Journal of Environmental Management 235 (2019) 178–185

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Journal of Environmental Management

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

Review

Recent advances in nitrogen removal from landfill leachate using biological T

treatments – A review
Lei Miaoa, Gangqing Yanga, Tao Taoa, Yongzhen Pengb,∗
a
School of Environmental Science and Engineering, Huazhong University of Science and Technology, Wuhan, PR China
b
National Engineering Laboratory for Advanced Municipal Wastewater Treatment and Reuse Technology, Engineering Research Center of Beijing, Beijing University of
Technology, Beijing, PR China

A R T I C LE I N FO A B S T R A C T

Keywords: Landfill leachate, generated from the wastes in a landfill, is a type of wastewater with high concentrations of
Nitrification ammonia and organics, causing a serious environmental pollution. Because of its complex and changing char-
Endogenous denitritation acteristics, it is difficult to remove nitrogen from landfill leachate economically and effectively. Hence, nitrogen
Anammox removal is a significant research priority of landfill leachate treatment in recent years. Biological processes are
Biofilm
known to be effective in nitrogen removal. In this work, the biological nitrogen removal treatments were divided
Landfill leachate
into the following processes: conventional nitrification-denitrification process, nitritation-denitritation process,
Nitrogen removal
endogenous denitritation process, and anaerobic ammonium oxidation (Anammox) process. This manuscript
summarized the theories and applications of these approaches in detail, and concluded that appropriate pro-
cesses should be selected in accordance with different characteristics of landfill leachate, in order to effectively
remove nitrogen from all stages of landfill leachate and reduce disposal costs. Finally, perspective on the
challenges and opportunities of biological nitrogen removal from landfill leachate was also presented.

1. Introduction process is most effective in removing nitrogen from landfill leachate;

With rapid economic development and increase in affluent life-
styles, municipal solid waste (MSW) generation has continued to grow
(Miao et al., 2019). There are several methods for disposing MSW, in-
cluding landfilling, composting, incineration, and others (Wiszniowski
et al., 2006). Sanitary landfilling is widely accepted around the world
due to its economic advantages (Renou et al., 2008). However, this
method produces a large amount of leachate, containing high con-
centrations of complex contaminants (Butt et al., 2014). As such,
landfill leachate can significantly pollute the environment if not treated
properly.
Because of its complex compositions and diverse characteristics,
landfill leachate cannot be treated thoroughly using only one process.
For example, the physic-chemical process can remove most of the
contaminants, however, it is expensive and produces secondary pollu-
tion (Di Iaconi et al., 2006). In contrast, the biological treatment is
widely applied for nitrogen removal from wastewater owing to its cost-
effectiveness (Jokela et al., 2002); however, it cannot effectively re-
move nonbiodegradable organics and heavy metals (Wiszniowski et al.,
2006). Given this, landfill leachate should be treated using processes
that couple physic-chemical and biological methods. The biological
therefore, it is important to develop novel biological nitrogen removal
processes.
2. Landfill leachate characteristics
Landfill leachate is defined as the liquid generated from the wastes
in a landfill and by rainwater percolation through wastes (Oliveiral
et al., 2014). Leachate characteristics can be described in terms of
quantity and quality. The quantity of landfill leachate is mainly affected
by rainfall, surface runoff, and groundwater infiltration. Climate is
another important factor, because it determines rainfall and evapora-
tion.
Characterizing landfill leachate quality is complicated by the pre-
sence of multiple contaminants (Boonnorat et al., 2018). In general, the
quality of landfill leachate is characterized using basic parameters such
as suspended solids (SS), COD, biochemical oxygen demand (BOD), pH,
and ammonia (NH4+-N). These parameters vary with rainfall, climate,
the waste ingredients, and landfill age. Landfill age is one of the most
important factors; leachate age is commonly classified into three stages
(Foo and Hameed, 2009): young (less than 5 years old), medium (5–10
years old), and mature (more than 10 years old). Young landfill

∗
Corresponding author.
E-mail address: pyz@bjut.edu.cn (Y. Peng).

https://doi.org/10.1016/j.jenvman.2019.01.057
Received 1 November 2018; Received in revised form 7 January 2019; Accepted 17 January 2019
0301-4797/ © 2019 Elsevier Ltd. All rights reserved.
L. Miao et al.
leachate usually contains high concentration of organics with a high
BOD/COD ratio (0.5–1.0). In mature leachate, COD concentration is
commonly less than 4000 mg/L, with a low BOD/COD ratio (< 0.1)
(Foo and Hameed, 2009). Ammonia is another important contaminant
in landfill leachate. High ammonia concentration is one typical char-
acteristic of mature leachate and it generally does not decline with
landfill age (Kjeldsen et al., 2002). Due to the high levels of ammonia
and organics, landfill leachate can result in serious pollution.
3. Biological nitrogen removal treatments
In general, there are two main methods treating landfill leachate:
physical-chemical treatments and biological treatments (Renou et al.,
2008). The physical-chemical treatments mainly include air stripping,
coagulation-flocculation, absorption, chemical oxidation, and mem-
brane separation (Wiszniowski et al., 2006). Physical-chemical treat-
ments have a relatively stable performance for most contaminants, and
are particularly good for removing refractory organics (Silva et al.,
2017). They are, however, expensive and generate some secondary
pollution (Renou et al., 2008). Biological treatments are widely used to
remove organics and nitrogen from wastewater because they are cost-
effectiveness (Jokela et al., 2002).
Existing nitrogen removal processes, including the conventional
nitrification-denitrification process (Ma et al., 2016), nitritation-deni-
tritation process (Yang et al., 2007), endogenous denitritation process
(Wang et al., 2013), and Anammox process (Li et al., 2014a), all belong
to the category of biological treatments. All these processes can be
applied to remove nitrogen from landfill leachate (Miao et al., 2014).
However, due to the variability in leachate characteristics, it is im-
portant to select the most suitable process to treat different landfill
leachates.
3.1. Conventional nitrification-denitrification process
In the conventional nitrification-denitrification process, NH4+-N is
converted into NO3−-N in aerobic condition; NO3−-N is then reduced to
N2 in the subsequent anoxic condition (Fig. 1). In the aerobic stage,
nitrification occurs in two processes (Bove et al., 2015). In the first
process, NH4+-N (1.0 mol) is oxidized by ammonia oxidizing bacteria
(AOB) to NO2−-N consuming 1.5 mol of oxygen. In the second process,
nitrite oxidizing bacteria (NOB) convert NO2−-N to NO3−-N, with only
0.5 mol of oxygen. After aeration, the denitrification of NO3−-N to N2
occurs during an anoxic stage by heterotrophic bacteria in the presence
of organics. Biodegradable organics are widely used as electron donors;
Fig. 1. Schematic diagram of nitrification and denitrification.
179
Journal of Environmental Management 235 (2019) 178–185
examples include methanol, alcohol, and organic acids (Remmas et al.,
2016).
Due to the presence of oxygen during nitrification, the organics in
the raw wastewater can be converted into CO2 by heterotrophic bac-
teria. It results in the absence of carbon source for subsequent deni-
trification (if no external organics are added). This creates a contra-
diction in the conventional nitrification-denitrification process,
resulting in wasted carbon sources and energy, and an increase in dis-
posal costs. Therefore, landfill leachate, especially mature landfill lea-
chate with a low COD/N ratio and a low BOD/COD ratio, is not suitable
for treatment with conventional nitrification-denitrification process
(Ibrahimpasic et al., 2010).
In general, the organic materials in landfill leachate should be
considered for nitrogen removal as much as possible. To prevent the
conversion of influent organics to CO2 by heterotrophic bacteria, some
modified systems have been deployed to treat landfill leachate with
high COD concentration (Kulikowska et al., 2011; Liu et al., 2018a).
Sun et al. (2010) introduced a two-stage system to treat landfill lea-
chate. This approach achieved COD, NH4+-N, and total nitrogen (TN)
removal efficiencies of 96.7%, 99.7%, and 98.3%, respectively. In this
system, ammonia was oxidized to nitrate in a sequencing batch reactor
(SBR), and the effluent was then recycled into an upflow anaerobic
sludge blanket (UASB) to remove TN by using the organics in the raw
landfill leachate. Liu et al. (2017) achieved COD and TN removals of
80.6% and 74.9%, respectively, through a two-stage anoxic/oxic com-
bined membrane bioreactor system treating landfill leachate; the effects
of different reflux ratios on the system were also investigated, and the
best performance was observed with a reflux ratio of 150% (Liu et al.,
2018b).
3.2. Nitritation-denitritation process
3.2.1. Overview of nitritation-denitritation process
During the nitrification-denitrification process, the conversions of
NO2−-N to NO3−-N and NO3−-N to NO2−-N increase energy con-
sumption and consume more carbon sources in the raw wastewater. If
nitrification could be controlled at the former stage, meaning that
ammonia is just oxidized to nitrite without further being oxidized to
nitrate, the subsequent denitrification could occur with nitrite. The
process controlling the conversion of ammonia only to nitrite is known
as partial nitrification (PN) or nitritation (Sun et al., 2015). The ni-
tritation-denitritation process has several advantages. First, aeration
consumption can be reduced by 25% (Yang et al., 2017), conserving
energy. Second, the process reduces the carbon source used for deni-
trification by 40% (Zhang et al., 2007), reducing costs. Third, nitrita-
tion and denitritation can both significantly decrease sludge yields,
facilitating sludge disposal (Ge et al., 2015). Fourth, the shorter reac-
tion time can reduce the reactor volume and save space.
The critical step in the nitritation-denitritation process is to achieve
stable nitritation. From a macro perspective, nitritation is achieved by
controlling the reaction conditions to cause nitrite accumulation and
prevent further nitrite oxidization. From a micro perspective, the nitrite
accumulation is maintained by accelerating AOB growth and activity,
and by restraining NOB proliferation and activity (Poot et al., 2016).
NOB has been reported having lower growth than AOB, so controlling
sludge retention time (SRT) could help decrease NOB population
(Regmi et al., 2014). Some studies have also found that AOB has a
higher affinity for oxygen than NOB, which means that the low DO
concentration could inhibit NOB growth and activity (Ma et al., 2016).
Besides, some other parameters such as high pH, high temperature and
high influent ammonia concentration, all can accelerate AOB activity
rather than NOB, which is beneficial for nitrite accumulation (Park
et al., 2010).
3.2.2. Application of nitritation-denitritation process on landfill leachate
Nitritation has been widely used to treat landfill leachate, mainly
L. Miao et al. Journal of Environmental Management 235 (2019) 178–185

due to the high ammonia concentration (Miao et al., 2014). High phosphorus release and accumulation. This species is known as gly-
concentration of ammonia may lead to high free ammonium (FA), and cogen accumulating organisms (GAOs) (Liu et al., 1994). PAOs and
then generate the high concentration of nitrite, which may lead to high GAOs, both capable of storing organics, usually coexist in EBPR systems
free nitrous acid (FNA). Both AOB and NOB are influenced by FA and which consist of anaerobic and aerobic stages. As a result, GAOs have
FNA, however, NOB is more sensitive (Chung et al., 2015). FA and FNA been regarded as competitors of PAOs (Zeng et al., 2003a).
are determined by ammonia (or nitrite) concentration, pH, and tem- The proportion of PAOs and GAOs depends on the operational
perature (Formula (1) and (2)) (Anthonisen et al., 1976). When pH and conditions of the EBPR. Operational factors include SRT, pH, tem-
temperature are held constant, high ammonia (or nitrite) concentration perature, carbon source, and influent P/COD ratio (Seviour et al., 2003;
leads to high FA (or FNA). It significantly inhibits NOB, and benefits Lopez-Vazquez et al., 2009). The P/COD ratio is the most important
nitrite accumulation (Ge et al., 2015). Wu et al. (2015) achieved stable parameter. Previous studies suggested that GAOs became the dominant
nitritation in an A/O reactor treating the mixture of municipal sewage species over PAOs at a low influent P/COD ratio (lower than 0.1) (Liu
and landfill leachate, with the help of FA inhibition. et al., 1997). GAOs could play a key role in endogenous denitrification
to achieve nitrogen removal with no need for external carbon sources
17 10 pH
CFA = CNH4+ N × × 6344 ) (Winkler et al., 2011). Landfill leachate, with a low P/COD ratio
14 ( 273
e + T + 10 pH (1) (< 0.002), facilitates GAOs accumulation (Miao et al., 2015c). There-
47 1 fore, nitrogen from landfill leachate may be removed through en-
CFNA = CNO1+ N × × −2300 dogenous denitrification by enriching GAOs.
14 e ( 273 + T ) × 10 pH (2)
As reported, pH usually serves as a real-time control parameter for
3.3.2. Application of nitritation-endogenous denitritation process
nitritation because there is significant variation in the pH profile during
Young and medium landfill leachate, which usually contain large
the nitrification process (Yang et al., 2007). At the beginning of the
amounts of biodegradable organics with a high COD/N ratio and low P/
aerobic phase, organics are first degraded due to the strong oxygen
COD ratio, can be effectively treated using the nitritation-endogenous
affinity of heterotrophic bacteria. The pH level increases when CO2
denitritation process.
stripping occurs. COD degradation is followed by ammonia oxidation.
The nitritation-endogenous denitritation process can take many
During this process, pH continues to decline, as the ammonia is oxi-
forms for treating landfill leachate; these forms are mainly based on a
dized to nitrite and the alkalinity is continuously consumed. After all
SBR system. Wang et al. (2013) changed SBR operation from a tradi-
the ammonia is converted into nitrite, pH increases again when CO2
tional mode to a modified mode, which was anaerobic-interval aerobic/
stripping occurs. This is because the alkalinity is no longer consumed
anoxic-anoxic operation. Using this modified operation, the TN removal
during the oxidation of nitrite to nitrate (Peng et al., 2008). The lowest
efficiency reached 95%. Li et al. (2014b) also developed a modified SBR
point in the pH profile is known as the “ammonia valley”. This “am-
system operated in an anaerobic-aerobic-anoxic mode to treat medium-
monia valley” indicates the termination of nitritation, preventing nitrite
aged leachate, and achieved TN and COD removal efficiencies of 98.3%
from being oxidized into nitrate (Wang et al., 2013). Sun et al. (2015)
and 89.8%, respectively.
established the nitrite pathway in a SBR, using process controls that
These findings support the conclusion that the modified SBR op-
included pH and DO, and maintained stable nitritation at temperatures
erations supported the ability of microorganisms to store carbon
below 15 °C. Moreover, nitritation has been applied in pilot and field
sources and fully utilize the organic substances in landfill leachate to
scale landfill leachate treatments (Sun et al., 2014; Chung et al., 2015).
remove nitrogen (Fig. 2). The anaerobic stage is the critical step for
Chen et al. (2016b) used a pilot-scale nitritation and denitritation
storing the carbon source. PHAs and glycogen were found to be the
process to treat landfill leachate, and DO level was controlled at
main carbon sources for endogenous denitritation and were used by
0.1–0.5 mg/L; a stable nitrite accumulation of 90% was achieved, and
microorganisms in order (Zhu et al., 2013a). In the anaerobic stage, the
the ammonia removal efficiency exceeded 95%.
biodegradable organics were partially stored as PHAs by microorgan-
isms, with concomitant glycogen consumption (Liu et al., 1996). PHAs
3.3. Nitritation-endogenous denitritation process storage minimized COD consumption, and became the available carbon
source for endogenous denitritation (Zeng et al., 2003b). Then PHAs
3.3.1. Theory of nitritation-endogenous denitritation (ED) process were oxidized with concomitant glycogen replenishment in the sub-
In the conventional nitrification-denitrification process, external sequent aerobic or anoxic stage (Oehmen et al., 2005). When PHAs
organic materials must be added to completely remove nitrogen, in- decreased to a low level, the electron donor was converted to glycogen.
creasing the cost and complexity of wastewater treatment. It highlights This is because the microorganisms are prone to use PHAs for cellular
the importance of developing new processes that fully use the carbon functions instead of denitritation (Qin et al., 2005). Therefore, in con-
sources in wastewater to remove nitrogen. The nitritation-endogenous trast with glycogen, PHAs appear to be more suitable for endogenous
denitritation treatment is a novel process based on intracellular stored denitritation (Miao et al., 2015c).
polymers. It can make good use of organic materials in the raw was- Fluorescence in situ hybridization (FISH) analysis indicated that
tewater and achieve nitrogen removal with no need to add external GAOs accounted for a definite proportion of the microorganism and
carbon sources.
In a common enhanced biological phosphorus removal (EBPR)
system, there is one species that absorbs and reserves organics in cells;
this species is referred to as phosphorus accumulating organisms
(PAOs) (Liu et al., 1996). PAOs take up carbon sources and convert
them into polyhydroxyalkanoates (PHAs) or glycogen under the anae-
robic condition, by utilizing the energy gained from hydrolysis of gly-
cogen and polyphosphate. Then, under aerobic condition, PHAs are
oxidized to produce energy for glycogen replenishment and phosphorus
uptake (Oehmen et al., 2005). The phosphorus can be removed through
sludge discharge.
Another species has been found that can store organics as PHAs in
the anaerobic condition, and then use PHAs for glycogen replenishment Fig. 2. Nitrogen removal mechanism via single modified SBR (Miao et al.,
and cell growth in the subsequent anoxic or aerobic condition, without 2015c).

180
L. Miao et al.
that they carried out the advanced nitrogen removal in endogenous
denitritation process (Miao et al., 2015c). The experiments also con-
firmed that low P/COD ratio and modified SBR operational mode
supported GAOs accumulation (Ahn et al., 2006) and facilitated carbon
source storage by GAOs (Miao et al., 2015c). Miao et al. (2016a) de-
tected 108 genera bacteria in endogenous denitritation system; about
46.21% of the total bacteria were the genera responsible for deni-
trification, mainly including Thauera, Ottowia, Paracoccus and Coma-
monadaceae_unclassified. In particular, Paracoccus and Comamonada-
ceae_unclassified could transform organic substances into PHAs and
carry out endogenous denitritation.
Some studies have found that the influent COD/N ratio and oxygen
supply in the aerobic stage are important for endogenous denitritation
performance (Zhu et al., 2013b; Miao et al., 2015a). Nitrogen removal
through endogenous denitritation was achieved when COD/N ratio
reached above 4, because that the deficient organics limited the amount
of carbon source stored at COD/N ratios of 1–3 (Miao et al., 2015c). A
stronger oxygen supply improved nitrogen removal, because it pre-
served more PHAs after aeration and then improved the endogenous
denitritation rate (Miao et al., 2015a). The carbon source type is an-
other important factor affecting endogenous denitritation. Miao et al.
(2016a) tested endogenous denitritation performance with acetate,
propionate and glucose used as carbon sources. The results showed that
the most efficient nitrogen removal was achieved with acetate, because
the acetate was most easily converted into intracellular polymers.
Glucose exhibited the lowest performance, because it was associated
with the lowest amount of PHAs storage. Moreover, the carbon source
type influenced PHAs composition (Fig. 3). All tests suggested that the
polyhydroxybutyrate (PHB) fraction in PHAs was present at higher le-
vels than polyhydroxyvalerate (PHV). PHV was only synthesized when
acetate served as the carbon source.
In summary, the nitritation-endogenous denitritation process can
fully use the organics in raw landfill leachate to remove nitrogen. While
this may result in a longer reaction period than with traditional pro-
cesses (Miao et al., 2015c), it can reduce the need for external carbon
sources and reduce energy consumption for aeration, lowering opera-
tional costs. Furthermore, the sludge production could also decline,
benefitting from long-term endogenous metabolism (Wang et al.,
2013).
Fig. 3. Mechanism of carbon source types impacting on intracellular stored
polymers during endogenous denitritation treating landfill leachate (Miao et al.,
2016a).
181
Journal of Environmental Management 235 (2019) 178–185
3.4. Anaerobic ammonium oxidation (Anammox) combination process
3.4.1. Theory of Anammox process
3.4.1.1. Overview of Anammox. Anammox was first discovered by
Mulder et al. (1995), and has been considered as an attractive process
with high cost-effectiveness and great potential (Jin et al., 2008). In
anaerobic conditions, ammonia is oxidized by nitrite (as an electron
acceptor) to nitrogen gas and a little nitrate. The specific chemical
formula (Formula (3)) is as follows (Kuenen, 2008).
NH+4 + 1.32NO−2 + 0.132HCO−3 + 0.512H+ → 1.02N2 + 0.26NO−3
+ CH2 O0.5 N0.15 + 2.19H2 O (3)
3.4.1.2. Ecophysiology of Anammox bacteria. Anammox bacteria belong
to a deep-branching lineage of the phylum Planctomycetales
(Shivaraman, 2003). Seven genera, including 22 species, have been
identified as Anammox bacteria (Kuenen, 2008; Kumar and Lin, 2010;
Khramenkov et al., 2013). Anammox bacteria are widely distributed,
generally existing in freshwater ecosystems, sewage treatment facilities,
and laboratory bioreactors (Zhu et al., 2013c; Wang et al., 2015).
Table 1 categorizes Anammox species based on their genera.
3.4.1.3. Physiology of Anammox bacteria. Anammox bacteria are
usually cultivated from soil and activated sludge (Lotti et al., 2015).
Anammox bacteria have been confirmed as a kind of chemoautotrophic
bacteria, and use CO2 as the carbon source (Kimura et al., 2011). The
bacteria can obtain energy from the oxidation of NO2−-N to NO3−-N
and fix carbon source through the acetyl coenzyme A (acetyl-CoA)
pathway (Schouten et al., 2004). Some Anammox bacteria (e.g. Ca.
Anammoxoglobus propionicus) can oxidize small organics, including
acetate and propionate, but do not incorporate these organics into
their biomass (Kartal et al., 2007, 2008). However, the slow cell growth
rate of Anammox, with a long doubling time (10–30 days), restricts its
application for wastewater treatment.
Several key parameters could affect Anammox activity. First,
Anammox bacteria are sensitive to oxygen (Egli et al., 2001). Conven-
tional views have held that Anammox bacteria could be temporarily
inhibited at DO concentration of 0.032 mg/L, and were later restored
after the oxygen has disappeared (Strous et al., 1999). Some studies also
found that no Anammox activity occurred after long exposure to high
DO concentrations (e.g. 18% of oxygen saturation) (Egli et al., 2001).
This indicates that low oxygen concentrations create a reversible in-
hibition for Anammox; exposure to high oxygen levels causes irrever-
sible inhibition. However, Anammox bacteria may adapt to high DO
concentrations when co-existing with aerobic bacteria (e.g. AOB) in a
step-wise culture (Liu et al., 2008b).
Second, temperature and pH influence bacteria growth and meta-
bolism (Tomaszewski et al., 2017). Some studies have found that the
optimal temperature for Anammox wastewater treatment ranges be-
tween 30 and 40 °C (Jin et al., 2012; Isanta et al., 2015); this narrow
range may limit Anammox application for wastewater treatment. The
physiological pH of Anammox bacteria is 6.7–8.3 (Strous et al., 1999).
pH determines the FA and FNA concentrations, and FA has been shown
to be an important inhibitor in the Anammox system. A low pH can lead
to low FA, which benefits Anammox bacteria activity (Jaroszynski
et al., 2011). However, other studies have suggested that Anammox was
primarily inhibited by pH rather than FA (Puyol et al., 2014). Extreme
pH values may affect proton transfer and other metabolic processes that
depend on pH gradients, such as energy production (Carvajal-Arroyo
et al., 2014).
Third, the affinity constants of Anammox bacteria for ammonium
and nitrite are both less than or equal to 0.1 mgN/L (Strous et al.,
1999). Therefore, a high substrate concentration could inhibit Ana-
mmox activity, especially at high nitrite concentrations (Lotti et al.,
2012). Fourth, organic matter is also considered as a significant factor
L. Miao et al. Journal of Environmental Management 235 (2019) 178–185

Table 1
Various known Anammox species.
Genus Species Origin Reference

Brocadia Ca. Brocadia anammoxidans Wastewaster treatment plants Kuenen and Jetten (2001)
Ca. Brocadia fulgida Wastewaster treatment plants Kartal et al. (2008)
Ca. Brocadia sinica Lab-scale bioreactor Hu et al. (2010)
Ca. Brocadia brasiliensis Araujo et al. (2011)
Ca. Brocadia caroliniensis Rothrock et al. (2011)
Kuenenia Ca. Kuenenia stuttgartiensis Biofilm trickling filter Schmid et al. (2000)
Scalindua Ca. Scalindua brodae Wastewater treatment for landfill leachate Schmid et al. (2003)
Ca. Scalindua wagneri Schmid et al. (2003)
Ca. Scalindua sinooifield high-temperature oil reservoirs Li et al. (2010)
Ca. Scalindua zhenghei marine ecosystem Hong et al. (2011)
Ca. Scalindua richardsii Fuchsman et al. (2012)
Ca. Scalindua sorokinii Kuypers et al. (2003)
Ca. Scalindua arabica Woebken et al. (2008)
Ca. Scalindua marina Brandsma et al. (2011)
Ca. Scalindua profunda Van De Vossenberg et al., 2013
Brasilis Ca. Brasilis concordiensis Lab-scale bioreactor Viancelli et al. (2011)
Jettenia Ca. Jettenia asiatica Biofilm reactor Quan et al. (2008)
Ca. Jettenia moscovienalis Lab-scale bioreactor Nikolaev et al. (2015)
Ca. Jettenia caeni Ali et al. (2015)
Anammoxoglobus Ca. Anammoxoglobus propionicus Lab-scale bioreactor Kartal et al. (2007)
Ca. Anammoxoglobus sulfate Rotating biological contactor Liu et al. (2008a)
Anammoximicrobium Ca. Anammoximicrobium moscowii filtrate of wastewater fermented sludge Khramenkov et al. (2013)

influencing Anammox (Anjali and Sabumon, 2014). The organics may
result in competition between Anammox and denitrifying bacteria
(Tomar et al., 2015). Li et al. (2015) found that NH4+-N and TN re-
moval efficiencies both exceeded 90% at COD concentration of 100 mg/
L. In contrast, Anammox accounted for only 69% of TN removal when
COD concentration reached about 300 mg/L. Although organics may
adversely impact Anammox, a low COD concentration or suitable COD/
N ratio may support the co-existence of Anammox and denitrification
(Ni et al., 2012; Miao et al., 2015b). This could subsequently promote
nitrogen removal efficiency.
3.4.1.4. Advantages and disadvantages of Anammox process. Compared
with nitrification-denitrification process, Anammox process has several
advantages. First, the process reduces oxygen consumption by 60% (Ma
et al., 2016), because only approximately 50% of influent ammonia
needs to be oxidized to nitrite instead of nitrate. Second, the process
saves 100% of the carbon source used to remove nitrogen, because
nitritation and Anammox are both autotrophic processes, without the
need for organics. Third, the slow growth of Anammox bacteria
significantly reduces sludge production, which also reduces sludge
disposal costs. Finally, nitrous oxide is not detected during Anammox
process, and studies have shown that Anammox bacteria does not
produce nitrous oxide in their metabolic processes (Kampschreur et al.,
2008; Kartal et al., 2010). This reduces greenhouse gas emissions.
Due to these advantages, Anammox process is considered a sus-
tainable way to treat wastewater, and has been applied for the treat-
ment of ammonia-rich wastewater (e.g. landfill leachate) (Miao et al.,
2016b). However, Anammox process also has some disadvantages. For
example, Anammox process requires a long accumulation time and
SRT. It prolongs Anammox process start-up (Chen et al., 2016a). As
another example, some factors adversely affect Anammox, including
oxygen and organics. As such, suitable reaction conditions are required
to support Anammox process and promote synergism between Ana-
mmox bacteria and other microorganisms.
3.4.2. Application of Anammox combination process
Several forms of Anammox have been used to treat wastewater; this
has been done in a granular sludge system or biofilm system, or in a
one-stage system or multi-stage system. A one-stage system combines
nitritation and Anammox in an integrated reactor, with simple opera-
tions. DO and organics levels are the major challenges for a one-stage
system, so controlling the operational conditions, such as DO, is
necessary (Zheng et al., 2016). Xu et al. (2010) successfully treated
landfill leachate in a single periodically aerated SBR with DO con-
centrations of 1.0–1.5 mg/L. Zhang et al. (2017) achieved simultaneous
nitritation, Anammox, and denitrification (SNAD) in a lab-scale SBR
with intermittent aeration, and the process obtained the highly efficient
Anammox with TN removal efficiency of 99%. The combination of ni-
trification, Anammox and denitrification has also been proved by de-
tecting the coexistence of nitrifiers, Anammox bacteria, and denitrifiers
though PCR-DGGE (Xiao et al., 2008).
Unlike the one-stage system, the multi-stage system joins nitritation
and Anammox together, proceeding in individual reactors. AOB and
Anammox bacteria are cultivated separately, protecting Anammox
bacteria from oxygen and organics, and promoting stable nitrogen re-
moval performance. Some researchers have applied coupled systems of
half-nitritation and Anammox to remove nitrogen from old-aged land-
fill leachate, however, the nitrogen removal efficiency was usually
lower than 90% (Anfruns et al., 2013; Li et al., 2017). It is likely be-
cause most of biodegradable organics were reduced in a half-nitritation
reactor and there was low biodegradability potential in the influent for
Anammox. Therefore, organics in the landfill leachate should be partly
introduced to Anammox system. Miao et al. (2014) achieved nitrogen
removal of 90% from landfill leachate through a three-stage SBR system
consisting of pretreatment, nitritation, and Anammox. In this system,
approximately 50 mg/L COD in the effluent of the pretreating SBR was
used for denitrification. Wang et al. (2016) developed a continuous-
flow system including partial nitrification and Anammox to treat
landfill leachate; the organics were introduced to the Anammox reactor
accompanied by the landfill leachate, and TN removal efficiency
reached about 94%.
The effect of organics in wastewater on nitrogen removal depends
on their concentrations (Ni et al., 2012). Low organics concentration,
particularly biodegradable organics, can improve nitrogen removal
without inhibiting Anammox bacteria activity (Miao et al., 2015b).
Anammox and denitrifying bacteria can also coexist and both con-
tribute to nitrogen removal in a wastewater treatment system (Kumar
and Lin, 2010). Therefore, promoting the synergy between Anammox
and denitrification is important in enhancing nitrogen removal per-
formance.
Some systems, such as biofilm and granular systems, have been
coupled with nitritation and Anammox processes to treat wastewater
(Lackner et al., 2014; Azari et al., 2017; Wen et al., 2017). Studies have
found that Anammox bacteria are prone to form granules or attach to

182
L. Miao et al.
carriers to form biofilm (Zheng et al., 2016; Miao et al., 2016b). This
aggregation, especially the formation of biofilm, has more microbial
populations with a smaller footprint (Van Hulle et al., 2010). Biofilm
also has a great ability to resist adverse factors from external environ-
ment (Nicolella et al., 2000), which contributes to a more stable eco-
logical system.
Because landfill leachate is complex wastewater with many biode-
gradable and refractory compounds, biofilms can act as a buffer to re-
duce toxic substance concentrations. Biofilms can also stabilize landfill
leachate treatment and improve nitrogen removal performance by al-
lowing Anammox and denitrifying bacteria to coexist (Miao et al.,
2018). It demonstrates that the biofilm system is appropriate for
treating landfill leachate. Phan et al. (2014) added a bio-carrier bed
into Anammox system to treat mature landfill leachate, and achieved
TN removal of 93 ± 1% at a nitrogen loading rate of 4.2 kgN/(m3·d).
Miao et al. (2016b) successfully operated an Anammox biofilm system
to treat landfill leachate, and improved nitrogen removal efficiency to
95%. This Anammox biofilm system could tolerate more organics in
comparison with the Anammox system coupled with floccule (Miao
et al., 2014).
As for biofilm, the microorganisms and complex high molecular-
weight extracellular materials produced by microorganisms are the
major components (Flemming and Wingender, 2010). These materials
are referred to as extracellular polymeric substances (EPS), which play
an important role in the formation of the biofilm and stability of mi-
crobial community (Li and Yang, 2007). EPS production changes in
different environments (Shammi et al., 2017), which could impact ni-
trogen removal performance. Miao et al. (2018) found that increasing
influent organics stimulated EPS production and made a significant
effect on the compositions of EPS. Anammox microorganism popula-
tions also increased significantly in the biofilm with increasing influent
organics. It indicates that EPS could promote Anammox bacteria en-
richment and provide nutrients and enzymes for microorganisms,
which facilitates effective nitrogen removal.
Furthermore, the microbial communities also affect EPS production.
Tang et al. (2011) found that Anammox bacteria have a strong tendency
to produce massive amounts of EPS. The variations of Anammox po-
pulations were similar to the variations of EPS compositions (Miao
et al., 2018). Furthermore, in the Anammox systems treating landfill
leachate, the dominant genus of Anammox bacteria was identified as
Candidatus Kuenenia (Miao et al., 2018; Zhang et al., 2018); Candidatus
Kueneniastuttgartiensis was the dominant species (Yapsakli et al., 2011;
Phan et al., 2017). This may be because the Candidatus Kuenenia has
high substrate affinity (Isanta et al., 2015) and adapts better to landfill
leachate.
4. Conclusions
This paper reviewed recent advances in biological nitrogen removal
treatments for landfill leachate. The conventional nitrification-deni-
trification process and nitritation-denitritation process are usually ap-
plied for young landfill leachate treatment. Nitritation-endogenous
denitritation process is preferred for medium landfill leachate, by fully
using organics in raw wastewater. The Anammox process is a promising
way for mature landfill leachate treatment, as it effectively removes
nitrogen and requires no external carbon source. In conclusion, ad-
vanced nitrogen removal from all stages of landfill leachate can be
achieved by selecting appropriate processes.
5. Research prospects
There have been many studies on biological nitrogen removal pro-
cesses for treating landfill leachate. Some processes have been ex-
tensively studied, while some processes would benefit from more re-
search. Recommended areas for future research are as follows:
183
Journal of Environmental Management 235 (2019) 178–185
(1) The nitritation-endogenous denitritation process has been shown to
perform well in treating landfill leachate; however, more researches
are needed to study the factors impacting the process, including DO
concentration, COD/N ratio, and influent substrate concentration.
(2) The lab-scale nitritation-endogenous denitritation process has been
successfully used for landfill leachate treatment; as such, testing the
performance of pilot-scale or full-scale processes is an important
next step.
(3) There are diverse microorganisms in the Anammox combined pro-
cesses, and the relationship and contributions among them are
unclear. This highlights the need for additional researches into the
mechanisms of Anammox combined processes.
(4) Because the characteristics of landfill leachate are special, it would
be useful to discover new species of Anammox bacteria that can
support landfill leachate treatment.
Acknowledgements
Financial support of the project Natural Science Foundation of
China (51808240) and Fundamental Research Funds for the Central
Universities (HUST) (grant number 2018KFYYXJJ134) and are grate-
fully acknowledged.
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