Suppression of nitrite-oxidizing bacteria under the combined conditions of high free ammonia and low dissolved oxygen concentrations for mainstream partial nitritation

https://doi.org/10.1016/j.eti.2020.101135Get rights and content

Highlights

  • Partial nitritation was achieved with suspended activated sludge in CSTR.

  • Nitrite-oxidizing bacteria were successfully suppressed in mainstream conditions.

  • Low dissolved oxygen combined with high free ammonia achieved partial nitritation.

  • A ratio of 0.003 mg O2 mg-1N immediately suppressed nitrite-oxidizing bacteria.

  • NO2--N/NH3-N ratio was 1.22 which was suitable for the subsequent Anammox reaction.

Abstract

The possibility of nitrite-oxidizing bacteria (NOB) suppression in mainstream wastewater (COD/N of 6.0, ammonia of approximately 50 mg N L−1) was investigated by controlling the ratio of dissolved oxygen to total ammoniacal nitrogen (DO/TAN) in a continuous suspended sludge reactor without prior COD removal. It was found that a combination of high free ammonia (FA) of approximately 20 mg N L−1 and low dissolved oxygen (DO) of 0.3 ± 0.1 mg L−1 (DO/TAN ratio of 0.003) established in the start-up period was an efficient way to inhibit the growth of NOB. The activity of NOB was more inhibited as FA concentration increased, particularly at the initial ammonia concentration of 175 mg N L−1 (FA of about 20 mg N L−1) Then, the influent ammonia was lower to 50 ± 2 mg N L1 (FA of 4.9 ± 0.2 mg N L−1), a typical nitrogen concentration of mainstream wastewater. After four months of operation, mainstream partial nitritation (PN) became stable with nitrite accumulation ratio over 87% and COD removal of above 94% (effluent COD of 18.4 mg L−1). Besides, the effluent NO2-N/NH3-N ratio was 1.22 ± 0.12, which was suitable for the subsequent anaerobic ammonium oxidation (anammox) reaction. The relative abundance of ammonia-oxidizing bacteria (AOB) (e.g., Nitrosomonas) was 15 times higher than NOB. The abundance (0.3%) of Nitrospira (k-strategist NOB) was insignificant, while Nitrobacter (r-strategist NOB) and Nitrotoga (NOB) were not detected during the steady state of PN under low nitrogen load because NOB was suppressed and washed out of the reactor by the condition of extremely low DO/TAN ratio in the start-up period. Thus, it was noted that NOB suppression for stable PN could be accomplished by providing high FA and low DO concentrations, especially in the early stage.

Introduction

In recent years, the high speed of urbanization and industrialization has not only caused great pressure on surface water bodies due to the excess of macronutrients like nitrogen and phosphorus. Nitrogen exists in many forms, of which ammonia, nitrate, and nitrite are the main concerns in the aquatic environments (Meays, 2019). In addition, 10 mg L−1 of nitrate nitrogen in drinking water are known to cause methemoglobinemia or blue baby syndrome (Lorna, 2004). In order to satisfy the discharge limit of nitrogen, nitrification-denitrification processes have widely been practiced in wastewater treatment plants (Rodrigo et al., 2018). However, the operation cost of the nitrification-denitrification processes was high as a large amount of oxygen was required for complete nitrification (3.43 g O2/g N) (Metcalf and Eddy, 2003) as shown in Eq. (1), and electron donors (carbon sources) are required for denitrification as shown in Eq. (2) (Dhananjai and George, 1978). Particularly, extra carbon (methanol, ethanol, acetate, peptone, glycerol, lactic acid, glucose, etc.) are needed to complete denitrification reactions in wastewater with low C/N ratio (Khanitchaidecha et al., 2010). Moreover, denitrification reactions release CO2 into the atmosphere. NH4++1.83O2+1.98HCO30.021C5H7NO2+0.98NO3+1.88H2CO3+1.041H2ONO3+1.08CH3OH+H+0.065C5H7NO2+0.467N2+0.76CO2+2.44H2O

In order to improve nitrogen removal efficiency and minimize energy consumption, new processes have been developed, including Nitritation-Denitrification (short-cut nitrogen removal) (Turk and Mavinic, 1986), ANaerobic AMMonium OXidation (ANAMMOX) (Mulder, 1989), Simultaneous Nitrification and Denitrification (SND) (Pochana and Keller, 1999), and Completely Autotrophic Nitrogen-removal Over Nitrite (CANON) (Third et al., 2001).

Among these techniques, anammox is drawing much attention since anammox bacteria directly oxidize ammonia to nitrogen gas (N2) with nitrite as an electron donor under strictly anaerobic conditions (Eq. (3)) (Hu et al., 2013). In order to anammox reaction to occur, a part of incoming ammonia needs to be converted to nitrite (partial nitritation, PN), and then remained ammonia is reacted with produced nitrite to form nitrogen gas. NH4++1.32NO2+0.066HCO3+0.13H+1.02N2+0.256NO3+0.066CH2O0.5N0.15+2.03H2O

A combined reaction of PN and anammox (PN/A) provides many advantages over traditional nitrification-denitrification processes, which include low sludge generation (0.12 g VSS g−1 NH3-Nremoved), no requirement for organic electron donors and low oxygen demand (1.84 g O2 g−1 NH3-Nremoved) (WERF, 2014) yielding low energy consumption (0.024 kWh per capita per day) (Siegrist et al., 2008). For PN/A processes, besides slow growth of anaerobic ammonia-oxidizing bacteria, major difficulties reside in PN step because nitrite is promptly converted to nitrate so that nitrite is hardly accumulated in nitrifying reactors (Siegrist et al., 2008). Therefore, the oxidation of nitrite must be restricted by washing out or suppressing NOB while AOB are growing.

The PN/A process for mainstream domestic wastewater (mainstream PN/A) are currently attracting much attention worldwide because of its economic benefits. However, it has also been well recognized that PN/A in mainstream wastewater encountered major technical challenges (Lotti et al., 2015a, Lotti et al., 2015b). The first challenge was the C/N ratio. The C/N ratios of mainstream municipal wastewater after primary settling were usually in the range of 7–12 g COD g−1 N (Metcalf and Eddy, 2003) while most of sidestream wastewater to which PN/A processes have often been applied contained COD/N ratio lower than 1 (Ma et al., 2015). The high C/N ratio created competitions for oxygen among heterotrophs and autotrophs in activated sludge, resulting in reduced population and activity of AOB and lowered nitrogen removal. Secondly, low ammonia concentrations in mainstream domestic wastewater ranging from 30 mg N L1 to 100 mg N L−1 (Metcalf and Eddy, 2003) hardly inhibited NOB because FA and FNA concentrations were too low to suppress NOB (Lackner et al., 2014, Ma et al., 2015, Xu et al., 2015). Therefore, PN/A processes have usually been applied to high-strength nitrogen wastewaters including old landfill leachate (Ganigué et al., 2009, Phan et al., 2014, Li et al., 2017), sludge rejected water from mainstream wastewater treatment plants (Trojanowicz et al., 2019) piggery wastewater (Hwang et al., 2005) and anaerobic digestion liquor (Lotti et al., 2019).

PN for the sidestream wastewater has commonly been achieved by controlling dissolved oxygen (DO) at low concentrations (Thanh et al., 2013, Corbalá-Robles et al., 2016) while free ammonia (FA) (Durán et al., 2014, Phan et al., 2017) and/or free nitrous acid (FNA) (Ganigué et al., 2009) concentrations are controlled to be high. These conditions have been established by adjusting pH (pH 7.2–8.2) and substrates concentrations (> 500 mg N L−1) of FA, real-time aeration control (Soliman and Eldyasti, 2018, Phan et al., 2017) and alternating anoxic and aerobic conditions.

To date, various strategies of NOB suppression including low DO (Blackburne et al., 2008), a combination of low DO and short hydraulic retention time (HRT) (Zeng et al., 2013), real-time aeration control (Peng et al., 2004), intermittent aeration (Kornaros et al., 2008, Liu et al., 2017), etc., have been explored. Among these methods, a limited DO level has been frequently applied to successfully attain PN in wastewater treatment systems (Ma et al., 2011). Besides, a method based on the DO/TAN ratio was considered as an effective method to achieve PN and was used by Isanta et al. (2015), Reino et al. (2016), Liu and Yang (2017) to treat low strength wastewater. Among these studies, Isanta et al. (2015) and Reino et al. (2016) were successful in achieving PN in granular sludge processes. A steep gradient of oxygen concentration in granules led to the competition for oxygen between AOB at the outer space and NOB in the inner layer. It was though that the activity of AOB was higher than that of NOB under low DO conditions because half-saturation constants of AOB for oxygen were lower than those of NOB (AOB: 0.2 to 0.4 mg L−1, NOB: 1.2 to 1.5 mg L−1) (Wiesmann, 1994, Liu et al., 2017). Therefore, NOB were easily suppressed in a granular reactor under low DO/TAN condition.

On the other hands, Liu and Yang (2017) did not achieve PN in a continuous suspended sludge reactor with DO/TAN ratio of 0.02, which was lower than that of other studies (Isanta et al., 2015, Reino et al., 2016). Moreover, Isanta et al. (2015) confirmed that NOB were not suppressed in conventional floccular sludge, where oxygen condition gradients were hardly established. From these results, it was noticed that the DO/TAN of 0.02 which was applied by Liu and Yang (2017) was not sufficiently low for NOB suppression in suspended sludge.

Therefore, it was assumed in this study that NOB would be suppressed by combined conditions of high FA and low DO (i.e., low DO/TAN). Thus, it was designed that the PN-CSTR received NH3-N of 175 mg N L−1 (FA of approximately 20 mg N L−1) and DO of 0.3 ± 0.1 mg L−1 in the start-up period to suppress NOB. Accordingly, an extremely low DO/TAN of 0.003 was established when the PN reaction occurred. Then, the concentration of NH3-N gradually lowered to 50 mg N L−1, a typical level of mainstream wastewater.

Section snippets

Bioreactor set up and operating conditions

A lab-scale CSTR with a working volume of 10 L and a settling tank of 5 L was operated at 27 ± 2 °C. The DO was controlled at 0.3–2.5 mg L−1 by adjusting aeration rates in the range of 8×10−3−2×10−2 vvm through a mass flow controller (MFC) (TSC-220, MPK, Korea) and the pH was adjusted at pH 8.1 ± 0.2. The reactor was inoculated with activated sludge obtained from a wastewater treatment plant in Suwon, Korea. The initial mixed liquor suspended solids (MLSS) concentration was approximately

Overall performance of PN-CSTR

As shown in Fig. 1, the PN process proceeded continuously for 115 days. In the period I (day 1–15), the influent concentration of NH3-N was 175 mg N L−1 (NLR of 0.12 kg N m−3 d1) and DO concentration in the reactor was maintained at 0.2 ± 0.1 mg L−1. FA concentrations in influent and effluent calculated by Eq. (4) were 19.9 mg N L−1 and 17.3 mg N L−1, respectively. The oxidation of ammonia to nitrite was insufficient with ARE of 16.24%, and effluent NO2-N concentration was lower than 2 mg N L

Conclusions

In this study, NOB was completely suppressed in a continuous suspended sludge reactor by maintaining high FA and low DO concentrations (DO/TAN ratio of 0.003). The mainstream partial nitritation was stable during four months of operation with nitrite accumulation above 87% and COD removal above 94%. Besides, the effluent NO2-N/NH3-N was 1.2 ± 0.2, which was similar to the anammox stoichiometric ratio. The inhibition of NOB activities appeared proportional to the increase of FA concentrations

CRediT authorship contribution statement

Linh-Thy Le: Conceptualization, Investigation, Writing - original draft. Sooyeon Lee: Formal analysis, Data curation. Xuan-Thanh Bui: Review & editing. Deokjin Jahng: Supervision, Writing - review & editing.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgment

This work was supported by National Research Foundation of Korea as “Individual Basic Science & Engineering Research Program” under Grant [number NRF-2018R1D1A1B07049694].

References (74)

  • LiuW. et al.

    Evaluating the feasibility of ratio control strategy for achieving partial nitritation in a continuous floccular sludge reactor: Experimental demonstration

    Bioresour. Technol.

    (2017)
  • LottiT. et al.

    Two-stage granular sludge partial nitritation/anammox process for the treatment of digestate from the anaerobic digestion of the organic fraction of municipal solid waste

    Waste Manage.

    (2019)
  • LottiT. et al.

    Faster through training: the anammox case

    Water Res.

    (2015)
  • MaY. et al.

    Achieving nitrogen removal via nitrite in a pilot-scale continuous pre-denitrification plant

    Water Res.

    (2009)
  • MaB. et al.

    The feasibility of using a two-stage autotrophic nitrogen removal process to treat sewage

    Bioresour. Technol.

    (2011)
  • Mosquera-CorralA. et al.

    Partial nitrification in a sharon reactor in the presence of salts and organic carbon compounds

    Process Biochem.

    (2005)
  • PedrousoA. et al.

    Nitrite oxidizing bacteria suppression based on in-situ free nitrous acid production at mainstream conditions

    Sep. Purif. Technol.

    (2017)
  • PishgarR. et al.

    Denitrification performance and microbial versatility in response to different selection pressures

    Bioresour. Technol.

    (2019)
  • PochanaK. et al.

    Study of factors affecting simultaneous nitrification and denitrification (SND)

    Water Sci. Technol.

    (1999)
  • ReinoC. et al.

    Kinetic and microbiological characterization of aerobic granulesperforming partial nitritation of a low-strength wastewater at 10 °C

    Water Res.

    (2016)
  • ThanhB.X. et al.

    Fouling characterization and nitrogen removal in a batch granulation membrane bioreactor

    Int. Biodeterioration Biodegrad.

    (2013)
  • ThirdK.A. et al.

    The CANON system (completely autotrophic nitrogen-removal over nitrite) under ammonium limitation: Interaction and competition between three groups of bacteria

    Syst. Appl. Microbiol.

    (2001)
  • WangJ. et al.

    Sludge bulking impact on relevant bacterial populations in a full-scale municipal wastewater treatment plant

    Process Biochem.

    (2014)
  • ZengW. et al.

    Population dynamics of nitrifying bacteria for nitritation achieved in Johannesburg (JHB) process treating municipal wastewater

    Bioresour. Technol.

    (2014)
  • ZengW. et al.

    Nitritation and denitrifying phosphorus removal via nitrite pathway from domestic wastewater in a continuous MUCT process

    Bioresour. Technol.

    (2013)
  • AonofrieseiF. et al.

    Activated sludge bulking episodes and dominant filamentous bacteria at wastewater treatment plant Contanta Sud (Romania)

    Proc. Rom. Acad.

    (2007)
  • APHA (American Public Health Association/American Water Works Association/Water Environment Federation), ., 1998....
  • BaeW. et al.

    Optimal operational factors for nitrite accumulation in batch reactors

    Biodegradation

    (2001)
  • BlackburneR. et al.

    Partial nitrification to nitrite using low dissolved oxygen concentration as the main selection factor

    Biodegradation

    (2008)
  • BockE. et al.

    Cell biology of nitrifying bacteria

  • Corbalá-RoblesL. et al.

    Analysing the effects of the aeration pattern and residual ammonium concentration in a partial nitritation-anammox process

    Environ. Technol.

    (2016)
  • DaimsH. et al.

    In situ characterization of Nitrospira-like nitrite-oxidizing bacteria active in wastewater treatment plants

    Appl. Environ. Microbiol.

    (2001)
  • DhananjaiB.S. et al.

    Kinetics of nitrification and denitrification reactions

    Biotechnol. Bioeng.

    (1978)
  • DowningL.S. et al.

    Effect of oxygen gradients on the activity and microbial community structure of a nitrifying, membrane-aerated biofilm

    Biotechnol. Bioeng.

    (2008)
  • DuránU. et al.

    Enhanced ammonia removal at room temperature by pH controlled partial nitrification and subsequent anaerobic ammonium oxidation

    Environ. Technol.

    (2014)
  • ElawwadA. et al.

    Long-term starvation and subsequent recovery of nitrifiers in aerated submerged fixed-bed biofilm reactors

    Environ. Technol.

    (2013)
  • FordD.L. et al.

    Comprehensive analysis of nitrification of chemical processing wastewaters

    J. Water Pollut. Control Fed.

    (1980)
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