The white-rot fungus Hypholoma fasciculare coexists with a bacterial community that uses low-molecular weight carbon sources provided by fungal, extracellular enzyme activities. Since fungal development on wood is limited by the availability of nitrogen (N), bacteria could contribute to the N supply. To prove or disapprove an interaction in terms of N transfer, N sources of the fungus and the coexisting bacterial isolates were investigated, and the bacterial N₂ fixation was quantified. Fungal, fungal—bacterial and bacterial wood decomposition was analysed by Fourier transform infrared spectroscopy (FTIR), mass loss and surface pH. Microbial N preferences were investigated by elemental analysis isotope ratio mass spectrometry (IRMS). In addition, diazotrophic activity was explored after cultivation under a ¹⁵ N₂/O₂ atmosphere. Decomposition was similar with and without bacteria and both H. fasciculare and coexisting bacteria preferred reduced N species, such as urea, ammonium and organic N. In most of the bacteria, the ¹⁵ N abundance in the biomass increased significantly but to a low extent if they were cultivated under a ¹⁵ N₂/O₂ atmosphere. This effect is considered an artefact and attributed to adsorption rather than to bacterial N₂ fixation activity. Hence, the bacteria coexisting with H. fasciculare rather competed for the same N sources than supported fungal N supply by diazotrophic activity.