SEARCH

SEARCH BY CITATION

Keywords:

  • Trifolium;
  • aeration;
  • aerenchyma;
  • clover;
  • pasture;
  • radial O2 loss ;
  • root porosity;
  • waterlogging

ABSTRACT

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Trifolium tomentosum and T. glomeratum are small (< 0·5 mg) seeded pasture legumes which are considered to be waterlogging tolerant and intolerant, respectively. The root porosity of the two species was compared for plants raised for 10 d in aerated nutrient solution and then transferred to either aerated (0·25 mol O2 m–3) or ‘hypoxic’ (0·031–0·069 mol O2 m–3) solutions for a further 7 and 21 d. After 21 d, T. tomentosum developed a root porosity of 11·2% in ‘hypoxic’ solution, which was significantly higher than the 6·1% developed by T. glomeratum. When grown in aerated solution, T. tomentosum also had a larger constitutive porosity (6·7%) than T. glomeratum (3·9%). Cylindrical root-sleeving O2 electrodes were used to measure the rates of radial O2 loss (ROL) from roots of the two species when in an O2-free medium. In general, roots previously grown in ‘hypoxic’ solution had higher rates of ROL than roots grown in aerated solution. Moreover, the rates of ROL along the main root of T. tomentosum were ≈ 5-fold faster than from equivalent locations along roots of T. glomeratum. Manipulations of the shoot O2 concentration resulted in rapid changes in ROL near the root tip of T. tomentosum plants raised in aerated or ‘hypoxic’ solutions, whereas for T. glomeratum ROL only increased for roots of plants raised in ‘hypoxic’ solution. Thus, the cortical air spaces in roots of both species raised in ‘hypoxic’ solution formed a continuous, low resistance pathway for O2 diffusion from the shoots to the roots. ROL from the lateral roots was also evaluated and it was 3-fold faster from T. tomentosum than from T. glomeratum. Moreover, ROL from lateral roots of T. tomentosum was 10–20-fold higher than from a position on the primary root axis the same distance from the root/shoot junction. Relatively, high rates of ROL were also recorded for young (40 mm in length) lateral roots of T. glomeratum which were previously grown in ‘hypoxic’ solution, but the ROL was low for the older lateral roots of this species. The substantial amounts of ROL from the lateral roots may limit O2 supply to the lower parts of the primary root axis, so that the laterals probably become the main functional root system in waterlogged soils.


INTRODUCTION

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Waterlogged soils usually contain very low levels of O2, so that the formation of aerenchyma is regarded as one trait essential for root function in these soils ( Armstrong 1979). Aerenchyma are cortical airspaces which provide a low resistance internal pathway for the movement of O2 from the shoots to the roots, where it is consumed in respiration and may also re-oxidize the rhizosphere ( Armstrong 1970, 1971, 1979). In legumes, aerenchyma may also be important for supplying O2 and N2 to root nodules ( Walker, Pate & Kuo 1983; Zook, Ewin & Stolzy 1986; James, Minchin & Sprent 1992; Pugh et al. 1995 ). The objective of the present work was to examine the response of root porosity and internal O2 movement to root-zone ‘hypoxia’ in two Trifolium species which contrast in waterlogging tolerance.

Trifolium tomentosum (L.) is more tolerant of waterlogged conditions than T. glomeratum (L.) ( Gibberd & Cocks 1997). The contrasting tolerance to waterlogging of these two small (< 0·5 mg) seeded pasture legumes makes them ideal species to examine to improve the understanding of traits associated with waterlogging tolerance in legumes.

The capacity to form aerenchyma, and the physical nature of these cortical air spaces, varies among plant genotypes ( Justin & Armstrong 1987). An earlier study of other species of Trifolium showed that roots produced by waterlogged T. fragiferum and T. repens had higher porosity than roots grown in drained soil ( Rogers & West 1993). The increased porosity in T. repens was associated with the lysigenous breakdown of cortical cells. The experiments reported here assessed the effect of root-zone ‘hypoxia’ on the growth and root porosity in T. tomentosum and T. glomeratum plants. Cylindrical root-sleeving O2 electrodes ( Armstrong & Wright 1975; Armstrong 1994) were also used to evaluate patterns of radial O2 loss (ROL) from roots of the two species when in an O2-free medium. Finally, the shoot O2 concentration was manipulated and the response of ROL behind the root tip was evaluated to test whether the root porosity formed a continuous low resistance pathway for O2 movement.

MATERIALS AND METHODS

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Experiment 1. Comparison of growth of T. tomentosum and T. glomeratum in aerated, ‘hypoxic’ and stagnant nutrient solutions

Plant materials and culture

Seeds of T. tomentosum (L.) (wooly clover) and T. glomeratum (L.) (cluster clover) were collected in February 1993 from a field near Goomalling (31°20′S, 116°50′E) in the southwest of Western Australia. This stock of seeds was used in all subsequent experiments. The seeds were scarified and then germinated in the dark over an aerated solution containing (mol m−3): Ca2+, 1·0; SO42−, 1·2; K+, 0·4; H3BO3, 0·5 × 10−3; and pH 5·8. The plants were grown in a phytotron at 20/15 °C (day/night) with natural light. After 3 d, seedlings were exposed to the light and transferred to a new 0·2-strength nutrient solution. At full strength the nutrient solution contained (mol m−3): K+, 1·0; Mg2+, 0·2; NH4+, 0·1; NO3, 0·7; SO42−, 0·71; Cl, 1·2; H2PO4, 7 × 10−3; Ca2+, 0·9; H3BO3, 5 × 10−3; Zn2+, 0·75 × 10−3; Cu2+, 0·2 × 10−3; Mn2+, 1 × 10−2; Mo, 0·02 × 10−3; Fe-sequesterine, 0·01. The pH was adjusted to 5·8 with KOH. On the 6th day the nutrient solution was increased to 0·5-strength. After 9 d, the seedlings were transplanted into airtight 4 × 10−3 m3 plastic pots with full-strength nutrient solution. Five seedlings were transplanted into each pot and were held in place using foam plugs (10 mm diameter; 8 mm length) placed around the hypocotyl, below the cotyledons. The solutions in all containers were bubbled with humidified air at 2 × 10−3 m3 min−1.

Treatments

Three treatments were applied to 10-d old seedlings for 7 d.

Aerated. The nutrient solution was renewed and aerated with humidified air at 2 × 10−3 m3 min−1. The O2 concentration remained at ≈ 0·25 mol m−3.

‘Hypoxic’. The nutrient solution was renewed and flushed with humidified N2 to an O2 concentration of 0·093 mol m−3 after which the pots were sealed. After 24 h the solutions were again flushed with N2 to reduce the O2 concentration to ≈ 0·031 mol m−3. The O2 in the solution was tested every 24 h, and after 48 h it had increased to 0·069 ± 0·017 mol m−3 (mean ± standard error). So, the ‘hypoxic’ pots were flushed again with N2 until the O2 concentration decreased to 0·031 mol m−3, and this was repeated every 2 d.

Stagnant agar solution. A stagnant solution containing agar was prepared by dissolving 1 kg of agar m−3 of nutrient solution at 95 °C (prior to the addition of Fe-sequesterine, P and trace elements). The agar solution was stirred until it cooled to room temperature, after which aliquots of Fe-sequesterine, P and trace element stocks were added. Stagnant agar solutions of this nature have been shown to prevent convection and thus restrict gas movements to diffusion, as occurs in waterlogged soils ( Wiengweera, Greenway & Thomson 1997).

The various treatment pots, with five replicate pots (five plants per pot) for each harvest date, were arranged in a randomized complete block design on a bench in a phytotron.

Harvests

Two harvests were taken, an initial harvest on day 10 (the day treatments commenced) and a second one after 7 d of the treatments. The length of the primary root axis and the number of lateral roots per plant were recorded. The plants were divided into roots and shoots and dried in an oven at 70 °C for 3 d, after which the dry weights were recorded.

Experiment 2. Evaluation of the effect of ‘hypoxia’ on the root porosity of the two species

The plant materials and culture were as described in expt 1, except that the stagnant agar treatment was omitted and the nutrient solutions were renewed every 7 d. Ten-day old plants were exposed to aerated or ‘hypoxic’ (0·031– 0·069 mol O2 m−3) solutions and the roots were sampled after 7 and 21 d of treatments. The root porosity (% gas space/volume) was determined as described by Raskin (1983) using the equations as modified by Thomson et al. (1990) . Root porosity was measured on the entire root system which was cut into 50 mm sections. Measurements were taken for plants grown in four replicate pots arranged in a randomized complete block design on a bench in a phytotron.

Experiment 3. Profiles of ROL along the primary root of the two species when transferred to an O2-free medium

The profiles of ROL along the primary roots of T. tomentosum and T. glomeratum were determined for plants raised in aerated or ‘hypoxic’ solutions and then transferred to an O2-free root medium for the measurements. The plants were grown as described in expt 1, and the aerated or ‘hypoxic’ treatments were imposed on 10-d old plants for 7 d.

Cylindrical root-sleeving O2 electrodes [2·25 mm internal diameter (i.d.), 5 mm in height] fitted with guides to hold the root along the central axis of the electrode, were used to measure the diffusion of O2 from the roots when in an O2-free medium, as described by Armstrong & Wright (1975). The root systems of intact plants were immersed in a deoxygenated solution in Perspex chambers (50 × 50 × 150 mm) containing (mol m−3): K+, 5·0; Ca2+, 0·5; Cl, 5·0; SO42−, 0·5; and 1 kg agar m−3. Plants with primary roots of ≈ 95 mm in length were inserted into the solution through a hole in a gas impermeable lid and the hypocotyl was sealed in place with Terostat (Terostat VII; Teroson, Heidelberg, Germany), so that the shoots were in air. The actual lengths of the roots are given in Fig. 2. ‘Steady-state’ ROL was measured with the centre of the electrode positioned at 10, 20, 30 and 40 mm behind the tip of the primary root. Root diameters were measured using a vernier microscope. The measurements were conducted at 20 °C in a controlled temperature room with a photon flux density of 78 μmol m−2 s−1 at shoot height.

image

Figure 2. . Radial O2 loss (ROL) from various positions on the main axes of primary roots when in an O2-free medium. The symbols represent Trifolium tomentosum grown in aerated (□) or ‘hypoxic’ (▪) solutions, and T. glomeratum grown in aerated (○) or ‘hypoxic’ (●) solutions for the final 7 d prior to being transferred into the O2-free medium. The data points are the means of five plants with standard errors. The average (n = 5) lengths (mm) of the roots examined were: T. tomentosum aerated = 100, ‘hypoxic’ = 94; T. glomeratum aerated = 95, ‘hypoxic’ = 84.

Download figure to PowerPoint

Experiment 4. Response of ROL near the root tip to an increased O2 concentration around the shoots

The plants were raised as described in expt 1, and treatments of ‘hypoxia’ or continued aeration were imposed on 10-d old plants for 7 d.

The plants were transferred to the Perspex chambers as described in expt 3. In addition, the shoots were sealed in a Perspex hood flushed with humidified air (20·5% O2) at 1 × 10−3 m3 min−1. ROL with the centre of the electrode positioned at 10 mm behind the root tip was monitored and it reached a ‘steady-state’ value after 2 h. The O2 concentration in the hood surrounding the shoots was then increased to 97·5%, and the response of ROL was monitored. When ROL had reached a new ‘steady-state’ value it was recorded, the O2 concentration around the shoots was again lowered to 20·5%, and the new ROL values were recorded. This cycle of manipulations was repeated. Root diameters at 10 mm behind the tip were measured using a vernier microscope.

Experiment 5. Profiles of ROL from intact lateral roots of the two species when in an O2-free medium

Lateral roots begin to emerge from the primary root of T. tomentosum and T. glomeratum when the plants are ≈ 15 d old. Lateral roots first emerge near the root/shoot junction and then sequentially down the primary root. Hence, a 21-d old plant can have numerous lateral roots which tend to be successively shorter with increasing distance from the root/shoot junction. Fourteen-day old T. tomentosum and T. glomeratum were exposed to aerated or ‘hypoxic’ conditions for 7 d prior to the ROL measurements. It was, therefore, possible to select lateral roots of different lengths and different points of origin along the primary root, with root tips approximately equidistant from the shoot/root junction. Three lateral roots were selected – lateral 1 the oldest lateral root emerged immediately below the root/shoot junction and was ≈ 60 mm in length; lateral 2 emerged 10 mm below lateral 1 on the primary root and was ≈ 50 mm in length; lateral 3 emerged 20 mm below lateral 1 and was ≈ 40 mm in length. The plants were set up in the Perspex chambers as described in expt 3 and ROL was measured with the centre of the electrode positioned at 10, 20 and 30 mm behind the tip of these lateral roots. ROL from the primary root axis was tested at a location 60 mm below the root/shoot junction (below the lateral roots).

RESULTS

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Experiment 1. Comparison of growth of T. tomentosum and T. glomeratum in aerated, ‘hypoxic’ and stagnant nutrient solutions

Root growth

At the commencement of the treatments, the primary roots of T. tomentosum plants were 49·6 ± 3·3 mm in length and for T. glomeratum the roots were 46·4 ± 1·6 mm (mean ± standard error). Over the next 7 d the primary roots of T. tomentosum and T. glomeratum in aerated solution grew at 9·4 and 6·8 mm d−1, respectively (averaged over the 7 d treatment period). ‘Hypoxic’ and stagnant conditions reduced the rate of root extension of T. tomentosum to 78% and 72% of the value in aerated plants, respectively; and for T. glomeratum in the same treatments the rates were only 60% and 40%, respectively ( Table 1). The root extension rates of T. tomentosum in ‘hypoxic’ solution and the stagnant agar solution were not statistically different, while for T. glomeratum root extension was lower in stagnant agar solution than in ‘hypoxic’ solution (P < 0·05). The relative growth rates of the whole root systems under the conditions tested closely reflected the trends described for the extension rates of the primary root of the two genotypes ( Table 1).

Table 1. . Growth of Trifolium tomentosum and T. glomeratum in aerated, ‘hypoxic’ and stagnant agar nutrient solutions. The plants were 10 d old at the commencement of the treatments which lasted for a further 7 d. The stagnant nutrient solution contained 1 kg agar m−3. The data given are means of five replicate pots with five plants per pot ± standard errors Thumbnail image of
Shoot growth

The relative growth rates of shoots of T. tomentosum grown in ‘hypoxic’ and stagnant agar solutions were, respectively, 94% and 86% of the values of plants in aerated solution ( Table 1). The relative growth rates of shoots of T. glomeratum grown in ‘hypoxic’ and stagnant solutions were, respectively, 96% and 81% of the values of plants in aerated solution. Hence, the large differences between the two genotypes in root growth under low O2 did not result in large differences in shoot growth over the experimental period.

Experiment 2. Evaluation of the effect of ‘hypoxia’ on the root porosity of the two species

For both harvest dates, the ‘hypoxic’ treatment resulted in a 1·6-fold increase in the root porosity of T. tomentosum and T. glomeratum (P < 0·05). Furthermore, the porosity of T. tomentosum roots was 1·4-fold higher than that of T. glomeratum after 7 d and 2-fold higher after 21 d ( Fig. 1). For example, after 21 d of hypoxia the root porosity of T. tomentosum was 11·2% while for T. glomeratum it was only 6·1% ( Fig. 1).

image

Figure 1. . Porosities (% gas space/volume) of Trifolium tomentosum and T. glomeratum roots grown in aerated (▪) or ‘hypoxic’ (□) solutions for the final 7 or 21 d. The plants were 10 d old at the commencement of the treatments. Data are means of four replicate pots with five plants per pot, bars are standard errors.

Download figure to PowerPoint

Experiment 3. Profiles of ROL along the primary root of the two species when transferred to an O2-free medium

ROL from the roots in an O2-free medium increased with distance behind the root tip for T. tomentosum (i.e. ROL increased as the electrode was moved closer to the root/shoot junction; Fig. 2). At all positions tested along the roots of T. tomentosum, with the exception of 40 mm behind the tip, ROL was faster (by ≈ 7·5 ng cm−2 min−1) for plants raised in ‘hypoxic’ compared with aerated solution. In contrast to T. tomentosum, ROL from roots of T. glomeratum raised in aerated solution was barely detectable, and for roots raised in ‘hypoxic’ solution it was only slightly higher than in plants grown in aerated solution.

To highlight the importance of the differences between the two species and treatments on root aeration in anoxia, the ROL values measured at 10 mm behind the root tip were converted to O2 concentrations at the root surface according to Armstrong & Wright (1975). When in the O2-free solution, the O2 concentrations at the surface of aerated and ‘hypoxic’ roots of T. tomentosum were 9·5- and 3·8-fold higher than for T. glomeratum, respectively ( Table 2). Thus, the internal O2 supply to the roots in the O2-free medium was significantly greater for T. tomentosum than for T. glomeratum (P < 0·05).

Table 2. . O2 concentrations (mmol m−3) on the root surface at 10 mm behind the tip of Trifolium tomentosum and T. glomeratum when the roots were in an O2-free medium. The plants were raised in either aerated or ‘hypoxic’ solutions for 7 d before being transferred to the O2-free medium. The O2 concentrations were calculated from the data presented in Fig. 2. Data are means (n = 5) ± standard errors Thumbnail image of

Experiment 4. Response of ROL near the root tip to an increased O2 concentration around the shoots

To test whether the porosity of T. tomentosum and T. glomeratum roots formed a continuous, low resistance pathway for gaseous movement, shoot O2 concentrations were manipulated and the response of ROL near the root tip was measured. Figure 3 shows the response of the diffusion current (generated by the reduction of O2 diffusing from the root at the Pt surface of the electrode) to manipulations of shoot O2 concentration for representative plants of both species from the two treatments. ‘Steady-state’ ROL values for five replicates of each plant type were determined and these data are given in Table 3.

image

Figure 3. . The response of the diffusion current measured at 10 mm behind the tip of roots in an O2-free medium to manipulations of the O2 concentration around the shoots of: (a) Trifolium tomentosum and (b) T. glomeratum grown in ‘hypoxic’ (broken line) or aerated (solid line) solutions for the final 7 d prior to the measurements. x and z, addition of 97·5% O2 around the shoots; y, 20·5% O2 around the shoots. The diffusion current is generated by the reduction of O2 diffusing radially from the root at the Pt surface of the electrode ( Armstrong & Wright 1975).

Download figure to PowerPoint

Table 3. . The effect of manipulating shoot O2 concentration on radial O2 loss (ROL) 10 mm behind the root tip of Trifolium tomentosum and T. glomeratum when the roots were in an O2-free medium. The plants were either raised in aerated or ‘hypoxic’ solutions for 7 d before being transferred to the O2-free medium. Data are means (n = 5) ± standard errors Thumbnail image of

At 20·5% O2 around the shoot, the rates of ROL from 10 mm behind the tip of T. glomeratum roots grown in aerated or ‘hypoxic’ solutions were significantly less than those for T. tomentosum roots ( Table 3). These findings are in agreement with the ROL values obtained in the previous experiment ( Fig. 2). When the O2 concentration around the shoots was increased to 97·5%, ROL from 10 mm behind the root tip increased in T. tomentosum plants raised in either aerated or ‘hypoxic’ solution, but for T. glomeratum it only increased for plants raised in ‘hypoxic’ solution ( Fig. 3). ROL from roots of T. glomeratum plants raised in aerated solution was hardly responsive to manipulations of the shoot O2 concentration ( Fig. 3 & Table 3), and this extremely small response was difficult to measure due to background O2 flux from the medium resulting in residual currents of 0·15 μA. Furthermore, for the plants raised in the ‘hypoxic’ solution the increase in ROL from near the root tip of T. glomeratum was not as rapid as the response in T. tomentosum ( Fig. 3); and the final rate of ROL reached a plateau at a value which was only 62% of the value for T. tomentosum. For all plants, lowering the O2 concentration around the shoots back to 20·5% decreased the rates of ROL near the root tip to values which were similar to the initial readings ( Table 3).

Experiment 5. Patterns of ROL from intact lateral roots of the two species when in an O2-free medium

For plants raised in ‘hypoxic’ solution, the rates of ROL from lateral roots of T. tomentosum were significantly greater than those from T. glomeratum, with the exception of the third, and shortest, lateral root ( Fig. 4). Moreover, the rates of ROL from near the tips of lateral roots of T. tomentosum grown in aerated and ‘hypoxic’ solutions were 10–20-fold faster than those from the primary root at a similar distance from the root/shoot junction ( Fig. 4).

image

Figure 4. . Radial O2 loss (ROL) at 10 mm behind the tip of three lateral roots and a position on the primary root all ≈ 60 mm below the root/shoot junction of 21-d-old plants when in an O2-free root medium. Lateral root 1, oldest lateral root ≈ 60 mm long originating immediately below the root/shoot junction; lateral root 2, ≈ 50 mm long and originating 10 mm below root 1; lateral root 3, ≈ 40 mm long and originating 20 mm below lateral 1. Data are means of five replicate plants with standard errors.

Download figure to PowerPoint

The patterns of ROL along the longest lateral roots ( Fig. 5) were similar to those already described for the primary root axes of the two species ( Fig. 2). However, there were no differences in ROL at the same positions along the longest lateral root of T. tomentosum between plants previously grown in aerated or ‘hypoxic’ solutions. In contrast, ROL at 20 and 30 mm behind the tip of the longest lateral root of T. glomeratum were faster for plants grown in ‘hypoxic’ solution than for the same species grown in aerated solution. The rates of ROL from the older lateral roots of T. tomentosum were ≈ 30-fold greater than those from T. glomeratum ( Fig. 5).

image

Figure 5. . Radial O2 loss (ROL) along the longest lateral root of 21-d-old Trifolium tomentosum grown in aerated (□) or ‘hypoxic’ (▪) solutions, and T. glomeratum grown in aerated (○) or ‘hypoxic’ (●) solutions, for the final 7 d prior to being transferred to the O2-free root medium. Treatments were imposed on 14-d-old plants for an additional 7 d. Data are means of five replicate plants with standard errors.

Download figure to PowerPoint

DISCUSSION

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Earlier studies have shown that T. tomentosum inhabits soils prone to waterlogging, whereas T. glomeratum is not found on these sites even though it is a dominant species in adjacent areas ( Gibberd & Cocks 1997). In the present study, exposure to low O2 treatments in solution culture resulted in a greater reduction of root extension in T. glomeratum compared with T. tomentosum. Tolerance of O2-deficient waterlogged soils in other plant species has been shown to be associated with the development of aerenchyma and internal O2 movement to the roots ( Armstrong 1971; Armstrong & Gaynard 1976; Armstrong, Healy & Webb 1982, 1983; Thomson et al. 1990 , 1992; Laan et al. 1990 ; Colmer et al. 1998 ). These traits were therefore assessed in these two species of Trifolium in order to evaluate the role of aerenchyma in waterlogging tolerance in pasture legumes.

Both T. tomentosum and T. glomeratum responded to root-zone hypoxia by increasing root porosity to 11·2 and 6·1%, respectively ( Fig. 1). Cross-sections of roots revealed that the increased root porosity in both species was due to a greater volume of cortical air spaces (data not shown). A study of four other species of Trifolium also showed differences in the response of root porosity to growth in low O2 conditions ( Rogers & West 1993). The porosity of roots existing prior to, or formed during, exposure to low O2, increased for T. fragiferum and T. repens, while in T. michelianum porosity only increased in new roots, and in T. subterraneum porosity remained low. The highest total root porosity was 7·6% for ‘hypoxic’ roots of T. michelianum, which was substantially lower than the porosity of T. tomentosum (11·2%) as determined in the present work. Nevertheless, a root porosity of 11·2% is at the lower end of values published for other non-wetland species. For example, 15% in wheat ( Thomson et al. 1990 ) and 15–20% in Zea mays ( Jackson et al. 1985 ). In some wetland species, root porosity can be as high as 35% in Rumex ( Laan et al. 1989 ) or 45% in rice ( Armstrong 1971; Colmer et al. 1998 ).

The present studies of ROL from roots of two Trifolium species in an O2-free medium showed that the increases in root porosity of plants preadapted in ‘hypoxic’ solutions resulted in a greater internal O2 supply to the roots ( Fig. 2). Moreover, the O2 concentration at the root surface just behind the tip of T. tomentosum roots was substantially higher than that for T. glomeratum ( Table 2). The gas spaces in roots of T. tomentosum grown in both aerated or ‘hypoxic’ solutions, and in roots of T. glomeratum grown in ‘hypoxic’ solution, provided a low resistance internal pathway for O2 diffusion, as evidenced by the rapid increase in ROL near the root tip when the shoots were exposed to 97·5% O2 ( Fig. 3). Thus, the superior internal aeration system in roots of T. tomentosum should result in greater root penetration by this species into waterlogged soils, compared with T. glomeratum (cf. Armstrong 1979).

ROL was shown to increase with distance behind the root tip, an observation recorded for other dryland plants such as wheat and triticale ( Thomson et al. 1992 ). In contrast, ROL decreases with distance behind the root tip in wetland plants such as rice ( Armstrong 1971; Colmer et al. 1998 ) and Pragmites australis ( Armstrong & Armstrong 1988) due to a physical barrier to ROL in the exterior of the root. Thus, in the two species of Trifolium studied, ROL will form a ‘sink’ for O2 over the entire length of the root, which in turn would decrease the maximum length of these roots in waterlogged soils (cf. Armstrong 1979).

Trifolium species tend to form lateral roots in response to waterlogging ( Rogers & West 1993; Gibberd & Cocks 1997). The laterals would form a sink for O2 which would consume O2 from the aerenchyma in the primary root and therefore decrease the supply of O2 to the primary root apex ( Fig. 5). Armstrong et al. (1983) also found that increases in the number of lateral roots reduced the apical O2 concentration in roots of pea, an effect which could be reversed by the excision of the lateral roots. The consumption of O2 by lateral roots, at the expense of its movement further down the primary root, would limit the final length of the primary root in anaerobic soil. Thus, the laterals probably become the main functional root system of T. tomentosum and T. glomeratum in waterlogged soils, especially as these two species did not form any adventitious roots in the present experiment, or in earlier experiments in waterlogged soil ( Gibberd & Cocks 1997). It should be noted, however, that flooding-tolerant T. semipilosum does form adventitious roots ( Shiferaw, Shelton & So 1992), a feature associated with waterlogging tolerance in other plants (e.g. in Rumex;Laan et al. 1989 ). However, the relatively shallow system of lateral roots of T. tomentosum and T. glomeratum may limit the capacity of these plant to recover once the waterlogging recedes and the soil profile dries.

CONCLUSIONS

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

Trifolium tomentosum and T. glomeratum responded to root-zone ‘hypoxia’ by increases in root porosity; although the porosity in roots of T. tomentosum was 1·8-fold higher than that in T. glomeratum. Moreover, the constitutive porosity in aerated roots of T. tomentosum was 1·7-fold greater than in T. glomeratum. The higher porosity in roots of T. tomentosum should be of adaptive advantage at the onset of, and during, waterlogging. The root porosity was shown to form a continuous, low resistance pathway for gaseous diffusion in both species. However, ROL was 3·8–9·5-fold higher from roots of T. tomentosum than from T. glomeratum when in an O2-free medium. Thus, the better root growth of T. tomentosum in low O2 conditions, when compared with T. glomeratum, was presumably associated with the greater porosity and enhanced internal O2 supply in roots of T. tomentosum.

ACKNOWLEDGMENT

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES

We thank the Grains Research and Development Corporation for financial support.

REFERENCES

  1. Top of page
  2. ABSTRACT
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. CONCLUSIONS
  8. ACKNOWLEDGMENT
  9. REFERENCES
  • Armstrong W. (1970) Rhizosphere oxidation in rice and other species: a mathematical model based on the oxygen flux component. Physiologia Plantarum 23, 623 630.
  • Armstrong W. (1971) Radial oxygen loss 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. (1994) Polarographic oxygen electrodes and their use in plant aeration studies. Proceedings of the Royal Society of Edinburgh 102B, 511 527.
  • Armstrong J. & Armstrong, W. (1988) Phragmites australis– A preliminary study of soil-oxidising sites and internal gas transport pathways. New Phytologist 108, 373 382.
  • Armstrong W. & Gaynard T.J. (1976) The critical oxygen pressures for respiration in intact plants. Physiologia Plantarum 37, 200 206.
  • Armstrong W., Healy M.T., Webb T. (1982) Oxygen diffusion in pea. I. Pore space resistance in the primary root. New Phytologist 91, 647 659.
  • Armstrong W., Healy M.T., Webb T. (1983) Oxygen diffusion in pea. II. Oxygen concentrations in the primary pea root apex as affected by growth, the production of laterals and radial oxygen loss. New Phytologist 94, 549 559.
  • Armstrong W. & Wright E.J. (1975) Radial oxygen loss from roots: the theoretical basis for the manipulation of flux data obtained by the cylindrical platinum electrode technique. Physiologia Plantarum 25, 21 26.
  • Colmer T.D., Gibberd M.R., Wiengweera A., Tinh T.K. (1998) The barrier to radial oxygen loss from roots of rice (Oryza sativa L.) is inducted by growth in stagnant solution. Journal of Experimental Botany 49, 1431 1436.
  • Gibberd M.R. & Cocks P.S. (1997) Effect of waterlogging and soil pH on the micro-distribution of naturalised annual legumes. Australian Journal of Agricultural Research 48, 223 229.
  • Jackson M.B., Fenning T.M., Drew M.C., Saker L.R. (1985) Stimulation of ethylene production and gas space (aerenchyma) formation in adventitious roots of Zea mays L. by small partial pressures of oxygen. Planta 165, 486 492.
  • James E.K., Minchin F.R., Sprent J.I. (1992) The physiology and nitrogen-fixing capability of aquatically and terrestrially grown Neptunia plena: the importance of nodule oxygen supply. Annals of Botany 69, 181 187.
  • Justin S.H.F.W. & Armstrong W. (1987) The anatomical characteristics of roots and plant response to soil flooding. New Phytologist 106, 465 495.
  • Laan P., Smolders A., Blom C.W.P.M., Armstrong W. (1989) The relative roles of internal aeration, radial oxygen losses, iron exclusion and nutrient balances in flood tolerance of Rumex species. Acta Botanica Neerlandica 38, 131 145.
  • Laan P., Tosserams M., Blom C.W.P.M., Veen B.W. (1990) Internal oxygen transport in Rumex species and its significance for respiration under ‘hypoxic’ conditions. Plant and Soil 122, 39 46.
  • Pugh R., Witty J.F., Mytton L.R., Minchin F.R. (1995) The effect of waterlogging on nitrogen fixation and nodule morphology in soil grown white clover (Trifolium repens L.). Journal of Experimental Botany 46, 285 290.
  • Raskin I. (1983) A method for measuring leaf, density, thickness and internal gas. Hortscience 18, 698 699.
  • Rogers M.E. & West D.W. (1993) The effects of root-zone salinity and hypoxia on shoot and root growth in Trifolium species. Annals of Botany 72, 503 509.
  • Shiferaw W., Shelton H.M., So H.B. (1992) Tolerance of some subtropical pasture legumes to waterlogging. Tropical Grasslands 26, 187 195.
  • Thomson C.J., 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 C.J., Colmer T.D., Watkin E.L.J., Greenway H. (1992) Tolerance of wheat (Triticum aestivum cvs. Gramenya and Kite) and Triticale (Triticosecale cv. Muir) to waterlogging. New Phytologist 120, 335 344.
  • Walker B.A., Pate J.S., Kuo J. (1983) Nitrogen fixation by nodulated roots of Viminaria juncea (Schrad. & Wendl.) Hoffmans. (Fabaceae) when submerged in water. Australian Journal of Plant Physiology 10, 409 421.
  • Wiengweera A., Greenway H., Thomson C.J. (1997) The use of agar nutrient solution to simulate lack of convection in waterlogged soils. Annals of Botany 80, 115 123.
  • Zook D.M., Erwin D.C., Stolzy L.H. (1986) Anatomical, morphological, and physiological responses of alfalfa to flooding. Plant and Soil 96, 293 296.