Evaluation of Intracellular Polyphosphate Dynamics in Enhanced Biological Phosphorus Removal Process using Raman Microscopy

A Raman microscopy method was developed and successfully applied to evaluate the dynamics of intracellular polyphosphate in polyphosphate-accumulating organisms (PAOs) in enhanced biological phosphorus removal (EBPR) processes. Distinctive Raman spectra of polyphosphates allowed for both identification of PAOs and quantification of intracellular polyphosphate during various metabolic phases in a lab-scale EBPR process. Observation of polyphosphate at individual cell level indicated thatthere are distributed states of cells in terms of polyphosphate content at any given time, suggesting that agent-based distributive modeling would more accurately reflect the behavior of an EBPR process than the traditional average-state based modeling. The results, for the first time, showed that the polyphosphate depletion or replenishment observed at the overall population level were collective results from shifts/transition in the distribution of abundance of PAOs with different amounts of polyphosphate inclusions during EBPR. Imaging construction based on simultaneous quantification of intracellular polyphosphate and protein revealed the spatial distribution of polyphosphate inside cells and showed that the polyphosphates accumulate in smaller or larger aggregates, rather than being evenly distributed within the cytoplasm. The results demonstated that Raman microscopy will allow for detailed cellular-level evaluation of polyphosphate metabolism and dynamics in EBPR processes and revealed mechanism insights, which otherwise would not be obtained using a traditional bulk measurement-based approach.