Microbial community of nitrogen-converting bacteria in anammox granular sludge

•The coexistence of AerAOB, HDB and AnAOB was confirmed by direct molecular evidence.•The anammox provided a high nitrogen removal as the main pathway for the ammonia degradation.•Dissimilatory nitrate reduction was mostly caused by HDB rather than by AnAOB.•2 OTUs of AnAOB belonging to the genus Candidatus Brocadia were found.•The activity of the AnAOB was 3.7–5.6 mg N g−1 VSS h−1.

The nitrogen removal pathways in Anaerobic ammonium oxidation (anammox) process where anaerobic ammonium-oxidizing (anammox) bacteria convert ammonium and nitrite directly to dinitrogen gas under anoxic conditions were investigated in this study. For this propose, microbial community of anammox granules from the lab-scale and full scale reactors were analyzed according to morphology, diversity and activity. The microbial diversity of anaerobic ammonia oxidizing bacteria (anAOB), heterotrophic denitrifying bacteria (HDB) and aerobic ammonium-oxidizing bacteria (AerAOB) was examined by clone libraries. The abundance of AnAOB and HDB was quantified by real time PCR (qPCR). The activities of partial nitrification, denitrification and anammox were estimated with batch tests and respirometry. The fluorescence in situ hybridization (FISH) showed an outer layer dominated by AnAOB whereas the core had a very low cell density with no AnAOB. The denaturing gradient gel electrophoresis (DGGE) results show the existence of genes related to the nitrogen-compounds-converting bacteria from AnAOB (16S rRNA), HDB (nirS) and AerAOB (amoA). In the context of phylogenetical analysis 35 AnAOB clones were analyzed belonging to the genus Candidatus Brocadia (2 Operational taxonomic units). On the other hand, amongst the 26 sequenced HDB clones had a high richness of different nirS genes (13 OTUs). In 38 clones of the amoA gene 7 OTUs were detected. The specific activity of the AnAOB was 3.7–5.6 mg N g−1 VSS h−1. Anammox was the main pathway for the ammonia degradation. The average HDB activity ranged around 0.9 mg N g−1 VSS h−1. With nitrate as a sole substrate for the HDB the average activity for the HDB without external carbon source was around 1.5 mg NO3–N g−1 VSS h−1. The activity of the AerAOB measured by respirometry test reached only 0.05 mg N g−1 h−1.