Molecular mechanisms of selection pressure-driven aerobic granulation in sequencing batch reactors

• Main Author: Xiong, Yanghui.
• Other Authors: Liu Yu
• Format: Theses and Dissertations
• Language: English
• Published: 2013
• Subjects: DRNTU::Engineering::Environmental engineering::Water treatment
• Online Access: http://hdl.handle.net/10356/51363
• Institution: Nanyang Technological University

Two major selection pressures, settling time and hydrodynamic shear force, have been widely reported to influence aerobic granulation in sequencing batch reactors (SBR). Big, round-shape, and dense aerobic granules can be developed at a short settling time or a high hydrodynamic shear force. However, the molecular mechanisms of aerobic granulation driven by selection pressure remain unclear. In phase 1 of this study, three SBR, namely R1, R2 and R3, were operated at different settling times of 3 min, 30 min and 60 min, respectively. A significant increase of the signaling molecule, AI-2, and enhanced production of extracellular proteins (PN), polysaccharides (PS), and deoxyribonucleic acid (eDNA) were found only in R1 which was operated at the shortest settling time of 3 min in the 45-d operation period. Enhanced PN, PS and eDNA, and AI-2 production were also observed after the settling times were shortened from 30 min to 1 min in R1 and from 60 min to 2 min in R2, respectively. These findings suggest that extracellular polymeric substances (EPS) and cellular communication contribute to settling time-induced aerobic granulation, but they played different roles. At the initial stage, secretion of EPS was triggered by a short settling time, which in turn initiated microbial aggregation leading to increased size and density of biomass, while AI-2-mediated quorum sensing (QS) was not significantly involved in initial aggregation as evidenced by a low AI-2 content. However, AI-2-mediated cellular communication mechanism was activated to regulate the growth of aerobic granules when the biomass density reached a critical value. It appears from this study that a short settling time in SBR would induce microbiological and physiological responses of bacteria which are required at different stages of aerobic granulation. In order to explore the molecular mechanisms of shear force-induced aerobic granulation, two SBR, namely R1 and R2 run at different superficial upflow air velocities of 0.4 cm s-1 and 2.9 cm s-1, respectively, were employed to develop aerobic granules. It was found that a higher superficial upflow air velocity at 2.9 cm s-1 would induce production of PN, PS and eDNA, and signaling molecules, AI-2 and AHLs. Increased PN, PS and eDNA, and signaling molecules AI-2 and AHLs were also observed after the superficial upflow air velocity in R1 was increased from 0.4 cm s-1 to 2.9 cm s-1. These suggest that both EPS and cellular communication would contribute to shear force-induced aerobic granulation, but with different roles in aerobic granulation, i.e., EPS contributed more than QS to the initiation of shear force-induced aerobic granulation, and QS was essential for structural development of aerobic granules. Reduced EPS and QS with disintegration of aerobic granules appeared after the superficial upflow air velocity in R2 was lowered from 2.9 cm s-1 to 0 4 cm s-1, suggesting that EPS and signaling molecules had essential roles in maintaining the structural stability and integrity of aerobic granules. These results offer insights into microbiological mechanisms of shear force-induced aerobic granulation in SBR. Although it was noted that PN, PS were involved in development and maintenance of the 3-D architecture of aerobic granules, some recent studies showed that PN would contribute more than PS to the formation, structure, and stability of biofilms and granules. In order to investigate the role of PN in maintaining the structural stability and integrity of aerobic granules, Proteinase K which efficiently hydrolyzes proteins, was used to remove PN from pre-cultivated mature aerobic granules. It was found that Proteinase K could effectively hydrolyze proteins, and further significantly disintegrate aerobic granules. A substantial reduction of PS was also observed concurrently with hydrolysis of PN by Proteinase K. This was probably due to collapse of extracellular matrices caused by hydrolysis of proteins. These suggest that both PN and PS are essentially involved in maintaining the structural integrity of aerobic granules. In addition, it was noted that signaling molecules, AI-2 and AHLs were inhibited by Proteinase K due to the fact that Proteinase K could degrade quorum sensing receptor proteins. This in turn provides a plausible explanation for the observed Proteinase K-triggered dispersal of aerobic granules.