A first study on the use of Chilean natural zeolite of different particle sizes (0.5, 1 and 2 mm in diameter) in laboratory-scale batch denitrificant reactors was carried out with the aim of assessing the microbial communities adhered to this material. Molecular techniques such as fluorescence in situ hybridization (FISH) and denaturing gradient gel electrophoresis (DGGE) fingerprints revealed a high microbial diversity with a strong presence of Gammaproteobacteria (70% of the total microorganisms) in reactors with zeolite 0.5 mm in diameter. Archaea were only detected in the reactors with zeolite 1 mm in diameter. In addition, the acclimatization and start-up of two UASB reactors 2 (one without zeolite and the other with added zeolite 1 mm in diameter) were conducted following three consecutive and progressive stages using upward velocities from 0.10 to