Enrichment, phylogenetic analysis and detection of a bacterium that performs enhanced biological phosphate removal in activated sludge

Activated sludge communities which performed enhanced biological phosphate removal (EBPR) were phylogenetically analyzed by 16S rRNA-targeted molecula r methods. Two anaerobic-aerobic sequencing batch reactors were operated wi th two different carbon sources (acetate vs. a complex mixture) for three y ears and showed anaerobic-aerobic cycles of polyhydroxybutyrate- (PHB) and phosphate-accumulation characteristic for EBPR-svstems. In situ hybridizati on showed that the reactor fed with the acetate medium was dominated by bac teria phylogenetically related to the Rhodocyclus-group within the beta-Pro teobacteria (81% of DAPI-stained cells). The reactor with the complex mediu m was also predominated by this phylogenetic group albeit at a lesser exten t (23% of DAPI-stained cells), More detailed taxonomic information on the d ominant bacteria in the acetate-reactor was obtained by constructing clone libraries of 16S rDNA fragments. Two different types of Rhodocyclus-like cl ones (R1 and R6) were retrieved. Type-specific in situ hybridization and di rect rRNA-sequencing revealed that R6 was the type of the dominant bacteria . Staining of intracellular polyphosphate- and PHB-granules confirmed that the Rb-type bacterium accumulates PHB and polyphosphate just as predicted b y the metabolic models for EBPR. High similarities to 16S rDNA fragments fr om other EBPR-sludges suggest that R6-type organisms were present and may p lay an important role in EBPR in general. Although the R6-type bacterium is closely related to the genus Rhodocyclus, it did not grow phototrophically . Therefore, we propose a provisional new genus and species Candidatus Accu mulibacter phosphatis.