454-Pyrosequencing Analysis of Bacterial Communities from Autotrophic Nitrogen Removal Bioreactors Utilizing Universal Primers: Effect of Annealing Temperature González Martínez, Alejandro Rodríguez-Sánchez, Alejandro Rodelas González, María Belén Abbas, Ben A. Martínez-Toledo, María Victoria Loosdrecht, Mark C. M. van Osorio Robles, Francisco González López, Jesús Juan Anaerobic ammonium oxidizing (Anammox) Molecular tools Temperature Identification of anaerobic ammonium oxidizing (anammox) bacteria by molecular tools aimed at the evaluation of bacterial diversity in autotrophic nitrogen removal systems is limited by the difficulty to design universal primers for the Bacteria domain able to amplify the anammox 16S rRNA genes. A metagenomic analysis (pyrosequencing) of total bacterial diversity including anammox population in five autotrophic nitrogen removal technologies, two bench-scale models (MBR and Low Temperature CANON) and three full-scale bioreactors (anammox, CANON, and DEMON), was successfully carried out by optimization of primer selection and PCR conditions (annealing temperature). The universal primer 530F was identified as the best candidate for total bacteria and anammox bacteria diversity coverage. Salt-adjusted optimum annealing temperature of primer 530F was calculated (47°C) and hence a range of annealing temperatures of 44–49°C was tested. Pyrosequencing data showed that annealing temperature of 45°C yielded the best results in terms of species richness and diversity for all bioreactors analyzed. 2015-10-02T07:41:55Z 2015-10-02T07:41:55Z 2015 info:eu-repo/semantics/article González-Martínez, A.; et al. 454-Pyrosequencing Analysis of Bacterial Communities from Autotrophic Nitrogen Removal Bioreactors Utilizing Universal Primers: Effect of Annealing Temperature. BioMed Research International, 2015: 892013 (2015). [http://hdl.handle.net/10481/38031] 2314-6133 2314-6141 http://hdl.handle.net/10481/38031 10.1155/2015/892013 eng info:eu-repo/semantics/openAccess Hindawi Publishing Corporation