Divalent cations were known to alleviate salinity stress on anammox bacteria. Understanding the mechanism of reducing the salinity stress on anammox granules is essential for the application of the anammox process for saline wastewater treatment. In this study, the effect of Ca²⁺ and Mg²⁺ augmentation on the recovery of the activity of freshwater anammox granules affected by salinity stress was evaluated. At the condition of a salinity stress of 5 g NaCl/L, the specific anammox activity (SAA) of the granule decreased to 50% of that of the SAA without NaCl treatment. Augmentation of Ca²⁺ at the optimum concentration of 200 mg/L increased the SAA up to 78% of the original activity, while the augmentation of Mg²⁺ at the optimum concentration of 70 mg/L increased the SAA up to 71%. EPS production in the granules was increased by the augmentation of divalent cations compared with the granules affected by salinity stress. In the soluble EPS, the ratio of protein to polysaccharides was higher in the granules augmented by Ca²⁺ than with Mg²⁺, and the functional groups of the EPS differed from each other. The amount of Na⁺ sequestered in the soluble EPS was increased by the augmentation of divalent cations, which seems to contribute to the alleviation of salinity stress. Ca. Kuenenia-like anammox bacteria, which were known to be salinity stress-tolerant, were predominant in the granules and there was no significant difference in the microbial community of the granules by the salinity stress treatment. Our results suggest that the alleviation effect of the divalent cations on the salinity stress on the anammox granules might be associated with the increased production of different EPS rather than in changes to the anammox bacteria.

Anaerobic ammonium oxidation (anammox) is a biological nitrogen removal process with attractive prospects, such as no carbon addition, less aeration, lower greenhouse gas generation, and lower sludge production. However, it is difficult to maintain a stable anammox process since the anammox bacteria have a slow growth rate and high sensitivity to many factors. Therefore, it is very important to analyze and maintain the anammox activity as a process indicator for its successful operation. The conventional method for measuring the concentration of nitrogen compounds, such as ammonium, nitrite, or nitrogen gas is inconvenient during the reaction time for specific anammox activity (SAA) analysis, which can result in an inaccurately determined SAA due to the substrate loss and temperature change. In this study, a respirometer was utilized to analyze the SAA. The SAA values from a respirometer (rSAA) showed a similar pattern to the SAA values (mSAA) from the conventional method. All of the SAA analyses showed the highest value at 35 °C with a granule size of 0.05. Therefore, the respirometer can be used as a simple and efficient tool for SAA analysis. Moreover, the operating maintenance and management of the anammox process can be improved due to the simple SAA analysis in the field.

The partial nitritation (PN)–anammox (PN/A) process offers a sustainable alternative to nitrogen management in wastewater treatment, addressing the high costs and increasing the low eco-friendliness associated with traditional nitrification/denitrification processes. Stable partial nitritation (PN) is critical for effective PN/A operation, and this study specifically focused on the need to suppress nitrite-oxidizing bacteria (NOB) to facilitate the enrichment of ammonia-oxidizing bacteria (AOB). Utilizing two sequencing batch reactors (SBRs), PN1 and PN2 with different free ammonia (FA) concentrations, this study aimed to evaluate the NOB suppression strategy while enriching AOB. The PN2 reactor, which operated with a higher initial FA concentration (50 mg/L), successfully maintained high nitritation activity, with 96.1% ammonium removal efficiency (ARE) and 95.1% nitrite accumulation efficiency (NAE) at reduced influent NH₄⁺-N concentrations (50 mg NH₄⁺-N/L, FA 10 mg/L). In contrast, PN1 showed inadequate NOB suppression due to lower FA concentrations (10 mg/L). These results suggest that initiating the nitritation process with higher FA concentrations can effectively suppress NOB, enhancing the stability and efficiency of PN/A processes in mainstream applications.