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Rice Science, 2021, 28(1): 43í57

Review

Flooding Tolerance in Rice: Focus on Mechanisms and

Approaches

Debabrata PANDA, Jijnasa BARIK

(Department of Biodiversity and Conservation of Natural Resources, Central University of Odisha, Koraput 764021, Odisha, India)

Abstract: Flooding is one of the most hazardous natural disasters and a major stress constraint to rice
production throughout the world, which results in huge economic loss. Approximately one-fourth of the
global rice crops (approximately 40 million hectares) are grown in rainfed lowland plots that are prone to
seasonal flooding. A great progress has been made during last two decades in our understanding of the
mechanisms involved in adaptation and tolerance to flooding/submergence in rice. In this review, we
summarized the physiological and molecular mechanisms that contribute to tolerance of flooding/
submergence in rice. We also covered various features of flooding stress with special reference to rice
plants, viz. different types of flooding stress, environmental characterisation of flood water, impact of
flooding stress on rice plant and their morphological, physiological and metabolic responses under
flooding. A brief discussion on the tolerance mechanism in rice exhibited to different types of flooding will
be focused for the future crop improvement programme for development of flooding tolerant rice variety.
Key words: flooding stress; submergence tolerance; Sub1; rice

Rice (Oryza sativa L.) is consumed as the primary
source of food in the globe, which provides food and
livelihood security to half (about 3.5 billion) of the
world population (Samal et al, 2018). It is a semi-
aquatic species, cultivated under different climatic
conditions, which are subjected to diverse biotic and
abiotic stresses. Among different abiotic stresses, flooding
is one of the major constraints for rice production
particularly in rainfed lowland areas, which threatens
global food security (Dar et al, 2017). This is becoming
a more serious issue concerning the global climate
change as the improved rice varieties are susceptible
to flooding (Afrin et al, 2018). A total of 22 million
hectares of world’s rice fields are unfavourably
submerged annually, which affects the livelihood of
more than 100 million people (Singh et al, 2016). The
population of world is alarmingly increasing and
likely to reach 9 billion by the year 2050 (Lee et al,
2014). Therefore, the present situation demands to
increase in food production to overcome food shortage
problem in near future (Wang et al, 2016). Due to
heterogeneity of flood prone ecosystem in tropical
Asian countries like India, few indigenous rice landraces
are still maintained and cultivated by poor farmers
(Ram et al, 2002; Barik et al, 2020). Although such
farming has poor yield capacity, this local landrace
has excellent adaptation to extreme water availability,
tolerant to different kinds of flooding and very important
for QTL mapping and gene discovery (Singh et al,
2017). Therefore, identification of new tolerant rice
genotypes having better performance than ‘FR13A’
(flooding tolerant genotype) is agronomically more
desirable (Sarkar et al, 2006). This needs exploration of
genetic diversity in the rice landraces for identification
of new genes and further improvement of germplasm
(Ahmed et al, 2016). Therefore, the present review
reported the effects of flooding and highlighted the
recent development of morphological, physio-biochemical

Received: 9 January 2020; Accepted: 9 May 2020

Corresponding author: Debabrata PANDA (dpanda80@gmail.com)
Copyright © 2021, China National Rice Research Institute. Hosting by Elsevier B V
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)
Peer review under responsibility of China National Rice Research Institute
http://dx.doi.org/10.1016/j.rsci.2020.11.006
http://dx.doi.org/


 44
and genetic basis of flooding tolerance in rice.
Flooding stress and flooding affected area
Although rice plants require large amount of water
during growth, flooding stress results in severe loss of
the crop as a consequence of anaerobic environment.
The crop loss would be more severe on account of
unpredictable changes in climatic conditions causing
floods (Vergara et al, 2014). The extensive rice
growing areas in South and South-East Asia especially
India, Bangladesh, Thailand, Vietnam, Myanmar and
Indonesia are exposed to flash flooding during
monsoon season (Sarkar et al, 2006). Nearly 22
million hectares of rainfed lowland areas of South and
South-East Asia get affected due to flooding, out of
which about 6.2 million hectares of rice lands are in
India (Azarin et al, 2017; Dar et al, 2017). Out of 22
million hectares, 15 million hectares of rainfed lowland
are affected only due to short term flash flooding
(Singh et al, 2017), and economic loss is estimated to
be 1 billion US dollar (Mackill et al, 2012). The flood
prone ecosystem comprises about 7% of global rice area
and produces 4% of world rice (Yang et al, 2017).
Types of flooding stress
Flooding during germination
Soil water logging normally takes place in case of
rainfall after sowing of seeds and particularly where
the lands are not properly levelled. The germination during
flooding is commonly referred as ‘ANAEROBIC
GERMINATION’ (AG), and the seeds have the
potential to germinate even under the conditions where
no air or oxygen is available. However, slow seed
germination, uneven and delayed seedling establishment
and high weed infestation are some of the constraints
that restrict rice large adaptation of direct seeding in
flood prone areas (Ismail et al, 2009, 2012). Almost
all varieties of rice are keenly susceptible to flooding
and get easily affected or damaged at the time of
germination (Yamauchi et al, 1993; Ismail et al, 2009;
Angaji et al, 2010; Miro and Ismail, 2013; Lee et al,
2014; Singh et al, 2017). AG is a complex trait
governed by various gene families which are linked to
many important processes like starch breakdown,
glycolysis, fermentation as well as other biochemical
and metabolic processes (Ismail et al, 2009, 2012).
Hence, AG is thus necessary for homogeneous
germination and better crop establishment under flooding
condition (Ismail et al, 2009; Magneschi and Perata,
2009; Septiningsih et al, 2013).

Rice Science, Vol. 28, No. 1, 2021
Flash flooding
Flash flooding can occur due to heavy rains or
overflowing of rivers and streams for a short duration
of 1–2 weeks and result in complete submergence of
plants (Vergara et al, 2014). Lands in low-lying land
areas and near the streams and rivers are generally
affected by such flash flooding. Depending on climatic
conditions, flash flooding can occur repeatedly in a
growing season, damaging the rice cultivation. The
flooding depth is not very deep in the flash flooding
condition. Flooding tolerance was defined as ‘the ability
of some rice varieties to survive 10–14 d of complete
flooding and renew its growth when the flood water
subsides’ by Catling (1992).
Stagnant flooding
In stagnant flooding, water depth ranges from 25 to 50
cm, and some parts of the shoot is normally seen over
the flood water. Rice production in such cases depends
upon the extent of submergence and may vary from
0.5 to 1.5 t/hm2 (Kuanar et al, 2017). In this condition,
the extended water stagnation remains from a few
weeks to several months. This is commonly seen in areas
where there is no proper drainage system, particularly
near the water channels leading to rivers during the
monsoon time, due to over flow of excessive rain
water. In West Bengal of India, 577 genotypes were
studied under stagnant flooding conditions, where
genotypic differences between survival and grain yield
were observed, but the flooding tolerant cultivars
(FR13A and FR43B) were not found to perform well
under the aforesaid condition. This suggests that the
flash flooding tolerant genotypes may not be the right
solution for stagnant flooding (Vergara et al, 2014).
Deep-water flooding
In deep-water flooding, water stagnation continues to
remain for a longer time. At times, the water level
may rise up to 4 m height (Singh et al, 2017). The
flooding stress depending upon the topography and
climatic conditions may continue for months together.
Adapted rice plants in this type accelerate their growth
in commensurate with the increase of flood water
level, so as to avoid complete submergence. In deep-
water flooding, a portion of plants successfully remains
above the water level. The deep-water rice escapes
complete flooding by fast internodal elongation
growth, which consumes high amount of carbohydrate.
Growth is achieved by about 25 cm per day as flood
water level increases (Singh et al, 2017) and can reach
Debabrata PANDA and Jijnasa BARIK. Flooding Stress in Rice
a height of 5 m with its panicle and top leaves under
these circumstances (Jackson and Ram, 2003). Most
of the deep-water rice growing areas in India are
normally found in several parts of Assam, Bihar,
Orissa, West Bengal and Uttar Pradesh (Bin Rahman
and Zhang, 2016).
Environmental parameters associated with
flooding stress
Flooding causes many complex abiotic stresses (Jackson
and Ram, 2003; Sarkar et al, 2006; Bailey-Serres et al,
2010), and the amount of harm that may be caused to
the inundated plants varies depending on flood water
characteristics like temperature, turbidity, water depth,
oxygen, carbon dioxide concentration and light intensity
(Das et al, 2009). These affect important plant processes
like chlorophyll retention, photosynthesis under water,
accumulation of carbohydrate, elongation and survival
(Das et al, 2009).
Gas diffusion
Gas diffusion under flooding has a key role for plant’s
survival since diffusion of oxygen under water takes
place slowly compared to open air (approximately 104
folds) (Panda et al, 2008). Thus, the oxygen depletion
creates hypoxia (low oxygen) and anoxia (no oxygen)
around plant tissues during flooding (Ito et al, 1999;
Damanik et al, 2012). This reduces the supplies of
carbon dioxide (CO2) and oxygen (O2) to the plants.
Such reduced supplies of O2 and CO2 limit respiration
and photosynthesis, respectively. Further endogenous
concentration of volatile hormone ethylene depends
strongly upon its outward diffusion, which is also
severely impeded under water. As a result, ethylene is
increased in submerged plant tissues. Thus, the reduced
diffusion of ethylene away from plants triggers chlorosis
and excessive elongation of leaves in intolerant cultivars
(Sarkar et al, 2006).
Light intensity
Poor light transmission under flood water is an
important limiting factor. Under these conditions,
light reaching the leaves of the flooded plant is
impaired by phytoplanktons, water, silt as well as
dissolved organic matter suspended in the flood water
(Das et al, 2009). Due to this, only a limited amount
of sun light reaches leaves and thus decreases the
potentiality of plants to photosynthesis (Sarkar et al,
2006). Light intensity is also very important for
maintaining O2 and CO2 concentrations, and therefore

45
greatly affects the physiological status of rice plants
under water. Light limitation has been shown to cause
severe injuries and accelerate plant mortality (Jackson
and Ram, 2003).
Temperature
Temperature is another important factor which plays a
vital role in rice plant’s survival under flooding. The
low temperatures (20 ºC) improve plant survival whereas
the high temperature (30 ºC) increases plant mortality.
The solubility of O2 and CO2 in flood water decreases
at high temperatures. High temperatures speed up
anaerobic respiration, resulting starvation and death of
the plants in a short duration (Ram et al, 2002).
pH
Plant survival is also greatly affected by pH values
during flooding. Lower flood water pH or enhanced
CO2 concentration increases photosynthesis during flooding
(Sarkar et al, 2006). Although this idea is not practical,
it indicates that the tolerant germplasms need to be
specific to the location to meet the local variations of
the flood water condition (Sarkar et al, 2006).
Toxic substances
Soil oxygen deficiency has indirect effects on plants
by promoting the growth of anaerobic bacteria. Some
anaerobes reduce ferric ion (Fe3+) to ferrous ion (Fe2+),
and due to greater solubility of Fe2+, Fe2+ concentrations can
rise to toxic level. Other anaerobic microorganisms may
reduce sulfate (SO42-) to hydrogen sulphide (H2S),
which is a respiratory poison. These toxic substances
cause severe damage to plants by reducing the redox
potential under flooding conditions and increase the
susceptibility to diseases (Ram et al, 2002). Also, the
acetic acids as well as butyric acids released by
anaerobic microorganisms are toxic to plants at high
concentrations.
Effects of flooding on morphology of rice
plants
Leaf traits
Flooding hinders formation of new leaf, and reduces
total leaf area and promotes leaf senescence (Kato et al,
2014). Singh et al (2014) observed that in susceptible
varieties, the dry weight of leaf is reduced sharply to
the extent of 70%, when it is only 30%–40% in
tolerant varieties under 17 d of flooding stress. They
found a strong relationship between the dry weight of
 46
leaf after flooding and the plant survival in critical
flooding conditions. It is noticed that there was no
perceptible reduction of leaf area index in tolerant
varieties after water recedes, suggesting their stronger
ability to recover original leaf area index after flash
flooding (Singh et al, 2014).
Plant height
Plant height is a key parameter which influences plant
productivity as well as its survival under flooding
stress. Most of rice genotypes elongate their shoots
under complete flooding (Fig. 1). Flash flooding
tolerant varieties reduce shoot growth while remaining
in flood, conserving their energy which is used for
survival soon after recession flood water (Jackson and
Ram, 2003; Bailey-Serres et al, 2010; Goswami et al,
2017; Bui et al, 2019). Particularly Sub1-incorporated
varieties show lower elongation of shoot in comparison
to other varieties (Sarkar and Bhattacharjee, 2011). A
study on 903 cultivars from International Rice Research
Institute Genebank collections confirmed the negative
relationship of shoot elongation and flash flooding
tolerance (Setter and Laureles, 1996). Kawano et al
(2002) observed that shoot elongation during flooding
utilises energy from stored carbohydrate. Singh et al
(2014) observed that shoot elongation increases
proportionately with flooding, and Sub1-introgressed
genotypes show slower shoot elongation rates compared
to intolerant genotypes. Thus, rice varieties having
slower elongation growth under water are preferred
for cultivation in the areas affected with flash flooding,
and the genotypes having faster elongation capacity
Fig. 1. Morphological, physiological, biochemical and molecular responses of rice plants under flooding stress (Mackill et al, 2012; Upadhyay,
2018).

Rice Science, Vol. 28, No. 1, 2021
are considered appropriate for deep water as well as
partially deep-water areas (Sarkar and Bhattacharjee,
2011; Vergara et al, 2014).
Root trait
Root health is vital for survival during flooding and
recovery. The roots of sensitive species suffer oxygen
deficiency in anoxic soils, which reduce respiration
and result in drastic energy crisis, whereas the tolerant
species can thrive (Bailey-Serres and Voesnek, 2008).
The concentration of potentially toxic compounds also
increases in the anaerobic soils, which can enter
through roots, damaging both shoot and root tissues.
The morphological and anatomical traits of roots
determine the root growth and functioning in anoxic
soils (Yamauchi et al, 2018). Long term retention of
leaf gas films during flooding will likely to improve
the supply of carbohydrates for growth and adventitious
root regeneration as well as enhance root and
rhizosphere aeration. Root porosity is also important
for root growth under anoxic soils, which is determined by
the extent and rate of aerenchyma formation and to
minimize resistance to O2 movement down the root.
The aeration of root will be improved significantly by
induction of barriers to radial oxygen loss, which
reduces oxygen loss, promotes aeration to active root
tips, and impedes the movement of toxin and gases in
anoxic soils (Yamauchi et al, 2018). The root elongation
was found to be minimal in all the rice genotypes
under flooding when compared with control condition.
The positive correlation between survival and root
growth is an indication of the capability of Sub1
Debabrata PANDA and Jijnasa BARIK. Flooding Stress in Rice
introgressed varieties to supply the required carbohydrates
needed for proper growth of its roots (Singh et al,
2014). Bui et al (2019) highlighted the importance of
Sub1 QTL in regulation of root physiology of rice under
flooding conditions and indicated that the tolerant varieties
show higher root activities like root tip viability and
root peroxidase, which result in minimum damage in
the roots as well as shoots under flooding.
Biomass
Flooding severely restricts O2 and CO2 gas exchange
between rice tissues and atmosphere, inhibits aerobic
respiration as well as photosynthesis, and accelerates
the consumption of energy reserves, leading to stunted
growth and death (Kato et al, 2014). Flooding inhibits
dry matter accumulation in susceptible genotypes
(Singh et al, 2014). Reduction in growth is a common
phenomenon when plants are flooded with water.
However, tolerant rice genotypes have the potentiality
to store sufficient amount of dry matter.
Aerenchyma formation and leaf gas film
The major adaptive features of rice to water logging
are the formation of aerenchyma. This aerenchyma
constitutes the gas spaces and gets interconnected, which
becomes the channel for continuous aeration between
roots and shoots. In rice, aerenchyma is well developed
in leaves, sheaths, roots and internodes (Steffens et al,
2011; Pradhan et al, 2017). Generally, the formation
of aerenchyma and its induction occurs within 1–3 d
of the anoxic treatment (Pradhan et al, 2017).
In flood water conditions, the presence of leaf gas
films enhances the internal aeration and makes the rice
tolerant to flood (Pedersen et al, 2009). The leaf gas
film facilitates the O2 entry from surrounding water
when in the dark, and the CO2 entry in the light for
photosynthesis. The role of gas films for flooding
tolerance has been confirmed through several studies
by artificially induced variation or loss of function
mutants (Pedersen et al, 2009). Kurokawa et al (2018)
reported the ‘Leaf Gas Film1’ (LGF1/OsHDS1) enhances
flooding tolerance in rice.
Survival
The extent of visible injury caused by flooding stress
is generally used as an indicator of sensitivity of
plants to flooding. However, 100% survival is
identified in tolerant cultivar FR13A after remaining
in flood water for 8 d while only less than 15% in
sensitive cultivar (Panda and Sarkar, 2013). In a study

47
among seven indica rice cultivars, after 4 d of
flooding treatment, the survival rate is more than 80%
in tolerant genotypes but only 22% in susceptible ones
(Panda et al, 2006). Survival after flooding seems to
be associated with the amount of non-structural
carbohydrates remaining in the shoots after flooding
(Pradhan et al, 2017).
Effects of flooding on physiology of rice
plants
Photosynthetic gas exchange
One of the crucial processes for plant growth is
‘Photosynthesis’. By this process, solar energy used
by green plants is converted into organic compound
releasing oxygen molecule. The adverse impact of
flooding on rice is mainly linked to water poor gas
exchange through impeding photosynthesis (Colmer
and Voesenek, 2009). Inundation of aerial organs under
complete flooding drastically reduces gas diffusion rates,
restricts uptake of oxygen, thus weakens photosynthesis
as well as transpiration and thereby the carbohydrate
reserves in the plant get exhausted, resulting in its
death (Das et al, 2009). Further turbid flood water can
almost completely block light accessibility and slow
down photosynthesis (Yang et al, 2017). Factors like
chlorophyll degradation, stomatal closure, lipid peroxidation
and less intracellular carbon dioxide concentration also
cause concomitant reduction in photosynthesis (Panda
et al, 2008). Senescence and plant death occur due to
decline in CO2 concentration and enhancement of leaf
reactive oxygen species (ROS) level because of continuing
of the light reaction. Panda and Sarkar (2013) observed
that flooding significantly inhibits the photosynthetic
rate along with stomatal conductance and rubisco
activity. The maximum reduction (95%) in photosynthetic
rate was found in the sensitive genotype IR42, and the
least reduction (74.8%) was found in the tolerant
genotype FR13A, when compared with non-submerged
control plants after 8 d of complete flooding (Panda et al,
2008). The maintenance of higher photosynthetic activity
in tolerant cultivars may be due to protection of
photosynthetic apparatus, higher amount of chlorophyll
and better stomatal conductance during and after
flooding compared to susceptible variety IR42 and
elongating variety Sabita (Panda and Sarkar, 2013).
Leaf photochemical activity
Photosystem II (PSII) is susceptible to various abiotic
stresses. When rice plants were completely submerged,
chloroplasts were disintegrated and leaf photosynthetic
 48
capacity was decreased (Panda et al, 2008). In vivo
chlorophyll fluorescence is used as a matter of
convenience and non-destructive tool to ascertain the
tolerance behaviour of different kinds of species under
different environments including flooding conditions
(Panda et al, 2006; Panda and Sarkar, 2011, 2012).
Changes in chlorophyll fluorescence emissions, mostly
from PSII, give all the information about photosynthetic
activity (Panda et al, 2008). Flooding affected the
photosynthetic apparatus which is evident from the
reduced values of chlorophyll fluorescence such as F0,
Fm and ratio of variable fluorescence to maximum
fluorescence (Fv/Fm) in both tolerant and intolerant
genotypes (Panda et al, 2006, 2008). A comparison
between chlorophyll fluorescence parameters and chlorophyll
contents indicates that the chlorophyll fluorescence
parameters are more sensitive to submergence (Panda
et al, 2006; Sarkar et al, 2006). Thus, various important
parameters can be quantified by the measurement of
chlorophyll fluorescence for identifying the tolerant
and intolerant genotypes within 4–6 d of flooding.
Chlorophyll pigments
Chlorophyll is one of the major plant pigments,
supporting the photosynthetic ability. Reduction in
chlorophyll content during flooding is common in rice
(Panda and Sarkar, 2012). Flooding promotes chlorophyll
degradation in sensitive and tolerant genotypes (Sarkar
et al, 2006), and this can be used as an index of
flooding tolerance. Ethylene stimulates the ‘chlorophyllase’
activity and it accelerates chlorophyll loss in the plants
under water. Chlorophyll retention by inhibition of ethylene
synthesis and action might improve photosynthesis
and supply energy reserve needed for maintenance
process (Pradhan et al, 2017). Singh et al (2014) also
indicated that the retention of chlorophyll content in
and after flooding becomes essential for plant survival,
as the same helps photosynthesis under water and
helps in faster recovery on desubmergence. Sarkar et al
(2006) observed that the chlorophyll content in Swarna is
reduced sharply after 10 d of complete flooding
compared to that in Swarna-Sub1. Further blocking
response of ethylene also reduces leaf chlorosis under
flooding (Singh et al, 2014).
Effects of flooding on biochemical
characteristics of rice plants
Carbohydrate
Non-structural carbohydrates are essential for plant
survival under various stresses (Panda and Sarkar,

Rice Science, Vol. 28, No. 1, 2021
2014). Carbohydrate concentration of stem prior to
flooding is considered as a major trait for survival of
rice plant under water (Panda and Sarkar, 2014). The
carbohydrates are consumed while remaining in
flooding state to provide requisite energy for the
plant’s survival (Nagai et al, 2010), and consumption
of carbohydrate by plants under water is essential to
tolerate the consequences of flooding. Rice seedlings
with high level of carbohydrates before flooding
accompanied with less shoot elongation and with the
capacity to retain chlorophyll are the essential traits
for flooding tolerance. Assessment showed that the
tolerant cultivars contain 30%–50% non-structural
carbohydrates than the sensitive ones (Sarkar et al,
2006). The genetic differences in flooding tolerance
need not be related to the carbohydrate status prior to
flooding, instead, it is related to the capability to
sustain higher amount of carbohydrates in flooding
conditions (Ram et al, 2002).
Anaerobic protein
Protein synthesis is known to be severely affected by
flooding. However, certain anaerobic proteins are induced
during flooding, and many of them are enzymes of
carbohydrate metabolism and alcoholic fermentation
(Sarkar et al, 2006). The anoxia tolerant and sensitive
cultivars may differ in level of production and number
of anaerobic polypeptides due to repression of most
aerobic protein synthesis in response to oxygen depletion.
Six of the inducible proteins are recognised to be
cytosolic enzymes in several crops including rice. They
are alcohol dehydrogenase, aldolase, glucose phosphate
isomerase, sucrose synthase, pyruvate decarboxylase
and glyceraldehyde phosphate dehydrogenase (Miro
and Ismail, 2013).
Plant hormone
Plant hormones like ethylene, gibberellic acid,
abscisic acid and their successive manipulations are
considered to have major roles through synergism and
antagonism actions for the survival of plants under
submergence (Huang et al, 2019). The enhanced
elongation growth of deep-water rice varieties is
mediated by the action of ethylene and gibberellic acid
(Jackson and Ram, 2003; Goswami et al, 2017). But
flash flooding tolerance is linked with a reduced level
of ethylene. Elongation of stem is not a desirable
criterion in flooding condition. Thus, synthesis of
gibberellic acid is blocked to reduce elongation and
preserve energy for proper growth. Ethylene enhances
Debabrata PANDA and Jijnasa BARIK. Flooding Stress in Rice
Fig. 2. Production and elimination of reactive oxygen species (ROS)
responding to adverse flood conditions (Ito et al, 1999; Upadhyay, 2018).
SOD, Superoxide dismutase; CAT, Catalase; APX, Ascorbate peroxidase;
GPX, Guaiacol peroxidase; MDAR, Monodehydroascorbate reductase;
DHAR, Dehydroascorbate reductase; GR, Glutathione reductase.
the responsiveness of internode tissue to gibberellic
acid by reducing production of abscisic acid, which is
a potent antagonist of gibberellic acid. The survival
rate is increased many folds when gibberellin biosynthesis
inhibitor is applied before flooding (Pradhan et al, 2013).
Regeneration capacity of rice plants after flooding
The capacity of plants to regenerate after flooding is
vital in order to give high yield. Prompt regeneration
of growth following flooding is an important
characteristic (Panda et al, 2008). Tolerant genotype
showed its ability to regenerate faster by formation of
fresh leaves with higher rate of stability to survive
(Panda et al, 2008). It was found that recovery becomes
faster in tolerant varieties due to efficient ROS
scavenging systems and lower in the process of lipid
peroxidation on being exposed to air (Kawano et al,
2002; Singh et al, 2014).
Effects of flooding on oxidative metabolism
When flood occurs, rice plants are required to adapt to
two extreme environmental conditions, hypoxic condition
when flood takes place and aerobic condition after recession
of flood (Panda and Sarkar, 2013). The damage of
plants suffered by flood is not perceptible in flooding
state but becomes distinct soon after desubmerge (Fig.

49
2). Plants all in a sudden are required to encounter an
environment with higher amount of light intensity and
oxygen completely different from the environment
prevailing under flooding condition. This post
oxygenation causes oxidative stress, resulting in
excess ROS production (Panda et al, 2006; Panda and
Sarkar, 2013). Such produced ROS acts as signalling
molecules inducing the plant to respond to the stresses
(Panda and Sarkar, 2013).
ROS are identified as secondary class of smaller
molecules which acts as a mediator to the responses of
biotic and abiotic stresses including flooding (Fig. 3).
ROS are capable of irreversible cellular damage by protein
oxidation, enzyme inactivation, changes in gene expression,
and decomposition of biological membranes etc. and
results in cell death (Damanik et al, 2012; Jajic et al,
2015). After desubmergence, the submerged rice
plants immediately exposed to an environment with
high light intensity and oxygen tension, resulting in
increased production of ROS and if not detoxified,
may cause acute harm to the cellular organisation and
plant death (Damanik et al, 2010). Genotypes that are
more efficient of mitigating the ROS are more
competent in sustaining plant growth. ROS comprises
of free radical (hydroxyl radical, superoxide anion etc.)
as well as non-radical forms (singlet oxygen, hydrogen
peroxide etc.) (Gill and Tuteja, 2010), which are
formed as byproducts of different metabolic processes
and generated by both enzymatic and non-enzymatic
methods (Sasidharan et al, 2018).
Superoxide radical (O2·ࡃ )
Superoxide radical is produced in different cell
Fig. 3. Reactive oxygen species (ROS) and their action in plants under
flooding stress (Upadhya, 2018).
 50
compartments like peroxisome, chloroplast, apoplast,
mitochondrial electron transport chain and plasma
membrane. The superoxide anion is mainly generated
in chloroplast by Mehler reactions, where oxygen is
reduced by electrons from photosynthetic electron
transport chain under different abiotic stress including
flooding (Jajic et al, 2015; Upadhyay, 2018). Superoxide
anion is short lived, which is moderately reactive free
radical with about 2–4 ms (milli second) of half-life,
and it cannot cross the membrane of chloroplast (Jajic
et al, 2015).
Hydrogen peroxide (H2O2)
Hydrogen peroxide plays a key role in plants under
flooding stress as a signalling molecule, which mediates
between various physiological processes (Fukao et al,
2019). It is also associated with the regulation of senescence
process (Peng et al, 2005), protection against disease
(Kumar et al, 2011), decreasing stress intensity at low
light (Zhang et al, 2011), acclimation to drought (Ishibashi
et al, 2011), and it can change the expression level of
hundreds of genes in plants (Yun et al, 2010). H2O2 is
the steadiest and firm, and less reactive ROS when
compared with other ROS and it can cross the membrane
easily. There are two possible pathways of hydrogen
peroxide production in plants, one is dismutation of
superoxide anion by superoxide dismutase (SOD) and
the other is via oxidases like amino and oxalate
oxidases. The balance between H2O2 scavenging
antioxidant enzymes and SOD in cells is believed to
be important for determination of the steady state level
of H2O2 under flooding (Upadhyay, 2018).
Hydroxyl radical (OH·)
Hydroxyl radical is among the most extremely
reactive ROS. It is produced from H2O2 and O2 in the
Fenton reaction in the neutral pH and ambient
temperature, and these toxic hydroxyl radicals can
penetrate the membrane and leaf mitochondrion (Gill
and Tuteja, 2010; Sharma et al, 2012). Hydroxyl radical
is considered to be responsible for mediating in vivo O2
toxicity in plants after flood water recedes (Upadhyay,
2018). In higher concentration, this super active
hydroxyl radical causes cell death due to lack of
enzymatic reaction for its detoxification under flooding
stress (You and Chan, 2015; Fukao et al, 2019).
Singlet oxygen (1O2)
This is the highly reactive, excited state of molecular
oxygen generated by the reaction of chlorophyll triplet
state and O2. Singlet oxygen is formed when one of

Rice Science, Vol. 28, No. 1, 2021
the unpaired electrons is promoted to a higher energy
orbital (Gill and Tuteja, 2010). Under control conditions,
singlet oxygen is formed during photosynthesis by the
photo activation of photosynthetizers, mostly chlorophylls
and their precursors. It is also produced under different
abiotic stresses including submergence (Springer et al,
2015; Upadhyay, 2018). It has dual role as oxidising
agent and signalling molecule under flooding stress
(Kim et al, 2008; Fukao et al, 2019). Since it is highly
reactive and their life span being short by about 3.1 to
3.9 s in pure water, it is able to interact with molecules
present mainly in its nearest environment (Jajic et al, 2015).
Lipid peroxidation under flooding stress
Lipid peroxidation produces malondialdehyde (MDA).
The amount of MDA indicates the extent of damage
resulted by ROS in the tissue (Damanik et al, 2012).
This is formed due to oxidative stress which induces
membrane damage. Significant negative co-relation
has been found between formation of MDA with
ascorbate concentration and survival rate in rice plants
(Kawano et al, 2002). Damanik et al (2012) indicated
the significant difference between MDA content in
rice seedlings under anoxic and aerobic conditions,
and the extended duration of anaerobic conditions
increased the lipid peroxidation. MDA content is
significantly higher in sensitive cultivars during flooding
and after exposure to air (Panda and Sarkar, 2013).
Effects of flooding on antioxidants defence
mechanism
Plant cells are able to neutralise the ROS with
efficient oxygen-scavenging machinery consisting of
several antioxidants (both non-enzymatic and enzymatic)
(Gill and Tuteja, 2010). Thus, induction of antioxidant
defense mechanism is essential for protection of plants
against different stresses. SOD, catalase (CAT), guaiacol
peroxidase (GPX), ascorbate peroxidase (APX), glutathione
reductase (GR), dehydroascorbate reductase (DHAR)
and monodehydroascorbate reductase (MDAR) are the
predominant antioxidant enzymes, which play important
roles in protecting plants from oxidative damages
(Upadhyay, 2018).
Enzymatic antioxidants
SOD
In aerobic organisms, SOD defends and protects from
being damaged by oxidative reactions. This is the first
antioxidant enzyme in the process of detoxification,
transforms superoxide anion to hydrogen peroxide
Debabrata PANDA and Jijnasa BARIK. Flooding Stress in Rice
(Gill and Tuteja, 2010; Steffens et al, 2013). SOD is
one form of metalloenzymes and it removes superoxide
anion by disproportionating it to hydrogen peroxide
and oxygen (Sharma et al, 2012). This enzyme exists
in three isoforms (Damanik et al, 2012; You and Chan,
2015). SOD activity is increased under stress condition
including flooding stress (Mishra et al, 2011). Over
production of SOD increases the plant’s tolerance to
oxidative stress (Sharma et al, 2012). Damanik et al
(2012) studied two Malaysian rice mutants along with
FR13A and observed the significant increase in SOD
activity in response to flooding.
CAT
CAT is identified as the first enzyme among all other
antioxidant enzymes (Sharma et al, 2012). CAT carries
out dismutation of hydrogen peroxide into H2O and
O2 (Gill and Tuteja, 2010; Sharma et al, 2012). CATs
possess high turnover rate, and each molecule of it
converts six million of hydrogen peroxide into water
and oxygen in 1 min (Gill and Tuteja, 2010). The
major production site of H2O2 is peroxisomes. The
actions of SOD and CAT when both taken together
become critical in elevating the adverse impact of
oxidative damage in plants. Damanik et al (2010)
observed that the CAT activity was 9-fold higher
under flooding as compared to control plants in case
of FR13A.
GPX
GPX is another antioxidant enzyme containing heme,
which prefers to oxidise guaiacol and pyragallol
instead of H2O2 (Gill and Tuteja, 2010). The enzymes
contain two structural Ca2+ and four disulfide bridges.
Various isoenzymes of GPX are located in different
tissues such as cell wall and cytosol. It plays vital
roles in various plant processes like decomposition of
hormone IAA, lignifications of cell wall, formation of
ethylene and lends protection against various stresses
including flooding (Gill and Tuteja, 2010; Upadhyay,
2018). Generally, it is known as stress enzyme due to
its effective quenching ability of harmful free radical
in stress condition (Sharma et al, 2012). GPX activity
in rice is increased under different duration of
flooding (Panda and Sarkar, 2012)
Ascorbate glutathione cycle
The Asada-Halliwell pathway or ascorbate glutathione
cycle plays an important role in scavenging ROS
during and after flooding (Damanik et al, 2012). This
pathway regenerates the reduced ascorbate and glutathione
as primary scavengers under flooding stress (Sarkar et al,

51
2006; Damanik et al, 2010). Glutathione reductase is a
key factor associated with recycles of reduced
glutathione, maintaining glutathione level in the cell,
resulting in supplying the total amount of glutathione
to mitigate the damages caused by submergence
(Panda and Sarkar, 2013). The ascorbate glutathione
cycle includes sequential reduction and oxidation of
ascorbic acid, glutathione and nicotinamide adenine
dinucleotide phosphate (NADPH) which is catalysed
by four antioxidant enzymes i.e. APX, MPAR, DHAR
and GR. This Asada-Halliwell pathway is located in
sub-cellular sites like cytosol, chloroplast, mitochondria
and peroxisome regulate flooding tolerance (Sharma
et al, 2012; Upadhyay, 2018).
Ascorbate peroxidase (APX)
APX is a major antioxidant enzyme of Asada-
Halliwell pathway and takes the lead role in reduction
of enhanced ROS level during and after flooding
stress (Sarkar et al, 2006; Upadhyay, 2018). APX
catalyses detoxification of hydrogen peroxide by
reducing H2O2 to H2O utilising the reducing power of
ascorbate (Damanik et al, 2010, 2012; Smirnoff,
2018). It is mostly found in both cytosol and
chloroplast, and is an important enzyme of glutathione-
ascorbate cycle, as its removes peroxides by transforming
ascorbic acid into dehydroascorbate (Damanik et al,
2010; Gill and Tuteja, 2010). This enzyme has a
greater affinity towards hydrogen peroxide compared
to catalase, making the enzyme an efficient scavenger
of H2O2 under flooding stress (Sarkar et al, 2006;
Upadhyay, 2018).
Monodehydroascorbate reductase (MDAR)
MDAR catalyses the ascorbic acid regeneration from
monodehydroascorbate radical using of NADPH. This
enzyme exists as cytosolic and chloroplast isozymes
(Gill and Tuteja, 2010; Suzuki et al, 2012). Presence
of this enzyme in peroxisome and mitochondria
decomposes the hydrogen peroxide present therein.
The ‘monodehydroascorbate’ is used as a substrate
only by this enzyme and the enzyme activity is
widespread in plants. The increased MDAR activity
was observed in several studies relating to the plants
subjected to different kinds of stresses including
flooding (Sharma et al, 2012).
DHAR
DHAR carries out the ‘dehydroascorbate’ reduction to
ascorbic acid by using the reducing agent glutathione
(Upadhyay, 2018). DHAR has a key role in maintaining
reduced ascorbate level. Dehydroascorbate has short
 52
life span and it can be either hydrolyse 2,3-
diketogluconic acid or convert to ascorbate via DHAR
(Sharma et al, 2012). Abiotic stresses such as flooding,
drought, chilling and salinity increase the DHAR
activity in plants (Chang et al, 2017).
GR
GR is dependent on NADPH, and it reduces the
oxidised glutathione (GSSG) to its reduced form
(GSH), thus maintaining high GSH/GSSG ratio (Rao
and Reddy, 2008). It is a flavo-protein and has an
essential disulfide group. Although GR is present in
other cell organelles as well as cytosol, its maximum
enzyme activity is its chloroplastic isoforms. Reduced
glutathione and GR in chloroplast together decomposes
hydrogen peroxide produced through Mehler reaction
(Sharma et al, 2012) and plays an important role in
combating abiotic stress in plants (Gill and Tuteja,
2010). The increased GR activity in plants under
stress has been studied by several workers. Damanik
et al (2010) revealed that the activity of GR serves as
a criterion for evaluation of flooding tolerance in rice.
GR sensitivity has been observed in tolerant rice varieties
Panikekoa and T1471 under flooding (Das et al, 2004).
Non-enzymatic antioxidants
Potent non-enzymatic antioxidants help in mitigation
of oxdative stress within the cell (Gill and Tuteja,
2010). These non-enzymatic antioxidants interact with
various cellular components and in addition to their
major roles in providing defense and as cofactors of
enzyme, they also involved in various processes
relating to plant growth and development (Sharma et al,
2012; You and Chan, 2015).
Ascorbic acid
Ascorbic acid is a powerful antioxidant with low
molecular weight and is abundantly available. It plays
a vital role in giving protection to plants from being
adversely affected by oxidative stress caused due to
submergence or flooding (Panda and Sarkar, 2013;
Steffens et al, 2013). It performs many important roles
in plant physiological processes, and is found in all
plant tissues, most abundantly in the tissues which
carry out photosynthesis. It is considered as a
powerful antioxidant because of its power to supply
electrons in various reactions catalyzed by both
enzymatic and non-enzymatic methods (Gill and
Tuteja, 2010). The ascorbate metabolism mostly takes
part inside mitochondria, which not only synthesise it
but also regenerates it from its oxidised form. Studies

Rice Science, Vol. 28, No. 1, 2021
showed that higher levels of reduced ascorbate play an
important role in creating defense to the plants to fight
against damage caused due to flooding (Kawano et al,
2002; Das et al, 2004). Over expression of enzymes
linked with ascorbic acid biosynthesis supports plant’s
endurance ability to overcome stress (Sharma et al,
2012). It reacts with ROS in both the photosystems I
and II through ASC-GSH as well as xanthophyll cycle
during flooding (Damanik et al, 2010).
Glutathione
Glutathione is a non-protein thiol which provides
intracellular protection against ROS-induced oxidative
damages (Gill and Tuteja, 2010; Sharma et al, 2012),
and located in all cell compartments, for example,
endoplasmic reticulum, vacuole, mitochondria, peroxisomes,
chloroplast, cytosol and apoplast. Because of its reducing
power glutathione plays crucial roles in diverse
physiological processes like regulation of sulfate
transport, cell growth and cell division, pathogen resistance,
conjugation of metabolites, signal transduction, enzymatic
regulation, formation, of protein and nucleic acid,
synthesis of phytochelatin, decomposition of xenobiotics
as well as the expression of stress responsive genes
(Mullineaux and Rausch, 2005). Glutathione acts as a
potential scavenger of harmful free radicals like hydrogen
peroxide, singlet oxygen and hydroxyl radical. It plays
an important role in antioxidant defense system by
regenerating ascorbic acid by ASC-GSH cycle (Gill
and Tuteja, 2010).
Genetic mechanism of flooding tolerance in rice
The genetic basis of flooding tolerance remained
ambiguous until the mid of 1990 (Bailey-Serres et al,
2010). Genetic mechanism of rice for tolerant to
flooding/submergence is shown in Fig. 4. ‘Quiescence’
and ‘Elongation’ are two major and opposite strategies
to encounter the effects of flooding during flash and
deep-water flooding, respectively (Fig. 4). The
‘escape’ type of adaptive response is mostly seen in
deep-water lowland rice, where vigorous shoot
elongation prevents the plant from complete flooding
as flood water level gradually increases. Two ethylene
responsive factor genes SNORKEL1 (SK1) and
SNORKEL2 (SK2) cloned from a deep-water rice
variety C9285 involve in the elongation of shoot in
these floating rice (Vergara et al, 2014; Tamang and
Fukao, 2015). Both the genes function in contradiction
with Submergence1A-1 (Sub1A-1). The Sub1A-1
empowers the plant to endure flash flooding by
Debabrata PANDA and Jijnasa BARIK. Flooding Stress in Rice
limited elongation until flooding water recedes (Xu et
al, 2006; Bailey-Serres et al, 2010). The tolerant rice
genotype adapted to flash flooding condition is
characterised by slow elongation growth and thus
conserves accumulated respirable biomass to resume
growth after the de-submergence (Bailey-Serres et al,
2012; Azarin et al, 2017). Allelic surveys showed that
SNORKEL (SK) genes are present only in deep-water
rice varieties which exhibit rapid internode elongation
in response to submergence (Tamang and Fukao,
2015). SKs promote the accumulation of bioactive
gibberellic acid in submerged internodes (Ayano et al,
2014). During flooding, ethylene accumulates,
triggering SK1/2 gene expression in C9285 (Minami et
al, 2018). It has been observed that ethylene increases
biosynthesis of gibberellic acid under flooding,
triggering the internode elongation in deep-water rice
(Fukao and Bailey-Serres, 2008). Kuroha et al (2018)
identified a rare allele of gibberellin biosynthesis gene
SD1 (SEMIDWARF1) which provides adaptation to
deep water. The SD1 protein directs increased
gibberellin synthesis, largely GA4, thus resulting in
internode elongation.
It is reported that a few anaerobic germination
tolerance QTLs were found on chromosomes 5 and 11
(Jiang et al, 2004). Five different QTLs were found on
chromosomes 1, 3, 7 and 9 identified by crossing IR64
with the tolerant donor Khao All Hlan On (Jiang et al,
2004; Angaji et al, 2010). The QTL qAG-9-2 or AG1
linked with local modulation of trehalose-6-phosphate
(T6P) is the most promising locus for breeding

53
Fig. 4. Genetic mechanism of rice for
tolerant to flooding/submergence.
A, Sub1A mediated response of submergence
tolerance in rice.
B, SNORKEL (SK) mediated escape
response for tolerance to deep-water
flooding in rice (Tamang and Fukao,
2015).
(Kretzschmar et al, 2015). T6P
(OsTPP7) gene is involved
in metabolism of T6P (Loreti
et al, 2018). In a population
obtained by crossing sensitive
line IR42 with tolerant variety
Ma-Zhan Red (Septiningsih
et al, 2013), another anaerobic
germination tolerance QTL
(qAG-7-1 or AG2) is found on
shoot arm of Chromosome
7. Zhang et al (2017) revealed
that LOC_Os06g03520 is linked
with anaerobic germination tolerance using 5291 single
nucleotide polymorphism markers.
Many studies considered Sub1 as a typical quantitative
trait. This QTL is identified from the rice landrace
FR13A on chromosome 9 through molecular mapping.
(Ram et al, 2002) indicated that 70% phenotypic
variance in flooding tolerance comes from this QTL
and the rest 30% were contributed by other secondary
minor QTLs located on chromosomes 1, 2, 5, 7, 10
and 11. At the molecular level, the Sub1 locus contains 3
ethylene responsive factors (ERFs) Sub1A, Sub1B and
Sub1C, which are upregulated under flooding (Xu et al,
2006). Sub1B and Sub1C are found in all rice varieties.
Sub1A is only responsible for transient flooding and it
is found at Sub1 QTL in FR13A and other tolerant
genotypes (Bailey-Serres et al, 2010). Xu et al (2000)
futher carried out fine mapping of Sub1 gene with
3000 F2 indivisual to a 0.16-cM region on Chromosome
9. Sub1A is recognised in two allele forms: Sub1A-1 is
located only in tolerant genotype and Sub1A-2 is
found in sensitive ones (Xu et al, 2006; Septiningsih
et al, 2012). Both the alleles encode similar proteins,
with the exception of Ser186 in Sub1A-1 and Pro186 in
Sub1A-2 alleles (Xu et al, 2006; Bailey-Serres et al,
2010). Sub1 controls shoot elongation, conserves reserve
carbohydrates, retains chlorophyll content etc., and as
a result, it controls plant survival and resumes tolerant
genotype growth when water recedes (Pucciariello and
Perata, 2013). Rice varieties having flooding inducible
Sub1A restricts/limits ethylene production which maintains
 54
mRNA and protein accumulation of gibberellic acid
signalling repressors during flooding (Singh et al,
2014; Fukao et al, 2019). The inhibition of gibberellic
acid responsiveness mediated by Sub1A represses genes
needed for catabolism of sucrose and starch, thus restricting
elongation growth in order to conserve precious
carbohydrates for its survival under water and recovery.
This quiescence is being very useful in plant breeding
(Neeraja et al, 2007; Septiningsih et al, 2013).
Marker-assisted breeding (MAB) approaches
for flooding tolerance in rice
The success of fine mapping of Sub1 from FR13A, a
sound QTL has enabled the MAB of modern rice
cultivars having the ability to withstand flash flooding
(Bailey-Serres et al, 2010). The polygenic QTL Sub1A
was introduced to eight rice cultivars using marker-
assisted backcrossing. International Rice Research
Institute has successfully introgressed Sub1 QTL into
high-yielding variety Swarna, which is presently
adapted in many states of India (Singh et al, 2013; Dar
et al, 2017). Swarna-Sub1 can endure two weeks of
flash flooding and recovers well after receding of
flood water. It was released in India in 2009 after few
years of successful evaluation in the farmers’ fields.
At the same time, Sub1 was also introgressed into
mega rice varieties grown in rainfed ecology in South
and South-East Asian countries (Neeraja et al, 2007).
The Sub1 incorporated popular varieties such as
IR64-Sub1, Samba Mahsuri-Sub1, Thadokkam 1-Sub1,
BR11-Sub1, BINA Dhan 11 and CR1009-Sub1 have
been recognized for enhancing rice productivity in
less favourable lowland ecosystem of eastern India,
Nepal and Bangladesh (Ismail et al, 2013; Ahmed et al,
2016). Singh et al (2016) also reported the transfer of
Sub1 to 10 highly popular regionally adapted Indian
rice varieties. The original and Sub1 introgressed
varieties show no apparent morphological variation in
numerous trials in farmer’s field (Singh et al, 2017).
Breeding for higher germination potential, increased
seedling strength and flooding tolerant rice varieties
during the period of seed germination was studied in
the past (Yamauchi et al, 1993, 1994; Biswas and
Yamauchi, 1997), but left with limited results. This is
due to lack of suitable genotypes with enhanced
tolerance, less available information on genetics of
anaerobic germination tolerance and non-availability
of clear simplified mechanism (Jiang et al, 2004).
Molecular breeding for AG tolerance in rice was
started using qAG9-2 (AG1) detected from Khao Hlan
On (Kretzschmar et al, 2015; Kato et al, 2019). AGL

Rice Science, Vol. 28, No. 1, 2021
was introgressed into some improved varieties or their
Sub1 derivatives such as IR64, IR64-Sub1, PSB
Rc18-Sub1 and PSB Rc82 (Septiningsih et al, 2013).
The Chinese traditional rice cultivar Ma-Zhan Red
was used as a donor in incorporation of qAG7-1 (AG2)
locus to mega varieties in several studies. Depending
upon the population of donor, the transfer of AG1 is
monitored by two or three markers (Azarin et al, 2017;
Kato et al, 2019).
Prospects
Although remarkable progress has been achieved
through marker assisted selection, we still have several
critical problems to overcome in molecular breeding
of flooding tolerant plants. Our understanding of the
whole-plant stress response mechanism is very limited.
Therefore, we need to investigate stress responses in
differentiated cell, tissues and organs and to connect
the data relevantly. Also, there is a need of finding
superior alleles of Sub1A/AG1/AG2 gene or other
novel genes which may provide better protection
against different types of flooding stress. The
summarized information of this review will facilitate
further investigations of signalling mechanisms of
flooding tolerance in rice and development of flooding
tolerant rice genotypes.
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