Insights into Biochemical Process and Associated Microbial Mechanisms in Fermentation Reactor of the Side-Stream Enhanced Biological Phosphorus Removal (S2ebpr) Process

Abstract

The side-stream enhanced biological phosphorus (P) removal (S2EBPR) process, which harnesses anaerobic hydrolysis and fermentation of sludge to produce additional internal carbon sources, has garnered growing interest for its efficient and stable P removal performance. However, current researches on the S2EBPR process lack a comprehensive understanding of its biochemical process in the side-stream sludge hydrolysis and fermentation reactor. In this study, simulated side-stream anaerobic fermentation tests were conducted to elucidate the biochemical process and associated microbial mechanisms by investigating the release profile of sludge hydrolysis and fermentation byproducts, the EBPR activity of the fermented sludge as well as changes in physiological activity and metabolic pathway of functionally relevant microorganisms at cellular and population level. The results showed that the majority of active P release, glycogen consumption and polyhydroxyalkanoates accumulation occurred during the first 60 h of fermentation, and EBPR activity also gradually increased, possibly due to enrichment of polyphosphate (poly-P) accumulating organisms (PAOs) and elimination of glycogen-accumulating organisms (GAOs). By utilizing single-cell Raman spectroscopy (SCRS), we found that poly-P and glycogen were utilized sequentially as the main energy source during the first 60 h, then poly-P was the primary energy source in the maintenance metabolism of PAOs. Fluorescence in situ hybridization (FISH)-Raman analysis suggested that Accumulibacter tended to utilize poly-P as the major energy source, while Tetrasphaera showed a preference for glycogen over poly-P. This research improves our mechanistic understanding of the S2EPBR process and will enhance our ability to effectively design and operate these systems for stable P removal.

Keywords: Side-stream EBPR, Biochemical metabolism, Polyphosphate-accumulating organisms, Single-cell Raman spectroscopy, FISH-Raman