BACKGROUND: Nitrogen removal by organic biofilters treating pig manure is incomplete and has to be optimized. This could be achieved by promoting the activity of denitrifiers, this being the limiting factor for nitrogen removal in this kind of process. However, changing the operating conditions could impair nitrification; no data are currently available about the activity and distribution of nitrifiers in organic biofilters. A better knowledge of the process of colonization by nitrifiers is needed to determine their potential to oxidize ammonia and whether they share the same physical space as denitrifiers or not. RESULTS: Data showed that nitrifiers were principally localized near the surface and that a microbiological gradient, in terms of nitrifying activity, was present from top to bottom. Moreover, in each section studied, the potential nitrifying activity measured was always higher than the observed nitrifying activity. These results suggest that the biomass had the capacity to oxidize more ammonia and that these transformations occurred near the surface of the biofilter. However, nitrifying activities were not limited by environmental factors but rather by the concentration of ammonia supplied to the process. CONCLUSIONS: Because denitrifiers and nitrifiers share the same physical space in this kind of process, particular precautions will have to be taken to maintain nitrification, while modifying conditions to promote denitrifier activity. (c) 2007 Society of Chemical Industry.