Journal of Water Process Engineering 45 (2022) 102461

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Journal of Water Process Engineering

journal homepage: www.elsevier.com/locate/jwpe

Achieving biological nutrient removal in an old sewage treatment plant

through process modifications – A simulation and experimental study
Reshma Mohan T a, *, H.N. Chanakya b, M.S. Mohan Kumar c, Lakshminarayana Rao b
a
Interdisciplinary Centre for Water Research, Indian Institute of Science, Bangalore, India
b
Centre for Sustainable Technologies, Indian Institute of Science, Bangalore, India
c
Department of Civil Engineering, ICWaR, IFCWS and RBCCPS, Indian Institute of Science, Bangalore, India

A R T I C L E I N F O A B S T R A C T

Keywords: Modifying old sewage treatment plants (STP) to meet revised stringent discharge standards is of great concern.
Wastewater treatment plant Process modeling can serve as an essential tool for this. This work reports a BioWin©-activated sludge model/
Biological nutrient removal anaerobic digestion for a 55 million litres per day STP located in South India. Fifty state variables and eighty
BioWin©
process equations were used in the model to achieve biological nutrient removal. The model was calibrated and
Process modification
Sensitivity analysis
was validated with plant data. The proposed modification was to incorporate an intermediate virtual anoxic zone
to achieve simultaneous nitrification-denitrification and total dissolved phosphorus (TDP as PO4) removal.
Several scenarios, namely, different hydraulic residence time (HRT) between 1.5 to 2.5 h, dissolved oxygen (DO)
between 2.5 to 4.5 mg/L, and mixed liquor suspended solid (MLSS) between 2500 to 4500 mg/L in the biore­
actor, were studied to identify optimum conditions. The optimum DO and MLSS levels were identified as 4 mg/L
and 4000 mg/L, respectively. The optimum HRT of 2 h in aeration zone, 1 h in anoxic, and 3 h in reaeration zone
was identified. Implementation of these modifications in the STP, with minimal operational interventions and no
capital costs, improved its performance as predicted by the model. The total nitrogen and TDP (as PO4) reduced
from 20 mg/L to 8 mg/L and 3.5 mg/L to 0.9 mg/L, respectively, and met the revised discharge standards. This
intervention gave a cost saving of approximately 5.6 million USD.

1. Introduction
The treated water from a sewage treatment plant (STP) is generally
discharged into a nearby flowing surface water body, wherein the water
improves its quality through natural processes before it is picked up by
another city downstream for its usage [1–3]. Urbanization has led to the
growth of cities, thereby reducing the distance the treated water travels
between adjacent cities. This, in turn, demands that the STPs treat the
water to higher standards. Authorities have continuously revised water
discharge standards of STPs, making it more stringent every decade,
requiring the STPs to be constantly upgraded (during their typical life of
about 25–30 years) to meet the new standards. Table S1 of supple­
mentary material tabulates how the treated water quality standards
have been made more stringent over the years in India. The up-gradation
can be achieved either by process modifications or retrofitting with
modern machinery. However, retrofitting an existing STP can be an
expensive proposition, both in terms of time and resources. Thus, it is
usually preferred to modify the treatment process to meet the new
standards without compromising the STP capacity. Mathematical
models are useful tools for process modifications and optimization of
STPs. Among the various mathematical models, activated sludge models
(ASM) are widely used for plant process optimization [4–6]. BioWin©
software [7] with built-in ASMs (such as ASM1, ASM2d, and ASM3) is
used for such process modeling. This study discusses process modifica­
tions using BioWin© and its implementation on an existing operational
STP to meet the new discharge standard. As revised standards demanded

Abbreviations: ASM, Activated Sludge Model; ASDM, Activated Sludge-Digestion Model; BNR, Biological Nutrient Removal; BOD, Biological Oxygen Demand;
CAS, Conventional Activated Sludge Process; COD, Chemical Oxygen Demand; DO, Dissolved Oxygen; EBPR, Enhanced Biological Phosphorous Removal; HRT,
Hydraulic Retention Time; MLD, Million Litres per Day; MLSS, Mixed Liquor Suspended Solids; NH4-N, Ammonical Nitrogen; NO3-N, Nitrate Nitrogen; NO2-N, Nitrite
Nitrogen; PAO, Phosphorous Accumulating Organisms; R ratio, Return Activated Sludge Ratio; SRT, Sludge Retention Time; STP, Sewage Treatment Plant; TDP, Total
Dissolved Phosphorous (as PO4); TKN, Total Kjeldahl Nitrogen; TN, Total Nitrogen.
* Corresponding author.
E-mail address: reshmat@iisc.ac.in (R. Mohan T).

https://doi.org/10.1016/j.jwpe.2021.102461
Received 11 July 2021; Received in revised form 18 November 2021; Accepted 20 November 2021
Available online 29 November 2021
R. Mohan T et al.
stringent biological nutrient removal (BNR), this study has focused on
the same.
The use of BioWin© ASM models for biological nutrient removal
(BNR) in wastewater treatment has been studied by several researchers
[8–13]. Dynamic BioWin© ASM models have been used in literature to
optimize conventional activated sludge (CAS) processes wherein they
have recommended either increasing the mean cell residence time
(MCRT) to 40–50 days or including ammonia stripping [14]. Bizukojc
et al. have identified kinetic and stoichiometric parameters required for
modeling [15,16] and have validated their models with literature data.
Even though several studies have focused on developing BioWin© based
models for process optimization, few studies have implemented the
model recommendations and observed real-time BNR removal in full-
scale STPs. Also, these studies focus only on a specific type of process
modifications, e.g., introducing anoxic zones, improving the aeration
capacity of blowers in activated sludge tanks, and varying the return
activated sludge ratio (R). However, a comprehensive study investi­
gating the combined effect of several process modifications on
improving the treatment and economic efficiency with minimal in­
terventions is lacking in the literature. In this work, it is hypothesized
that creation of virtual anoxic zones in the bioreactor coupled with
appropriate modifications of the operating parameters such as hydraulic
retention time (HRT), dissolved oxygen (DO) levels and R ratio in the
bioreactor would result in SNDN and EBPR. A BioWin©-ASM model was
developed for the 55 million litres per day (MLD) STP and was validated.
This validated model was used to identify the optimum conditions for
process modifications to achieve BNR. Several sensitivity studies were
also conducted to assess the combined effect of various operating con­
ditions, including DO concentrations, mixed liquor suspended solid
(MLSS) concentrations, HRT, and R ratio on the BNR process. Process
economics and the potential advantages of following a similar approach
on many existing old STPs are briefly discussed.
1.1. Biological nutrient removal (BNR): theory and mechanism
Biological nitrogen removal is a two-step simultaneous nitrification-
denitrification (SNDN) process [17]. During nitrification, the influent
ammonia (NH3) is oxidized into nitrite (NO2) by the ammonia-oxidizing
bacteria (AOB). The main reactions describing this conversion of NH3 to
NO2 are given in Eqs. (1) and (2). As shown in Eqs. (1) and (2), NH3 is
converted into intermediate hydroxylamine (NH2OH) and is further
converted to NO2.
NH3 + O2 + 2e− →NH2 OH + H2 O (1)
NH2 OH + H2 O→NO−2 + 4H+ + 4e− (2)
The NO2 thus formed is further oxidized into nitrate (NO3) by the
nitrite-oxidizing bacteria (NOB). The reaction describing the conversion
of NO2 to NO3 is given in Eq. (3).
NO−2 + 0.5 O2 →NO−3 (3)
Denitrification includes the biochemical reduction of nitrate‑ni­
trogen (NO3-N) into dinitrogen gas under anoxic conditions by the
heterotrophic bacteria in the presence of an organic carbon source
(COD). Eq. (4) shows the complete redox reaction of denitrification.
2NO−3 + 10e− + 12 H+ →N2 + 6H2 O (4)
Thus, one can achieve simultaneous nitrification and denitrification
(SNDN) in a bioreactor under appropriate process conditions.
Removal of total phosphorous (TP) in STPs is achieved either by the
addition of phosphorus reacting chemicals or by enhanced biological
phosphorous removal (EBPR) process [19]. In the EBPR process, a group
of specialized bacteria known as the luxury phosphorous accumulating
organisms (PAO) removes phosphorous in two stages [20]. In the first
stage, PAOs uptake volatile fatty acids (VFA) in the wastewater under
2
Journal of Water Process Engineering 45 (2022) 102461
anaerobic conditions and store them as polyhydroxyalkanoate (PHA).
They also release phosphorus (orthophosphate) in this stage to provide
the energy for the uptake of the VFAs. Under aerobic conditions, the
stored PHA is metabolized to provide energy for cell growth in the
second stage. Simultaneously, the soluble orthophosphates are
consumed and stored, resulting in a net phosphorous removal from the
wastewater [20]. The NMR spectroscopy method is the most commonly
used method to quantify these polyphosphates released and consumed
in EBPR process. The quantification using spectroscopy involves six
steps: pre-centrifugation of sludge, pre-extraction using EDTA – NaOH
solution, centrifugation and separation of sludge pellet, primary
extraction, concentration by rotary evaporation, and measurement
using NMR spectroscopy [22]. A dominant factor in the EBPR by the
PAOs is the amount of readily biodegradable COD and the VFAs present
in the influent [20]. Some typical operating conditions affecting EBPR
are given in detail in the Supplementary Section S3.
Several studies have examined the possibilities of modifying the
conventional activated sludge (CAS) process to improve BNR, i.e.,
removal of nitrogen and phosphorus (N and P) biologically
[1,19,20,24,25]. The approach of retrofitting existing STPs to reduce N
and P by controlling the aeration and non-aeration periods was inves­
tigated by Hanhan et al. and Habermeyer et al. [26,27]. They identified
that operating STPs with optimum dissolved oxygen (DO) levels be­
tween 1.5 and 1.7 mg/L coupled with intermittent aeration, i.e., 180
min of aeration followed by 160 min of no-aeration, resulted in
maximum N and COD reduction. There have been only a few studies that
focus solely on EBPR since it requires strict anaerobic conditions. Ber­
nard et., al. (2017) has observed that it was possible to achieve EBPR by
creating deep anaerobic zones. Using slow-speed top entry mixers of ~2
W/m3 capacities in aeration tanks, they have maintained DO <0.5 mg/L
and have achieved EBPR [19]. Khiewwijit et al. (2015) have inferred
from their numerical studies that 80% of P recovery and 35% reduction
in CO2 emission (in CAS) was possible by including bio-flocculation and
partial nitrification conditions [29]. In the present study, a BioWin©
ASM-based model is used to optimize BNR in the 55 MLD plant.
2. Model development and theory
2.1. BioWin© model theory
The BioWin© uses a general activated sludge/anaerobic digestion
(ASDM) model for all the process simulations. The general ASDM model
has fifty state variables and eighty process expressions for the growth
and decay of microorganisms. The basic expression in the ASDM model
for the time rate of change of microbial biomass (X) is based on Monod
kinetic equation and mass balance is given as [17]:
dX
= Microbial growth − Microbial Decay (5)
dt
dX
(
S
) (
S
)
= μmax X0 + ημmax X0 − bX0 (6)
dt S + Ks S + Ks
where, X0 is the microbial biomass concentration at time t0, μmax is the
maximum growth rate of this microorganism, S is the concentration of
the limiting substrate for growth, Ks is the half-saturation coefficient, η is
the correction factor for anoxic growth and b is the decay coefficient for
the biomass. The first term in the R.H.S of Eq. (6) represents the mi­
crobial growth under aerobic condition, the second term represents the
microbial growth under anoxic condition and the third term represents
the microbial decay. Eq. (6) can be modified depending on the type of
microbial mass, the substrate that is being utilized and the environ­
mental conditions (pH, DO etc.) that limits its growth.
The substrate (S) utilization rate by the microbes is given as [17]:
( )
dS μmax X0 S
= (7)
dt Y S + Ks
R. Mohan T et al.
where, Y is the yield coefficient. The substrate concentration with time
also varies with hydrolysis, ammonification, and denitrification pro­
cesses. To include these, additional terms are added to Eq. (7). The
processes that are included in the ASDM model for the growth, decay of
microorganisms and the substrate synthesis in an activated sludge (AS)
system are given in Table S2. There are several more processes that are
incorporated in BioWin©, but for the present study, for the proposed
modification with aeration and anoxic zones, the processes listed in
Table S2 were employed. More detailed explanation of the process
theory is available in the BioWin© help manual [18].
The mass transfer of air and gases to the liquid phase is modeled in
BioWin© using the fundamental theory of surface renewal. A few as­
sumptions are made in the BioWin© mass transfer model, including the
assumption of completely mixed gas and liquid phases with uniform
concentration, and a constant gas holdup throughout the control volume
(being modeled). The mass balance for the liquid and gas phase using
the mass transfer theory is given respectively as [18]:
dMjT [ in in ] [ (
*
) ]
= Q .Cjt − Q.Cjt + αF.(kL aL )j . C∞,j − CUN,j .VL + Rxj (8)
dt
dmj [ in in ] [ ( ) ]
*
= q .cjt − q.cjt + αF.(kL aL )j . C∞,j − CUN,j .VL (9)
dt
where, Mj,T is the mass of component j in the liquid phase, Qin and Q is
the liquid flow into and out of the control volume respectively, Cjtin and
Cjt is the influent and effluent concentration of component j respectively,
(kLaL)j is the overall liqid mass transfer coefficient for component j, α is
the ratio of diffuser for process water to clean water, F is the ratio of
diffuser for process water to given time of diffuser to process water, C∞,j*
is the saturated concentration of component j at the gas/liquid interface,
CUN,j is the bulk liquid concentration of unionized component j, VL is the
volume of the liquid phase and Rxj is the net production of component j
by reaction. For Eq. (9), mj is the mass of component j in the gas phase,
qin and q is the gas flow into and out of the control volume respectively,
cjtin and cjt is the influent and effluent concentration of gas component j
respectively. Eqs. (8) and (9) are simultaneously solved to include the
mass balance and mass transfer of liquid and gas phases.
The BioWin© simulator has two numerical modules for solving,
namely the steady state and dynamic interactive modules. Steady state
module is applied for analysing systems based on constant influent
loading and operating parameters. The steady state solver uses a com­
bination of Newton- Raphson (NR) second order method and decoupled
linear search (DLS) method for the solution [18]. The second module is
the dynamic interactive module, which is specifically suitable for time
varying inputs and time varying operating conditions. The convergence
of dynamic simulations is determined by the allowable minimum rela­
tive error. BioWin also has an option for providing concentration limits
for parameters (default value - 0.001 mg/L). The lower the set concen­
tration limits, the higher is accuracy of simulation [18]. For the present
study, all simulations were carried out using the dynamic interactive
module, since all the influent and operating conditions were varying
with time. Fifty-nine differential process equations were solved to find
the time rate change of substrate and biomass using the BioWin© model.
The flow balance in the BioWin© model is based on the fundamental
principles of continuity, momentum and energy conservation. The HRT
of each unit (aeration/anoxic tanks) is calculated by dividing the volume
‘V’ of the tank (m3) by the amount of discharge ‘Q’ (m3/h) coming to the
unit. The initial conditions that are to be provided for the model includes
the quantity of water treated per day (MLD), the volume of each unit of
the STP (m3), the inlet raw sewage quality (COD, BOD, TN, NH4-N, NO3-
N, TDP as PO4 and pH), DO and MLSS concentration in aeration zones,
return activated (R) sludge ratio, calibrated kinetic and stoichiometric
parameters and wastage ratio (W).
The assumptions while using the BioWin – ASDM model and the
differences from the field condition were [18]:
3
Journal of Water Process Engineering 45 (2022) 102461
• The bioreactor was well-mixed, and the concentration of all com­
ponents (including DO in aeration and reaeration zone) was uniform
throughout the reactor. In actual field conditions, the bioreactor was
well-mixed (in aeration and reaeration zones) due to the aeration/
mixing provided by the surface blowers.
• All scenarios in the study were modeled at a constant temperature of
26 ◦ C. While the average field temperature during sampling (pre and
post-modification) was 26 ± 3 ◦ C.
• The model was simulated for the maximum flow of 55 MLD. How­
ever, in the field conditions, due to variation in the quantity of the
incoming sewage the actual flow was 53 ± 2 MLD.
• For modeling purposes, the microbial growth and decay were
assumed to follow a first-order kinetics.
• For modeling purposes, the microbial growth and substrate utiliza­
tion were assumed to follow Monod kinetics.
• The oxygen transfer from air to liquid (wastewater) was modeled
using simple surface renewal theory.
2.2. Wastewater treatment plant process description
The Koramangala and Challaghatta (K&C) Valley wastewater treat­
ment plant, which was commissioned in 1979 and located in the Ban­
galore Urban district in Karnataka, India was chosen as the study STP.
This K&C Valley plant has three CAS-STPs (30 MLD, 60 MLD and 218
MLD) with a total design capacity of 318 MLD. The secondary treated
water from the K&C Valley plant is pumped into a network of run-off
channels, which eventually fill up 134 tanks, rejuvenating by indi­
rectly recharging the depleted groundwater aquifers in the parched
districts of Kolar and Chikkaballapur districts in Karnataka. In the 218
MLD STP of the K&C Valley unit, after the primary treatment (coarse
screens, fine screens, and grit chambers), the incoming wastewater is
divided into 5 streams (three 36 MLD streams and two 55 MLD streams).
The present study focuses on one of the 55 MLD wastewater stream. The
55 MLD treatment unit consists of a primary clarifier, an aeration tank
equipped with equally spaced surface aerators and two secondary clar­
ifiers as shown in Fig. 1 (a). The details of the individual treatment
processes of the 55 MLD unit, the dimensions and the operating condi­
tions are given in Table 1. The raw sewage, primary settled sewage and
treated water parameter values for the 55 MLD unit is given in Table 3.
2.3. Description of BioWin© model and model calibration
BioWin© 5.3 [7] was used to create a plant wide ASM model for the
55 MLD unit. The existing conditions before process modifications were
modeled as the baseline scenario. For the baseline scenario, the model
included the primary clarifier, the aeration tank and the two secondary
clarifiers of the 55 MLD unit as shown in Fig. 1. The stoichiometric and
kinetic parameters were calibrated against the baseline scenario ac­
cording to STOWA protocol [19] as shown in Table 4.
The total COD in wastewater was divided into biodegradable (bCOD)
and non-biodegradable (nbCOD) COD. The biodegradable fraction was
further divided into readily biodegradable (rbCOD) and slowly biode­
gradable (sbCOD) COD. The non-biodegradable fraction consisted of
non-biodegradable soluble COD (nbsCOD) and non-biodegradable par­
ticulate COD (nbpCOD). Using respirometric tests of the endogenous
activated sludge and influent sample, the bCOD (mg/L) and rbCOD (mg/
L) was determined as [20]:
CO
bCOD = (10)
1 − YH
COs
rbCOD = (11)
1 − YH
where ‘CO’ and ‘COs’ were the consumed oxygen of sample and
consumed oxygen in soluble (filtered sample) sample (mg/L) respec­
tively, and ‘YH’ was the heterotrophic yield coefficient (g cell COD
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Fig. 1. Layout for the 55 MLD K&C Valley plant for (a) baseline scenario (before modification), (b) after process modification; S1 to S4 are wastewater sam­
pling locations.

characteristic fractions in the model were identified [19] and were

Table 1
adjusted to match the measured treated water values specifically for
55 MLD unit individual tank dimensions and operating conditions.
TKN, NH4-N, NO3-N and TN values. The final adjusted values and their
Treatment unit Dimensions Operating conditions references are shown in Table 4. The sludge retention time value was
Circular tank, calculated based on the BioWin© model prediction values. The cali­
Primary clarifier Diameter 45 m, HRT: 2.1 h brated model was validated for the baseline scenario and the results of
Water depth 3 m. the model validation are discussed in Section 3.1. The DO concentration,
DO concentration: 2.5 mg/L
Air flow rate:10,051 m3/h
MLSS concentrations and the R-ratio in the aeration tank were main­
Rectangular tank,
Aerators: Equally spaced tained at 2.5 mg/L, 2800 mg/L and 0.69 (1600 m3/h of RAS), respec­
Length 78 m,
Aeration tank
Width 47.2 m,
surface aerators tively. The kinetic and stoichiometric parameters as shown in Table 4
MLSS concentration: 2800 were used for simulation. The STOWA protocol recommends using data
Water depth 3.9 m
mg/L
that is clearly different from the one used in calibration and the same
HRT: 6.0 h
Circular Tank, was followed in the present study.
Secondary clarifier Diameter 42.5 m, HRT:2.5 h
Water depth 3.5 m 3. Methods
Return activated sludge 6 number – 15 kW Total RAS flow rate:
(RAS) pump house pump 1200–1600 m3/h
3.1. Process modification and sensitivity studies using validated BioWin
model
formed (g COD oxidized)− 1). The value of heterotrophic yield coefficient
‘YH’ was assumed as 0.72 (g cell COD formed (g COD oxidized)− 1) at 26o To achieve BNR removal, a few modifications in the treatment pro­
C as specified in the ASM 1 model theory [21]. cess were conceived and simulated first on the validated BioWin model.
The modification was to provide a virtual anoxic zone between two
nbsCOD = CODS − rbCOD (12)
aerobic zones to improve the SNDN and BPR process by selectively
switching OFF the surface aerators present in this section of the biore­
nbpCOD = COD − bCOD − nbsCOD (13)
actor. The presence of aeration zones followed by anoxic zones is known
where ‘COD’ and ‘CODs’ were total and soluble (filtered sample) COD to assist the nitrification- denitrification process for nitrogen removal
(mg/L), respectively. [23]. Also, it is hypothesized that due to the width of the tank (47.2 m),
The non-biodegradable soluble TKN (nbsTKN) typically passes inside the anoxic zone, there are pockets of anaerobic regions which act
through the treatment system untouched. Assuming that the soluble as the anaerobic selector for PAOs. A few literatures have explored the
biodegradable organic nitrogen gets degraded completely within the growth of PAOs under anaerobic conditions inside anoxic tanks [24,25].
treatment process, nbsTKN (mg N/L) was calculated as [22]: The aim was to achieve the modifications without taking up any civil
works in the STP. This was implemented in the validated BioWin model,
nbsTKN = fTKN − NH4 (14) by dividing the aeration tank into three compartments- aeration zone,
anoxic zone and re-aeration zone as shown in Fig. 1 (b).
where ‘fTKN’ was the effluent filtered TKN (mg N/L), and ‘NH4’ was the The total phosphorous (TP) is the sum of total particulate phospho­
effluent ammonia (mg N/L). The PCOD/VSS ratio was calculated by rous (TPP) and total dissolved phosphorus (TDP as PO4). A major per­
dividing particulate COD (PCOD – mg/L), which was the residual COD centage (65–70%) of phosphorus that gets accumulated in natural water
after filtration (through a 0.45 μm filter) by the volatile suspended solids bodies through effluent discharge and cause eutrophication is the TDP
(VSS - mg/L) fraction [22]. (as PO4) [26]. Generally, TPP is removed through sedimentation process
For the present study, the BioWin© model was first run with default (primary and secondary) as sludge, while TDP (as PO4) removal requires
kinetic and stoichiometric values. Then the most sensitive wastewater either chemical or biological phosphorous removal methods. Hence, for

4
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

the present study, the TDP (as PO4) removal efficiency of the 55 MLD
plant was considered and hence reported.
To perform sensitivity analysis, several scenarios were simulated
using the modified plant model as shown in Table 2. The first scenario
was to vary the DO concentration in the aeration and re-aeration zones.
The DO concentrations were varied between 3 and 4.5 mg/L insteps of
0.5 mg/L. The change in the wastewater characteristics as predicted by
the model was studied. The second scenario included varying the mixed
liquor suspended solids (MLSS) concentration between 3000 and 4500
mg/L in the aeration zone and the re-aeration zone. The third scenario
was to optimize the hydraulic retention time (HRT) of wastewater in the
three zones. Finally, in the fourth scenario, the return activated sludge
(R) ratio defined as the ratio of activated sludge recycle flow rate to the
flow rate of inlet sewage to the aerator was varied from 0.03 to 0.69.
3.2. Implementation of the model suggested modifications on the 55 MLD
STP
The optimum process modifications as suggested by the validated
BioWin model were implemented on the 55 MLD K&C valley CAS pro­
cess. The aeration tank operating conditions were modified to achieve
aeration zone, anoxic zone and re-aeration zone by controlling the
operation of surface aerators. The central anoxic zone was created by
switching off the surface aerators. The DO levels in the aeration and re-
aeration zone were controlled by controlling the speeds of the surface
aerators. The required HRT in each zone was achieved by properly
selecting the surface aerator’s locations and the MLSS concentrations
were controlled by adjusting the recycle flow rate. Post modifications,
the wastewater characteristics were monitored for a period of 6 months.
The flow diagram for the development, validation and implementation
of BioWin© model before and after the process modification is shown in
Fig. S3. A paired ‘t’ test was conducted to determine whether there is
statistical evidence that the mean difference between pre and post-
modification values is significantly different from zero. The details of
the paired ‘t’ test is given in Supplementary Section S4.
• S4 at the final outlet (treated water) after secondary clarification as
shown in Figs. 1 & 2 and
• S31 at the outlet of aeration zone
• S32 at the outlet of the anoxic zone
• S33 at the outlet of the re-aeration zone in Fig. 1
All wastewater samples were collected between 9 AM and 12 Noon,
and samplings were carried out for one week, both pre-modification and
post-modification. The weather was favourable during the sampling
procedure and the plant operated at dry weather conditions at an
average air temperature of 26 ± 3 ◦ C. The DO concentrations at each
point of treatment were measured on the field using an optical DO probe
(Lovibond-740800, luminophore-covered membrane), and a few sam­
ples were analyzed in the laboratory using the modified Winkler DO
titration procedure [27] to cross-check the probe readings. The pH
readings at each point of treatment were also determined using a probe
(Hach 5475020). The BOD of the samples was determined using the
standard 5 day BOD test [27], and the COD was determined using the
closed reflux, titrimetric method with 0.1 M ferrous ammonium sulphate
(FAS) as the titrant [27]. For the nitrogen components, the total Kjeldahl
nitrogen (TKN) in the samples was determined in the lab using the
macro Kjeldahl method [27], which involves the digestion and distil­
lation of samples to NH4-N. The NH4-N in the samples were then pre
distilled and determined using the standard sulfuric acid titration
method [27]. The NO3-N in the samples were analyzed using UV spec­
troscopy as the difference in sample absorbances at 220 and 275 nm
[27]. The NO2-N was also analyzed using UV spectroscopy at 543 nm
absorbance. The total phosphorous (TP) in the treated water was
analyzed using the Vanadomolybdo - Phosphoric Acid Method [27]. The
phosphorous in the treated water was first converted into orthophos­
phates using the sulfuric acid- nitric acid digestion method then deter­
mined colorimetrically using a spectrophotometer at 880 nm
absorbance [27]. The TS, TSS, TDS, and VSS in the wastewater samples
were determined using gravimetric analysis following oven drying [27].
The detailed procedure for determining all these parameters is given in
the APHA manual of standard methods for the examination of water and
wastewater [27].

3.3. Wastewater characterization

Wastewater sample from different locations of the 55 MLD K&C
valley STP were collected and analyzed. The samples collected were:
• S1 at the inlet of the plant (raw sewage),
• S2 at the primary clarifier outlet,
• S3 at aeration tank outlet,
Table 2
Summary of sensitivity analysis conditions.
Scenario 1:
Varying DO concentration in aeration zones.
MLSS constant at 4000 mg/L, R ratio of 0.05 and
HRT of 2.5 h aeration, 1.5 h anoxic and 2 h re-
aeration
Scenario 2:
Varying MLSS concentration in aeration zones
DO constant at 4 mg/L, R ratio of 0.05 and HRT of
2.5 h aeration, 1.5 h anoxic and 2 h re-aeration
Scenario 3:
Varying HRT in aeration and anoxic zones
DO constant at 4 mg/L, MLSS of 4000 mg/L and R
ratio of 0.05
Scenario 4:
Varying R ratio in aeration zones
DO constant at 4 mg/L, and HRT of 2.5 h aeration,
1.5 h anoxic and 2 h re-aeration
Case 1: DO - 3 mg/L
Case 2: DO - 3.5 mg/L
Case 3: DO - 4 mg/L
Case 4: DO - 4.5 mg/L
Case 1: MLSS - 3000 mg/L
Case 2: MLSS -3500 mg/L
Case 3: MLSS - 4000 mg/L
Case 4: MLSS - 4500 mg/L
Case 1: 3 h aeration, 2 h
anoxic, 1 h reaeration
Case 2: 2 h aeration, 2 h
anoxic, 2 h reaeration
Case 3: 2 h aeration, 1 h
anoxic and 3 h reaeration
Case 1: R ratio - 0.03
Case 2: R ratio - 0.09
Case 3: R ratio - 0.21
Case 4: R ratio - 0.44
Case 5: R ratio - 0.69
3.4. Microbiological analysis
The type of microbial biomass is a crucial factor in SNDN and EBPR
efficiency in the treatment unit. The bacterial community structure and
diversity of the aeration and the anoxic zones were analyzed to achieve
this. The samples (aerobic and anoxic) for the microbiological analysis
were collected from the plant and stored in a deep freezer at − 40 ◦ C. The
samples were then sent to Eurofins Scientific Co. Ltd. (Bangalore, India)
for DNA extraction, PCR amplification, and cluster generation and
sequencing. The metagenomic DNA was isolated from the samples using
the Nucelospin kit (BIOKE, Netherlands), and the quality of the isolated
metagenomic DNA samples was analyzed using Nanodrop 2000 UV–vis
spectrophotometer (Thermo Scientific, Wilmington, USA). The ampli­
con libraries were prepared using the Nextera XT index kit (Illumia Inc.)
as per the 16S metagenomic sequencing library preparation protocol
(Part #15044223 Rev.B). The primers used in the study were (synthe­
sized at Eurofins Genomics) 16S rRNA F (GCCTACGGGNGGCWGCAG)
and 16S rRNA R (ACTACHVGGGTATCTAATCC). The amplicon library
was purified by AMPure XP beads and quantified using a Qubit fluo­
rometer. The data analysis was conducted using the QIIME software
platform provided by Eurofins Scientific Co. Ltd. (Bangalore, India).
For understanding bacterial diversity, the rarefaction curve was
analyzed. The rarefaction curves allow the calculation of species rich­
ness for a given number of samples. The rarefaction curve for the present
samples is shown in Fig. 2. As seen from Fig. 2, the steep slope, in the
beginning, indicates the presence of large diversity in bacterial species
in the given samples.

5
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Table 3
K&C Valley 55 MLD unit wastewater characteristics.
Sl. Parameters Units K&C Valley 55 MLD unit raw K&C Valley 55 MLD unit primary K&C Valley 55 MLD unit Treated water
no sewage characteristics settled sewage characteristics treated water characteristics discharge standards
[28]

1 pH Value – 8.5 ± 1.0 7.5 ± 1.0 7.0 ± 1.0 6.5–9.0

2 Total suspended solids mg/L 230 ± 10 80 ± 5 22 ± 5 <10
3 Biochemical oxygen 193 ± 5 69 ± 3 6.4 ± 1 <10
demand (5 day at 25 ◦ C)
4 Chemical oxygen demand 448 ± 10 167 ± 5 39 ± 5 <50
5 Total nitrogen 39.2 ± 3 36.8 ± 5 20 ± 2 <10
6 Ammonical nitrogen 28.2 ± 2 26 ± 2 5.1 ± 3 <5
7 Dissolved P 15.3 ± 2 12.6 ± 1 3.5 ± 2 <2
8 Fecal coliform (MPN/ 10E− 06 ± 1E− 06 – 190 ± 10 <230
100 ml)

simulations. Fig. 3 shows the comparison of experimental and BioWin©

Fig. 2. Rarefaction plots for aeration and anoxic samples.
4. Results and discussion
4.1. Model validation for the baseline scenario
To develop a validated model, the ‘as-is’ (existing) operating con­
ditions of the 55 MLD STP, as shown in Table 1, was used for
simulation values at different stages of treatment for this baseline sce­
nario (before modification). The BioWin model for this baseline scenario
consists of a primary clarifier, aeration zone, and two secondary clari­
fiers (as shown in Fig. S3). The inlet BOD to the model is 193 mg/L, and
after primary clarification, the BOD is reduced to 55 mg/L.The water
after aeration zone passes through a secondary clarifier and the water
quality at the outlet of the secondary clarifier is shown as treated water.
As can be seen from Fig. 3, the simulation values matched very closely
with that of experimentally measured values for all the locations i.e.,
primary clarifier outlet, aeration tank outlet and treated water. In order
to quantify the accuracy of the model, the relative error between the
simulation and experimental values were computed, specifically for the
treated water [30]. The relative error between the simulation value and
experimental value was 4.9%, 5.6, 2.0, 6.0, 5.0, 6.1 and 2.5% for COD,
BOD, DO, TN, NH4-N, NO3-N and TDP (as PO4), respectively. The
maximum relative error observed was 6.1% for NO3-N readings and was
< the acceptable deviation of 10% [31]. This was deemed acceptable
considering the error in analytical measurements and the fluctuating
operating conditions during sampling. Thus, the developed BioWin©
model was considered validated for the baseline scenario and this vali­
dated model was used for all other process simulations.
The BOD and COD at the primary clarifier outlet, aeration tank
outlet, and treated water were measured following the standard
methods for the examination of water and wastewater [27]. The total
BOD is measured using 5 - day BOD test, and the total COD is measured
using the titrimetric method [27]. The standard method used for

Fig. 3. Comparison of experimental and BioWin© simulation values (baseline scenario) at (a) Primary clarifier outlet (b) Aeration tank outlet and (c) Treated water.

6
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

determination of all the parameters is given in Table S1. As shown in Table 4

Fig. 3, though the COD and BOD5 concentrations were within the treated Calibrated kinetic and stoichiometric parameters in BioWin© 5.3.
water discharge limits (shown in Table 2) [28]. The TN and TDP (as PO4) Parameter BioWin© Calculated/ Remarks References
concentration of treated water was 20 mg/L and 3.6 mg/L, respectively. default adjusted
Both TN and TDP (as PO4) concentrations were above the discharge limit value value
of 10 mg/L and 2 mg/L, respectively [28]. Efforts were made to reduce Raw wastewater characteristics
the TN and TDP (as PO4) concentrations in the treated water which are Non-colloidal 0.75 0.64 Varied as per Seiffert et al.,
discussed in the following sections. slowly literature 2010
biodegradable
– Fxsp [gCOD/
4.2. Identification of process modifications through sensitivity analysis g of slowly
degradable
As seen from the experimental observations shown in Table 4, the TN COD]
Non- 0.05 0.052
and TDP (as PO4) values were higher the permissible discharge limits. To
biodegradable
improve these parameters BNR was considered a variable process soluble - Fus
modification. Hence to achieve BNR via SNDN and TDP (as PO4) [gCOD/g of
removal by EBPR, the biological reactor was divided virtually into total COD]
Non- 0.13 0.08
aeration zone, anoxic zone and re-aeration zones and simulations were
biodegradable
run by varying different parameters one at a time. The model layout particulate -
used is as shown in Fig. 1 (b). Fup [gCOD/g
of total COD]
4.2.1. Scenario 1: effect of DO concentration in aeration zones Ammonia Fna 0.66 0.65
[gNH3-N/
The DO concentration in the aeration and re-aeration zone was
gTKN]
varied from the baseline value of 3.0 mg/L up-to 4.5 mg/L in steps of 0.5 Soluble non- 0.02 0.016
mg/L. For all these cases, the MLSS concentration was kept constant at biodegradable
4000 mg/L and the HRT was maintained as 2.5 h in aeration zone, 1.5 h TKN - Fnus
[gN/gTKN]
in anoxic zone followed by 2 h in re-aeration zone.
The simulation values of TN, NH4-N, NO3-N, NO2-N, and Organic N Calibrated kinetic parameters of autotrophic bacteria
concentrations after each treatment stage and at different DO concen­ Max. spec. 0.9 0.82 Reduced - 0.9 Lopez-
growth rate to 0.82 d− 1 Vazquez,
tration is shown in Fig. 4. Also shown in Fig. 4 are the stage-wise per­
[1/d] Literature Mithaiwala,
centage reduction (PR) (calculated between each stage of treatment- reported Moussa and
calculation given in Section S3) of TN, NH4-N and NO3-N values. range is Brdjanovic,
As can be seen from Fig. 4, simulation results shows that when the between 1.0 2013
operating DO levels in the aeration and reaeration zone was increased and 0.45 d− 1
Aerobic decay 0.17 0.24 Increased Elawwad,
from 3.0 mg/L, the TN and TDP (as PO4) of the treated water reduced. rate [1/d] from 0.17 to 2018
This improvement was seen both at DO levels of 3.5 and 4.0 mg/L. 0.24 d− 1 to
However, operating at 4.5 mg/L, this benefit of higher DO levels was not match treated
seen. As shown in Fig. 4, when the DO level was increased from 3.0 to water
ammonia
4.0 mg/L, the PR of TN in the first aeration zone increased from 52% at
values.
DO of 3.0 mg/L to 57% when DO was at 4.0 mg/L. With an increase in Follows study
DO level from 3.0 to 4.0 mg/L, there was an improvement in the TN where value
removal due to the improvement in nitrification reactions by autotrophs was increased
in the aeration zone which is indicated by the reduction in NH4-N to 0.23 d− 1

concentration and increase in NO3-N concentration at the outlet of the Calibrated kinetic parameters of heterotrophic bacteria
aeration zone. A similar trend of drop in NH4-N concentration and in­ Max. spec. 3.2 4.0 Adjusted from Elawwad,
growth rate 3.2 to 4 d− 1 to 2018
crease in NO3-N concentration was observed in the last re-aeration zone
[1/d] match treated
when DO was increased from 3.0 mg/L to 4.0 mg/L. However, when the water COD
DO increased from 4.0 to 4.5 mg/L, the PR of TN instead of further values.
increasing, reversed its trend and reduced to 52% as shown in Fig. 4. Range in
This was attributed to inadequate HRT in the anoxic zone to denitrify the literature is
between 1
higher NO3-N formed which would remain in the treated water.
and 8 d− 1
Fig. 5a, shows the overall percentage reduction PRo (from raw Substrate half 5 8.5 Adjusted from Liwarska-
sewage to treated water – calculation given in Section S2) of TN and TDP sat. [mgCOD/ 5 to 8.5 Bizukojc and
(as PO4) with DO. As shown in Figs. 4 and 5a, the PRo, in TN concen­ L] mgCOD/L. Biernacki,
In agreement 2010
tration increased until DO of 4 mg/L and this was attributed to the
with Hulsbeek, J
improved SNDN by the autotrophs and the heterotrophs. However, as literature and Kruit, Jans
shown in Figs. 3 and 6a, at higher DO of 4.5 mg/L, the PR of TN studies where and Roeleveld,
decreased, and this was attributed to inadequate HRT in the anoxic zone the observed Paul and van
to denitrify the higher NO3-N formed which remains in the treated ranges is in Loosdrecht,
between 5 2002
water.
and 22.5
As shown in Fig. 5a, the TDP (as PO4) also followed a similar trend as mgCOD/L
that of TN with DO, i.e., with an increase in DO, the PRo of TDP (as PO4)
Calibrated stoichiometric parameters
increased until DO of 4 mg/L and operating at higher DO levels >4.0 Particulate inert 1.6 1.54 Decreased Elawwad
mg/L, the PRo of TDP (as PO4) decreased [4]. The synthesis of TDP (as COD:VSS ratio from 1.6 to et al., 2019
PO4) by PAOs can occur under aerobic and anaerobic conditions. The [mgCOD/ 1.54 mgCOD/
simulation results showed that the orthophosphate (PO4-P) was mgVSS] mgVSS
consumed in the aerobic tank while PO4-P was released in the anoxic/ (continued on next page)

7
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Table 4 (continued ) nitrification activities by the ammonia-oxidizing bacteria reduce after a

Parameter BioWin© Calculated/ Remarks References pH of 9.6 [42]. While the denitrification rates are depressed at pH below
default adjusted 6.0 and above 8.0, and the highest rate of denitrification occurs at a
value value range of 7.0 to 7.5 [42]. The optimum temperature for both nitrification
Agrees with and denitrification are observed between 5 and 35 ◦ C, with a maximum
literature nitrification rate observed at 30 ◦ C [42]. The nitrification rates also
studies where increase with DO concentration. It has been reported that a DO con­
the observed
centration above 4.0 mg/L is required to achieve maximum nitrification
range is in
between 1.6 rates, while other investigators have found that only 0.5 to 3.0 mg/L is
and 1.42 required [43]. Another crucial factor is the type of biomass; the presence
mgCOD/ of ammonia-oxidizing bacteria improves nitrification, and the presence
mgVSS of denitrifiers improves denitrification. A detailed microbiological
Particulate 1.6 1.54
substrate COD:
analysis of the aeration and anoxic samples is given in the Supplemen­
VSS ratio tary Section 4.3.
[mgCOD/ As seen from Fig. 6, for all the cases, there was an improvement in the
mgVSS] TN concentration with each stage of treatment. With an increase in the
MLSS concentration from 3000 mg/L to 4000 mg/L (Case 1 to Case 3),
the TN in the treated water reduced from 7.8 to 6.9 mg/L. The reduction
anaerobic tank. In the anoxic/anaerobic zone, the DO should be main­
in TN was due to the reduction in NH4-N concentration. This result was
tained at a very low concentration (almost zero) for effective EBPR. The
attributed to optimum pH (8.0 to 9.0), temperature (26 ± 3 ◦ C), DO
BioWin model was developed with a zero DO concentration in the
concentration (4 mg/L), and higher microbial count (nitrifiers) available
anoxic/anaerobic zone. The optimum DO maintained in the aeration
at higher MLSS. With DO availability, the higher MLSS would result in
zone was 4 mg/L. The recirculating aeration sludge might have
an increased rate of microbial activity (ammonia-oxidizing bacteria),
increased DO levels. However, the HRT maintained in the aeration (2.5
leading to a higher rate of SNDN [34].
h) and anoxic/anaerobic (1.5 h) zone was observed to be sufficient
When MLSS was increased from 3000 mg/L to 4000 mg/L (Case 1 vs
enough so that the DO of wastewater in the anoxic/anaerobic zone was
Case 2 and 3), in the aeration zone, the observed reduction in TN was
significantly less (less than 0.5 mg/L). However, at higher DO levels
57%. However, this effect was reduced slightly (2% reduction in TN
(aerobic conditions), the efficiency of TDP (as PO4) synthesis by PAOs is
conversion in aeration zone) when MLSS was further increased to 4500
lower since high DO reduces the uptake of PO4-P [4]. This is also known
mg/L. This small drop was attributed to increase in NH4-N and Organic
to potentially leave lower BOD for anaerobic processing. This was
N from 0.35 to 0.45 mg/l and 1.7 to 2.1 mg/L respectively. Even though
considered as the reason for higher TDP (as PO4) retained at DO of 4.5
the particulate Organic N gets assimilated and removed through sludge,
mg/L.
it is believed that the soluble Organic N increases with MLSS and re­
From all these observations, the optimum DO concentration in the
mains in the treated water [33,34], which is justified by the higher
aeration zone and re-aeration zone was identified as 4 mg/L. This value
Organic N concentration at MLSS of 4500 mg/L. Also, it is observed in
was used for further simulation studies.
several studies that around 15–85% of soluble organic N remains in
Operating the plant at higher DO levels means higher air flow rates.
treated water [44].
The air demand increased from 10,051 m3/h for the baseline scenario
The TDP (as PO4) concentration followed a similar pattern (shown in
with 2.5 mg/L DO to 13,500 m3/h for Case 4 with 4.5 mg/L. This in­
Table S4). Between Case 1 to Case 3, the TDP (as PO4) concentration
crease in the aeration demand increases the air blower operating cost by
reduced in treated water. However, simulations under Case 4 conditions,
15% from the baseline scenario. For the optimum DO concentration of 4
with an MLSS of 4500 mg/L, the TDP (as PO4) concentration for treated
mg/L, the aeration demand was 13,050 m3/h and the increase in the air
water was higher as compared to previous cases. One of the re­
blower operating cost from the base line scenario was 8%. The air de­
quirements for PAOs for EBPR in anaerobic conditions is to have a
mand was calculated using the aeration model in BioWin [32].
minimum BOD5 to TDP (as PO4) ratio of 20:1 [35]. The typical operating
The simulation results of all other parameters at different DO con­
conditions affecting EBPR are given in detail in the Supplementary
centrations are shown in Table S3. The model predicted improved water
Section S3. However, this ratio was 18:1 due to lower BOD5 concen­
quality with increase in DO levels as shown in Table S3. The COD and
tration in Case 4 (Table S4) thereby reducing the efficiency of P removal.
BOD concentration reduced from 43 to 35 mg/L and 6.5 to 3.6 mg/L as
This explained the higher TDP (as PO4) concentration observed for Case
DO increased from 3 to 4.5 mg/L, respectively. This was attributed to the
4.
higher biological degradation of soluble substrate (COD) by the het­
The PRo of TN and TDP (as PO4) at different MLSS concentrations is
erotrophs in aeration and reaeration zones at higher DO levels [4].
shown in Fig. 5b. As seen from the Fig. 5b, the maximum PRo for TN and
TDP (as PO4) was observed at MLSS of 4000 mg/L. Hence, the optimum
4.2.2. Scenario 2: effect of MLSS concentration in aeration zone
MLSS concentration which resulted in minimum TN and TDP (as PO4)
In this scenario, different MLSS concentrations of 3000 mg/L, 3500,
concentrations in treated water was identified as 4000 mg/L and was
4000 and 4500 mg/L were simulated in the aeration zone. For these
used for further studies. The simulation results of all other parameters
simulations, the DO concentrations in the aeration and re-aeration zone
are given in Table S4. The model predicted improved water quality with
were held constant at 4.0 mg/L. The HRT of 2.5 h in aeration zone, 1.5 h
increase in MLSS concentrations as shown in Table S4. The COD and
in anoxic zone and 2 h in reaeration zone was simulated for all the cases.
BOD concentration reduced from 43 to 35 mg/L and 4.5 to 3.5 mg/L as
For all the cases simulated, the values of TN, NH4-N, NO3-N, NO2-N and
MLSS increased from 3000 to 4500 mg/L. This improvement in water
Organic N concentrations after each treatment stage and the stage-wise
quality was attributed to increased microbial activity at higher MLSS
PR of TN, NH4-N and NO3-N values are shown in Fig. 6. The total ni­
concentrations.
trogen concentration and its fractions are (organic N, NH4-N, NO3-N,
and NO2-N) are given in Tables S4–S6. The nitrogen balance after pro­
4.2.3. Scenario 3: effect of varying the hydraulic retention time (HRT) in
cess modification is given in Supplementary Section S5 and Table S9.
the three zones
Several factors affect SNDN, including pH, temperature, DO con­
With DO and MLSS fixed at 4.0 mg/L (optimum value) and 4000 mg/
centration, and type of biomass. Generally, it is observed that the opti­
L (optimum value) respectively, the HRT of wastewater in the aeration
mum pH for nitrification ranges between 8.0 and 9.0 [42], and the
zone, anoxic zone and reaeration zone were varied. For all the cases

8
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Fig. 4. Simulation results of variation of (1) TN, (2) NH4-N, (3) NO3-N, (4) NO2-N and (5) Organic N at each stage of treatment and at different DO concentration; PR:
Percentage reduction (calculated between each stage of treatment) of TN, NH4-N and NO3-N.

simulated, the values of TN, NH4-N, NO3-N, NO2-N and Organic N
concentrations after each treatment stage and the stage-wise PR of TN,
NH4-N and NO3-N values are shown in Fig. 7. The simulation results for
all other parameters are given in Table S5.
As shown in Fig. 7, for Case 1, the HRT was chosen so that the water
would have equal HRT of 2 h in all three virtual zones i.e., 2 h each in
aeration, anoxic and re-aeration zone. The TN and NH4-N concentration
in the treated water were 7.2 mg/L and 1.2 mg/L, respectively (Fig. 7
and Table S5). And the NO3-N concentration in the treated water was 4
mg/L, which accounted to be 55% of the TN indicating inadequate HRT
for SNDN specifically in the aeration zone. To increase SNDN in aeration
zone, higher HRT Case i.e., Case 2 was simulated. For Case 2, the HRT
was 2.5 in the aeration zone, 1 h in the anoxic zone, and HRT in re-
aeration zone was kept same as that of Case 1 i.e., 2 h. As can be seen
from Fig. 7, when HRT in aeration zone was increased from 2.0 to 2.5 h
(Case 1 vs Case 2), the PR in TN increased from 55% to 57% primarily
due to increased SNDN in aeration zone which is evident from higher
NH4-N reduction from 80 to 83%. However, the reduction in the HRT of
0.5 h in the anoxic zone only showed a marginal drop of 1% in PR of TN
between Case 1 and Case 2. This result showed that even 1.5 HRT in
anoxic zone was still adequate to achieve denitrification in anoxic zone.
However, the NH4-N at the anoxic zone outlet was still high and to
address this, the Case 3 which had higher HRT in the re-aeration zone
was simulated.
Case 3 with HRT of 2 h of aeration in followed by 1 h of anoxic
conditions and 3 h of reaeration was simulated. As seen from Fig. 7,
when the HRT in the reaeration zone was increased from 2.0 h to 3 h
(Case 2 vs Case 3), the PR of TN and NH4-N, in the re-aeration zone,

9
R. Mohan T et al.
Fig. 5. PRo: Percentage reduction (raw sewage to treated water) of TN and TDP (as PO4) with (a) DO concentration (b) MLSS concentration in aeration and rea­
eration zones.
increased from 23 to 32% and from 90 to 92%, showing increased SNDN
in re-aeration zone.
Also, Case 3 showed lowest TN value of 6.7 mg/L in treated water
relative to all other cases as shown in Table S5 and this was attributed to
the adequate reaction times for both nitrification and denitrification in
the three zones. The intermittent anaerobic and aerobic condition with
optimum HRT is known to improve the nitrification- denitrification ef­
ficiency [23].
The TDP (as PO4) was minimum in Case 3 (Table S5). It is well un­
derstood that, treating wastewater under anaerobic conditions, followed
by prolonged aerobic conditions acts a selector and favours the growth
of PAOs A selector is a zone in a biological treatment process with
specific environmental conditions that promote the growth of a specific
group of microorganisms [35–37]. It is hypothesized that the virtual
anoxic zone acted as a selector for the PAOs and gave them a selective
advantage over other microorganisms in the system [35–37]. This
explained the minimum TDP (as PO4) in Case 3. Also, the COD and BOD
concentrations reduced from 42 to 34 mg/L and 5 to 2.9 mg/L respec­
tively, from Case 1 to Case 3. Hence, from all these observations, Case 3
was identified as the optimum case in scenario 3.
4.2.4. Scenario 4: effect of return activated sludge (R) ratio
To further optimize the process, the R ratio was varied between 0.03
and 0.69. The optimum DO concentration of 4 mg/L was considered for
simulation in the aeration and re-aeration zones. The optimum HRT of 2
h in aeration zone, 1 h in anoxic zone and 3 h in re-aeration zone was
simulated.
The variation of MLSS in the aeration and re-aeration zone, the
aeration requirement and the treated water COD with R ratio is shown in
Fig. 8. As seen from Fig. 8, with increase in R ratio from 0.03 to 0.69, the
MLSS in the aeration and re-aeration zone increased from 1052 to 6451
mg/L, respectively. Similarly, as shown in Fig. 8, the air requirement to
maintain a fixed DO concentration of 4 mg/L in aeration and re-aeration
zones also increased with R ratio. This increase in air requirement was
attributed to higher microbial count at higher R ratio. Since at higher R
ratio, the microorganism count will be higher, the amount of air to be
provided to maintain a fixed DO concentration will also be higher.
Analysing the treated water COD variation with R ratio, as shown in
Fig. 8 and Table S6 (Case 1 to Case 5), it was observed that, COD levels
decreased until R ratio of 0.21 and then it increased with R ratio. This
increase was attributed to the decrease in food to microorganism ratio
(F/M) with increasing R, which leads to starvation and decay of the
microorganisms, increasing the dissolved organic matter and thereby
the total COD. This result was in agreement with the observations re­
ported by [38].
The increase in the energy cost with increase in air requirement and
R ratio is shown in Fig. S4.
The variation of TN with R ratio for treated water is shown in Fig. 9.
10
Journal of Water Process Engineering 45 (2022) 102461
The TN concentration reduced until R ratio of 0.21 and then increased
steadily with R. The NH4-N also showed a similar trend. This showed
that at R ratios above 0.21, the HRT provided was inadequate to drive
the nitrification reaction leading to increased NH4-N in the system. The
NO3-N and NO2-N concentration in the treated water reduced rapidly
with increase in the R ratio till 0.21, indicating a higher denitrification
process. However above 0.21, this reduction tends to saturate. Several
researchers have observed similar results indicating an increase in
denitrification with increase in R ratio for fixed DO concentration pro­
vided that the availability of organic matter (COD) is not limited
[38,39]. With an increase in the R ratio, the concentration of microbial
mass recycled into the process increases. To identify the specific type of
microbial mass, molecular biological techniques are necessary. Johnston
et al. [40] has monitored microbial community in three full-scale STPs
and observed the abundance of ammonia-oxidizing bacteria without any
seasonal fluctuations. Wiginton et al. [41] have also identified the
abundance of amoA (ammonia-oxidizing – 45%) and nosZ (denitrifiers –
25%) in an activated sludge system using 16sRNA sequencing. Some
potential abundant denitrifiers have also been identified by using
culture-independent methods in wastewater by Thomson et al. [42].
They observed that denitrifiers constitute 20–30% of the total bacterial
composition in an activated sludge system. But irrespective of the type of
microbial mass, the Organic N, composed of protein, urea, amino acids,
and humic substances, increases with biomass concentration. This
explained the higher organic N concentration in the treated water with
increasing R ratio.
Organic carbon content is also an essential factor in accomplishing
effective denitrification since it is an energy source for bacteria [43]. A
C/N ratio of 3.5–4.5 g COD/g N is considered as optimum for effective
SNDN [44]. The C/N ratio of each zone for all the scenarios is given in
the modified supplementary material, Table S7. As seen from Table S7,
the C/N ratio for all the cases in the scenario was observed to be within
the acceptable range, indicating effective denitrification efficiency.
The TDP (as PO4) concentration decreased with R ratio until 0.21
and then increased (Table S6). It is observed in several studies that P
removal efficiency in the anoxic zones increased with R ratio (specif­
ically due to nitrate recycling) [45,46]. S But one of the limiting factors
for EBPR is the availability of organic content or the BOD: P ratio (COD:
P ratio) of minimum 20:1. But for R ratio beyond 0.21, (for Cases 4 and
5) the BOD: P ratio was less than 20:1, thereby reducing the efficiency of
P removal. Hence, it was inferred that the optimum R ratio of 0.21
produces minimum TN, and TDP (as PO4) concentrations for the fixed
DO of 4 mg/L.
4.3. Microbiological analysis results
The wastewater samples from the aeration and anoxic zones were
analyzed to identify the microbial community. The composition of the
R. Mohan T et al.
Fig. 6. Simulation results of variation of (1) TN, (2) NH4-N, (3) NO3-N, (4) NO2-N and (5) Organic N at each stage of treatment and at different MLSS concentration;
PR: Percentage reduction (calculated between each stage of treatment) of TN, NH4-N and NO3-N.
bacterial community and the similarity and differences between the two
samples were investigated. A total of 12 phyla were observed in the two
samples. Proteobacteria was the most abundant community (30–40%),
which is consistent with the samples from activated sludge systems
[47,48]. Bacteroidetes (12–15%), Planctomycetes (10–12%), Chlorofexi
(9–12%), Actinobacteria (7–8.2%) and Nitrospirae (4–5%) were the other
important groups. The dominant class of bacteria and its percentage
composition in the aeration and anoxic tanks are given in Fig. 10. As
seen from Fig. 10, in the aeration tank samples, the ammonia-oxidizing
bacterial classes observed were, Nitrosovibrio (6.99%), Nitrosomonas
(5.52%), Nitrosolobus (5.03%) and, Nitrosospira (4.87%). These ammo­
nium oxidizing bacteria belong to the beta subdivision of the Proteobac­
teria [49]. Nitrobacter (9.19%), Nitrospira (4.87%), and Pseudomonas
(3.5%) were the nitrite-oxidizing bacterial classes found abundant in the
aeration samples. The nitrite-oxidizing bacteria usually belong to the
alpha and gamma subdivisions of the Proteobacteria [50]. The
11
Journal of Water Process Engineering 45 (2022) 102461
denitrifying bacteria (shown in Fig. S6) observed in the anoxic samples
were Achromobacter (6.07%), Proteus (5.5%), Alcaligenes (5.25%),
Pseudomonas (4.32%), and, Spirillum (3.91%), all belonging to the beta
and gamma subdivisions of Proteobacteria. Other denitrifying bacterial
species were Micrococcus (3.5%) belonging to the Actinobacteria phylum
and Bacillus (3.27%) belonging to the Firmicutes phylum [22].
As seen from the microbiological analysis results, there was a clear
difference in the bacterial community in the aeration and anoxic sam­
ples. The aeration samples were abundant in ammonia and nitrite-
oxidizing bacteria, whereas the anoxic samples were rich in denitrify­
ing bacterial classes. The classes observed in both the samples were
Pseudomonas (Proteobacteria) and Nitrospira (Nitrospirae). These are the
bacterial classes that survive in both aerobic and anoxic conditions [48].
Also, as seen from Fig. 10, PAOs, including Pseudomonas (4.32%) and
Amaricoccus (4.1%) belonging to the Proteobacteria phylum, Brevi­
bacterium (3.91%) and Tetrasphaera (3.5%) belonging to the
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Fig. 7. Simulation results of variation of (1) TN, (2) NH4-N, (3) NO3-N, (4) NO2-N and (5) Organic N at each stage of treatment and at different HRT; Percentage
reduction (calculated between each stage of treatment) of TN, NH4-N and NO3-N.

Actinobacteria phylum and Anaerolineae (2.51%) belonging to the While to implement DO of 4 mg/L in the aeration and reaeration
Chloroflexi phylum were identified in the anoxic samples. The presence zones, the air flow rate was maintained at 13,050 m3/h. Diffuse air
of these anaerobic PAOs confirms the hypothesis that the proposed blowers were repositioned into these zones to maintain this airflow rate.
modification has been successful in EBPR. The optimized HRT of 2 h of aeration, followed by 1-h of anoxic con­
dition and 3 h of reaeration was also implemented. The optimum R ratio
was identified as 0.21 with an MLSS of 5000 mg/L from the simulation
4.4. Plant performance after process modifications
results. However, the original plant R ratio of 0.69 and the corre­
sponding MLSS of 6451 mg/L were not modified due to site conditions.
Based on the sensitivity analysis results, only the optimized process
This optimum R ratio is planned to be implemented in the second phase
modifications were implemented in the 55 MLD STP at the K&C Valley
of process modification in the 55 MLD unit. It is assumed that imple­
STP and compared with the experimental results. To implement the
menting the optimum R ratio will bring down the recycling pump cost by
process modification of converting the existing aeration tank into three
15%. The plant was run for 6 months continuously based on the opti­
virtual zones, namely aeration, anoxic, and reaeration, changes in the
mized parameters and the process conditions were stabilized over this
plant’s operating conditions were necessary. Some selective surface
period. The sludge-based parameters including mixed liquor suspended
aerators were operated in a Switch ON/Switch OFF mode to create a
solids (MLSS), mixed liquor volatile suspended solids (MLVSS) the TP
virtual anoxic zone such that anoxic conditions and DO levels less than
concentration of sludge and the sludge volume index (SVI) after the
0.5 mg/L was achieved in these sections of the bioreactor. The return
process modification is given in Table S9.
activated sludge saturated with DO could negatively affect the denitri­
Fig. 11 shows the comparison of BioWin© simulation and experi­
fication process in the virtual anoxic zone [1]. To prevent this, the DO
mental values at different stages of treatment after the process modifi­
levels in the anoxic zone were frequently monitored using DO sensors,
cation by implementing the optimized operating conditions (Case 5
and it was observed that the DO concentration was less than 0.5 mg/L at
Scenario 4). As can be seen from Fig. 11, the simulation results matched
all times.

12
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Fig. 8. Variation of MLSS in aeration zone, treated water COD and aeration requirement with R ratio.

Fig. 9. Variation of TN, NH4-N, NO3-N, NO2-N and Organic N with R ratio in treated water.

very closely with that of the experimental values for all the water sample
locations i.e., aeration zone outlet, anoxic zone outlet, re-aeration zone
outlet and treated water. The higher BOD at the aeration zone outlet is
due to the high return activated sludge flow of 38 MLD (R ratio 0.69)
back into the aeration zone. The relative error between the simulation
and experimental values were computed for the treated water values to
quantify the accuracy of the model [31]. The relative error for treated
water values was 6.1, 4.9. 4.1, 2.5, 3.2, 6.7 and 5.5% for COD, BOD, DO,
TN, NH4-N, NO3-N and TDP (as PO4), respectively and was lower than
the acceptable upper limit of 10% [31]. The treated water COD con­
centration was 29 mg/L which was within the treated water discharge
limit of 50 mg/L. Similarly, BOD5 of the treated water was 5.5 mg/L
which was also within the upper limit of 10 mg/L for the treated water.
The TN value of treated water was 8.8 mg/L which was within the upper
limit of 10 mg/L. This validated that the introduction of nitrification –
denitrification zones (aeration- anoxic-reaeration) with optimized
operating conditions was successful to meet the recently revised
discharge standards.
The TP concentration in the treated water after the modification was
observed to be 4.25 ± 0.5 mg P/L which was within the effluent
discharge limit of 4.5 mg P/L as laid down by the CPHEOO [51]. The
TDP (as PO4) value of the treated water as per simulation was 2.9 mg/L.
However, TDP (as PO4) of the treated water measured in the STP was <1
mg/L. This reduction in the TDP (as PO4) may be because of several
reasons. The modestly improved phosphorus reduction observed in STP
was a result of improvement in the BNR, specifically due to the presence
of low nitrate concentrations in anaerobic zones [1] and can be the first
cause. Secondly, the anaerobic mass fraction of luxury PAOs was high

13
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

Fig. 10. Microbial class composition (in percentage) in (a) aerobic and (b) anoxic samples.

Fig. 11. Comparison of experimental and BioWin© simulation values (after modification of plant) of (a) Aeration zone outlet (b) Anoxic zone outlet (c) Reaeration
zone outlet (d) Treated water.

14
R. Mohan T et al. Journal of Water Process Engineering 45 (2022) 102461

(18% - as given in Section 4.3), which is justifiable for the HRT in anoxic Table 5
zone and R ratio [52] of 0.69. The third reason for reduction in TDP (as Estimated, actual cost and savings (in dollars) for the implementation of pro­
PO4) could be the creation of deep anaerobic conditions achieved by posed modification.
switching off the surface aerators. This is known to reduce the entry of Sl. Particulars Estimated Actual Savings (in
oxygen through the liquid surface, thereby creating a pseudo- fermen­ no. amount (in amount (in dollars)
tation zone that assists the growth of the PAOs. Another major dollars) dollars)

contributing factor for the TDP (as PO4) reduction can be the accumu­ 1 Civil works including $ 20,310 0 $ 20,310
lation of phosphorous in the biomass by luxury PAOs in the form of piping, erection, testing
and commissioning
polyphosphates under endogenous conditions, which is the final phase
2 Supply of mechanical and $ 608,300
of growth of activated sludge culture under starvation (F/M ratio < 1; electrical $ 608,300 0
Mulbarger MC et al., 1971) and needs to be experimentally confirmed. $ 2,038,540
(a) Addition of $ 2,038,540 0
4.5. Economic comparison submersible diffusers
(b) Addition of blowers for $ 147,490 0 $ 147,490
air supply $ 33,960 0 $ 33,960
The process modifications in the 55 MLD unit STP was shown to be (c) Addition of pipes and
successful in improving the BNR removal and treated water quality to valves $ 2,828,290 0 $
meet emerging standards. The modifications were implemented in the (d) Addition of 110 KW 2,828,290
motor feeder for
plant with a minimum process intervention, resource investment and
blowers
without shutting down the plant. The time for the implementation and Total
the stabilization of the proposed modification was 8 months. With the 3 Chemical cost for a year
optimization of process modification using the BioWin© ASDM model, $ 700,000 0 $ 700,000
the cost for implementation of the modification was only US$ 0.4 (a) Alum $ 1,902,490 0 $ 1,902,490
(b) Polyelectrolyte $ 2,602,490 0 $
million as against an initial estimate of US$ 6 million which also Total 2,602,490
involved shutting down of the plant operation, constructing new re­ 4 Operating cost for a year $ 548,910 $ 400,000 $ 148,910
actors and making significant changes to plant operating conditions. A 5 Total cost $ 6,000,000 $ 400,000 $
significant improvement in the treated water quality was achieved in a 5,600,000
short period of time with minimum capital costs. The treated water
quality can be further improved by implementing the optimized R ratio its implementation on existing operational STPs to meet the new
of 0.21 as observed in the simulation results. A lower R ratio of 0.21 can discharge standard is an economical alternative for improving the
also reduce the recycle pump operating cost by 40% and the air pump treatment efficiency of STPs in India and other developing countries.
cost (to maintain DO of 4 mg/L) by 29% which leads to a reduction in
the total energy cost of the treatment plant by 35%. Hence, this modi­
fication of R ratio is proposed to be implemented to further improve the Declaration of competing interest
efficiency of the plant. The details of estimated cost, actual cost and
savings for the implementation of the proposed modification is given in The authors declare that they have no known competing financial
Table 5. interests or personal relationships that could have appeared to influence
For developing countries, it is important to modify existing sewage the work reported in this paper.
treatment plants without major interventions to improve the quality of
treated water, using mathematical models and simulating several sce­ Acknowledgement
narios. Adaptable onsite modifications with minimal capital cost as
proposed, modeled, and validated in this work can serve as a solution for The authors would like to gratefully acknowledge the strong support
improving the treatment efficiency of several STPs in India and in other provided by Bangalore Water Supply and Sewerage Board (BWSSB).
developing countries.
Appendix A. Supplementary data
5. Conclusions
Supplementary data to this article can be found online at https://doi.
In this work, a plant-wide BioWin© ASM model was developed for org/10.1016/j.jwpe.2021.102461.
the 55 MLD treatment unit in the K&C Valley sewage treatment plant to
make the plant compliant with the revised discharge standards. The ‘as- References
is’ conditions of the STP were simulated, and the model kinetic and
stoichiometric parameters were calibrated. The existing bioreactor was [1] U.S. EPA, Case studies on implementing low-cost modifications to improve nutrient
reduction at wastewater treatment plants. EPA-841-R-15-004, in: United States
divided virtually into aeration, anoxic and re-aeration zones. Modeling Environ. Prot. Agency, Off. Wetl. Ocean. Watersheds, Off. Sci. Technol. Off.
efforts identified the optimum DO level to be 4 mg/L and MLSS con­ Wastewater Manag., vol. Draft Vers, no. August, 2015, p. 22.
centration to be 4000 mg/L. Modeling also suggested maintaining a [2] R.T. Mohan, M.S.M. Kumar, L. Rao, Numerical modelling of oxygen mass transfer
in diffused aeration systems: a CFD-PBM approach, J. Water Process Eng. 40
hydraulic residence time in the virtually created aeration, anoxic and re- (January) (2021) 101920.
aeration zones as 2 h, 1 h, and 3 h, respectively. The water quality results [3] P.S.G. Subramanian, et al., Decentralized treatment and recycling of greywater
after process modification showed drastic improvements. The total ni­ from a school in rural India, J. Water Process Eng. 38 (August) (2020), 101695.
[4] M. Henze, W. Gujer, T. Mino, M. van Loosedrecht, Activated Sludge Models ASM1,
trogen and total dissolved phosphorous concentrations were reduced ASM2, ASM2d and ASM3 5, 2000 no. 0.
from 20 mg/L to 8 mg/L and 3.5 mg/L to 0.9 mg/L before and after the [5] R. Vitanza, I. Colussi, A. Cortesi, V. Gallo, Implementing a respirometry-based
process modification. The COD and BOD5 concentrations were also model into BioWin software to simulate wastewater treatment plant operations,
J. Water Process Eng. 9 (2016) 267–275.
reduced to 33 and 5.5 mg/L, respectively. The treated water from STP
[6] A. Elawwad, M. Matta, M. Abo-Zaid, H. Abdel-Halim, Plant-wide modeling and
thus met the revised standards. These efforts resulted in a capital cost optimization of a large-scale WWTP using BioWin’s ASDM model, J. Water Process
saving of US $ 5.6 million and avoided other operational disturbances of Eng. 31 (December 2018) (2019) 100819.
plant shutdown. This study shows that using mathematical modeling [7] C. EnviroSim Associates Ltd, BioWin 5.2, 2018.
[8] I. Jones, Richard, Takács, Modeling the impact of anaerobic digestion on the
tools such as BioWin© is highly beneficial to achieve process modifi­ overall perfomance of biological nutrient removal in wastewater treatment plants,
cations in old operating STPs. Process modifications using BioWin© and Proc. Water Environ. Fed. (2004) 244–257.

15
R. Mohan T et al.
[9] D. Brdjanovic, M. Mithaiwala, M.S. Moussa, G. Amy, M.C.M. van Loosdrecht, Use
of modelling for optimization and upgrade of a tropical wastewater treatment plant
in a developing country, Water Sci. Technol. 56 (7) (2007) 21–31.
[10] J. Barat, Ramon, J. Serralta, Vicky Ruano, Emérita Jiménez, Josep Ribes,
Aurora Seco, Ferrer, Biological Nutrient Removal Model No. 2 (BNRM2): a general
model for wastewater treatment plants, Water Sci. Technol. 67 (2013).
[11] C. Duran, D. Ozdes, A. Gundogdu, M. Imamoglu, H.B. Senturk, Tea-industry waste
activated carbon, as a novel adsorbent, for separation, preconcentration and
speciation of chromium, Anal. Chim. Acta 688 (1) (2011) 75–83.
[12] M. Moussa, Moustaf, A. Rojas, C. Hooijmans, Huub Gijzen, van Loosdrecht, Model-
based evaluation of nitrogen removal in a tannery wastewater treatment plant,
Water Sci. Technol. 50 (2004) 251–260.
[13] M. Kim, G. Nakhla, M. Keleman, Modeling the impact of food wastes on wastewater
treatment plants, J. Environ. Manag. 237 (2019) 344–358.
[14] A. Elawwad, Optimized biological nitrogen removal of high-strength ammonium
wastewater by activated sludge modeling, J. Water Reuse Desalin. 8 (3) (2018)
393–403.
[15] E. Liwarska-Bizukojc, R. Biernacki, Identification of the most sensitive parameters
in the activated sludge model implemented in BioWin software, Bioresour.
Technol. 101 (19) (2010) 7278–7285.
[16] A. Elawwad, M. Matta, M. Abo-Zaid, H. Abdel-Halim, Plant-wide modeling and
optimization of a large-scale WWTP using BioWin’s ASDM model, J. Water Process
Eng. 31 (March) (2019), 100819.
[17] Y. Liu, Overview of some theoretical approaches for derivation of the Monod
equation, Appl. Microbiol. Biotechnol. 73 (6) (2007) 1241–1250.
[18] Envirosim, in: BioWin Help Manual, 2017, p. 828.
[19] S. Seiffert, C. Köhler, M. Plattes, E. Henry, P. Schosseler, Comparison of three
calibration protocols for activated sludge models based on case studies, in:
Integrating Water Systems - Proceedings of the 10th International on Computing
and Control for the Water Industry, CCWI 2009, 2010, 2014, pp. 485–491. March.
[20] IWA WERF Report 99-WWF-3, Methods for Wastewater Characterization in
Activated Sludge Modelling, 2004.
[21] M. Henze, W. Gujer, T. Mino, M. van Loosedrecht, Activated sludge models ASM1,
ASM2, ASM2d and ASM3, Water Intell. Online 5 (2015), 9781780402369.
[22] Metcalf, Eddy, Wastewater Engineering Treatment & Reuse, 2004.
[23] O. Hanhan, N. Artan, D. Orhon, Retrofitting activated sludge systems to
intermittent aeration for nitrogen removal, Water Sci. Technol. 46 (8) (2002)
75–82.
[24] Z. Hu, M.C. Wentzel, G.A. Ekama, Anoxic growth of phosphate-accumulating
organisms (PAOs) in biological nutrient removal activated sludge systems, Water
Res. 36 (19) (2002) 4927–4937.
[25] G. Xin, H.L. Gough, H.D. Stensel, Effect of anoxie selector configuration on sludge
volume index control and bacterial population fingerprinting, Water Environ. Res.
80 (12) (Nov. 2008) 2228–2240.
[26] M. Huber, K. Athanasiadis, B. Helmreich, Phosphorus removal potential at sewage
treatment plants in Bavaria – a case study, Environ. Challenges 1 (November)
(2020), 100008.
[27] APHA, Standard Methods for the Examination of Water and Wastewater, 2017.
[28] CPHEEO, Chapter 7: Recycling and reuse of sewage, in: Man. Sewerage Sew. Treat.
Syst, 2013, pp. 7.1–7.53.
[29] C.M. Lopez-Vazquez, M. Mithaiwala, M.S. Moussa, M.C. van Loosdrecht,
D. Brdjanovic, Optimisation, Coupling ASM3 and ADM1 for wastewater treatment
process and biogas production in a developing country: case-study Surat, India,
J. Water Sanit. Hyg. Dev. 3 (1) (2013) 12–25.
[30] H. Park, L.A. Stefanski, Relative-error prediction, 1998.
[31] J. Cao, O.J. Oleyiblo, Z. Xue, Y.M. Otache, Q. Feng, Achieving low effluent NO3-N
and TN concentrations in low influent chemical oxygen demand (COD) to total
16
Journal of Water Process Engineering 45 (2022) 102461
Kjeldahl nitrogen (TKN) ratio without using external carbon source, Chin. J.
Oceanol. Limnol. 33 (4) (2015) 1039–1052.
[32] R. Bashar, K.G. Karthikeyan, D.R. Noguera, Simulation-based analysis of full-scale
implementation of energy neutral wastewater treatment plants, J. Water Process
Eng. 40 (January) (2021), 101875.
[33] G.F. Parkin, P.L. McCarty, Production of soluble organic nitrogen during activated
sludge treatment, J. Water Pollut. Control Fed. 53 (1) (1981) 99–112. Nov.
[34] R. Joshi, M. Kasi, T. Wadhawan, E. Khan, Investigating organic nitrogen
production in activated sludge process: size fraction and biodegradability, Sci.
Total Environ. 773 (2021), 145695.
[35] K. Curtin, S. Duerre, B. Fitzpatrick, P. Meyer, N. Ellefson, Biological Nutrient
Removal Editing and Graphic Design, 2011 no. August.
[36] H. Zhang, A. Sayyar, Y. Wang, T. Wang, Generality of the CFD-PBM coupled model
for bubble column simulation, Chem. Eng. Sci. 219 (2020), 115514.
[37] Wisconsin Department of Natural Resources, Nutrient Removal -Total Phosphorus,
2015 no. August.
[38] T.W. Tan, H.Y. Ng, Influence of mixed liquor recycle ratio and dissolved oxygen on
performance of pre-denitrification submerged membrane bioreactors, Water Res.
42 (4–5) (2008) 1122–1132.
[39] Z. Brown, Jeanette, Carl Koch, Erdal, Biological Nutrient Removal (BNR) Operation
in Wastewater Treatment Plants, 2005.
[40] J. Johnston, T. LaPara, S. Behrens, Composition and dynamics of the activated
sludge microbiome during seasonal nitrification failure, Sci. Rep. 9 (1) (2019)
1–15.
[41] S.K. Wigginton, et al., Nitrifying and denitrifying microbial communities in
centralized and decentralized biological nitrogen removing wastewater treatment
systems, Water (Switzerland) 12 (6) (2020) no. August.
[42] T.R. Thomsen, Y. Kong, P.H. Nielsen, Ecophysiology of abundant denitrifying
bacteria in activated sludge, FEMS Microbiol. Ecol. 60 (3) (2007) 370–382. Jun.
[43] K. Pochana, J. Keller, Study of factors affecting simultaneous nitrification and
denitrification (SND), Water Sci. Technol. 39 (6) (1999) 61–68.
[44] S.H. Isaacs, M. Henze, Controlled carbon source addition to an alternating
nitrification-denitrification wastewater treatment process including biological P
removal, Water Res. 29 (1) (1995) 77–89.
[45] Y. Chen, C. Peng, J. Wang, L. Ye, L. Zhang, Y. Peng, Effect of nitrate recycling ratio
on simultaneous biological nutrient removal in a novel anaerobic/anoxic/oxic
(A2/O)-biological aerated filter (BAF) system, Bioresour. Technol. 102 (10) (2011)
5722–5727.
[46] G.A. Ekama, Recent developments in biological nutrient removal, Water SA 41 (4)
(2015) 515–524.
[47] D. Zhao, et al., Pyrosequencing analysis of bacterial community and assembly in
activated sludge samples from different geographic regions in China, Appl.
Microbiol. Biotechnol. 98 (21) (Nov. 2014) 9119–9128.
[48] T. Zhang, M.F. Shao, L. Ye, 454 Pyrosequencing reveals bacterial diversity of
activated sludge from 14 sewage treatment plants, ISME J. 6 (6) (2012)
1137–1147.
[49] Y.-H. Ahn, Sustainable nitrogen elimination biotechnologies: a review, Process
Biochem. 41 (8) (2006) 1709–1721.
[50] A. Teske, E. Alm, J.M. Regan, S. Toze, B.E. Rittmann, D.A. Stahl, Evolutionary
relationships among ammonia- and nitrite-oxidizing bacteria, J. Bacteriol. 176 (21)
(Nov. 1994) 6623–6630.
[51] CPHEOO, Chapter 5: design and construction of sewage treatment facilities, in:
Manual on Sewerage and Sewage Treatment Systems, 2018, p. 219.
[52] P. Dold, D. Conidi, Achieving enhanced biological P removal: Have we forgotten
how to design a bioP plant?, in: WEFTEC 2019 - 92nd Annu. Water Environ. Fed.
Tech. Exhib. Conf, 2019, pp. 1452–1466.