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In order to obtain highly enriched cultures of phosphate-accumulating organisms (PAOs), activated sludge from a local sewage treatment plant was cultivated in lab-scale sequencing batch reactors (SBR) with alternating acetate and propionate as carbon sources. After operating for over a month, the anaerobic phosphorus release rate, as well as aerobic phosphorus uptake rate, of the system were increased from 0.60-0.75 mg/(g·h) to over 8.5 mg/(g·h). Meanwhile, the phosphorus removal efficiency of the system was increased from 10% to over 90%. As assessed by fluorescence in situ hybridization (FISH) quantification, the proportion of PAOs in all bacteria was 75±5%, which indicated that the highly enrichment of PAOs was achieved.Then, the activated sludge highly enriched of PAOs were seeded, and cultivated with mixed carbon sources of different acetate to propionate ratio, to obtain granule-based enhanced biological phosphorus removal (EBPR) system. The long-term effects of acetate to propionate ratio on the granulation progress of EBPR system were investigated. Furthermore, the microbial community of the granule-based EBPR systems cultivated with mixed carbon sources of different acetate to propionate ratio were investigated by molecular biotechnology such as polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), 16S rDNA clone, and etc.After operating for 90 d, the results showed that the effects of acetate to propionate ratio on the granule growth rate, granule sludge characteristics and morphology, and phosphorus removal efficiency were all of regularity. The granule growth rate was obviously decreased as the acetate to propionate ratio increased, which leading to a gradient of granule volume average particle diameter of different granule-based EBPR systems. The volume average particle diameters of the mature granules were 745.08μm,680.99μm,642.38μm,599.41μm, and 550.64μm in the treatment of 0%,25%,50%,75% and 100% acetate in mixed carbon sources, respectively. And the sludge volume index (SVI) of the mature granules were 75 mL/g,60 mL/g,50 mL/g,40 mL/g and 30 mL/g in the treatment of 0%,25%,50%,75% and 100% acetate in mixed carbon sources, respectively. The granules of all systems were formed after operation for 20 d, and grew mature after 90 d. In the systems with relatively higher proportion of propionate in the mixed carbon sources than acetate, the mature granules were tended to have white color, smooth surface and ellipsoidal shape. In the systems with relatively lower proportion of propionate in the mixed carbon sources than acetate, the mature granules were tended to have buff color, dense microstructure and spheroidal shape. The phosphorus removal of granule-based EBPR system showed significant differences among the treatments, which were 97.0%,85.9%,77.4%,56.5% and 31.5% in the treatment of 0%,25%,50%,75% and 100% acetate in mixed carbon sources, respectively.At the beginning of the treatment, variety of microbial community was detected in the seeding sludge, which were about 22 different bacteriums. PAOs was dominant in the seeding sludge, including Acinetobacter, Uncultured alpha proteobacterium, Uncultured Chlorobi bacterium, Uncultured bacterium and Uncultured Rhodocyclaceae bacterium. GAOs existed in the seeding sludge was Candidatus Competibacter phosphatis. Besides that, there was certain Denitrifying bacterium in the seeding sludge as well.After operating for 30 d, the similarities of the microbial community were 48.5%,43.0%,47.6%,41.4% and 43.5% in the treatment of 0%,25%,50%,75% and 100% acetate in mixed carbon sources, respectively, comparing to the beginning of the treatment. It indicated that the microbial community structures of all systems were changed significantly during the sludge granulation progress. Uncultured bacterium, a type of PAOs that was dominant in the seeding sludge, was disappeared in all systems. Uncultured Rhodocyclaceae bacterium, some Candidatus Competibacter phosphatis and some Denitrifying bacterium were disappeared as well, while Acinetobacter, Uncultured alpha proteobacterium and Uncultured Chlorobi bacterium were still dominant in the systems.The granules grew mature in all systems after operating for three months; meanwhile the microbial community structure reached stable state. In the mature granule-based EBPR system, the microbial community was mainly including Candidatus Competibacter phosphatis, Uncultured alpha proteobacterium, Uncultured Chlorobi bacterium, Uncultured bacterium and Denitrifying bacterium. Comparing to the non-granule-based EBPR system, the microbial community of granule-based EBPR system was relatively pure.The differences among the granule-based EBPR system cultivated with mixed carbon sources of different acetate to propionate ratio, was mainly manifested as the quantity variance of Candidatus Competibacter phosphatis (GAOs) and Uncultured Chlorobi bacterium (PAOs). As the proportion of acetate in the mixed carbon sources increased, Candidatus Competibacter phosphatis proliferated in the system, which was responsible for the decline of phosphorus removal efficiency. Moreover, Uncultured Chlorobi bacterium was found to proliferate in the system with higher proportion of propionate in the mixed carbon sources, which should be another reason of the higher phosphorus removal efficiency in those systems.Additionally, the fully coupled activated sludge model No.3 (FCASM3) and a generalized model for aerobic granule-based SBR were adjusted and employed for describing the biological reaction and mass transfer progress in the granule-based EBPR system, respectively. Based on those two models, a preliminary model for granule-based EBPR system was conceived.
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