Denitrification is a major biological process contributing to nitrate and nitrite reduction. However, this process remains poorly understood at alkaline pH although such conditions can be encountered in natural (e.g. soda lakes) or industrial environments (e.g. geological waste repositories with cementitious materials). To investigate the nitrate reduction (NR) rate for pH > 9.5 in a cementitious environment, several batch reactors were implemented, with cement leachate or with hardened cement paste (HCP). In the experiments carried out with cement leachate, NR dropped from 0.72 mM/h at pH 9.5 to 0.17 mM/h at pH > 11, while the concentration of nitrite increased. The NR was inhibited at pH close to 12, as was the nitrite reduction at pH above 11. In the reactor containing HCP, the NR rate was 0.75 mM/h at pH close to 10. Calcite precipitated on the HCP surface. Epifluorescence microscopy observations coupled with DNA labelling suggested the presence of microorganisms attached to the HCP surface. This was confirmed by biological growth coupled with NR activity after the transfer of the HCP into a new medium, considered to be sterile. The bacterial community analysis showed that the highly selective culture conditions led to the selection of two species: Halomonas sp. and a species known for its versatile metabolism and ability to form biofilms, i.e. Thauera sp.