Mechanistic Evaluation Of The Adaptation of Nitrifiers and PAOs To Low DO Operation

Introduction - Biological nutrient removal (BNR) under conventional DO concentrations (>2 mg/L) is implemented to ensure that high nitrification and phosphorus removal efficiency is maintained. However, implementing low DO BNR could provide extensive benefits. Required aeration energy is decreased, less DO is recycled to anoxic zones, less supplemental carbon and alkalinity addition are required, and when utilizing advanced aeration with an ammonia residual there could be a decreased chlorine disinfection demand along with the possibility of implementing AvN aeration control for to the implementation of shortcut N removal. Low DO nitrification and phosphorus removal are possible at moderate to high SRTs in bench scale reactors (Bellucci et al., 2011; Izadi et al., 2022; Liu & Wang, 2013) and pilot-scale activated sludge processes (Keene et al., 2017), with DO concentrations below 0.5 mg/L being achieved. Gradually decreasing DO concentration was suggested as a method to maintain nutrient removal while adapting polyphosphate accumulating organisms (PAOs) and nitrifying biomass to low DO operation (Izadi et al., 2022; Keene et al., 2017). It is proposed that as PAOs and nitrifiers adapt to low DO they adopt lower oxygen half-saturation coefficients due to a higher affinity for oxygen. A possible shift and enrichment in the microbial community has been seen with adaptation in which a shift of the dominating microorganisms in the community could occur (Izadi et al., 2022; Liu & Wang, 2013). Additionally, simultaneous nitrification and denitrification (SND) in low DO aerobic zones is a possibility based on the availability of carbon and nitrate for ordinary heterotrophic organisms (OHOs) (Keene et al., 2017; Klaus & Bott, 2020). The main question surrounding low DO BNR operation is whether it is possible to achieve without losing treatment capacity. Conventional thinking and models suggest that low DO operation requires increasing the aerobic SRT to achieve the same effluent NH4 and PO4 concentrations, leading to an effective loss of treatment capacity. The objective of this project is to assess whether PAO and nitrifier adaptation to low DO can occur without requiring an increase in aerobic SRT and to determine how best to achieve that adaptation to maintain growth rates that are consistent with expectations at DO concentrations > 2 mg/L. Hypothesis and Objectives –
It was hypothesized that nitrifying (AOB and NOB) and phosphorus removing (PAO) organisms adapt to gradually decreasing DO concentrations over time, adopting lower oxygen half saturation coefficients while maintaining their maximum specific growth rates. Furthermore, OHOs also can adapt to low DO conditions, which may negatively impact SND rates. Glycogen accumulating organisms (GAOs) may also adapt to low DO conditions, competing for carbon with PAOs in the anaerobic zone, but based on previous studies (Carvalheira et al., 2014; Oehmen et al., 2007), we hypothesize that PAOs have the potential to outcompete GAOs at lower operating DO. AOB, NOB, OHO, and PAO kinetics were measured over the course of a controlled reduction in operating DO in a BNR pilot scale process. SND was monitored during adaptation by measuring aerobic denitrification rates. Floc settleability was closely monitored since heterotrophic filamentous biomass may grow at low DOs when readily biodegradable carbon is available and cause sludge bulking. Changes in floc morphology were monitored with microscopy. Lastly, the microbial community was characterized before, during, and after the adaptation to determine any population shifts and changes in gene expression. Nitrifier oxygen kinetics for nitrifying activated sludge at the HRSD VIP main plant were measured over the course of a gradual decrease in DO. A respirometric test and substrate utilization test were conducted simultaneously for direct comparison. Methods – The pilot plant was fed raw sewage from the HRSD VIP treatment plant and included preliminary treatment, primary clarification, and BNR. Preliminary treatment included 6mm mechanical screens, grit removal, 2mm mechanical screens, and temperature control to 20 degrees C. Primary effluent was pumped into the BNR stage which was set up to run as an A/O BNR activated sludge process consisting of an anaerobic tank followed by five aerobic tanks in series (Figure 1). The flow into the pilot plant was set for 1 gpm. The aerobic portion of the process was initially operated at a conventional DO concentration of 2 mg/L, with an aerobic SRT of 4-6 days. The DO concentrations in the aerobic tanks were then aggressively decreased until the maximum target effluent ammonia (1 mg/L-N) and total phosphorus (1 mg/L-P) concentrations were approached. At this point the DO was gradually decreased over time with gradual increases in SRT as needed to maintain the target effluent concentrations. Weekly batch tests were conducted for biomass activity, oxygen half-saturation coefficients, and substrate half-saturation coefficients as the low DO adaptation occurred in the pilot plant. Other kinetic testing included batch tests for SND rates, denitrifying PAOs (DPAOs), denitrifying GAOs (DGAOs), and internal carbon storage. Daily sludge volume index (SVI) measurements were taken using a settleometer to monitor settleability. Daily influent and effluent sampling (composite samples) of the BNR process were analyzed for VFA, NH3, NO2, NO3, OP, tCOD, sCOD, HACH TNT kits (HACH, CO, USA), BOD, TSS, VSS. Grab samples for nutrient profiles along the reactors were also used. Online sensors and analyzers included pH, DO, MLSS, NH3, NO2, NO3, N2O, OP, and temperature. Floc imaging was conducted to determine floc morphology changes. Weekly microbial samples were also taken for metagenomic analysis.
Results The results from tracking changes in the oxygen half saturation constants for the nitrifiers in HRSD's VIP plant are shown in Figure 3, which indicate that the oxygen half saturation constants decreased over time as the average operating DO was gradually decreased. The nitrifier substrate uptake rates from the respirometric and substrate utilization tests used were fit to Monod curves and plotted together for direct comparison. An example of this can be seen in Figure 2, which shows consistent curve fits and Ko estimates for both tests. As the DO is gradually decreased in the pilot, the oxygen half saturation constants are expected to decrease with DO, similar to the VIP plant. This shift in oxygen half saturation constant at the same SRT would mean less DO being required to achieve the same effluent concentrations, which can be seen in the effluent ammonia example in Figure 4. Acknowledgements- This material is based upon work supported by the U.S. Department of Energy's Office of Energy Efficiency and Renewable Energy (EERE) under the Advanced Manufacturing Office, Award Number DE-EE0009509 'Transforming Aeration Energy in Water Resource Recovery Facilities (WRRFs) through Suboxic Nitrogen Removal' to Carollo Engineers, Inc and partners