Enhanced biological phosphorus removal (EBPR) activated sludge was operated in a laboratory-scale sequencing batch reactor (SBR) fed with acetate as the sole carbon source. The microbial community of the sludge was analyzed using the polymerase chain reaction (PCR) – denaturing gradient gel electrophoresis (DGGE) method for about 2 months of start-up period. As a result, the number of major bands decreased during the enrichment, indicating that the microbial community structure was getting simpler. Since the phosphate removal activity was maintained at a high level, the bacteria which still remained at the end can be considered as the important bacteria playing key roles in the present EBPR sludge, maybe polyphosphate accumulating organisms (PAOs). The dominant band in the last sample on the DGGE gel was excised and the DNA recovered from it was sequenced. The sequence was closely related to one of the putative PAOs group which Crocetti et al. (2000) and Hesselmann et al. (1999) have proposed. This PAOs group is closely related to the Rhodocyclus group (b-Proteobacteria). The fluorescence in situ hybridization (FISH) method with the probe specific for this PAOs group and the DAPI staining at a phosphate-probing concentration indirectly showed that these Rhodocyclus related bacteria really accumulated polyphosphate.

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