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Ann Bot. 2009 December; 104(7): 1435–1444.
Published online 2009 October 23. doi:  10.1093/aob/mcp257
PMCID: PMC2778401

Photosynthetic acclimation is important for post-submergence recovery of photosynthesis and growth in two riparian species

Abstract

Background and Aims

Concomitant increases in O2 and irradiance upon de-submergence can cause photoinhibition and photo-oxidative damage to the photosynthetic apparatus of plants. As energy and carbohydrate supply from photosynthesis is needed for growth, it was hypothesized that post-submergence growth recovery may require efficient photosynthetic acclimation to increased O2 and irradiance to minimize photo-oxidative damage. The hypothesis was tested in two flood-tolerant species: a C3 herb, Alternanthera philoxeroides; and a C4 grass, Hemarthria altissima. The impact of low O2 and low light, typical conditions in turbid floodwater, on post-submergence recovery was assessed by different flooding treatments combined with shading.

Methods

Experiments were conducted during 30 d of flooding (waterlogging or submergence) with or without shading and subsequent recovery of 20 d under growth conditions. Changes in dry mass, number of branches/tillers, and length of the longest internodes and main stems were recorded to characterize growth responses. Photosynthetic parameters (photosystem II efficiency and non-photochemical quenching) were determined in mature leaves based on chlorophyll a fluorescence measurements.

Key Results

In both species growth and photosynthesis recovered after the end of the submergence treatment, with recovery of photosynthesis (starting shortly after de-submergence) preceding recovery of growth (pronounced on days 40–50). The effective quantum yield of photosystem II and non-photochemical quenching were diminished during submergence but rapidly increased upon de-submergence. Similar changes were found in all shaded plants, with or without flooding. Submerged plants did not suffer from photoinhibition throughout the recovery period although their growth recovery was retarded.

Conclusions

After sudden de-submergence the C3 plant A. philoxeroides and the C4 plant H. altissima were both able to maintain the functionality of the photosynthetic apparatus through rapid acclimation to changing O2 and light conditions. The ability for photosynthetic acclimation may be essential for adaptation to wetland habitats in which water levels fluctuate.

Key words: Aerenchyma, Alternanthera philoxeroides, flooding, growth, Hemarthria altissima, low light, photosynthesis, shade, submergence, waterlogging, wetland plant

INTRODUCTION

Flooding is detrimental to many terrestrial plants. Soil flooding disrupts the metabolism of mesophytic plants by displacing O2 from soil pores and promoting O2 depletion by roots and soil microbes (Drew, 1990). Partial to complete submergence imposes further stress by dramatically reducing gas exchange in above-ground parts of plants, exacerbating O2 shortage (Vartapetian and Jackson, 1997; Blom, 1999; Colmer and Pedersen, 2008a). In addition, low irradiance is another important factor which affects survival of submerged plants. Turbid floodwater considerably lowers light energy reaching submerged plants, causing severe energy limitation (Vervuren et al., 2003; Mommer et al., 2005a). For example, at a depth of 25 cm in highly turbid floodwater, light intensity can already be below the photosynthetic light compensation point (Setter et al., 1987). As oxygenic photosynthesis of chloroplasts provides not only carbohydrates but also O2 in submerged plants (Mommer et al., 2007), low light doubly restricts the underwater performance of plants by decreasing the availability of energy as well as O2.

Although certain morphological responses are induced by both submergence stress and shade stress, such as increased specific leaf area, vertical leaf orientation or petiole elongation, acclimation and adaptation to submergence are directed predominantly to an improved capacity for gas exchange rather than light capture (Mommer et al., 2005a). Altered morphology and anatomy of submerged leaves can enhance underwater gas exchange (Mommer et al., 2005a, b; Voesenek et al., 2006; Colmer and Pedersen, 2008b). Moreover, submerged plants of some species show stimulated elongation of shoot organs to restore contact with the atmosphere above the water surface (Benschop et al., 2005; Jackson, 2008). Formation of longitudinally interconnected gas spaces in roots and shoots (aerenchyma, sometimes also pith cavity), which facilitates internal gas diffusion, is found in many flood-tolerant species (Colmer, 2003; Evans, 2003; Jung et al., 2008).

Along with the abilities to cope with limited O2, CO2 and energy availability during flooding, the capacity to quickly resume normal (aerobic) physiological and metabolic activities after the removal of floodwater is another important criterion for flooding tolerance (Gibbs and Greenway, 2003; van Eck et al., 2004). Sudden re-exposure to ambient O2 levels can result in increased production of reactive oxygen species (Wollenweber-Ratzer and Crawford, 1994; Benschop et al., 1998) and acetaldehyde (Zuckermann et al., 1997; Tsuji et al., 2003) in plant tissues acclimated to low O2 conditions. For submerged plants, this large increase in O2 concentration can be accompanied by an even larger and abrupt increase in light intensity, which threatens their photosynthetic machinery accustomed to low light conditions. High-light-induced damage to the photosynthetic apparatus (photoinhibition; Osmond, 1994) could impede post-submergence recovery of photosynthesis. Upon returning to terrestrial conditions, plants must overcome these problems of concomitant increase in O2 and irradiance to grow and store sufficient carbohydrates before the next flooding event.

Although our understanding of the mechanisms of flooding survival in plants has advanced remarkably (Gibbs and Greenway, 2003; Bailey-Serres and Voesenek, 2008), much less is known about post-flooding recovery of growth and photosynthesis in flood-tolerant plants, even though these carbon acquisition and accumulation processes are essential for species performance during flood intervals, and further, for survival of the future flooding. We hypothesized that rapid growth recovery after de-submergence may require the ability to acclimate the photosynthetic apparatus to increased O2 and irradiance to minimize photo-oxidative damage. This hypothesis was tested in two flood-tolerant perennial species: a C3 herb, Alternanthera philoxeroides; and a C4 grass, Hemarthria altissima. These species were chosen to see whether C3 and C4 pathways affect photoacclimatory responses during and after flooding. The impact of low O2 and low light on growth and photosynthesis recovery was assessed by using three different water levels for flooding (well-drained, soil flooding and complete submergence) with or without shading.

MATERIALS AND METHODS

Plant materials

Alternanthera philoxeroides (Mart.) Griseb. is an exotic invasive perennial C3 weed in China, which can establish in aquatic, semi-aquatic and terrestrial environments including dry farmland (Allen et al., 2007; Gao et al., 2008). Hemarthria altissima (Poir.) Stapf & C.E. Hubb. is a perennial C4 grass with long spreading stolons and short rhizomes. Plants of H. altissima prefer moist soils and occur along river banks, but are also capable of growing in dry land (Wang et al., 2005; Yang et al., 2007). Both species are found in the zone of water-level fluctuations of the Three Gorges Reservoir area of China. For the present study, plants were collected in the Three Gorges Reservoir area and brought to the institute Phytosphäre, Forschungszentrum Jülich, Germany. Plants of A. philoxeroides were propagated from cuttings and those of H. altissima from tillers. Young plants were then planted in pots (2·2 L, one plant per pot) containing TYP ED 73 soil (mixture of clay and white peat, with pre-mixed fertilizer enough for 2 to 3 months of plant cultivation; Einheitserdewerk, Fröndenberg, Germany) and allowed to grow for 1 month in the glasshouse prior to the experiments.

Plant cultivation and experimental conditions

All experiments were conducted under semi-controlled conditions in the glasshouse. Daily air temperature and relative humidity ranged between 20–22 °C and 40–60 % during the experimental period. Illumination in the glasshouse (SON-T AGRO 400, Philips, Eindhoven, the Netherlands) was automatically turned on when the ambient light intensity outside the glasshouse became <400 µmol photons m−2 s−1 between 0600 and 2200 h local time. Under such conditions, the photosynthetically active radiation measured at the plant level was typically 200–300 µmol photons m−2 s−1. Plants were watered daily during cultivation.

Experimental design

For the experiments, 114 and 130 plants of A. philoxeroides and H. altissima, respectively, were randomly selected. Three different flooding treatments were applied: ‘Control’ (no flooding), waterlogging (‘Waterlogged’, water level at the soil surface) and submergence (‘Shaded-submerged’, fully submerged under 1-m-deep water and covered with a neutral shade cloth). The shade cloth was used to mimic low-light environments typically found in turbid floodwater; the light intensity beneath the shade cloth was 3–8 µmol photons m−2 s−1. The same shading treatment was also applied in combination with Control (‘Shaded-control’) and Waterlogged treatments (‘Shaded-waterlogged’) to assess the impact of low irradiance on growth and photosynthesis. Flooding treatments were conducted in plastic tanks (72 and 480 L in volume for waterlogging and submergence, respectively), with six plants per tank. Care was taken to keep plants completely underwater for Shaded-submerged treatment. Following 30 d of different flooding treatments with or without shading, plants were transferred back to the growth conditions and recovery was monitored for 20 d.

Physico-chemical properties of floodwater

Physico-chemical properties of floodwater were checked in the morning (1000–1200 h) at the beginning of the experiment (day 0) and during the submergence treatment (day 20). Water samples were taken immediately before the measurements at the half depth of the floodwater (at approx. 50 cm deep) in three replicate submergence tanks. The inorganic carbon concentrations, including dissolved CO2, HCO3 and CO32−, were estimated by using the titration method described by Klee (1998). The level of dissolved O2 was measured with an O2 electrode (AM 39, Sensortechnik Meinsberg, Germany). Temperature and pH were also determined to calculate CO2 and O2 levels. Mean values are summarized in Table 1.

Table 1.
Physico-chemical properties of floodwater in submergence tanks

Light microscopy of transverse sections

Transverse sections of root and stem/culm tissues were hand-sectioned immediately before observation. The positions of the sections were 3 cm above and 3 cm below the soil surface for stem/culm and root, respectively. Sections were observed under a light microscope Zeiss Axiophot2 (Carl Zeiss, Jena, Germany) and images were taken by a digital camera (D200, Nikon, Tokyo, Japan).

Growth analyses

Dry weight data were collected at the beginning and at the end of the flooding treatments (day 0 and 30, respectively) as well as after 10 and 20 d of recovery under the growth conditions (day 40 and 50, respectively). At each sampling point, 8–10 plants were harvested for every treatment. The plant materials were put in an oven at 75 °C until a constant mass was reached. The parameters used for growth analyses were: above- and below-ground dry weight, total number of branches/tillers, and length of the longest internodes and main stem.

Chlorophyll a fluorescence measurements

Chlorophyll (Chl) a fluorescence analysis can provide information about photoinhibition, photochemical efficiency and photoprotective energy dissipation of photosystem II (PSII) (Schreiber and Bilger, 1993; Maxwell and Johnson, 2000). Measurements were performed in the glasshouse in the morning (0900–1100 h) every other day during the experiments. For Shaded-submerged plants, no measurements were taken during submergence. Fluorescence was measured on youngest, fully expanded leaves (i.e. mature leaves closest to the growth zone of the shoot) by using a Handy PEA instrument (Hansatech Instruments, King's Lynn, UK). The maximal and minimal fluorescence intensity of dark-adapted leaves (Fm and F0, respectively) were determined after 30 min of dark adaptation by using leaf clips. The maximal and steady-state fluorescence intensity of light-adapted leaves (Fm′ and Fs, respectively) were determined after 4·5 min illumination at 800 µmol photons m−2 s−1 with the built-in red light source of the PEA instrument. The intensity and duration of the saturation pulse applied to determine Fm and Fm′ were 3500 µmol photons m−2 s−1 and 1 s, respectively. The maximal quantum yield of PSII in the dark (Fv/Fm) was calculated as Fv/Fm = (FmF0)/Fm, the effective quantum yield of PSII in the light (ΔF/ Fm′) as ΔF/ Fm′ = (Fm′Fs)/Fm′, and the non-photochemical energy quenching in PSII (NPQ) as NPQ = (FmFm′)/Fm′ (Maxwell and Johnson, 2000).

Statistical data analysis

Growth data were checked for normal distribution and equal variance between populations by using SigmaStat 2·0 (SPSS Inc., Chicago, IL, USA). The data were then statistically tested for differences between treatments (having the same sampling size) by analysis of variance (one-way ANOVA) or for differences between sampling times (having slightly different sampling sizes) by the Kruskal–Wallis test (both in SigmaStat 2·0). As the difference in sampling size was minimal in our experiment (n = 8–10), the Kruskal–Wallis test gave essentially the same results as one-way ANOVA.

RESULTS

Changes in physico-chemical properties of floodwater during the submergence treatment

Shaded-submerged treatment decreased the pH value of floodwater (Table 1). Accordingly, the HCO3 concentration in the floodwater decreased slightly; by contrast, the CO2 concentration increased. When CO2 and HCO3 levels were added together, there was a minor decrease in the total inorganic carbon from day 0 to day 20 (from 467 to 440 µmol L−1). Amounts of CO32− detected in the floodwater were negligible (<1 µmol L−1, data not shown). The dissolved O2 concentration changed little in the submergence tanks.

Shoot and root anatomy

Both A. philoxeroides and H. altissima are terrestrial plants which can grow well in wetland. Transverse sections prepared from shoot/culm and root tissues of Shaded-submerged plants showed a well-developed pith cavity and aerenchyma (Fig. 1), suggesting structural adaptation to wetland habitats. The samples were taken at 30 mm above and 30 mm below the soil surface for stems/culms and roots, respectively. These tissues were formed during cultivation in the glasshouse, i.e. they are not aquatic shoots/culms or roots which grew during Shaded-submerged treatment. In fact, different flooding treatments did not visibly alter the number and/or size of pith cavity and aerenchyma during the 30-d flooding treatments, and similar anatomy was also observed in well-drained Control plants (data not shown). Thus, it seems that these plants have porous shoots and roots constitutively.

Fig. 1
Transverse sections of stem/culm (A, B) and root tissues (C, D) of Shaded-submerged plants of A. philoxeroides (A, C) and H. altissima (B, D). Sections were made at 30 mm above and 30 mm below the soil surface for stems/culms and roots, respectively. ...

Growth responses to flooding with or without shading

Growth responses to different flooding treatments with or without shading were compared in A. philoxeroides and H. altissima during 50 d of the flooding and recovery experiments. Above- and below-ground dry mass increased in Control plants of A. philoxeroides >20-fold during the experiment (Fig. 2A and B, respectively). Waterlogged treatment resulted in a significantly smaller dry mass increase compared with Control. Plants of A. philoxeroides have a repent growth form, and the contact of shoots with the floodwater in the waterlogging tank stimulated the formation of adventitious roots at stem nodes. On day 30 these adventitious roots amounted to 7–10 % of the total root fresh weight for Waterlogged plants (data not shown). When shaded (Shaded-control, Shaded-waterlogged and Shaded-submerged), A. philoxeroides virtually did not accumulate dry mass, with or without flooding. Yet the dry mass increase started between day 30 and 50, with the recovery in Shaded-submerged plants being slower than in other shaded plants both above and below ground. The root-to-shoot ratio increased in Control plants of A. philoxeroides with increasing plant size (Fig. 2C), presumably due to ‘ontogenic drift’ (Geng et al., 2007). The ratios strongly decreased in Shaded-control and Shaded-waterlogged plants from day 0 to 30, whereas they did not change significantly in Waterlogged and Shaded-submerged plants. The ratio then increased during the recovery period in all treatments (except for Control) to reach Control-like values by day 50 in all but Shaded-submerged plants.

Fig. 2
Changes in above-ground dry mass (A, D), below-ground dry mass (B, E) and root-to-shoot ratio (C, F) of A. philoxeroides (A–C) and H. altissima (D–F) following the different flooding treatments with or without shading. Each value is a ...

The Control plants of H. altissima accumulated much less dry mass than Control plants of A. philoxeroides: <10-fold for above-ground and <6-fold for below-ground (Fig. 2D and E, respectively) dry mass. As in A. philoxeroides, dry mass accumulation of Waterlogged plants was less than in Control plants, and shading allowed no increment of above- and below-ground dry mass, with or without flooding. Accumulation of dry mass started in shaded plants after day 30, but the major increase occurred between day 40 and 50. The apparent lack of change in above-ground dry mass of Shaded-submerged plants between day 30 and 40 (Fig. 2D) actually contained growth of some new leaves which replaced mature leaves lost shortly after de-submergence. Unlike in A. philoxeroides, the root-to-shoot ratio decreased in Control plants of H. altissima from day 0 to 30 (Fig. 2F). While retarding the dry mass increase, Waterlogged treatment did not affect the root-to-shoot balance in H. altissima. Shading slowed the decrease in root-to-shoot ratio, but the values for Shaded-control and Shaded-waterlogged plants approached those of Control and Waterlogged plants within the first 10 d of recovery. Only Shaded-submerged plants maintained higher values throughout the experiment. In general, the root-to-shoot ratio of H. altissima was about half that of A. philoxeroides on day 50 (Fig. 2C, F), indicating that relative dry mass allocation to shoots was greater in H. altissima than in A. philoxeroides, and vice versa for relative allocation to roots, during the experiments.

The responses of above-ground growth patterns to different treatments were further characterized by comparing the number of branches (A. philoxeroides) or tillers (H. altissima). The total number of branches increased from day 0 to 30 in Control and Waterlogged plants of A. philoxeroides by about four- and three-fold, respectively (Fig. 3A). Shading suppressed the formation of new branches, but Shaded-control and Shaded-waterlogged plants were able to produce many new branches after the end of the treatments. These plants had as many branches as Control and Waterlogged plants by day 50 even though their above-ground dry mass was still much less (Fig. 2A). Recovery of branching was far slower in Shaded-submerged plants, having only half as many branches as Control plants at the end of the recovery period (Fig. 3A). The above-ground growth of H. altissima was characterized by a large increase (about eight-fold in Control plants) in total tiller number (Fig. 3B). Tiller formation in this grass species responded very sensitively to flooding as well as to shading treatments; it was strongly impaired in Waterlogged treatment and was completely halted until day 40 in Shaded-control and Shaded-waterlogged treatments. Shaded-submerged plants did not grow new tillers during 20 d of the recovery period.

Fig. 3
Changes in the total branch/tiller number of A. philoxeroides (A) and H. altissima (B) following the different flooding treatments with or without shading. Each value is a mean of 8–10 plants (±s.e.). All plants were under the Control ...

Although suppressing the accumulation of dry mass and formation of new branches, 30 d of submergence strikingly enhanced elongation of internodes in A. philoxeroides (Fig. 4A). Even though dry mass accumulation was similarly inhibited in all shaded plants, only Shaded-submerged plants had longer internodes than Control plants (by approx. 20 %) on day 30. By elongating the existing internodes (i.e. without formation of new internodes) Shaded-submerged plants increased the total length of the main stem by about 40 cm (or 200 % of the initial length on day 0), while the corresponding increment in Shaded-control and Shaded-waterlogged plants was only about 15 cm (Fig. 4B). The main stems of Control and Waterlogged plants grew much longer than those of shaded plants, but this was achieved by making new internodes (data not shown). In marked contrast, variations in the internode length among different treatments were minimal for H. altissima (Fig. 4C). The main culms grew longer only in Control and Waterlogged plants, which produced several new internodes during the period (Fig. 4D).

Fig. 4.
Length of the longest internodes (A, C) and the main stem/culm (B, D) for A. philoxeroides (A, B) and H. altissima (C, D) at the end of the different flooding treatments with or without shading. Each value is a mean of 8–10 plants (±s.e.). ...

Photosynthetic responses to flooding with or without shading

The above results demonstrated the ability of A. philoxeroides and H. altissima to resume growth and dry mass accumulation relatively quickly after de-submergence. According to the present hypothesis, the photosynthetic apparatus of these plants must be able to cope with extreme changes in O2 and light. However, growth recovery was retarded in Shaded-submerged plants compared with Shaded-control and Shaded-waterlogged plants (Figs 2 and and3),3), which may reflect slower acclimation and recovery of photosynthesis due to detrimental effects of submergence on the photosynthetic apparatus.

The maximal quantum yield of PSII (Fv/Fm) indicated no significant photoinhibition in Waterlogged plants of A. philoxeroides throughout the experiment (Fig. 5A). Shaded-submerged plants had slightly lower Fv/Fm values shortly after de-submergence, but Fv/Fm fully recovered in these plants on day 39. Shading quickly increased Fv/Fm to the maximal level (approx. 0·84) in both Shaded-control and Shaded-waterlogged plants, whereas the transfer back to the growth conditions transiently decreased Fv/Fm to the values found in Shaded-submerged plants (Fig. 5B). Thereafter, Fv/Fm fully recovered in Shaded-control and Shaded-waterlogged plants by day 37, i.e. 2 d earlier than in Shaded-submerged plants. In contrast to the situation in A. philoxeroides, Waterlogged treatment markedly decreased Fv/Fm in H. altissima in the first 20 d, suggesting some photoinhibitory damage to PSII (Fig. 5C). Yet Fv/Fm started to recover in the last 10 d of Waterlogged treatment to become comparable with that of Control plants 3 d after draining the soil. Fv/Fm values of Shaded-submerged plants during the recovery period were as high as those of Control plants. The shading responses of Fv/Fm in H. altissima were generally the same as described for A. philoxeroides, albeit with less pronounced changes (Fig. 5D). Full recovery of Fv/Fm was found on day 37 in Shaded-control and on day 39 in Shaded-waterlogged treatment.

Fig. 5.
Changes in the maximal quantum yield of photosystem II (Fv/Fm) in dark-adapted leaves of A. philoxeroides (A, B) and H. altissima (C, D) during and after different flooding treatments with or without shading. Data from Shaded-submerged plants are shown ...

Whereas Fv/Fm indicated little PSII photoinhibition in all but Waterlogged plants of H. altissima, the effective quantum yield of PSII (ΔF/Fm′) revealed a strikingly reduced capacity of shaded plants to utilize light energy under illumination (Fig. 6). Waterlogged treatment had no significant effect on ΔF/Fm′ in A. philoxeroides (Fig. 6A). The shading treatment caused a rapid and dramatic decrease in ΔF/Fm′ in Shaded-control and Shaded-waterlogged plants to a level as low as that measured in Shaded-submerged plants (Fig. 6B). Recovery of ΔF/Fm′ started in all shaded plants of A. philoxeroides within 3 d after the end of the treatments, although it took approx. 10 d to recover fully. In comparison, H. altissima generally had much lower photochemical efficiency than A. philoxeroides (Fig. 6C, D). The pronounced negative effect of Waterlogged treatment was also visible in ΔF/Fm′ of this species (Fig. 6C). Unlike Fv/Fm, ΔF/Fm′ of H. altissima was substantially reduced to approx. 60 % of Control plants at the beginning of Shaded-control and Shaded-waterlogged treatments, or at the end of Shaded-submerged treatment (Fig. 6D). For both species, recovery of ΔF/Fm′ in Shaded-submerged treatment was not slower than in Shaded-control and Shaded-waterlogged treatments.

Fig. 6.
Changes in the effective quantum yield of photosystem II (ΔF/Fm′) in leaves of A. philoxeroides (A, B) and H. altissima (C, D) during and after different flooding treatments with or without shading. Values were obtained after 4·5 ...

When a large fraction of absorbed light energy becomes excessive due to low ΔF/Fm′, leaves may up-regulate NPQ to protect the photosynthetic apparatus and minimize photoinhibition. Shading substantially diminished the NPQ capacity in both A. philoxeroides and H. altissima, but rapid recovery was observed after day 30 (Fig. 7). The changes in NPQ paralleled the large variations in Fv/Fm and ΔF/Fm′ in Waterlogged plants of H. altissima (Fig. 5C). Again, the NPQ recovery of Shaded-submerged plants did not differ from that in other shaded plants for both species (Fig. 7B, D).

Fig. 7.
Changes in non-photochemical energy quenching (NPQ) in leaves of A. philoxeroides (A, B) and H. altissima (C, D) during and after different flooding treatments with or without shading. Values were obtained after 4·5 min of illumination at 800 ...

DISCUSSION

Post-submergence recovery of growth and photosynthesis

During the 20-d recovery period the total dry mass of Shaded-submerged plants increased by 255 and 148 % in A. philoxeroides and H. altissima, respectively. This shows that A. philoxeroides could achieve a greater relative growth rate than H. altissima after de-submergence. Similar increases in dry mass, ranging between 22·1 and 305·6 %, have been reported for some species found in low-elevation grassland along the Rhine River during 3 weeks of recovery after 30 d of submergence (van Eck et al., 2004).

Post-submergence growth recovery, as observed in dry mass (Fig. 2) and formation of new branches and tillers (Fig. 3), was preceded by recovery of ΔF/Fm′ and NPQ in both A. philoxeroides and H. altissima (Figs 557). Yet the slowest growth resumption of Shaded-submerged plants was not accompanied by a corresponding delay in photosynthetic recovery; the fluorescence parameters indicated comparable recovery in all shaded plants after day 30 (Figs 557). This means that the retarded growth recovery of Shaded-submerged plants was not a result of slow photosynthetic acclimation. The leaves of these wetland plants were able to quickly adjust the capacities of photosynthetic electron transport and dissipation to prevailing light environments (Figs 6 and and7),7), and there was no sign of severe photoinhibition after the end of the submergence and/or shading treatments (Fig. 5). The ability for flexible photosynthetic acclimation may be essential for wetland plants living in fluctuating water, in which the availability of light, CO2 and O2 changes dramatically and periodically.

There was no significant difference in post-submergence recovery patterns of growth and photosynthesis between the C3 plant A. philoxeroides and the C4 plant H. altissima in the present experiments. Thus, both the C3 and the C4 pathway seem to be compatible with the metabolic and morphological adjustment involved in flooding tolerance. The only differences found between the two species were the generally lower ΔF/Fm′ in H. altissima and its higher sensitivity to waterlogging. It is likely that the lower PSII efficiency in the light (Fig. 6C, D) contributed to the smaller relative dry mass increase in H. altissima compared with A. philoxeroides during recovery. Decreased stomatal conductance under soil flooding, as has been demonstrated in some woody species (Mielke et al., 2003), may have restricted leaf gas exchange in Waterlogged plants of H. altissima, resulting in photoinhibition under the ambient light in the glasshouse (Fig. 5C).

What, then, could be the reason for the slower growth recovery in Shaded-submerged plants? Shaded-submerged plants may have consumed more carbohydrates and energy than Shaded-control and Shaded-waterlogged plants during the 30-d treatments, leading to severely reduced carbohydrate availability for growth at the beginning of the recovery period. Additionally, or alternatively, limited O2 supply may have deteriorated root functionality (Vartapetian and Jackson, 1997) to delay growth recovery and cause loss of leaves upon de-submergence. Post-submergence injuries due to accumulation of reactive oxygen species (Wollenweber-Ratzer and Crawford, 1994; Benschop et al., 1998) and acetaldehyde (Zuckermann et al., 1997; Tsuji et al., 2003) are also potential factors contributing to growth retardation. These possible causes of post-submergence growth retardation in A. philoxeroides and H. altissima will be investigated in future studies.

Flooding tolerance in A. philoxeroides and H. altissima

Flood-tolerant species can grow at low elevations along the floodplain while less flood-tolerant species are confined to higher elevations of the floodplain gradient (van Eck et al., 2004). The two riparian species examined here, A. philoxeroides and H. altissima, occur in low-elevation sites of river banks and are capable of enduring long-term submergence (Wang et al., 2008a, b). However, different survival rates, 50 and 90 % for A. philoxeroides and H. altissima, respectively, have been reported after 180 d of complete submergence (Wang et al., 2008a, b). Although the recovery of dry mass accumulation after 30 d of submergence was greater in A. philoxeroides than in H. altissima in the present study, the better success of H. altissima than A. philoxeroides under very long submergence, as in the previous studies by Wang et al. (2008a, b), may be explained by the contrasting growth responses of these two plants during submergence.

Submergence enhanced stem elongation in A. philoxeroides (Fig. 4A, B), a morphological response observed in certain flood-tolerant species (Mommer et al., 2005b; Jackson, 2008). Rapid stem elongation serves to regain its contact with the atmosphere or well-oxygenated upper water layers. Once contact with the atmosphere is established in relatively shallow water, these plants can perform efficient photosynthesis and survive prolonged submergence (Voesenek et al., 2004). Strongly elongated stems and petioles are also found in plants under shade environments (‘shade avoidance’; Ballaré et al., 1999; Valladares, 2003; Franklin and Whitelam, 2005). However, plants of A. phioloxeroides in Shaded-control and Shaded-waterlogged treatments did not exhibit strong stem elongation in the present study (Fig. 4A, B). This shows that the elongation response of A. phioloxeroides was induced by submergence, not by shading, probably via ethylene signalling (Benschop et al., 2005; Mommer et al., 2005a; Cox et al., 2006; Jackson, 2008). It has been demonstrated that submergence-induced elongation is different from shade-induced elongation; the former can occur at relatively high light intensities (e.g. 100 µmol photons m−2 s−1) and high red/far-red ratios (Pierik et al., 2005) whereas the latter is induced by low red/far-red ratios (Ballaré et al., 1999; Valladares, 2003; Franklin and Whitelam, 2005).

The escape strategy requires not only contact with the atmosphere but also porous shoots and roots to allow internal gas diffusion (Fig. 1A, C; Colmer, 2003; Evans, 2003; Jung et al., 2008). The importance of petiole porosity for the escape strategy has been recently demonstrated in species of Rumex (Pierik et al., 2009); whereas R. palustris, having sufficient aerenchyma in the petiole, accumulated biomass in shoots during submergence, R. acetosa, a congeneric species without such anatomy, did not. In shallow floodwater, in which elongated shoots of A. philoxeroides can reach the atmosphere, this species may outperform H. altissima by the ‘escape’ strategy, even though the submergence-induced stem elongation may be costly under energy limitation. With increasing depth of floodwater and decreasing availability of light, the advantage of the escape strategy may be outweighed by its cost.

Plants of H. altissima did not elongate internodes in response to submergence (Fig. 4C). Instead of trying to escape, this plant may have adopted a ‘quiescence’ strategy to survive long submergence (Geigenberger, 2003; Voesenek et al., 2006). There were indications that H. altissima is not only flood-tolerant but also shade-tolerant, a beneficial trait for a quiescence strategy (Geigenberger, 2003; Voesenek et al., 2006). For example, the capacity of PSII electron transport measured in H. altissma was less than half that of A. philoxeroides (Fig. 6). Further studies are needed to elucidate the physiological mechanisms of the pronounced long-term flood tolerance of H. altissima.

CONCLUDING REMARKS

The C3 plant A. philoxeroides and the C4 plant H. altissima are both able to maintain the functionality of the photosynthetic apparatus after de-submergence through rapid acclimation to changing O2 and/or light conditions. The ability for photosynthetic acclimation may be essential for adaptation to wetland habitats in which the water level fluctuates. Although limited light availability had the major impact on photosynthesis and growth in our experiments, submergence delayed growth recovery more strongly than shading or the combination of soil flooding and shading. Physiological and metabolic factors determining the extent of post-submergence growth retardation in flood-tolerant species deserve further investigation.

ACKNOWLEDGEMENTS

F.-L. L. was supported by a PhD scholarship from the Deutsche Akademische Austausch Dienst. We thank Beate Uhlig (Institut für Phytosphäre, Forschungszentrum Jülich GmbH) for help with plant cultivation and harvesting. Two anonymous referees and the Handling Editor (Tim Colmer) provided valuable comments.

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