte-cyanobacteria associations to cope with instant and long-term changes in water availability. Our data suggest that both extreme precipitation and drought have a negative impact on N fixation of cyanobacteria-bryophyte associations.