Journal of Experimental Botany, Vol. 62, No. 1, pp. 39–57, 2011
doi:10.1093/jxb/erq271 Advance Access publication 16 September, 2010
REVIEW PAPER
Ion transport in seminal and adventitious roots of cereals
during O2 deficiency
Timothy David Colmer* and Hank Greenway
School of Plant Biology, Faculty of Natural and Agricultural Sciences, The University of Western Australia, 35 Stirling Highway, Crawley,
WA 6009, Australia
* To whom correspondence should be addressed: E-mail: [email protected]
Received 14 June 2010; Revised 5 August 2010; Accepted 12 August 2010
Abstract
O2 deficiency during soil waterlogging inhibits respiration in roots, resulting in severe energy deficits. Decreased
root-to-shoot ratio and suboptimal functioning of the roots, result in nutrient deficiencies in the shoots. In N2flushed nutrient solutions, wheat seminal roots cease growth, while newly formed adventitious roots develop
aerenchyma, and grow, albeit to a restricted length. When reliant on an internal O2 supply from the shoot, nutrient
uptake by adventitious roots was inhibited less than in seminal roots. Epidermal and cortical cells are likely to
receive sufficient O2 for oxidative phosphorylation and ion transport. By contrast, stelar hypoxia–anoxia can develop
so that H+-ATPases in the xylem parenchyma would be inhibited; the diminished H+ gradients and depolarized
membranes inhibit secondary energy-dependent ion transport and channel conductances. Thus, the presence of
two transport steps, one in the epidermis and cortex to accumulate ions from the solution and another in the stele to
load ions into the xylem, is important for understanding the inhibitory effects of root zone hypoxia on nutrient
acquisition and xylem transport, as well as the regulation of delivery to the shoots of unwanted ions, such as Na+.
Improvement of waterlogging tolerance in wheat will require an increased capacity for root growth, and more
efficient root functioning, when in anaerobic media.
Key words: Aerenchyma, anoxia, Hordeum vulgare, hypoxic stele, hypoxia, ion transport, soil waterlogging, Triticum aestivum,
xylem, Zea mays.
Introduction
Waterlogged soils are usually anaerobic (Ponnamperuma,
1984). When water displaces gases in the soil, the inward
flux of atmospheric O2 is inhibited by approximately
320 000 times, owing to the 104-fold slower diffusivity and
the 32-fold lower solubility of O2 in water than in air
(Armstrong and Drew, 2002). The rate of soil O2 depletion
following waterlogging depends upon the metabolic activity
of microorganisms and plant roots, so O2 depletion will be
more rapid at warmer temperatures and with high organic
matter in the soil (Drew, 1992). When O2 is lacking,
oxidative phosphorylation ceases, causing severe energy
deficits with adverse effects on growth and nutrient uptake,
and even resulting in root death. Following the onset of
anoxia, root cells of many plant species begin to die within
a few hours, or at most days (Gibbs and Greenway, 2003).
In addition to O2 deficiency, and depending on soil
type, waterlogging can also pose other challenges to roots;
Mn2+, Fe2+, S2–, carboxylic acids, CO2 and ethylene
(C2H4) can accumulate owing to soil microbial activity
(Ponnamperuma, 1984). In submerged tissues, endogenous
Abbreviations: COPR, critical O2 pressure for respiration; COPE, critical O2 pressure for root extension; Lp, hydraulic conductivity; Lpc, hydraulic conductivity of root
cortical cells; Lpr, hydraulic conductivity across roots, to the xylem; NORC, non-selective outward-rectifying channel; ROL, radial O2 loss; SKOR channel, selective K+
outward-rectifying channel.
Definitions: Anoxia, zero O2. Hypoxia, O2 below the critical O2 pressure for respiration (COPR), but not zero. Anaerobiosis, strictly defined as a condition without O2, but
has been used to describe N2-flushed solutions in which O2 would be close to zero, but are not sure to be anoxic. Hypoxic–anoxic stele, stele which receives
inadequate O2 for oxidative phosphorylation, but with some uncertainties that the entire stele is truly anoxic (Darwent et al., 2003).
ª The Author [2010]. Published by Oxford University Press [on behalf of the Society for Experimental Biology]. All rights reserved.
For Permissions, please e-mail: [email protected]
40 | Colmer and Greenway
ethylene typically increases to between 0.1 and 1 Pa, partial
pressures at which maximum physiological effects are often
elicited (Jackson, 2008). In waterlogged soils CO2 can
increase to 5–40 kPa, with adverse effects on roots of some
species (Greenway et al., 2006). ‘Soil toxins’ can affect ion
transporters, however, this aspect is not discussed further
here (for a recent review see Pang and Shabala, 2010).
In addition, there may be adverse interaction between
waterlogging and other chemical constraints such as salinity
(Barrett-Lennard, 2003) and high Al (Setter et al., 2009).
Nevertheless, no such complexities were found in a series
of experiments on wheat (Triticum aestivum L.) lasting for
12 d: the response by wheat to anaerobic (i.e. N2-flushed)
solutions (Trought and Drew, 1980a) simulated, to a large
extent, the response to soil waterlogging in a pot experiment
(14 C, sandy soil with 3.3% organic matter; Trought and
Drew, 1980c). The similarities included slower relative
growth rates of roots and shoots, dramatically reduced
nutrient transport to the shoots resulting in nutrient
deficiencies, and premature leaf senescence (Trought and
Drew, 1980a, b, c). Similar to wheat, barley (Hordeum
vulgare L.) also soon became nutrient deficient upon soil
waterlogging (Drew and Sisworo, 1979).
Roots in anoxic media depend on internal gas-phase
diffusion of O2 from the shoots, via aerenchyma, to
enable respiration (Armstrong, 1979). Aerenchyma provides
large interconnected gas channels that greatly enhance
O2 movement from the shoots into, and along, roots. The
importance of aerenchyma to enable respiration in the roots
of maize (Zea mays L.) seedlings when in an anoxic medium
was demonstrated by the adenylate energy charges in the
5 mm root tips of intact plants with roots in an anoxic
medium. The energy charges were 0.7 and 0.4 in roots with
13% and 4% gas-filled porosity, respectively, compared with
0.9 when the roots were in air (Drew et al., 1985). A gasfilled porosity of 13% represents relatively modest aerenchyma formation; this value for maize compares with root
porosity of up to 41% in rice (Oryza sativa L.) (Colmer,
2003a). Even when there is some O2 in the root medium (i.e.
hypoxia¼suboptimal, but not zero O2), longitudinal gasphase O2 diffusion within roots is an important component
of O2 supply (Armstrong and Beckett, 1987).
For roots receiving O2 via aerenchyma, the internal
concentration will decrease in a curvilinear gradient with
distance down the roots, since movement occurs via diffusion
and O2 is consumed along the diffusion pathway (Armstrong,
1979). Radial gradients in O2 also occur, the steepness of
these gradients depends upon resistances to O2 diffusion and
O2 consumption rates. So, apical regions and the stele of
roots may receive a suboptimal supply of O2, since these
tissues are typically of low gas-filled porosity, both have
relatively high O2 demands, and occur at the ends of
longitudinal (gas phase) and radial (predominantly liquid
phase) diffusion paths (Armstrong, 1979; Armstrong and
Beckett, 1987; Armstrong et al., 1991; Gibbs et al., 1998a).
The focus of this review is on growth and function of
seminal and adventitious roots of wheat in O2-deficient
media, but data for maize (Zea mays L.) and barley
(Hordeum vulgare L.) are included for topics not yet wellstudied in wheat, and some comparisons are also made with
rice (Oryza sativa L.). Inhibition of nutrient uptake in
O2-deficient roots received early attention as a major
component of waterlogging damage in wheat, at least for
the first 12 d (key papers by Trought and Drew, 1980a,
1981) and other crops, including barley and maize (reviewed
by Drew, 1988). Subsequent work, as reviewed here, has
also identified poor root functioning as a key limitation for
waterlogging tolerance in dryland crops (wheat, barley, and
maize). An overview of the responses of wheat to soil
waterlogging and associated root zone hypoxia–anoxia is
given in Fig. 1. The present review highlights the links
between root O2 supply, energy metabolism, and energydependent ion transport, particularly the inhibition of xylem
loading when the stele becomes anoxic, or at least ‘severely
hypoxic’, the preferred term by Darwent et al. (2003), based
on their detailed radial O2 profiles in maize roots. Broader
information on waterlogging tolerance in wheat (and barley)
is available in the review by Setter and Waters (2003).
Recent reviews, that complement the focus here, have
discussed the influence of waterlogging on soil and plant
parameters of a nutrient uptake model (Elzenga and van
Veen, 2010) or have dealt more broadly with the adverse
effects of, and plant adaptations to, soil waterlogging and
complete submergence of the shoots (Bailey-Serres and
Voesenek, 2008; Colmer and Voesenek, 2009).
Techniques for studying plant responses to
hypoxia using nutrient solutions
Most studies of the function of roots in O2-deficient
conditions have used nutrient solutions, as O2 and nutrient
levels can be controlled and measured, and roots can be
accessed and studied, more easily than in soils. ‘Anaerobic’
treatment was commonly imposed by flushing a nutrient
solution with N2 gas, but in many studies anaerobiosis
(i.e. zero O2) was not confirmed (Trought and Drew,
1980a). These solutions might still have contained some
O2 (dissolved O2 was 0.003 mM in N2-flushed solutions in
experiments by Kuiper et al., 1994); O2 supply to roots from
N2-flushed solutions will depend upon the purity of the N2
gas, pot design, and the flushing rates used. Even when
O2 concentration in the external solution is known, the O2
status of root tissues can still be uncertain, since internal O2
will depend upon respiration rates, turbulence as determined by rates of bubbling or stirring, pot size and design,
root density in the solution, root diameters, diffusion
resistances across unstirred boundary layers of various sizes
and across tissues of various diameters. As these factors
differ amongst studies and species, comparisons of results
can be difficult across experiments.
An alternative to continuous N2 flushing is the use of
nutrient solutions containing 0.1% (w/v) agar, which can be
de-oxygenated by N2 flushing prior to use. The dilute agar
prevents convective movements in the solution so this
stagnant solution better mimics the changes in gas
Ion transport in hypoxic roots | 41
Soil waterlogging
Death of distal portions
of seminal roots
Anoxia
Growth of seminal
roots ceases
Decreased root:shoot ratio
Increased ethylene
Altered balance
of hormones
(ethylene, auxin,
cytokinins, etc.)
Uppermost part of seminal roots receive
some O2 internally and survive
Solute loss and
entry of soil toxins
Small nutrient uptake
Leaf senescence
Leaf necrosis
Increased number of adventitious roots
Aerenchyma forms, but relatively low porosity,
lack of ROL barrier, hexagonal cell packing,
large stele, impede O2 diffusion to stele and apex
Short adventitious roots - growth
restricted by O2 diffusion to apex
Root:shoot ratio only partially restored
Some altered hormonal
balance persists
Moderate nutrient uptake
Shoots remain small
Death
Very little
shoot growth
Partial mitigation of
nutrient deficiency
Grain yield depends also
on recovery of roots and shoots
when water recedes
Fig. 1. Simplified overview of the responses to soil waterlogging of wheat seminal and adventitious roots, and resulting restrictions for
shoot growth. Intolerance of roots to anoxia, and the poor capacity for internal O2 transport impacts severely on seminal roots. Although
adventitious root numbers can increase, and these form more aerenchyma than seminal roots, suboptimal characteristics related to O2
transport result in modest growth and suboptimal functioning in waterlogged soils. ROL, radial O2 loss.
composition found in waterlogged soils; i.e. increases in
CO2 and ethylene in addition to low O2 in the continuously
N2-flushed solutions (Wiengweera et al., 1997). The advantage of using stagnant agar solution is that accumulation of
ethylene enhances the development of aerenchyma
(Shiono et al., 2008), so that roots of wheat in stagnant
agar solution were of higher porosity than those in N2flushed solution (Wiengweera et al., 1997). On the
other hand, the N2-flushed treatment can have the advantage, depending on the experimental objective, of only
changing one factor (i.e. O2 concentration) from the
condition in aerated controls. An overview of responses, of
seminal and adventitious roots of wheat to these two
methods of imposing O2 deficiency in the root zone, is given
in Fig. 2.
Adverse effects of O2 deficiency on growth
and mineral nutrient status
Declines in shoot growth of plants in anaerobic root media
may be caused by: (i) reduced development of roots, causing
a suboptimal root-to-shoot ratio (root:shoot), and/or (ii)
impaired functioning of roots. Decreased root:shoot may
lead to an altered balance of phytohormones that might
slow shoot growth and stimulate adventitious root growth,
keeping root:shoot within certain limits (Drew, 1983).
In addition, the combination of a reduced size of the root
system and suboptimal root functioning would compromise
the provision of nutrients, and perhaps water, to the
shoots. The reduced shoot and root growth is unlikely to
be associated with a shortage of sugars for catabolism, or
for incorporation into biomass, because during root
zone O2 deficiency sugars accumulate in tissues as growth
is inhibited relatively more than photosynthesis (Setter
et al., 1987). In wheat with roots in N2-flushed solution
for 12–14 d, sugars increased by 20–30% in shoots and by
55–70% in roots (Barrett-Lennard et al., 1988). Sugar
accumulation was also reported in roots of barley when
growth was inhibited by low O2 supply (Limpinuntana and
Greenway, 1979). In view of these large increases in
sugar concentrations, ethanol-insoluble dry weight (i.e.
structural biomass with soluble sugars removed) rather than
total dry weight has been used by some authors to
assess growth (Barrett-Lennard et al., 1988). Thus, respiration in roots in N2-flushed solution would be inhibited by
low O2 availability (see below), rather than shortage of
substrate.
Responses of wheat to anaerobic nutrient solution at the
early tillering stage, from the key study by Trought and
Drew (1980a), are provided in Tables 1 and 2, and these are
discussed in more detail below.
O2 deficiency decreases root-to-shoot ratio and has
more severe effects on seminal, than on adventitious,
root growth
In the key study on wheat at the early tillering stage (Trought
and Drew, 1980a), ratios of root-to-shoot (root:shoot,
dry weight basis) were 0.57 and 0.18 in aerated and N2flushed nutrient solutions, respectively (Table 1). Other
42 | Colmer and Greenway
Adventitious roots
(moderate aerenchyma)
Seminal roots
(none or little aerenchyma)
N2 flushed (turbulent,
low O2 solution)
Stagnant agar solution
(deoxygenated)
In both cases, aerenchyma forms, but more in stagnant solution
(Enhanced by entrapped ethylene)
Stagnant agar solution
(deoxygenated)
N2 flushed (turbulent,
low O2 solution)
Epidermis and cortex
receive some O2 from
solution, stele severely
hypoxic/anoxic
Low O2
inhibits growth
No O2 from
external solution
Death of
root tips
Nutrient uptake possible
in epidermis, but xylem
loading inhibited
Short roots
Only most basal few
cm receive some O2
via internal diffusion
from shoot base
Decreased capacity for nutrient
uptake and inhibited delivery to shoots
Shoot nutrient deficiency
Inhibition of shoot growth
Root growth ceases at length supported by internal O2
diffusion, distal portion of stele and apex severely hypoxic, but
survive
Aerobic epidermis and cortex, but stele
hypoxic/anoxic in distal portions
Nutrient uptake continues into roots, but active xylem
loading almost abolished, so ion movement to xylem
depends upon the gradient from the ‘aerobic cortex’
Most O2 derived from ROL
is ‘stripped away’
Stelar hypoxia-anoxia
possibly more prevalent
Aerobic ‘rhizosphere’
develops where ROL is high
Greater internal O2 supply reduces stelar
hypoxia-anoxia in larger portion of root
In both cases, adventitious root functioning is superior to that of
seminal roots, although diminished nutrient delivery via xylem
to shoots is still significant
Moderate shoot growth
Fig. 2. Oxygen status and associated impacts on ion transport processes in seminal and adventitious roots of wheat when in two
systems used to impose O2 deficits in nutrient solution experiments. N2-flushed solutions contain very low O2 and turbulence reduces
boundary layers so facilitates any O2 uptake from the solution, whereas 0.1% agar solutions are stagnant and can be deoxygenated prior
to use. As ethylene accumulates in stagnant agar solution, this method is more analogous to the changes in gas composition in
waterlogged soils. The contrasting responses of seminal and adventitious roots (low and moderate aerenchyma, respectively) and
treatment methods (N2-flushed versus stagnant agar) are described. ROL, radial O2 loss.
experiments on wheat (e.g. Kuiper et al., 1994) also found
similar decreases in root:shoot, despite large differences in
control plant growth and inhibitory effects of the N2
treatment on shoot relative growth rates between these
experiments conducted at different temperatures (see footnote of Table 1). Root:shoot also decreased, albeit to
a smaller extent, when stagnant agar was used (Watkin
et al., 1998) rather than N2 flushing to impose the
‘anaerobic’ treatment.
These decreases in root:shoot are mainly due to the
cessation of seminal root growth. By contrast, adventitious
roots emerge (also termed nodal or crown roots) which at
least partially replace the damaged seminal roots (e.g. for
wheat, Trought and Drew, 1980a; for other species see
reviews by Drew, 1983; Jackson and Drew, 1984). These
adventitious roots typically develop more aerenchyma than
the pre-existing seminal roots, and so have an improved
capacity for internal O2 supply (discussed below). So, after
periods of anaerobiosis lasting longer than 2 weeks, the
adventitious root system is the largest fraction of roots for
wheat (Trought and Drew, 1980a; Barrett-Lennard et al.,
1988). However, these roots cannot grow to their full
lengths, so the increased numbers of adventitious roots,
from 7.2 to 11 at 14 d of exposure to N2-flushed solution,
only partially compensated for the reduced rates of
elongation; the relative growth rate (ethanol-insoluble dry
weight basis) of adventitious roots was 0.173 d1 in N2-
flushed solution compared with 0.188 d1 in aerated
solution (Barrett-Lennard et al., 1988). Increases in numbers of adventitious roots during hypoxia are usually found
and sometimes can be quite large; for example, in a tolerant
barley cultivar adventitious root number increased from 14
in drained soil to 33 in waterlogged soil (Pang et al., 2007).
As examples of reduced root lengths, the main axes of
adventitious roots of wheat in N2-flushed nutrient solutions
only grew to 9 cm (Barrett-Lennard et al., 1988) or 12 cm
(Trought and Drew, 1980a); extension ceases when roots
reach a length beyond which internal O2 diffusion cannot
sustain growth (Armstrong, 1979). Although growth ceased,
the tips of the adventitious roots remained alive as shown
by resumed extension upon re-aeration (Trought and Drew,
1980a; Barrett-Lennard et al., 1988).
Evidence for nutrient deficiency in shoots of wheat with
O2-deficient roots
A persuasive case for nutrient deficiency in shoots of wheat,
when grown with roots in N2-flushed nutrient solution, was
made in the studies by Trought and Drew (1980a). After 14 d
in the N2-flushed treatment, uptake of all macronutrients
was reduced (Table 2). Decreases in nutrient concentrations
in the shoots can be quite severe, as one example the N
concentration (mmol g1 dry weight) was reduced to 24% of
that in shoots of the aerated control (Trought and Drew,
Ion transport in hypoxic roots | 43
Table 1. Responses of wheat to N2-flushed nutrient solutiona
Plants (cv. Capelle Desprez) were raised in aerated solution for
13 d, with aeration then continued or replaced by N2 flushing for
another 14 d, all at 14 C. Data from Trought and Drew (1980a);
values are means 6standard errors.
Parameter
Aerated
N2-flushed
% of
Control
Shoot N concentration
(mmol g1 dry weight)
Shoot fresh weight (g)
Shoot dry weight (mg)
Seminal root dry weight (mg)
Adventitious root dry
weight (mg)
Root:shoot (dry weight basis)
Adventitious root numberb
2.9960.06
0.7260.07
24
2.5760.11
357619
175610
2763
0.8260.04
210612
2061
1762
32
59
11
63
0.57
Not given, but not
different between
treatments
2161
0.18
32
0
Not specified
10
25
Longest adventitious root (cm)
Adventitious root aerenchyma
(%)
Towards tip
Near base
1261
57
Table 2. Effect of N2-flushed treatment on nutrient uptake and
transport to shoots of wheat
Values were estimated based on changes in shoot content and total
transpiration between samplings, assuming no recirculation from
shoots to roots via phloem. Wheat (cv. Capelle Desprez) was raised
in aerated solution for 14 d, with aeration then continued or replaced
by N2 flushing for another 14 d, all at 14 C. Data from Trought and
Drew (1980a). Overall, data were similar for 0–4, 4–8, and 8–14 d of
N2 flushing and the data presented here are for 8–14 d.
Element
Nitrogena
Phosphorus
Potassiumb
Calcium
Magnesium
Nutrient
solution
Calculated (mM) for xylem
stream of plants with roots
in
(mM)
Aerated
N2
flushed
0.50
0.25
0.75
1.00
0.50
9.52
1.28
4.92
0.51
0.29
2.27
0.24
0.55
0.26
0.07
–
–
a
The example given is from the key study by Trought and Drew
(1980a), with the results being considered typical for wheat. Similar
inhibitory effects on roots, as described above by Trought and Drew
(1980a), were also seen for cv. Gamenya grown at 15/20 C night/day
in Kuiper et al. (1994). Reductions in shoot growth were less when
wheat was in N2-flushed solution with high nutrient concentrations
(Trought and Drew, 1981; Barrett-Lennard et al., 1988), in which
seminal root growth was still severely inhibited.
b
Numbers were not reported by Trought and Drew (1980a).
In some studies adventitious root numbers have been reported to
increase in N2-flushed conditions (e.g. from 7 to 11 per plant after 14 d
of hypoxia in cv. Gamenya; Barrett-Lennard et al., 1988).
1980a). Decreased nutrient concentrations in the shoots
were not only due to reduced root:shoot, but were also
caused by poor functioning of the root system; i.e. the
inhibition of ion transport to the shoot per unit of root, as
shown by greatly reduced concentrations of macronutrients
in the xylem (Table 2).
The hypothesis that nutrient deficiencies retard the
growth of wheat in waterlogged soil can be tested by the
response to increased nutrient supply. Some success with
this approach has been achieved, though few of the experiments restored the growth of plants with O2-deficient roots
to that in aerated/drained controls. Shoot N concentrations
were increased in wheat in both aerated and N2-flushed
solutions when high external NO
3 (10 mM) was supplied;
but growth was only improved when all nutrients in the
medium were increased 4-fold (Trought and Drew, 1981).
Then, shoot fresh weight (the best indicator available of
growth in that experiment) of plants in N2-flushed solution
was reduced by 45% compared with 60% when only NO
3
was increased, however, there was no significant improvement in root growth (Trought and Drew, 1981). Improved
root growth at high mineral nutrition was indicated in
another experiment with wheat in waterlogged sand culture;
Nutrient uptake rates
(mmol g1 dw root d1) in
a
Nitrogen
Phosphorus
Potassiumb
Calcium
Magnesium
0.50
0.25
0.75
1.00
0.50
Aerated
N2-flushed
284
57
202
22
12
113
15
39
17
4
Water uptake rates
(ml g1 dw root d1) in
a
Aerated
N2-flushed
47
52
NO
3.
Supplied as
Potassium concentration in solution is uncertain as the value of
0.75 in Table 3 in Trought and Drew (1980a) and reproduced here,
differs from the total of 5.0 mM specified in the composition of the
nutrient solution in the Materials and methods. dw, dry weight.
b
when Hoagland nutrient solution was increased from halfto full-strength (7–14 mM NO
3 ) the decrease in root dry
weight relative to drained controls of a tolerant variety was
less severe (50% reduction at 14mM NO
3 compared with
75% reduction at 7mM NO
;
Huang
et
al.,
1994). That N
3
deficiency contributed to some extent to the adverse effects
of waterlogging was shown by applying a 0.1 M foliar urea
spray to wheat seedlings during the first 3 d of a 14 d
waterlogging treatment; reductions in shoot fresh weight at
the end of 14 d were decreased from 40% to 32% and in
chlorophyll concentrations from 76% to 64% (Trought and
Drew, 1980b). High nutrition may also aid recovery after
a return to aerated conditions. Even when the fresh weight
of wheat shoots was not improved in N2-flushed solution
containing 5 mM rather than 0.1 mM NO
3 , transference to
aerated solution containing 0.5 mM NO
3 , increased the
44 | Colmer and Greenway
fresh weight of the shoots, relative to the weight at the time of
transfer, by 60% and 140% for plants exposed to 0.1 mM and
5 mM NO
3 , respectively, during the preceding 14 d in N2flushed solution (Trought and Drew, 1981).
Experiments with foliar nutrient sprays on barley support
that nutrient deficiency contributes to decreased growth of
dryland cereals during the first 14 d of soil waterlogging.
Daily foliar sprays of complete Hoagland nutrient solution
to barley in pots of waterlogged soil improved shoot and
root growth, to varying degrees, of six genotypes; root dry
weight in one of the two tolerant genotypes became as high
as in the drained pots (Pang et al., 2007); i.e. foliar sprays
were most effective when the particular genotype has an
inherent high tolerance to waterlogging. The improvement
of root growth is particularly noteworthy and needs verification. The foliar spray also increased N and K+ concentrations (% dry weight) in shoots and roots, reaching levels in
the plants in drained soil for one of the tolerant genotypes.
Foliar nutrient application also increased the number of
adventitious roots by 30–40%, while the percentage of
chlorotic leaves decreased. The experiment of Pang et al.
(2007) lacked a treatment of foliar application of nutrients to
the shoots of plants in drained soil, so whether the response
was specific to plants in waterlogged soil was not determined.
Together, these experiments (Trought and Drew, 1980b,
1981; Pang et al., 2007) support the notion of nutrient
deficiency as a major factor limiting the shoot growth of
wheat and barley in waterlogged soils. Nevertheless, adverse
factors other than nutrient deficiencies probably also
contribute to the inhibition of shoot growth when roots are
in hypoxic nutrient solution. Barrett-Lennard et al. (1988)
found that wheat with high nutrient levels in solution (e.g.
with NO
3 at 15 mM) still showed small (7–10%) inhibitions
of shoot relative growth rate in a N2-flushed treatment, even
though shoot tissue nutrient concentrations remained above
‘sufficiency levels’. We suggest these remaining reductions
in shoot growth of wheat in N2-flushed solution with high
nutrients may be associated with the regulation of
root:shoot, as the root system in the N2-flushed solutions
was still relatively small (see below). Possible regulation of
root:shoot via hormonal changes in plants in O2-deficient
media, for example, inhibited production of IAA, cytokinins, and gibberellins, or increased ABA transport to the
shoot, was reviewed by Armstrong and Drew (2002).
Summing up; attempts to relieve inhibitions of growth of
wheat in N2-flushed solutions by increasing nutrient supply
have only been partially successful. One interesting limitation to this approach was shown in the experiments by
Huang et al. (1994): an increase from half- to full-strength
Hoagland nutrients reduced the percentage of aerenchyma
in adventitious roots, in the tolerant cultivar from 30% to
24% (Huang et al., 1994); therefore improvements in
nutrition may have been partly counteracted by reduced O2
transport from shoots to roots. So, although some improvement might be achieved by improved management (i.e. foliar
sprays and/or higher fertilizer use), the key approach remains
the development of cultivars with improved tolerance to
waterlogging. At least two characteristics need attention,
growth of a reasonably-sized root system in waterlogged
soil, particularly in terms of root length and number of
adventitious roots, and improvement of root function in
O2-deficient media. In the sections below, key limitations to
root functioning in hypoxic–anoxic media, are discussed.
Aerenchyma and internal O2 supply to
seminal and adventitious roots of wheat
Internal O2 supply is crucial to both growth and ion
transport in hypoxic roots. As much of this review concerns
the response of wheat to N2-flushed treatments in nutrient
solutions, a summary is given of what is known of the O2
status of the roots studied. Furthermore, limitations to
internal O2 transport are considered for seminal and
adventitious roots of wheat, as compared with rice.
In the experiments by Trought and Drew (1980a) on
wheat in N2-flushed solution, transverse sections of adventitious roots showed aerenchyma occupied 10% of the crosssection near the tip and 25% near the root–shoot junction.
Further information on O2 supply to adventitious roots of
wheat was obtained for intact plants grown with roots in
N2-flushed nutrient solutions (Barrett-Lennard et al., 1988),
in which adventitious root porosity was 14%. O2 concentrations in the cortical gas spaces of these adventitious roots
were assessed using root-sleeving O2 electrodes (Armstrong
and Wright, 1975); this technique requires transferring the
roots from the N2-flushed solution into O2-free stagnant
agar. Assessed O2 partial pressures near the root tip were
1 kPa for 90 mm long roots and 2 kPa for 60 mm long
roots, whereas in basal root zones near the root–shoot
junction the levels were 4 and 8 kPa, respectively (BarrettLennard et al., 1988). No O2 data for the seminal roots were
obtained, but the proximal sections had only 3% porosity,
compared with 14% in the adventitious roots (BarrettLennard et al., 1988), so internal O2 concentrations would
almost certainly be much lower in the seminal than in the
adventitious roots.
Adventitious root elongation for wheat in stagnant agar
nutrient solution increased in response to raising O2 around
the shoots in short-term experiments, confirming root
growth in this stagnant medium was limited by low O2
supply (Wiengweera and Greenway, 2004). Raising O2
around the shoots from 20 kPa to 40 kPa for 2 h and then
to 80 kPa, increased root elongation by 5-fold for 11–12 cm
long roots, and by 2-fold for both 5–6 and 1–2 cm long
roots (Wiengweera and Greenway, 2004). The large stimulation in the 11–12 cm roots is consistent with the predicted
low O2 near the apex for roots of this length with 15–20%
porosity (Armstrong, 1979). Thus, when O2 around the
shoots was increased, more O2 could diffuse to the root
apex and elongation resumed. Similarly, in 5–7-d-old
seminal roots of wheat which, contrary to older seminal
roots can develop porosity (namely 12%) during hypoxia,
there were also large stimulations of elongation when O2
around the shoots was increased to 100 kPa (Thomson
et al., 1990).
Ion transport in hypoxic roots | 45
The surprising result in Wiengweera and Greenway
(2004) was the substantial stimulation of the short adventitious wheat roots, indicating O2 deficiency even in the tips
of these short roots which are close to the potential O2
source at the shoot base. The crucial importance of this
limitation is highlighted by the rather slow extension of
these young roots when in stagnant agar solution, at ;50%
of aerated controls, with ambient O2 around the shoots and
hence we will discuss some reasons why the growth of even
these short adventitious roots was suboptimal. O2 partial
pressure around the shoots required to stimulate root
extension in wheat exceeds by ;20-fold the 4 kPa required
around shoots of rice for optimum extension of 5–7 cm long
adventitious roots (Armstrong and Webb, 1985); i.e. with
ambient O2 concentration around the shoots, O2 provision
to the tips of rice roots in an anoxic medium was more than
ample, but clearly suboptimal for roots of wheat. The
rice was grown in waterlogged potting mix prior to being
washed out and transferred into stagnant agar; the roots
were reported to be aerenchymatous, but percentage
porosity or aerenchyma was not given. Assuming porosity
in the rice roots of up to 41% (e.g. stagnant agar-raised
plants; Colmer, 2003a), the much lower porosity of 15%
for wheat adventitious roots grown in stagnant agar
(Wiengweera et al., 1997) would only partially explain the
higher shoot O2 requirement to sustain extension in the
short adventitious roots of wheat. Other possible reasons
include: the 15% porosity reported by Wiengweera et al.
(1997) was for the entire adventitious root system, but short
roots might have had lower porosity than the longer roots
(TD Colmer, personal observations for young adventitious
roots of wheat). In addition to the lower amount of
aerenchyma (i.e. lower maximum porosity) formed in roots
of wheat, other factors presumably also contribute to the
spectacular differences in O2 diffusion to the root apex, as
compared with rice. Firstly, radial O2 loss (ROL) from
basal parts of wheat adventitious roots with aerenchyma
can be substantial (Thomson et al., 1992), whereas roots of
rice possess a barrier against ROL in basal zones
(Armstrong, 1971; Colmer, 2003a), although even in rice
the barrier is less pronounced in short than in long roots
(Colmer et al., 2006a). The barrier to ROL restricts radial
losses, thus promoting longitudinal diffusion towards the
apex (Armstrong, 1979). Secondly, as aerenchyma typically
terminates a few cm behind root tips (e.g. rice, Armstrong,
1971; wheat, Thomson et al., 1992), diffusion to the apex
depends on porosity in the intervening tissue, which is
almost certainly less porous in wheat, than in rice. Root
cortex cells in rice occur in a cubical arrangement (Justin
and Armstrong, 1987), whereas in wheat the pattern is
hexagonal (assessed from micrographs in Huang et al.,
1994; TD Colmer, personal observations). These different
cell packings can result in substantial differences in tissue
porosity. In rice roots with cubic packing, porosity was
;9% near the non-aerenchymatous tip (Armstrong, 1971),
whereas it can be as low as 1% in roots with hexagonal cell
arrangements (Justin and Armstrong, 1987). Thus, very low
porosity in the younger tissues in adventitious roots of
wheat, resulting in high resistance to internal O2 diffusion,
would explain the finding of Wiengweera and Greenway
(2004) that, even in short roots, O2 supply from the shoot is
suboptimal for root growth in an O2-free medium.
Taking rice as a model species is useful but also has
limitations for comparisons with dryland cereals. Adaptation of rice to flooded conditions involves a range of
characteristics (traits in wetland plants were reviewed by
Bailey-Serres and Voesenek, 2008; Colmer and Voesenek,
2009), which amplifies the challenge to achieve a large
improvement in waterlogging tolerance of dryland species
such as wheat. In any case, paddy rice may not be the best
model, since dryland cereals such as wheat would typically
experience transient waterlogging, while even 3 d waterlogging can severely retard development after drainage
occurs (Malik et al., 2002). So upland and/or rain-fed rice,
in which final growth might be in drying soils, might
provide a better model for dryland cereals. Recovery upon
drainage following waterlogging would be especially important for wheat, as rapid growth of deep roots following
drainage will be required to obtain sufficient water later in
the season in Mediterranean climates. Some traits of
importance in rice, such as a barrier to ROL (Armstrong,
1971; Colmer, 2003a) might be counterproductive for
species faced with transient or intermittent flooding; possible reductions in water and nutrient uptake for roots of
wetland plants with a barrier to ROL were suggested by
Koncalova (1990). However, the few experiments conducted
to address this question have not found such drawbacks
(e.g. NO
3 uptake by rice roots, Rubinigg et al., 2002; water
uptake by Hordeum marinum roots, Garthwaite et al.,
2006). The exception is a strong barrier induced in rice
roots in response to soil toxins, then water permeability
declined and toxin entry was impeded (Armstrong and
Armstrong, 2005a). Even when the barrier in roots of rice
was induced by growth in stagnant solution, toxin (namely
Cu2+) entry was impeded (Kotula et al., 2009). One
drawback of a barrier to ROL, however, could be restricted
O2 entry into roots when the soil is not waterlogged,
causing suboptimal O2 supply to persist in some root parts
even in a drained soil. Such trade-offs, if any do occur,
would be reduced in species with inducible barriers to ROL,
as compared with those that constitutively develop a barrier
(reviewed by Colmer, 2003b). Inducible barriers to ROL
occur in rice (Colmer, 2003a) and in some wild relatives of
cultivated dryland species in the Triticeae (McDonald et al.,
2001; Garthwaite et al., 2003); however, in rice once the
barrier has formed, it persists even upon re-aeration, while
new root growth again lacks a barrier (MCH Cox and TD
Colmer, unpublished data).
Comparison of rice to wheat also emphasizes that in the
search for more tolerant wheat cultivars, a combination
of higher aerenchyma percentage and a larger root system
containing aerenchyma should be sought in wheat
(Thomson et al., 1992). Genetic diversity in capacity to
form a large system of adventitious roots was recently
reported for barley (Pang et al., 2007); after 14 d of soil
waterlogging a tolerant genotype produced 1.7-times more
46 | Colmer and Greenway
adventitious roots than an intolerant genotype. Increased
porosity of seminal roots would also be beneficial. As stated
earlier, young seminal roots of wheat formed 12% gas-filled
porosity when raised in hypoxic nutrient solution (Thomson
et al., 1990), and it would be beneficial if that capacity was
enhanced in older seminal root systems. The capacity to
form aerenchyma in seminal roots might differ between
wheat genotypes, as aerenchyma did form in seminal roots
of wheat in sand culture when waterlogging was imposed on
14-d-old plants; with the genotype Savanna forming aerenchyma along the entire root length and reaching 21% near
the root–shoot junction. By contrast, in seminal roots of
a less tolerant genotype no aerenchyma formed in the
critical section near the root–shoot junction, although some
aerenchyma formed in the lower sections (Huang et al.,
1994). Yet, although during waterlogging growth in Savannah was better than in the second genotype, the dry weight
of the shoots of Savannah at two levels of N supply (3 and
6 mM NO
3 ) was still only 50% and 70% of the drained
controls, respectively (Huang et al., 1994). Experiments are
needed to test the capacity for aerenchyma formation in
seminal roots of a larger range of wheat genotypes.
Similarly, tests are required for much older plants; when
wheat in the booting stage (153-d-old) was waterlogged at
1.5–2 cm above the soil surface, cross-sections showed large
aerenchyma formed within 96 h in 10–15 cm long adventitious roots (Jiang et al., 2010). Furthermore, development
of constitutive aerenchyma in roots, both in adventitious
and seminal systems, might enhance the tolerance of
transient waterlogging, as aerenchyma takes time to develop
fully; 72–102 h for wheat adventitious roots (Malik et al.,
2003) and 48 h for 50–80 mm long seminal roots, as judged
by O2 transport from the shoots to root tips (Thomson
et al., 1990).
In summary, for wheat in anaerobic media, the bulk of
the seminal roots would receive very little O2 owing to low
tissue porosity, while after exceeding a certain length or age,
the seminal roots might not form aerenchyma (at least in
one genotype, Thomson et al., 1990). Adventitious roots
develop aerenchyma, so their internal O2 supply is superior
to that of pre-existing seminal roots, but the low-to-moderate
porosity and lack of a barrier against ROL (cf. Thomson
et al., 1992) means the more distal portions (i.e. further from
the root–shoot junction) of wheat adventitious roots still
suffer from O2 deficits. These O2 deficits in the distal portions
of the seminal and adventitious roots of wheat, including
towards the apex and also within the stele, as reviewed
below, will impact on functioning in anaerobic media.
O2 gradients in roots: occurrence of stele
hypoxia–anoxia
O2 concentrations can differ markedly across roots. An
example of an O2 profile, from the external medium to the
centre of an excised, primary and non-aerenchymatous
maize root, is shown in Fig. 3. Even in a flowing medium,
a considerable boundary layer resistance was evident as
a steep O2 gradient between bulk solution and epidermis.
O2 also declined steeply across tissues of low porosity,
namely the outer few cell layers of the root and again in the
stele, whereas within the porous cortex the gradient was
very gradual. Such concentration gradients are inherent
when supply is via diffusion between a source and sink, as
determined by resistance to diffusion and O2 consumption
along the diffusion path (Armstrong and Beckett, 1987).
The O2 concentration in the pericycle lining the protoxylem
and metaxylem vessels of the roots shown in Fig. 3 has been
Fig. 3. Profile of O2 concentration across an excised primary root of Zea mays in a flowing nutrient solution containing 0.05 mM O2, at 25
C. An O2 microelectrode was moved in steps towards, and then through, the root (75 mm behind the apex, total length 135 mm). The flow
rate of the medium was 1.8 mm s1, calculated using the depth of the vessel as 10 mm (H. Greenway, personal communication), rather
than the erroneous 100 mm stated by Gibbs et al. (1998). The line with symbols is the in-track, and that without symbols is the out-track.
Reproduced from Gibbs et al. (1998) in Australian Journal of Plant Physiology with kind permission of CSIRO Publishing.
Ion transport in hypoxic roots | 47
assessed at 3 lM or lower. No ready comparisons with
other tissues of high O2 demands can be made, the best
available data is for COPE of rice roots of 1.25–2.5 lM
(assessed by Gibbs et al., 1998a from Armstrong and Webb,
1985). These rice root tips presumably had relatively low O2
requirements, as extension rates of 0.6 mm h1 (;14 mm d1)
(Armstrong and Webb, 1985) were relatively low compared
with up to 26 mm d1 (Colmer, 2003a), as the tips might
have been low in sugars (Gibbs and Greenway, 2003). Thus,
the low O2 concentration of 3 lM in the xylem parenchyma
indicates severe hypoxia, since the cells loading ions into the
xylem can be expected to have high rates of respiration (de
Boer and Volkov, 2003).
Gradients in O2 across roots were inferred by the classical
studies of root respiration by Berry and Norris (1949); these
authors hypothesized that as O2 was depleted from the
bulk medium, hypoxia–anoxia would occur in the core of
tissues, while more peripheral cells still received adequate
O2 for oxidative phosphorylation. When the O2 concentration was below the critical O2 pressure for respiration
(COPR) of excised onion root segments, the respiratory
quotient (CO2 produced/O2 consumed) increased, implying
a switch to ethanolic fermentation in cells of the innermost
core (Berry and Norris, 1949). This interpretation of the
onset of O2 deficits in the core of tissues when the external
concentration declines below the COPR is consistent with
mathematical modelling (Armstrong and Beckett, 1987;
Armstrong et al., 2009); with progressive expansion of the
volume of the anoxic core as external O2 supply decreases
below the COPR (i.e. the distance decreases to which O2 can
diffuse into the tissue before all is consumed).
The length of root over which the stele becomes O2
deficient depends upon several factors, these have already
been alluded to in the Introduction; applying these general
principles of Armstrong and Beckett (1987) to the stele, the
factors are: (i) the concentration of O2 in the cortical gas
spaces (determined by external O2 concentration, O2
diffusion in any aerenchyma and/or in the porous cortex,
rates of ROL along the root, and distance from the root–
shoot junction), (ii) the resistance to radial O2 diffusion
through the endodermis and stelar tissues, (iii) the rate of
O2 consumption in the stele, (iv) the radius of the stele,
determining its absorbing surface at the endodermis, subsequent diffusion-path distance to the centre, and total
volume. So, one adaptation to restricted O2 supply in roots
might be a narrower stele (Armstrong et al., 2000), reducing
its volume-to-surface area ratio, and diffusion-path length
for O2 to reach the parenchyma cells near the xylem. Species
differ in the percentage of adventitious root cross-sectional
area occupied by the stele, with wheat at 16% (assessed
from micrographs in Huang et al., 1994) compared with
only 5% in rice (McDonald et al., 2002). Morphological
acclimation might also occur, the proportional area of stele
in cross-sections of barley adventitious roots decreased by
20% in waterlogged soil, compared with drained controls
(Pang et al., 2004).
The O2 profile in Fig. 3 was for an excised maize root
immersed in solution, so there was no other O2 source than
the root medium. The O2 provision to roots of intact plants
is less well-defined than for excised roots. In studies with
intact maize, severe O2 deficiency in the stele was indicated
for the ;20 mm root tip of a 105 mm long nonaerenchymatous root; by contrast, in roots with aerenchyma only the stele in the 10 mm root tip was low in O2
(Darwent et al., 2003). These authors discuss uncertainties
in interpretations of values obtained with the O2 microelectrodes and conclude some O2 consumption might have
occurred concurrently with anaerobic catabolism, hence
they suggest referring to ‘severely hypoxic’ rather than
‘anoxic’ steles (Darwent et al., 2003). In view of these
uncertainties, data on O2 concentrations need to be
complemented with metabolic evidence.
A hypoxic stele was indicated by the higher pyruvate
decarboxylase activity in the stele versus the cortex from the
intact primary root of intact maize seedlings in anoxic
medium (Thomson and Greenway, 1991). Furthermore,
ATP:ADP in roots of intact maize in N2-flushed solution
(Table 3) are consistent with the notion that the cores of
tissues can become O2 deficient (suboptimal respiration),
whereas the outer layers receive adequate O2 to meet their
respiratory demands. Near the root–shoot junction (the
source of O2 for roots in anoxic medium), ATP:ADP in
roots of ;14% porosity was more similar to that in aerated
Table 3. ATP-to-ADP ratio (i.e. ATP:ADP) in intact adventitious
roots of Zea mays pretreated with different O2 supplies so as to
obtain a range of root porosity values, prior to transfer into N2flushed solution, with shoots in air
Treatments were imposed on 12-d-old plants for 11 d, in solutions
bubbled with 40% O2 (;4% root porosity), 5% O2 (;10% root
porosity), or non-turbulent solution (;14% root porosity). ATP:ADP
was measured on roots 75 min after transfer of the roots into the
N2-flushed solution. Data from Drew et al. (1985).
Segments at distance
(mm) from the root–
shoot junction
0–10 (basal segment)
100–110
135–145
145–150
(tips of roots with ;14%
porosity)
200–205
205–210
(tips of roots with ;4 and
;10% porosity)
ATP:ADPa in roots
with porosity of
;4%
;10%
;14%
3.860.03
1.160.04
n.a.b
n.a.
3.660.3
1.560.1
n.a.
n.a.
4.460.05
3.260.5
1.760.02
2.060.15
0.560.02
0.760.02
1.760.05
1.560.2
n.a.
n.a.
a
For comparison with the values in the table, ATP:ADP in root
segments when excised and in aerated solution was 5.5 and when in
N2-flushed was 0.66.
b
n.a. ¼ not applicable. n.a. entries in the table result from roots
being of different lengths following pretreatments; roots from nonturbulent solution were shorter than those pretreated with 5% or 40%
O2, so root tips were different distances from the root–shoot junction
as indicated in the table.
48 | Colmer and Greenway
roots, whereas in roots with ;4% and ;10% porosity
ATP:ADP had declined, and the ratio then remained rather
steady between 100 mm and 205 mm from the root–shoot
junction (Table 3). The very low values for ATP:ADP of
0.5–0.7 in the distal part of the roots with ;4% porosity
indicates these were, not unexpectedly, anoxic in the cortex
as well as in the stele. The pattern down the roots and
values in distal regions for the roots with ;10% porosity are
to be expected when the porous cortex has sufficient O2 for
oxidative phosphorylation, while the adjoining stele is at
least severely hypoxic (models of Armstrong and Becket,
1987). Consistent with the O2 profiles from Darwent et al.
(2003), roots with ;14% porosity only dropped to low
ATP:ADP further than 110 mm from the root–shoot
junction. ATP:ADP of 1.7–2 in the distal parts of these
roots of ;14% porosity (Table 3) were presumably the
composite of high values in the ‘aerobic cortex’ and low
values in the ‘hypoxic–anoxic stele’. For an anoxic stele
these values were assessed at 2.4–1.8 (see Appendix 1). An
anoxic stele would have a large effect on ATP:ADP in the
whole roots, since stelar cells are more cytoplasmic than
those of the cortex (discussed in relation to ion transport by
de Boer and Volkov, 2003; and based on differences in O2
consumption rates between stele and cortex, Armstrong and
Beckett, 1987; Armstrong et al., 2000). Confirmation of
these deductions may be obtained by separating stele from
cortex for determinations of nucleotides, as was successfully
done for the active–inactive state of PDC; i.e. the separation
and liquid N2 freezing of the tissues was fast enough to
prevent the expected rapid metabolic changes (Thomson
and Greenway, 1991).
The above evidence strongly indicates that resistances to
diffusion limit O2 availability to the interior of roots when
external O2 decreases below the COPR (Armstrong et al.,
2009); however, an alternative hypothesis has been proposed of the down-regulation of respiration by plant tissues
‘sensing’ declines in O2 availability (Gupta et al., 2009).
This alternative hypothesis, however, appears not to have
accommodated the influence of diffusion limitations causing
declines in O2 consumption rates by tissues when external
O2 is below the COPR. In any case, the comprehensively
documented occurrence of a severely hypoxic–anoxic stele
in maize roots when external O2 declines below the COPR is
adequate to account for the observed decreases in respiration of roots in hypoxic solutions (Atwell et al., 1985). The
consequences of O2 deficiency in the stele, or part thereof,
for ion transport by roots is considered in some detail later
in this review.
Water uptake and possible bypass flow of
nutrients, in O2-deficient roots
The possible inhibition of water transport is considered
before reductions in ion transport, since flow rates of xylem
sap can, under certain circumstances, determine the rates of
ion transport to the shoots, for example, in plants of high
ion status a 5-fold decrease in transpiration reduced K+
flow to the shoots by 4-fold (Russell and Barber, 1960).
Effects of hypoxia–anoxia on root hydraulic conductivity
(Lp)
Short-term studies, of less than 1 h exposure, show that O2
deficits reduced hydraulic conductivity of wheat root
cortical cells (Lpc) by 45%, while root hydraulic conductivity for flow to the xylem (Lpr) was reduced by 2–45%,
presumably due to partial closure of aquaporins, either in
the cortical cells or in the endodermis (Bramley et al., 2010).
The partial closures of aquaporins may be part of a more
general reduced permeability of membrane channels under
hypoxia–anoxia, particularly since some aquaporins can
also conduct several other small neutral solutes (Bramley
and Tyerman, 2010). Reduced conductivity of ion channels
has been shown for animal cells (Hochachka, 1991) and
indicated for K+ channels in anoxic rice coleoptiles (Colmer
et al., 2001). Furthermore, transcript abundances of most
aquaporin genes decreased in Arabidopsis exposed to
0.03 mM O2, but transcripts of NIP 2:1 increased (Liu
et al., 2005); of interest since the NIP 2:1 protein might
transport small metabolites (Bramley and Tyerman, 2010).
Presumably, the NIP protein also increased in abundance,
although neither the turnover rates of aquaporins nor the
functionality of this aquaporin under hypoxic conditions is
known. Whether reductions in Lpr due to closure of
aquaporins persist over longer periods remains uncertain.
Any long-term reductions of Lpr are more likely to be
associated with any of the following factors: cell death,
modifications of cell walls, and plugging of the xylem
vessels (Bramley and Tyerman, 2010).
For maize, two studies have assessed Lpr of hypoxic roots
for several hours both using excised roots. One study is
relevant to roots with low gas-filled porosity in waterlogged
soil; i.e. with time the roots became severely hypoxic or
anoxic. In this study, the excised roots showed the first
reductions in Lpr after 5 h of commencement of continuous
N2 flushing, when O2 in the solution had decreased below
2 kPa; O2 would have continued to decline to very low
levels and the Lpr remained reduced by ;60% for the last
17 h (Everard and Drew, 1987). A second study is relevant
to aerenchymatous roots in waterlogged soil, since the
excised maize roots were exposed to a steady O2 concentration of 0.05 mM so that the cortex would have been aerobic
whereas the stele was hypoxic (cf. Fig. 3). These roots
initially showed a ;25% reduction in Lpr, followed by at
least partial recovery within 1.5–4 h (Gibbs et al., 1998b).
Consistently, in the case of wheat in N2-flushed solution for
14 d, water uptake (root dry weight basis) was not reduced
when compared with aerated controls (Table 2), and was even
50% higher in the N2-flushed treatment during the first 4 d
(Trought and Drew, 1980a). Importantly, most of this flow
must have been via the, putative, hypoxia-intolerant seminal
root system, since at 4 d these roots still contributed ;90% to
the total root mass (Trought and Drew, 1980a).
Ion transport in hypoxic roots | 49
In summary, closure of aquaporins during the first few
hours of root zone hypoxia remains interesting, but at
present appears to be of little relevance to longer-term
exposures of roots to hypoxia and anoxia. Longer-term
studies of wheat are needed to confirm these deductions,
and should evaluate possible acclimation in aquaporin
regulation over a few days of hypoxic treatment, for both
seminal and adventitious roots. Furthermore, it would be
interesting to evaluate the cellular responses in the cortex
and stele of the adventitious roots, as these are expected to
differ in O2 supply when in an anoxic medium (aerobic
cortex and hypoxic–anoxic stele can occur, as described
earlier in this review).
Ion transport across roots by mass flow?
Trought and Drew (1980a) proposed that for wheat in N2flushed solution, water (and nutrients) might simply enter
via mass flow through damaged roots in response to
transpiration; an assessment based on comparisons of ion
concentrations estimated in the xylem with those in the
external solution (Table 2). We question this notion because
energy-dependent transport of anions is indicated by similar
phosphate and higher N in the xylem than in the external
solution (Table 2). The more likely mechanism for the
persisting, albeit greatly reduced, ion uptake by roots of
wheat in N2-flushed nutrient solutions (Table 2) is secondary energy-dependent transport via the aerenchymatous
adventitious roots, and through the basal portions of the
seminal roots, which would still receive O2 from the shoots
(Armstrong, 1979; Barrett-Lennard et al., 1988). Application of inhibitors of respiration to roots in N2-flushed
solutions could be used to test whether the main pathway of
the nutrient uptake is by mass flow, or by energy-dependent
transport via the root portions still receiving O2 from the
shoots. Furthermore, the possible occurrence of mass flow
of external solution (also termed by some ‘by-pass flow’)
can be tested by the addition to the nutrient solution of an
apoplastic tracer such as PTS, as used so effectively in
studies of rice (Yeo et al., 1987). By-pass flow was ;0.5% of
total water flow in aerated wheat roots (Garcia et al., 1997),
so any increase in such flows during root hypoxia should be
easily detected.
Nutrient uptake into roots, and transport into
the xylem, during hypoxia
A much larger effect of hypoxia on ion transport to the
xylem than on net uptake into the root cells will be
discussed using the scheme in Fig. 4 and also the crucial
information discussed earlier that, depending on a number
of root characteristics, low external O2 concentrations may
lead to severe hypoxia in the stele over much of the root
length, while the cortex and epidermis still receive sufficient
O2 for oxidative phosphorylation.
In fully-aerobic roots, ions taken up from the external
solution are loaded into the xylem with energy provided by
their plasma membrane H+-ATPases with subsequent ion
release into the xylem (de Boer and Volkov, 2003). For
example, K+ flux into the xylem is through SKOR channels;
i.e. highly K+-selective outward rectifying channels (de Boer
and Volkov, 2003; Pang and Shabala, 2010). This process is
also dependent on the accumulation of high K+ in the
xylem parenchyma cells (de Boer and Volkov, 2003), as
supported by X-ray microanalysis, showing particularly
high K+ concentrations in the xylem parenchyma cells
bordering the protoxylem and early-maturing metaxylem
vessels (Drew et al., 1990). During stelar hypoxia–anoxia,
H+-ATPases in the xylem parenchyma would be inhibited,
so the sophisticated regulatory mechanism of optimizing
transport to the xylem is disrupted, as SKOR channels
would close. We suggest that ions accumulated by the
epidermis and cortex from the external solution might still
flow via plasmodesmata to the xylem parenchyma by
diffusion, facilitated by cytoplasmic streaming, while ions
in the xylem parenchyma could still be released through
non-selective ion channels, most likely via NORC (nonselective outward-rectifying channels, de Boer and Volkov,
2003; Pang and Shabala, 2010); these outward rectifying
channels are strongly voltage dependent (Pang and Shabala,
2010). This, albeit reduced ion movement to the xylem,
would be facilitated by the depolarization of the membranes
of the xylem parenchyma, favouring passive cation flow
into the xylem vessels. Overall, both the total flow of ions to
the xylem and its K+/Na+ selectivity will be reduced in
hypoxic roots.
Electrochemical gradients for both anions and K+ transfer to the xylem vessels are likely to be favourable. Data on
the effect of decreased O2 in the medium on trans-root
electrical potential (i.e. electrical potential difference between external bathing medium and xylem sap) are available for Plantago, when O2 was lowered to 10 kPa the transroot electrical potential became more negative from –20 mV
in aerated roots to –60 mV in the hypoxic roots (de Boer
et al., 1983), presumably because the stele became hypoxic.
During hypoxia, the xylem parenchyma cells are likely to
depolarize to a value less negative than –100 mV; as
measured for cortical cells of seminal roots of wheat
(–90 mV at 0.008 mM O2, Buwalda et al., 1988b; –80 mV
at 0.02–0.03 mM O2, Zhang and Tyerman, 1997). So the
potential difference still favours anion release, but maintains, albeit less pronounced, an electrical gradient favouring K+ fluxes from the xylem into the parenchyma.
Nevertheless, the free energy gradient for K+ fluxes may be
from the parenchyma to the xylem, since the electrical
component of the free energy can be more than balanced by
a steep concentration gradient across the xylem parenchyma–
xylem interface. The free energy gradient for diffusive ion
movement between xylem parenchyma and xylem vessels can
be assessed from the flux ratio equation (equation 3.24 in
out
o
i zjFEM/RT
Nobel, 1974; Jin
), where Jin
j /Jj ¼cj /(cje
j is the influx
out
of ion j, Jj is the efflux of ion j, coj and cij are outside and
inside concentrations of ion j, e is the exponential, zj is the
valence of ion j, F is Faraday, EM is the membrane potential,
R is the gas constant, and T is the temperature in oK).
50 | Colmer and Greenway
A Fully-aerobic root
Xylem
parenchyma
Epidermis
and cortex
-120 mV
ATP
H+
Na+
H+
Anion
B Aerobic epidermis/cortex and anoxic stele
H+
E
n
d
o
d
e
r
m
i
s
-120 to
-160 mV
Na+
retrieval
ATP
Epidermis
and cortex
-20 to
- 40 mV
-120 mV
Xylem
parenchyma
ATP
HKT
H+
H+
Na+
Ion
K+
Ions
Gated
Cation
Xylem
H+
H+
H+
SKOR
Anion
NORC
Ions
-100 mV
-60 mV
E
n Reduced Na+
d retrieval
HKT
o
ATP
d
H+
e
r
m
Ion
i
H+
s
Gated
SKOR
Cations
& Anions
Cation
Anion
Xylem
NORC
Anion
[ion]
[ion]
Hypothetical [ion]
gradients across roots
Ext.
Epidermis
and cortex
Xylem
parenchyma
Xylem
Ext.
Epidermis
and cortex
Xylem
parenchyma
Xylem
Fig. 4. Diagram of ion movements through roots to the xylem, based on de Boer and Volkov (2003), Munns and Tester (2008), and
Pang and Shabala (2010), simplified by us to postulate on the consequences of a hypoxic–anoxic stele (see text). (A; Left half) In aerobic
roots, ATP produced in respiration is available to membrane H+-ATPases, ‘energized membranes’ enable ion transport, so ions are
accumulated to high concentrations in xylem parenchyma and released into the xylem (‘active loading’). (B; Right half) In roots with
anoxic stele but aerobic cortex, active xylem loading is inhibited, and ion movement to the xylem depends upon ion levels accumulated
in the aerobic cortex, with diffusion across the symplast to the xylem facilitated by cytoplasmic streaming. Below each main diagram,
graphs show hypothetical K+ gradients across roots (note: Ext., external solution). Numerous ion transporters and channels are not
shown in this simplified scheme. Channels of particular interest are: (i) SKOR which releases K+ into the xylem (highly selective for K+
over Na+) as part of the loading system, but which would close owing to the inhibition of the H+-ATPase and thus inhibition of energydependent transport in the hypoxic–anoxic stele, and (ii) NCOR a non-selective channel, transporting K+, Na+, and anions, that might be
gated in aerobic cells but could open when membranes depolarize during hypoxia–anoxia. Na+ retrieval from the xylem is also shown via
a HKT-type Na+ uniport (Munns and Tester, 1980); in a hypoxic–anoxic stele although retrieval would be reduced, some could possibly
still occur, but Na+ would then need to move to the aerobic cortex and be compartmentalized in vacuoles (dependent upon transtonoplast H+ difference and Na+/H+ antiport; Munns and Tester, 2008).
Assuming –100 mV, in the xylem parenchyma cells and –60 mV
in the vessels, this equation shows the free energy gradient for
K+ fluxes would be into the xylem provided the concentration
in the parenchyma cells is at least five-times greater than in the
xylem. That requirement can easily be achieved even at a high
xylem K+ concentration of 10 mM, since K+ in the cytoplasm
of xylem parenchyma would probably be 100 mM or higher
(10-fold greater than in the xylem; Drew et al., 1990). The rate
of delivery via this mechanism in roots with a hypoxic–anoxic
stele would, however, be substantially lower than in aerobic
roots, which have xylem loading. Consistent with the view that
transport to the xylem can still occur, even though the loading
process in the xylem parenchyma is inhibited, comes from
a SKOR-minus mutant (i.e. SKOR being a central component
of the loading mechanism); the mutant retained 50% of K+
transport to the shoot compared with the wild type (de Boer
and Volkov, 2003).
The available data for tissue ion concentrations, ion
uptake and xylem transport in roots of cereals (discussed
below) support the argument that xylem loading is inhibited
more than net ion uptake by O2-deficient roots. After 14 d
of N2-flushed treatment, there were only 0–15% decreases of
P or N concentrations in either adventitious or seminal
roots, compared with reductions of about 30% in the shoots
(Buwalda et al., 1988a). These favourable nutrient concentrations in the hypoxic roots are not unique for the high
external concentrations used in the experiment by Buwalda
et al. (1988a). At a lower external K+ of 0.4 mM, K+
concentration in the shoots was reduced by 45% whereas
the reduction was only 20% in adventitious roots, while in
seminal roots K+ increased by ;25% (Kuiper et al., 1994).
Effects on K+:Na+ selectivity will be discussed in a later
section.
Similarly to the reduction for other ions, wheat tissue Cl–
concentrations were also decreased by ;50% in the shoots,
but increased by 20–30% in the seminal roots and by 10–
30% in the adventitious roots, as compared with plants in
aerated solution (Buwalda et al., 1988a, Cl– external at
1 mM). The, albeit, small increases of Cl– concentration in
the adventitious roots are particularly noteworthy since the
Ion transport in hypoxic roots | 51
relative growth rates of these roots was only 10% lower
than in the aerated treatment (Barrett-Lennard et al., 1988).
The decreased Cl– concentration in the shoots, despite
increases in the roots, again support the notion that net ion
uptake by the roots is less vulnerable to hypoxia than
transport to the shoots.
The above-described observations have the defect that the
transport to the shoot was through a combination of
adventitious and seminal roots, of different gas-filled
porosity, and the internal O2 concentrations are either
unknown, or based on deductions from measurements
with cylindrical O2 electrodes. Such defects were avoided in
studies using excised primary roots of maize in a flowing
solution at 0.05 mM O2 (Gibbs et al., 1998a; Fig. 3 for
radial O2 profile). Cl– was at 0.4 mM, a suitable concentration to study energy-dependent anion uptake. Anion uptake
from low external concentrations must require energy since
both the root cells and the xylem have negative electrical
potentials relative to the medium.
In these excised maize roots the amounts of Cl– transported to the xylem were, in % of aerated roots, 60% and
30% at 0.05 and 0.02 mM O2, respectively (Gibbs et al.,
1998a). Despite these reductions in total Cl– transport, the
remaining flux still required at least one energy-dependent
transport step between external solution and the xylem, as
shown by the ratios of [Cl–]xylem:[Cl–]medium in the root
exudate at a range of O2 concentrations in the medium of:
93 at 0.27 mM O2, 70 at 0.05 mM O2, and 48 at 0.02 mM
O2, but only 0.6 in anoxia (Gibbs et al., 1998a). These
data are consistent with the development of, at least,
a severely hypoxic stele (Fig. 3), which would inhibit the
xylem loading step in the xylem parenchyma, but maintain
Cl– accumulation in the epidermis and cortex. For these
excised roots with the only O2 source in the external
solution, sufficient O2 supply to the epidermis is ensured
(Fig. 3).
The above data for Cl– transport in excised maize roots
(Gibbs et al., 1998a) also allow an estimate of residual
transport to the xylem when H+-ATPase activity in the
stele is diminished; i.e. xylem loading is inhibited.
A conservative estimate of this residual ion movement is
30% of that in fully aerobic roots; i.e. the value given in the
preceding paragraph for Cl– transport to the xylem at
external O2 of 0.02 mM (as % of aerated roots). High ion
concentrations in the cortex would provide a substantial
diffusion gradient towards the xylem, and since plasmodesmata remain conductive in hypoxic roots (Cleland et al.,
1994), cell-to-cell ion movement could occur in the symplast, facilitated by cytoplasmic streaming. Further testing
of the hypothesis on the changes in uptake by the root cells
and transport to the xylem, when the stele becomes
hypoxic–anoxic, would include measuring the initial unidirectional Cl– influx over the first 60 min after supply of
36 –
Cl as this represents the influx into the cytoplasm (cf.
Tregeagle et al., 2010); the hypothesis predicts this flux
would not be greatly affected by changing O2 between 0.27
mM and 0.02 mM in the flowing medium bathing excised
roots.
K+:Na+ selectivity
Stelar hypoxia might, in addition to decreasing nutrient ion
fluxes to the shoots, also impact on K+:Na+ selectivity of
transport to the shoots. The present review only considers
K+:Na+ selectivity for external Na+ at 10 mM or lower (i.e.
low concentrations not considered as saline). In hypoxia–
anoxia, net K+:Na+ selectivity to shoots is reduced and, as
will be shown shortly, this is partly due to a substantial
increase of Na+ flow to the shoots. This increase occurs,
even though Na+ in aerated solutions is thought to be
transported to the xylem by a SOS1 antiport (Munns and
Tester, 2008), which presumably gets inhibited during O2
deficiency. We suggest any decrease in intake via this
antiport is more than compensated for by opening of the
non-selective NORC channels, as these open due to the
strong membrane depolarization of xylem parenchyma cells
(Pang and Shabala, 2010). Transport to the shoot would be
mitigated by Na+ retrieval from the xylem and this
possibility will also be discussed.
For wheat in external K+ and Na+ both at 0.4 mM, shoot
+
K :Na+ decreased from 150 to 4.5 after 18 d in N2-flushed
solution, while adventitious root K+:Na+ decreased from 10
to 2; both these declines were mainly due to increased Na+
(Kuiper et al., 1994). Similar decreases in K+:Na+ were
found for 10–15-d-old maize; for example, at 0.025 mM O2
in the external solution with K+ at 1 mM and Na+ at
10 mM, the ratio decreased from 96 to 7 in the shoots, but
only from 19 to 9.5 in the roots (Drew and Lauchli, 1985).
At first sight it seems enigmatic that substantial decreases
in K+:Na+ occur for the aerenchymatous roots of wheat, as
described in Kuiper et al. (1994); i.e. for ion pools which are
presumably mainly located in the ‘aerobic cortex’. However,
it is possible that a substantial portion of the Na+ pool in
the cortical cells was associated with retrieval of Na+ from
the xylem. Since the cortical cells would have sufficient
energy to sequester the Na+ into vacuoles, the crucial
question is whether retrieval from the xylem sap is credible
in these hypoxic roots. In roots with a substantial length of
‘aerobic’ stele towards the root–shoot junction (as indicated
by models, Armstrong and Beckett, 1987; and shown in
Darwent et al., 2003), Na+ retrieval from the xylem would
presumably be important in mitigating Na+ flow to leaves
(Johanson and Cheeseman, 1983), most likely via HKT
transporters of subfamily 1 which function as a Na+ uniport
(Munns and Tester, 2008). In most cases, where the root
portions near the root–shoot junction still receives sufficient
O2 to maintain oxidative phosphorylation in the stele as
well as in the cortex, the driving force for Na+ retrieval
would be very strong; taking –120 mV in the stelar
parenchyma cells and –20 mV in the xylem of these upper
portions receiving O2, the flux ratio equation (Nobel, 1974)
indicates that Na+ retrieval would stop only when the Na+
concentration in the xylem parenchyma was 52 times higher
than in the xylem. With an anoxic stele, potential for Na+
retrieval from the xylem via a HKT uniport is weaker; yet
still likely, since the negative potential relative to the xylem
of –40 mV (de Boer et al., 1983), will still permit a 5-fold
52 | Colmer and Greenway
higher Na+ concentration in the xylem parenchyma than in
the xylem before retrieval would cease. In xylem sap of
barley, expressed from the cut end of the shoots of control
plants grown in supported hydroponics without Na+ added
(i.e. ‘background’ Na+ not defined), Na+ concentrations
were about 2–3 mM (Shabala et al., 2010) and these
concentrations would presumably decrease when there
was rapid transpiration. So, taking reasonable concentrations of 0.5–2.5 mM Na+ in the xylem, Na+ retrieval
would cease only when the symplast concentration reached
2.5–12.5 mM, these values are above the expected level if
Na+ withdrawn from the xylem moves to the cortical cells
and is sequestered into vacuoles. This suggestion needs
further testing with focused experiments to elucidate for
hypoxic roots whether the Na+ accumulated is derived
straight from the medium, or by entry and subsequent
retrieval of Na+ from the xylem.
Summing up, the data reviewed here on O2 profiles,
respiration, and ion transport in roots, some of which still
need more rigorous testing, are all consistent with the
dominant influence of development of a hypoxic–anoxic
stele under hypoxia, rather than the result of downregulation of metabolism involving an O2-sensing system
(as suggested by Geigenberger, 2003; Gupta et al., 2009).
The one observation which does not fit well with the model
of an ‘aerobic cortex’ and ‘hypoxic–anoxic stele’ is the
depolarization by 34 mV in the cortical cells of wheat upon
exposure to 0.02–0.03 mM O2 (Zhang and Tyerman, 1997),
when these cells can be expected to be still ‘aerobic’, i.e.
depolarization in the still aerobic cortex could be used as an
argument in favour of the down-regulation of metabolism.
However, depolarization in the cortex might result from
‘electrical coupling’, as the expected depolarization in the
stele might be conveyed to cortical cells. This is possible
even though the coupling ratio between cortical cells in
the wheat roots was only 6% (Zhang and Tyerman, 1997),
as the possible quantitative importance of coupling can not
be readily gauged from the coupling ratio alone (Solocar,
1977).
Ion uptake by seminal and adventitious roots
of wheat in O2-deficient media
Ion uptakes by seminal and adventitious roots of wheat in
N2-flushed versus aerated solutions were measured separately, in split-root experiments (Kuiper et al., 1994). The
net uptakes (root fresh weight basis), expressed as a percentage of the aerated controls, were 50% for K+ and 35% for
NO
3 in seminal roots, and 82% and 55% in adventitious
roots (average for 26 d and 33 d in N2-flushed solution;
Kuiper et al., 1994). This difference between root types is
less than expected in view of the much larger gas-filled
porosity of 14% in the adventitious roots than the 3% in
seminal roots (reported for the same cultivar by BarrettLennard et al., 1988). The seminal roots presumably
benefited from the low O2 concentration of 0.003 mM in
the N2-flushed solution, probably enabling respiration at
least in the epidermis and thus ion uptake by this outermost
cell layer. By contrast, when in stagnant agar nutrient
solution, the lack of turbulence would negate O2 supply
from the nutrient solution and hence the internal O2 supply
will solely depend on diffusion via the aerenchyma, then
net uptake of K+ and P by seminal roots (root fresh
weight basis), never exceeded 20% of the aerated controls
(Wiengweera and Greenway, 2004). This low uptake
occurred even though K+ concentration in the shoots would
have been 35% lower than in the aerated plants
(Wiengweera et al., 1997). By contrast, in the adventitious
roots, O2 supply to the epidermis was demonstrated by
ROL (Barrett-Lennard et al., 1988) and, accordingly, net
K+ uptakes by the adventitious roots, as a percentage of the
aerated controls, were 70% between 4–8 d and 115% during
the final 8–12 d (Wiengweera and Greenway, 2004). Thus,
overall, in media without any O2 supplied externally,
adventitious roots are capable of greater ion transport than
seminal roots, presumably owing to the higher porosity
enabling more O2 movement from the shoot to the root to
sustain respiration in the adventitious roots.
Surprisingly, measurements of O2 and ion uptake rates
for the same roots, at various O2 supply, are generally
lacking. The exception being the pioneering study of barley
seedling roots in hypoxia (0.02 mM O2, 0.05% agar
solution) by Pang et al. (2006) using ion-specific and O2
microelectrodes. At 10 mm behind the tip of 70 mm long
roots, a position just behind the elongation zone, O2 influx
was reduced by 80% in a hypoxia-tolerant cultivar and by
93% in a hypoxia-intolerant cultivar, as compared with
rates when bulk medium O2 was 0.28 mM (Pang et al.,
2006). It is not clear why the varieties differed in O2 uptake;
root lengths and diameters were the same, and although
root porosity values were not given, potential differences in
internal O2 supply from the shoots were probably not
a factor, since excised roots showed similar responses as
intact roots (Pang et al., 2006). No response curve was
determined for O2 uptake versus O2 concentration.
Net K+ uptake from 0.2 mM external K+ concentration
(also at 10 mm behind the root tip) was reduced by hypoxia
by 35–65% in the intolerant cultivar, but was not affected in
the tolerant cultivar (Pang et al., 2006). K+ net uptake by
this tolerant cultivar at 0.02 mM O2 was 2.1 lmol g1 fresh
weight h1, essentially the same as for seminal roots of
wheat in turbulent solutions at 0.003 mM O2 (Kuiper et al.,
1994); the Pang et al. (2006) data were converted by us from
surface area to a fresh weight basis (average of Figs 6 and 7
in Pang et al., 2006). The persistence of a reasonably high
net uptake in the barley was despite a reduction of 80% of
O2 uptake in stagnant solution at 0.02 mM external O2
compared with that by roots in stagnant air-saturated
solution. Net H+ efflux in the mature zone was changed
into a net influx in the presence of vanadate, the inhibitor of
the plasma membrane H+-ATPase. Pang and Shabala
(2010) discussed these findings as being consistent with the
possible engagement of the H+-ATPase under anoxia, citing
the case for anoxic maize root tips (Xia and Roberts, 1996).
However, the two situations are quite different, the barley
Ion transport in hypoxic roots | 53
roots received O2 (see above) whereas the maize root tips
were in anoxia. O2 uptake by the barley roots would have
enabled oxidative phosphorylation in the epidermis and
perhaps also in the outer cortical cells, so accordingly, ATP
should have been available to the H+-ATPases in the root
periphery. Although the outer cell layers would have
received some O2, cells towards the interior would presumably have been severely hypoxic or even anoxic.
Recovery of nutrient transport in
O2-deficient roots upon re-aeration
Recovery of nutrient uptake and transport to the xylem
upon re-aeration following anaerobiosis has been evaluated
on only a few occasions. To our knowledge, only one
experiment has evaluated net ion uptake in the first hours
after re-aeration of previously hypoxic wheat roots (Kuiper
et al., 1994); in this experiment it took ; 4 h of re-aeration
before net K+ uptake accelerated for both seminal and
adventitious roots. These lags indicate that the membrane
transport processes in the roots exposed to anaerobic
solution were impaired, and not simply suffering a simple
energy limitation, so that repairs were needed; whether the
lags were associated with general metabolism (e.g. lag in
recovery of mitochondria), to membranes, or to the ion
uptake machinery, remains to be elucidated. The lag in recovery following re-aeration might be associated with the
production of oxygen radicals after re-aeration, for example, 1–10 mM H2O2 caused pronounced membrane depolarization and massive K+ effluxes in roots of four barley
cultivars (Chen et al., 2007). Kuiper et al. (1994) also
assessed the recovery of K+ and NO
3 net uptakes during
the first 23 h of re-aeration; rates were 1.3–3.5-fold higher
for both seminal and adventitious roots, compared with
roots that continued in the N2-flushed solution.
In contrast to wheat, roots of rice grown in N2-flushed
solution had, at most, minor reductions in ion uptake
compared with continuously aerated plants (John et al.,
1974). Somewhat unexpectedly, upon transfer of rice from
N2-flushed to aerated solutions during the first 2 h there
were substantial ‘overshoots’ (i.e. increases above rates in
continuously aerated plants) for P, Cl–, and K+ uptakes by
roots and transport to the shoots (John et al., 1974; rates
determined by tracer uptake). Confirmation of these
responses is needed using more refined techniques, but
the ‘overshoots’ imply that, in rice, there may be acclimative
changes in nutrient-transport systems, in addition to the
well-known enlargement of aerenchyma.
More studies of recovery following re-aeration are
needed, using techniques with high temporal resolution
(e.g. unidirectional influxes using isotopes cf. Davenport
et al., 2005; ion-specific microelectrodes, cf. Pang et al.,
2006). Use of elevated O2 around the shoots would enable
recovery to be assessed with time-course measurements
continuing in stagnant agar (i.e. maintaining the same
conditions external to the roots, especially the unstirred
medium for the microelectrode net flux measurements). No
experiments to examine the influence of improved internal
aeration on nutrient uptake by roots of cereals are available, but in young alder trees illumination of the stem
improved O2 supply to the roots (Armstrong and
Armstrong, 2005b); and the improved O2 status restored
+
net uptake of NO
by roots in N2-flushed
3 , P and K
solution above the rates in aerated solution (Grosse and
Meyer, 1992). The improvements due to improved O2
supply in this experiment with alder seem in little doubt,
but the extent of the restoration of nutrient uptake remains
uncertain because of the high variability of the data by
Grosse and Meyer (1992).
Conclusions and perspectives
Improved root growth and functioning will be required to
alleviate shoot nutrient deficiencies, a main cause for
reduced growth of wheat, barley, and maize in anaerobic
solutions and very likely also in many waterlogged soils.
Improved root functioning should also aid tolerance of ‘soil
toxins’ in anaerobic soils, such as reduced metal ions and
salinity, as well as promote recovery following transient
waterlogging. Priority traits for improving the performance
of wheat roots in waterlogged soils would be an ability to
develop aerenchyma in seminal roots to prevent their death,
as well as a larger volume of more efficient aerenchyma in
adventitious roots. Another beneficial trait would be cubic
cell packing in the root cortex to increase porosity of nonaerenchymatous tissue, as occurs in rice, whereas the three
dryland cereals have hexagonal cell packings (few genotypes
examined). Stelar hypoxia, which inhibits the sophisticated
system of xylem loading, may not be alleviated merely by
more aerenchyma but could be avoided with a smaller
radius stele. In addition to improved internal aeration,
other traits might also improve the functioning of hypoxic
roots, such as protection against free radicals of oxygen.
Oxidative damage might occur after cell damage (Smirnoff,
1995), so lipid peroxidation associated with membrane
damage is possibly a secondary effect, nevertheless, studies
of animal cells indicate that radicals formed can further
accelerate the pathology (Arthur et al., 2008). So antioxidant systems should have ameliorating effects and might be
important during recovery, which would be particularly
relevant to survival following the transient waterlogging of
seminal roots of low gas-filled porosity. Genetic differences
in reactive-oxygen scavenging systems were important
during the recovery of rice following de-submergence (Ella
et al., 2003). A broad programme exploiting differences
within and across species is needed to combine the necessary,
complex characteristics; trait screening, physiological-based
breeding and selection including development and use of
molecular markers (Setter and Waters, 2003), and, where
possible, wide hybridizations and cytogenetic approaches to
make use of tolerant ‘wild’ relatives (Colmer et al., 2006).
One of the exciting challenges for future physiological
research is to elucidate more fully the functioning of roots
with the intriguing combination of an aerobic cortex
54 | Colmer and Greenway
(epidermis and cortex with sufficient O2 for oxidative
phosphorylation) and a hypoxic–anoxic stele. The persistence of substantial, albeit reduced, ion transport to the
xylem of roots with a hypoxic–anoxic stele might be viewed
as a primitive system before evolution of the sophisticated
system of xylem loading. The postulated different concentration gradients between epidermis and xylem may be
tested using X-ray microanalysis (e.g. for salinity, see
Lauchli et al., 2008). Further, physiological co-operation of
the aerobic cortex and hypoxic–anoxic stele seems likely,
such as consumption in the cortex of products of anaerobic
metabolism in the stele and, crucially, a possible transfer of
high energy compounds produced in the aerobic cortex to
the hypoxic–anoxic stele (suggested by Gibbs and Greenway, 2003). Physiological experiments need to be more
focused and less descriptive, using some techniques which
have already shown promise, such as measurements of both
net uptake by the root cells and transport to the xylem,
while measuring the trans-root electrical potential and O2
profiles across roots. Further net flux measurements along
roots, using ion-selective microelectrodes with concurrent
O2 fluxes also measured, would contribute to the elucidation of ion transport and K+:Na+ selectivity as affected by
hypoxia. Recovery following re-aeration (or elevation of O2
supply around shoots) should also be included in future
experiments, with measurements taken as soon as possible
after the O2 supply is increased. Such experiments will
address whether transport activity is merely limited by low
energy associated with low O2 supply, or also by other
lesions in metabolic capacity including transport capacity or
loss of membrane integrity.
the stele than in the cortex plus epidermis, a value based on the
ratio of O2 uptake rates in the stele versus the cortex (Armstrong
and Beckett, 1987). Then, the fraction of the nucleotide pool in the
cortex plus epidermis¼(0.7631)/[(0.7631)+(0.2435.5)]¼0.36, and
for the stele¼(0.2435.5)/[(0.7631)+(0.2435.5)]¼0.64. Thus the
ATP:ADP in a root segment as a whole with a hypoxic–anoxic
stele, becomes (0.3635.5)+(0.6430.66)¼2.4. The assessment of the
nucleotide concentration in stele versus cortex is not sure,
however, even if we take the unlikely high value of 10-times higher
concentration in the stele than cortex, and repeating the calculation above, then the ATP:ADP is 1.8; i.e., not all that much lower
than when assuming a 5.5-times difference.
Acknowledgements
Armstrong W, Beckett PM, Justin SHFW, Lythe S. 1991.
Modelling, and other aspects of root aeration by diffusion. In: Jackson
MB, Davies DD, Lambers H, eds. Plant life under oxygen deprivation.
The Hague: SPB Academic Publishing, 267–282.
For incisive criticisms on a final draft: Sergey Shabala for
the whole review; Helen Bramley for the water transport
section; Bill Armstrong on the hypoxic–anoxic stele; and
Natasha Teakle for useful comments on the whole review.
The Grains Research and Development Corporation in
Australia is thanked for supporting research by TDC on
waterlogging tolerance in wheat and wild species in the
Triticeae.
Appendix
Here we assess that the ratio of ATP-to-ADP (i.e. ATP:ADP) in
a mature root segment of maize, with an anoxic stele and ‘aerobic
cortex’ that still receives sufficient O2 for oxidative phosphorylation, will be in the range 2.4 to 1.8. This assessment is based on the
following values for maize: (i) proportion of the root occupied by
stele and cortex are 0.24 and 0.76 (Gibbs et al., 1998); (ii)
ATP:ADP in aerobic tissues of 5.5 and in anoxic tissues of 0.66
(Drew et al., 1985). These values are for roots exposed to 1 mM
NaF, since Drew et al. (1985) used this inhibitor of glycolysis in
addition to an anoxic medium in their studies with roots of intact
plants; (iii) assumed values for proportions of the nucleotide pool
contained in the cortex and stele, based on their proportional
volumes and expected differences in cytoplasm content. As an
example, we assume 5.5-times higher nucleotide concentration in
References
Armstrong J, Armstrong W. 2005a. Rice: sulphide-induced barriers
to root radial oxygen loss, Fe2+ and water uptake, and lateral root
emergence. Annals of Botany 96, 625–638.
Armstrong W. 1971. Radial oxygen losses from intact rice roots as
affected by distance from the apex, respiration, and waterlogging.
Physiologia Plantarum 25, 192–197.
Armstrong W. 1979. Aeration in higher plants. Advances in Botanical
Research 7, 225–332.
Armstrong W, Armstrong J. 2005b. Stem photosynthesis not
pressurized ventilation is responsible for light-enhanced oxygen supply
to submerged roots of alder (Alnus glutinosa). Annals of Botany 96,
591–612.
Armstrong W, Beckett PM. 1987. Internal aeration and the
development of stelar anoxia in submerged roots. A multishelled
mathematical model combining axial diffusion of oxygen in the cortex
with radial losses to the stele, the wall layers and the rhizosphere. New
Phytologist 105, 221–245.
Armstrong W, Cousins D, Armstrong J, Turner DW, Beckett PM.
2000. Oxygen distribution in wetland plant roots and permeability barriers
to gas-exchange with the rhizosphere: a microelectrode and modelling
study with Phragmites australis. Annals of Botany 86, 687–703.
Armstrong W, Drew MC. 2002. Root growth and metabolism under
oxygen deficiency. In: Waisel Y, Eshel A, Kafkafi U, eds. Plant roots:
the hidden half. New York: Marcel Dekker Inc, 729–761.
Armstrong W, Webb T. 1985. A critical oxygen pressure for root
extension in rice. Journal of Experimental Botany 36, 1573–1582.
Armstrong W, Webb T, Darwent M, Beckett PM. 2009. Measuring
and interpreting respiratory critical oxygen pressures in roots. Annals
of Botany 103, 281–293.
Armstrong W, Wright EJ. 1975. Radial oxygen loss from roots: the
theoretical basis for the manipulation of flux data obtained by the
cylindrical platinum electrode technique. Physiologia Plantarum 35, 21–26.
Arthur PG, Grounds MB, Shavlakadze T. 2008. Oxidative stress as
a therapeutic target during muscle wasting: considering the complex
interactions. Current Opinion in Clinical Nutrition and Metabolic Care
11, 408–416.
Ion transport in hypoxic roots | 55
Atwell BJ, Thomson CJ, Greenway H, Ward G, Waters I. 1985.
A study of impaired growth of roots of Zea mays seedlings at low
oxygen concentration. Plant, Cell and Environment 8, 179–188.
Colmer TD, Flowers TJ, Munns R. 2006b. Use of wild relatives to
improve salt tolerance in wheat. Journal of Experimental Botany 57,
1059–1078.
Bailey-Serres J, Voesenek LACJ. 2008. Flooding stress:
acclimations and genetic diversity. Annual Review of Plant Biology 59,
313–339.
Colmer TD, Huang S, Greenway H. 2001. Evidence for downregulation of ethanolic fermentation and K+ effluxes in the coleoptile of
rice seedlings during prolonged anoxia. Journal of Experimental
Botany 52, 1507–1517.
Barrett-Lennard EG. 2003. The interaction between waterlogging
and salinity in higher plants: causes, consequences and implications.
Plant and Soil 253, 35–54.
Barrett-Lennard EG, Leighton PD, Buwalda F, Gibbs J,
Armstrong W, Thomson CJ, Greenway H. 1988. Effects of
growing wheat in hypoxic nutrient solution and of subsequent
transfer to aerated solutions. I. Growth and carbohydrate status
of shoots and roots. Australian Journal of Plant Physiology 15,
585–598.
Berry LJ, Norris WE. 1949. Studies on onion root respiration.
I. Velocity of oxygen consumption in different segments of roots at
different temperatures as a function of partial pressure of oxygen.
Biochimica et Biophysica Acta 3, 593–606.
Bramley H, Turner NC, Turner DW, Tyerman SD. 2010. The
contrasting influence of short-term hypoxia on the hydraulic properties
of cells and roots of wheat and lupin. Functional Plant Biology 37,
183–193.
Bramley H, Tyerman SD. 2010. Root water transport under
waterlogged conditions and the roles of aquaporins. In: Mancuso S,
Shabala S, eds. Waterlogging signalling and tolerance in plants.
Heidelberg: Springer, 151–180.
Buwalda F, Barrett-Lennard EG, Greenway H, Davies BA. 1988a.
Effects of growing wheat in hypoxic nutrient solution and subsequent
transfer to aerated solutions. II. Concentrations and uptake of nutrients
and sodium in shoots and roots. Australian Journal of Plant Physiology
15, 599–612.
Buwalda F, Thomson CJ, Steigner W, Barrett-Lennard EG,
Gibbs J, Greenway H. 1988b. Hypoxia induces membrane
depolarization and potassium loss from wheat roots but does not
increase their permeability to sorbitol. Journal of Experimental Botany
39, 1169–1183.
Chen Z, Cuin TC, Zhou M, Twomey A, Naidu BP, Shabala S.
2007. Compatible solute accumulation and stress-mitigating effects in
barley genotypes contrasting in their salt tolerance. Journal of
Experimental Botany 58, 4245–4255.
Cleland RE, Fujiwara T, Lucas WJ. 1994. Plasmodesmal-mediated
cell to cell transport in wheat roots is modulated by anaerobic stress.
Protoplasma 178, 81–85.
Colmer TD. 2003a. Aerenchyma and an inducible barrier to radial
oxygen loss facilitate root aeration in upland, paddy and deepwater
rice (Oryza sativa L.). Annals of Botany 91, 301–309.
Colmer TD. 2003b. Long-distance transport of gases in plants:
a perspective on internal aeration and radial oxygen loss from roots.
Plant, Cell and Environment 26, 17–36.
Colmer TD, Cox MCH, Voesenek LACJ. 2006a. Root aeration in
rice (Oryza sativa L.): evaluation of oxygen, carbon dioxide, and
ethylene as possible regulators of root acclimatizations. New
Phytologist 170, 767–778.
Colmer TD, Voesenek LACJ. 2009. Flooding tolerance: suites of
plant traits in variable environments. Functional Plant Biology 36,
665–681.
Darwent MJ, Armstrong W, Armstrong J, Beckett PM. 2003.
Exploring the radial and longitudinal aeration of primary maize roots by
means of Clark-type oxygen microelectrodes. Russian Journal of Plant
Physiology 50, 722–732.
Davenport R, James RA, Zhakrisson-Plogander A, Tester M,
Munns R. 2005. Control of sodium transport in durum wheat. Plant
Physiology 137, 807–818.
de Boer AH, Prins HBA, Zanstra PE. 1983. Bi-phasic composition
of trans-root electrical potential in roots of Plantago species:
involvement of spatially separated electrogenic pumps. Planta 157,
259–266.
de Boer AH, Volkov V. 2003. Logistics of water and salt transport
through the plant: structure and functioning of the xylem. Plant, Cell
and Environment 26, 87–101.
Drew MC. 1983. Plant injury and adaptation to oxygen deficiency in
the root environment: a review. Plant and Soil 75, 179–199.
Drew MC. 1988. Effects of flooding and oxygen deficiency on plant
mineral nutrients. In: Lauchli A, Tinker PB, eds. Advances in plant
nutrition, Vol. III. New York: Praeger, 115–159.
Drew MC. 1992. Soil aeration and plant root metabolism. Soil Science
154, 259–268.
Drew MC, Lauchli A. 1985. Oxygen-dependent exclusion of sodium
ions from shoots by roots of Zea mays (cv. Pioneer 3906) in relation to
salinity damage. Plant Physiology 79, 171–176.
Drew MC, Saglio PH, Pradet A. 1985. Larger adenylate energy
charge and ATP/ADP ratios in aerenchymatous roots of Zea mays in
anaerobic media as a consequence of improved internal oxygen
transport. Plant Physiology 165, 51–58.
Drew MC, Sisworo ET. 1979. The development of waterlogging
damage in young barley plants in relation to plant nutrient status and
changes in soil properties. New Phytologist 82, 301–314.
Drew MC, Webb J, Saker LR. 1990. Regulation of K+ uptake and
transport to the xylem in barley roots: K+ distribution determined by
electron probe X-ray microanalysis of frozen-hydrated sections.
Journal of Experimental Botany 41, 815–825.
Ella ES, Kawano N, Ito O. 2003. Importance of active oxygenscavenging system in recovery of rice seedlings after submergence.
Plant Science 165, 85–93.
Elzenga JTM, van Veen H. 2010. Waterlogging and plant nutrient
uptake. In: Mancuso S, Shabala S, eds. Waterlogging signalling and
tolerance in plants. Heidelberg: Springer, 23–35.
Everard JD, Drew MC. 1987. Mechanisms of inhibition of water
movement in anaerobically treated roots of Zea mays L. Journal of
Experimental Botany 38, 1154–1165.
56 | Colmer and Greenway
Garcia A, Rizzo CA, Ud-Din J, Bartos SL, Senadhira D,
Flowers TJ, Yeo AR. 1997. Sodium and potassium transport to the
xylem are inherited independently in rice, and the mechanism of
sodium:potassium selectivity differs between rice and wheat. Plant,
Cell and Environment 20, 1167–1174.
Johanson JG, Cheeseman JM. 1983. Uptake and distribution of
sodium and potassium by corn seedlings. Role of mesocotyl in sodium
exclusion. Plant Physiology 73, 153–158.
Garthwaite AJ, von Bothmer R, Colmer TD. 2003. Diversity in root
aeration traits associated with waterlogging tolerance in the genus.
Hordeum. Functional Plant Biology 30, 875–889.
Justin SHFW, Armstrong W. 1987. The anatomical characteristics
of roots and plant response to soil flooding. New Phytologist 106,
465–495.
Garthwaite AJ, Steudle E, Colmer TD. 2006. Water uptake by roots
of Hordeum marinum: formation of a barrier to radial O2 loss does not
affect root hydraulic conductivity. Journal of Experimental Botany 57,
655–664.
Koncalova H. 1990. Anatomical adaptations to waterlogging in roots
of wetland graminoids: limitations and drawbacks. Aquatic Botany 38,
127–134.
Geigenberger P. 2003. Response of plant metabolism to too little
oxygen. Current Opinion in Plant Biology 6, 247–256.
Gibbs J, Greenway H. 2003. Mechanisms of anoxia tolerance in
plants. I. Growth, survival and anaerobic catabolism. Functional Plant
Biology 30, 1–47.
Gibbs J, Turner DW, Armstrong W, Darwent MJ, Greenway H.
1998a. Response to oxygen deficiency in primary roots of maize.
I. Development of oxygen deficiency in the stele reduces radial solute
transport to the xylem. Australian Journal of Plant Physiology 25,
745–758.
Gibbs J, Turner DW, Armstrong W, Sivasithanparam K,
Greenway H. 1998b. Response to oxygen deficiency in primary roots
of maize. II. Development of oxygen deficiency in the stele has limited
short-term impact on radial hydraulic conductivity. Australian Journal
of Plant Physiology 25, 759–763.
Greenway H, Armstrong W, Colmer TD. 2006. Conditions
leading to high Pco2 (5–40 kPa) in waterlogged-flooded soils and
possible effects on root growth and metabolism. Annals of Botany 98,
9–32.
Grosse W, Meyer D. 1992. The effect of pressurised gas transport
on nutrient uptake during hypoxia of Alder roots. Botanica Acta 105,
233–236.
Gupta KJ, Zabalza A, van Dongen JT. 2009. Regulation of
respiration when the oxygen availability changes. Physiologia
Plantarum 137, 383–391.
Hochachka PW. 1991. Metabolic strategies of defence against
hypoxia in animals. In: Jackson MB, Davies DD, Lambers H, eds. Plant
life under oxygen deprivation. The Hague: SPB Academic Publishing,
122–128.
Huang B, Johnson JW, Nesmith S, Bridges DC. 1994. Growth,
physiological and anatomical responses of two wheat genotypes to
waterlogging and nutrient supply. Journal of Experimental Botany 45,
193–202.
Jackson MB, Drew MC. 1984. Effects of flooding on growth and
metabolism of herbaceous plants. In: Kozlowski TT, ed. Flooding and
plant growth. New York: Academic Press, 47–128.
Jackson MB. 2008. Ethylene-promoted elongation: an adaptation to
submergence stress. Annals of Botany 101, 229–248.
Jiang Z, Song X-F, Zhou Z-Q, Wang L-K, Li J-W, Deng X-Y,
Fan H-Y. 2010. Aerenchyma formation: programmed cell death in
adventitious roots of winter wheat (Triticum aestivum) under
waterlogging. Functional Plant Biology 37, 748–755.
John CD, Limpinuntana V, Greenway H. 1974. Adaptation of rice
to anaerobiosis. Australian Journal of Plant Physiology 1, 513–520.
Kotula L, Ranathunge K, Steudle E. 2009. Apoplastic barriers
effectively block oxygen permeability across outer cell layers of rice
roots under deoxygenated conditions: roles of apoplastic pores and of
respiration. New Phytologist 184, 909–917.
Kuiper P, Walton CS, Greenway H. 1994. Effects of hypoxia on ion
uptake by nodal and seminal wheat roots. Plant Physiology and
Biochemistry 32, 267–276.
Lauchli A, James RA, Huang CX, McCully M, Munns R. 2008.
Cell-specific localization of Na+ in roots of durum wheat and possible
control points for salt exclusion. Plant, Cell and Environment 31,
1565–1574.
Limpinuntana V, Greenway H. 1979. Sugar accumulation in barley
and rice grown in solutions of low concentrations of oxygen. Annals of
Botany 43, 373–381.
Liu H-Y, VanToai T, Moy LP, Bock G, Linford LD,
Quackenbush J. 2005. Global transcription profiling reveals
comprehensive insights into hypoxic response in Arabidopsis. Plant
Physiology 137, 1115–1129.
Malik AI, Colmer TD, Lambers H, Schortemeyer M. 2003.
Aerenchyma formation and radial O2 loss along adventitious roots of
wheat with only the apical root portion exposed to O2-deficiency.
Plant, Cell and Environment 26, 1713–1722.
Malik AI, Colmer TD, Lambers H, Setter TL, Schortemeyer M.
2002. Short-term waterlogging has long-term effects on the growth
and physiology of wheat. New Phytologist 153, 225–236.
McDonald MP, Galwey NW, Colmer TD. 2001. Waterlogging
tolerance in the tribe Triticeae: the adventitious roots of Critesion
marinum have a relatively high porosity and a barrier to radial oxygen
loss. Plant, Cell and Environment 24, 585–596.
McDonald MP, Galwey NW, Colmer TD. 2002. Similarity and
diversity in adventitious root anatomy as related to root aeration
among a range of wet- and dry- land grass species. Plant, Cell and
Environment 25, 441–451.
Munns R, Tester M. 2008. Mechanisms of salinity tolerance. Annual
Review of Plant Biology 59, 651–681.
Nobel PS. 1974. Introduction to biophysical plant physiology. San
Francisco: WH Freeman.
Pang JY, Newman I, Mendham N, Zhou M, Shabala S. 2006.
Microelectrode ion and O2fluxes reveal differential sensitivity of barley
root tissues to hypoxia. Plant, Cell and Environment 29, 1107–1121.
Pang J, Ross J, Zhou M, Mendham N, Shabala S. 2007.
Amelioration of detrimental effects of waterlogging by foliar nutrient
sprays in barley. Functional Plant Biology 34, 221–227.
Ion transport in hypoxic roots | 57
Pang J, Shabala S. 2010. Membrane transporters and waterlogging
tolerance. In: Mancuso S, Shabala S, eds. Waterlogging signalling and
tolerance in plants. Heidelberg: Springer, 197–219.
Pang J, Zhou M, Mendham N, Shabala S. 2004. Growth and
physiological responses of six barley genotypes to waterlogging and
subsequent recovery. Australian Journal of Agricultural Research 55,
895–906.
Ponnamperuma FN. 1984. Effects of flooding on soils. In: Kozlowski
TT, ed. Flooding and plant growth. New York: Academic Press, 9–45.
Rubinigg M, Stulen I, Elzenga JTM, Colmer TD. 2002. Spatial
patterns of radial oxygen loss and nitrate net flux along adventitious
roots of rice raised in stagnant or aerated solutions. Functional Plant
Biology 29, 1475–1481.
Russell R, Barber DA. 1960. The relationship between salt uptake
and the absorption of water of intact plants. Annual Review of Plant
Physiology 11, 127–140.
triticale (Triticosecale cv. Muir) to waterlogging. New Phytologist 120,
335–344.
Thomson CJ, Greenway H. 1991. Metabolic evidence for stellar
anoxia in maize roots exposed to low O2 concentrations. Plant
Physiology 96, 1294–1301.
Tregeagle JM, Tisdall JM, Tester M, Walker RR. 2010. Cl– uptake,
transport and accumulation in grapevine rootstocks of differing
capacity for Cl–-exclusion. Functional Plant Biology 37, 665–673.
Trought MCT, Drew MC. 1980a. The development of waterlogging
damage in young wheat plants in anaerobic solution cultures. Journal
of Experimental Botany 31, 1573–1585.
Trought MCT, Drew MC. 1980b. The development of waterlogging
damage in wheat seedlings (Triticum aestivum L.). II. Accumulation
and redistribution of nutrients by the shoot. Plant and Soil 56,
187–199.
Setter TL, Waters I. 2003. Review of prospects for germplasm
improvement for waterlogging tolerance in wheat, barley and oats.
Plant and Soil 253, 1–34.
Trought MCT, Drew MC. 1980c. The development of waterlogging
damage in wheat seedlings (Triticum aestivum L.). I. Shoot and root
growth in relation to changes in the concentrations of dissolved gases
and solutes in the soil solution. Plant and Soil 54, 77–94.
Setter TL, Waters I, Atwell BJ, Kupkanchanakul T, Greenway H.
1987. Carbohydrate status of terrestrial plants during flooding. In:
Crawford RMM, ed. Plant life in aquatic and amphibious habitats.
Oxford: Blackwell Scientific, 411–443.
Trought MCT, Drew MC. 1981. Alleviation of injury to young wheat
plants in anaerobic nutrient solution in relation to the supply of nitrate
and other inorganic nutrients. Journal of Experimental Botany 32,
509–522.
Setter TL, Waters I, Sharma SK, et al. 2009. Review of wheat
improvement for waterlogging tolerance in Australia and India: the
importance of anaerobiosis and element toxicities associated with
different soils. Annals of Botany 103, 221–235.
Watkin ELJ, Thomson CJ, Greenway H. 1998. Root development
and aerenchyma formation in two wheat cultivars and one Triticale
cultivar grown in stagnant agar and aerated nutrient solution. Annals of
Botany 81, 349–354.
Shabala S, Shabala S, Cuin TA, Pang J, Percey W, Chen Z,
Conn S, Eing C, Wegner LH. 2010. Xylem ionic relations and salinity
tolerance in barley. The Plant Journal 61, 839–853.
Wiengweera A, Greenway H. 2004. Performance of seminal and
nodal roots of wheat in stagnant solution: K+ and P uptake and effects
of increasing O2 partial pressures around the shoot on nodal root
elongation. Journal of Experimental Botany 55, 2121–2129.
Shiono K, Takahashi H, Colmer TD, Nakazono M. 2008. Role of
ethylene in acclimations to promote oxygen transport in roots of plants
in waterlogged soils. Plant Science 175, 52–58.
Smirnoff N. 1995. Antioxidant systems and plant responses to the
environment. In: Smirnoff N, ed. Environment and plant metabolism:
flexibility and acclimation. Oxford: BIOS Scientific Publishers, 217–243.
Socolar SJ. 1977. The coupling coefficient as an index of junctional
conductance. Journal of Membrane Biology 34, 29–37.
Thomson CJ, Armstrong W, Waters I, Greenway H. 1990.
Aerenchyma formation and associated oxygen movement in seminal
and nodal roots of wheat. Plant, Cell and Environment 13, 395–403.
Thomson CJ, Colmer TD, Watkins ELJ, Greenway H. 1992.
Tolerance of wheat (Triticum aestivum cvs Gamenya and Kite) and
Wiengweera A, Greenway H, Thomson CJ. 1997. The use of agar
nutrient solution to simulate lack of convention in waterlogged soil.
Annals of Botany 80, 115–123.
Xia JH, Roberts JKM. 1996. Regulation of H+ extrusion and
cytoplasmic pH in maize root tips acclimated to a low-oxygen
environment. Plant Physiology 111, 227–233.
Yeo AR, Yeo ME, Flowers TJ. 1987. The contribution of an
apoplastic pathway to sodium uptake by rice roots in saline conditions.
Journal of Experimental Botany 38, 1141–1153.
Zhang WH, Tyerman SD. 1997. Effect of low oxygen concentration
on the electrical coupling of cortical cells of wheat roots. Journal of
Plant Physiology 150, 567–572.