A systematic insight into a single-stage deammonification process operated in granular sludge reactor with high-loaded reject-water: characterization and quantification of microbiological community

Abstract

Reject-water from sludge dewatering was treated in a single-stage deammonification reactor. The aims were to characterize the microbiological community within deammonification granules. In situ mapping of the intact granular sludge were made with fluorescent in situ hybridization (FISH). The Planctomycetes community in the destroyed granular sludge was characterized by FISH, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), cloning and sequence analysis. AOB within Betaproteobacteria were concentrated within the first 50-100 μm of the granule, and Planctomycetes in the first 100-200 μm were responsible for nitrogen elimination. PCR-DGGE verified the higher diversity of Planctomycetes in the deammonification reactor than the anaerobic cultivation. The sequence analysis after PCR-DGGE and cloning identified the dominant Planctomycetes species for anammox reaction as Ca. Brocadia fulgida (accession no. EU478693). FISH detection using the universal probe AMX368 specific for all anammox bacteria including Ca. Brocadia fulgida failed; however, the probe BFU613 specific for Ca. Brocadia fulgida gave clear positive FISH signals. The three-dimensional structure of the ribosome may hinder binding of the universal probe to the corresponding 16S rRNA region. A combination of multiple methods for the analysis of the microbiological community was necessary. Oligonucleotide probes should be carefully selected, and a negative FISH analysis has to be verified by other molecular biological techniques.