Changes in specific area and energy of root surface of cereal plants in Al-solution cultures. Water vapor adsorption studies

Surface areas and energetic properties of the shooting stage roots of rye (Secale L.), triticale (Triticale), barley (Hordeum L.) and four wheat (Triticum L.) varieties were estimated from experimental water vapor adsorption data. Roots stressed during 10 days at pH 4 with aluminium concentrations ranging from 0 to 40 mg dm^–3 were studied. Roots grown continuously at pH 7 were taken as controls. The surface properties of the roots grown at pH 4 without Al addition were apparently the same as those of the control roots. With the increase of the concentration of the aluminium treatment the surface area of the roots increased for all of the plants, beginning at 5 mg Al dm^–3 for barley, at 10 mg Al dm^–3for wheat and triticale, and at 40 mg Al dm^–3 for rye. The average water vapor adsorption energy of the root surface decreased in general with the increase of Al stress concentration for all plants but triticale, for which this increased. The sensitive cereal varieties seem to have greater amount of high energy adsorption centers (more polar surface) than the resistant ones (lower surface polarity), however more data is needed to justify this hypothesis. For Al-sensitive roots, fraction of high energy adsorption sites decreased and fraction of low energy sites increased under the Al stress. Smaller changes in adsorption energy sites were noted for roots of Al-resistant plants.     

Changes in micropore system of roots of wheat and triticale seedlings under aluminum stress as determined using water vapor adsorption—desorption

Plant root structure including the pore system is severely injured in Al-toxic environments. Experimental water vapor desorption isotherms estimated using vacuum chamber method were used to characterize micropore build-up of roots of wheat (Lanca) and triticale (Debo) 4-day seedlings and its changes after 24 h (8 mg dm⁻³) aluminum stress. The micropore volumes and average pore radii increased after the stress whereas pore fractal dimensions decreased. The above changes were more pronounced for these roots which were not able to grow further after the stress when compared to the control roots, the total micropore volume was almost twice higher and the fractal dimension reached a value of 2 which is characteristic for flat, two-dimensional surface. The observed alterations of pore system indicate damage of root tissue and possible strong binding of aluminum on cell wall surfaces.     

Several lanthanides activate malate efflux from roots of aluminium‐tolerant wheat

Many elements of the lanthanide series exist as trivalent cations in solution below pH 6. The present study was carried out to investigate whether lanthanides could stimulate malate efflux from wheat (Triticum aestivum L.) roots, as has been found for trivalent aluminium (Al) cations. Excised root apices treated with 100 µm of each of seven different lanthanide elements (lanthanum, praseodymium, europium, gadolinium, terbium, erbium, and ytterbium) stimulated malate efflux, with five‐ to fifty‐fold more malate being released from an Al‐tolerant wheat line than from a near‐isogenic Al‐sensitive line. As erbium stimulated the greatest malate efflux of the lanthanides tested, this response was characterized further. The characteristics of the erbium‐activated efflux were similar to the Al‐activated efflux described previously suggesting that both of these ions activate the same transport mechanism. The capacity for erbium‐activated malate efflux cosegregated with Al tolerance in wheat seedlings derived from a cross between Al‐sensitive and Al‐tolerant near‐isogenic lines. This is the first study to identify cations, other than Al, which can activate malate release from wheat roots. It also provides additional evidence that malate efflux from root apices is the primary mechanism for Al tolerance in wheat.     

The effects of lime and phosphorus on the function of wheat roots in acid top soils and subsoils

Two wheat varieties with differing aluminium tolerance were grown in pots of acid soil. Liming did not change significantly the amounts of chemically extractable P and K, but caused improved vegetative growth, increased inflow of P and K and reduced uptake of Al. Without lime, roots had a higher content and concentration of P than shoots; liming reversed this. Without lime the sensitive variety with a shorter root length had an Al inflow ten times that of the tolerant one: tolerance involves a mechanism for exlcuding Al. The inflow of P per unit inflow of Al (mol ratio) without lime was three times greater for the tolerant variety which therefore has more P to counteract the effects of Al. The same varieties were grown in two-layer soil columns, with a low P status and a limed topsoil and acid subsoil. Liming the subsoil improved plant growth but this was still restricted by low P availability. Addition of P to the topsoil caused good growth regardless of subsoil acidity: root growth increased in both layers and P (labelled with³²P) taken up from the topsoil was translocated to roots in the subsoil. This P inactivated root Al and allowed the roots to grow and take up more P from the acid subsoil with however a reduction in inflow. The sensitive variety was affected more by the acid subsoil and low P availability, had a similar ability to translocate P to subsoil roots but could not attain the growth rate of the tolerant wheat even with P and lime.     

Aluminium tolerance in triticale,wheat and rye as measured by root growth characteristics and aluminium concentration

Summary Screening large populations of plant species for Al tolerance requires simple and rapid tests. In this study, root characteristics of 12 cultivars of triticale (X Triticosecale, Witt Mack), wheat (Triticum aestivum L.), and rye (Secale cereale L.) were measured in nutrient solution with 0 or 6 ppm Al added. Aluminum injury to roots of triticale and wheat was characterized by decreases in root length, increases in the number of roots, and in Al-sensitive Redcoat and Arthur wheats by decrease in root weight. Root length and number of roots were correlated in triticale (r=−0.73^*) and in wheat (r=−0.85^*). Root length was also correlated with root weight in wheat (r=0.65^*); there was no relationship between the number of roots and weight. Differences in Al tolerance of cultivars of the three species were greater when the solution was adjusted to pH 4.8 only on the first day of the experiment than when pH was maintained at pH 4.8 throughout the growing period. Triticale and rye cultivars low in ability to increase solution pH gradually overcame Al toxicity by increasing the nutrient solution pH between 12 and 22 days. Aluminum sensitive triticale and wheat accumulated more Al in roots than tolerant cultivars when the solution pH was not adjusted daily; but no differences in Al accumulation were obtained between wheat cultivars at constant pH value. This study indicated that root length and number of roots can be reliably used for screening triticales for Al tolerance within 12 days of exposure to Al. Root length, Al concentration, and dry weight after 22 days of Al treatment were also reliable criteria for evaluating differential Al tolerances among triticale cultivars.     

Solution pH modifies the response of Norway spruce seedlings to aluminium

Norway spruce (Picea abies) was exposed to nutrient solutions containing a range of aluminium (Al) concentrations at several pH levels (3.2, 4 and 5). Root growth was reduced by 100 µM and 400 µM Al at pH 4 and 5, but at pH 3.2 only by 400 µM Al. The Al content of the roots increased with increasing pH. The Al content of the roots was higher at the root tips than at the older root parts at all pH values. Using X-ray microanalysis it could be shown that higher levels of Al at increased pH were mainly due to increased Al contents in root cortex cell walls. In seedlings, mycorrhizal with Pisolithus tinctorius or Lactarius rufus, the Al concentration of cortex cell walls was higher when nitrate (NO₃) rather than ammonium (NH₄) was the nitrogen (N) source.     

Effect of phosphorus supply to the surface roots of wheat on root extension and rhizosphere chemistry in an acidic subsoil

Seedlings of two cultivars of wheat (Triticum aestivum L.) differing in tolerance to aluminium (Al) were grown using a split-root sand/soil culture technique. Each culture tube was divided horizontally into a surface (0–150 mm) compartment and a subsurface (150–250 mm) compartment separated by a root-permeable paraffin wax barrier. Thus phosphorus (P) supplied to surface roots could not percolate or diffuse into the soil in the subsurface compartment. The soil in the subsurface compartment was divided into ‘rhizosphere’ and ‘non-rhizosphere’ zones using a porous (5 μm) membrane. Root growth of both cultivars into the subsurface zone was enhanced by increased P supply to surface roots, but did not conform to known relationships between root growth and soil pH, extractable-Al, or pH, Al or P concentrations in soil solution. Concentrations of Al in soil solution in the rhizosphere were greater than those in solution in the bulk soil. Concentrations of Al reactive with pyrocatechol violet (30s-RRAI) in the rhizosphere soil solution were generally greater than those in non-rhizosphere soil. With the Al-sensitive cultivar, root dry weight and length increased as concentrations of RRAl in the rhizosphere soil solution increased. Increased concentrations of Al in rhizosphere soil solutions were not related to the presence of organic ligands in solution. The effect of P in promoting root penetration into the acidic subsurface stratum was not related to differential attainment of maturity by the plant shoots, but appeared to be related to the effect of P in enhancing the rate of root growth. Thus, suboptimal supply of P to the surface roots of a plant, even at levels sufficient to preclude development of nutritional (P) stress symptoms, may seriously reduce tolerance to Al, and hence diminish the ability of roots to penetrate into acidic subsoils.     

Changes in the microbial community in a forest soil amended with aluminium in situ

Considerable knowledge exists about the effect of aluminium (Al) on root vitality, but whether elevated levels of Al affect soil microorganisms is largely unknown. We thus compared soils from Al-treated and control plots of a field experiment with respect to microbial and chemical parameters, as well as root growth and vitality. The field experiment was established in a 50-year-old Norway spruce (Picea abies L.) stand where no Al or low concentrations of Al had been added every 7–10 days during the growth season for 7 years. Analysis of soil solutions collected using zero tension lysimeters and porous suction cups showed that Al treatment lead to increased concentrations of Al, Ca and Mg and lower pH and [Ca + Mg + K/Al] molar ratio. Corresponding soil analyses showed that soil pH remained unaffected (pH 3.8), that exchangeable Al increased, while exchangeable Ca and Mg decreased due to the Al treatment. Root in-growth into cores placed in the upper 20 cm of the soil during three growth seasons was not affected by Al additions, neither was nutrient concentration or mortality of these roots. The biomass of some taxonomic groups of soil microorganisms, analyzed using specific membrane components (phospholipid fatty acids; PLFAs), was clearly affected by the imposed Al treatment, both in the organic soil horizon and in the underlying mineral soil. Microbial community structure in both horizons was also clearly modified by the Al treatment. Shifts in PLFA trans/cis ratios indicative of short term physiological stress were not observed. Yet, aluminium stress was indicated both by changes in community structure and in ratios of single PLFAs for treated/untreated plots. Thus, soil microorganisms were more sensitive indicators of subtle chemical changes in soil than chemical composition and vitality of roots.     

Influence of root-induced pH on the solubility of soil aluminium in the rhizosphere

Increase in solubility of soil aluminium (Al) as a result of root-induced decrease of soil pH was studied. Soil samples of known distances from the roots of NH₄-N fertilized Ryegrass were analyzed for pH and aluminium extractable with 0.01 M CaCl₂. Results showed that though no Al was found in bulk soil (pH 6.8), its concentration in the vicinity of roots increased to 0.023 mM with a concomitant decrease of soil pH from 6.8 to 4.4.     

The response of tomato seedling roots to infection by Verticillium albo-atrum or Fusarium oxysporum f. sp. lycopersici

The response of seedling roots of near-isogenic tomato varieties to infection by Verticillium albo-atrum or Fusarium oxysporum f. sp. lycopersici was investigated. Studies of the infection of seedling roots not artificially damaged indicated that there was an extra-vascular expression of resistance towards V. albo-atrum but not to F. oxysporum. Roots of resistant tomato seedlings infected by V. albo-atrum contained the fungus in the epidermis and outer cortex while susceptible roots became heavily colonised. Observations made by transmission electron microscopy showed that the fungus appeared to be abnormal in growth and appearance in the epidermal and cortical cells of resistant seedling roots but normal in susceptible roots. Two preformed antifungal terpenoids were detected in seedling roots in greater amounts in resistant that in susceptible varieties. The possible mechanisms of seedling root resistance to vascular wilts are discussed.     

Physiological aspects of aluminium tolerance associated with the long arm of chromosome 2D of the wheat (Triticum aestivum L.) genome

Aluminum (Al) uptake in roots of wheat nearisogenic lines having differing tolerances to aluminium toxicity was studied using roots and root segments immersed in a nutrient solution at a controlled pH and temperature. At low Al concentrations a mechanism preventing root tips from accumulating too much Al was observed in an Al-tolerant isoline and a BH1146 euploid. This mechanism was more efficient when divalent cations of calcium or magnesium were present in the nutrient medium. Al accumulation steadily increased in root tips of the Al-sensitive wheat isoline during all 24 h of incubation, and the presence of divalent cations in the medium even increased Al concentration in root tissue. However, at higher Al concentrations in the medium the mechanism preventing the root tips of Al-tolerant genotypes from accumulating too much Al was not observed, and in effect Al concentration in root tips of both Al-tolerant and Al-sensitive isolines increased. It is concluded that genetical factors are located on the long arm of chromosome 2D from the BH1146 euploid that control the mechanism preventing root apical meristems from accumulating too much Al at low Al concentrations in the medium. However, there must be other genetical factors also located on this chromosome segment that control Al detoxication in root tips of Al-tolerant lines at higher external Al concentrations.     

Root phosphate exudation and pH shift in the rhizosphere are not responsible for aluminum resistance in rice

A series of hydroponic experiments and an agar culture experiment were carried out to investigate aluminum (Al) accumulation and translocation in two rice (Oryza sativa L.) cultivars (Kasalath and Koshihikari) that differ in Al resistance. Al-resistance mechanisms, including Pi exudation under Al stress and pH shifts in the rhizosphere, were also studied. Al content in rice shoots was 41 mg kg⁻¹ on average and did not differ between the two cultivars, which demonstrated that the rice cultivars were not Al accumulators. The majority of Al (95–97%) accumulated in roots. Al content in roots in the resistant cultivar (Koshihikari) was lower than that in the sensitive cultivar (Kasalath), which indicated that Al-exclusion mechanisms were mainly acting in rice. However, the rate of Pi exudation from the whole root or root tips was very low in both cultivars and was not significantly influenced by Al exposure, and thus seemed not to be the main Al-resistance mechanism. On the other hand, experiments with pH-buffered solution and color changes following culture in agar medium containing bromocresol purple revealed that the Al-induced pH increase could not explain the high Al resistance of rice. In addition, the Al content in shoots of Koshihikari was lower after the formation of iron plaque on the root surface, whereas that of Kasalath was not lower. These results suggested that rice roots cell wall components or root surfaces such as iron plaque, rather than pH changes and/or root exudates including organic acids and phosphate, play important roles in Al resistance in rice.     

MECHANISMS OF ALUMINUM TOLERANCE IN TRITICUM AESTIVUM L. (WHEAT). I. DIFFERENTIAL PH INDUCED BY WINTER CULTIVARS IN NUTRIENT SOLUTIONS

Twenty winter cultivars of Triticum aestivum L. (wheat) were grown in solution culture with and without aluminum (Al) (74 μM, 2.0 mg L^-1) for 14 days. Exposure to Al increased root growth of the most tolerant cultivar, while both root and shoot growth were depressed in all other cultivars. On the basis of a root tolerance index (RTI = weight of roots grown with Al/weight of roots grown without Al), cultivar tolerance to Al ranged 9-fold, from 0.13 ± 0.01 to 1.16 ± 0.10. Symptoms of Al toxicity were most evident on roots. Aluminum-affected roots were relatively short and thick and had numerous undeveloped laterals. Leaves of some cultivars showed chlorosis resembling iron deficiency, and others showed purple stems typical of phosphate deficiency. Plants of all cultivars grown with and without Al depressed the pH of nutrient solutions, presumably until NH₄⁺ was depleted, at which point the pH increased. Cultivar tolerance, expressed both as the root tolerance index and a shoot tolerance index, was negatively correlated with the negative log of the mean hydrogen ion (H⁺) concentration, the minimum pH, and the slope of the pH decline, each calculated from pH data collected during the first 9 days of the experimental period before any sharp rises in pH occurred. These results are consistent with the hypothesis that the Al tolerance of a given cultivar is a function of its ability to resist acidification of the nutrient solution and hence to limit the solubility and toxicity of Al.     

Sensitivity to H- and Al ions limiting growth and distribution of the woodland grass Bromus benekenii

One pH experiment and two aluminium experiments were conducted in order to investigate the effects of H- and Al ions on growth of Bromus benekenii. Continuously flowing solution cultures were used with ion concentrations simulating natural soil solutions. In all experiments, treatment effects were more pronounced on root than on shoot growth. In the pH experiment, root growth decreased with decreasing pH within the pH range 4.5 to 3.5. The critical pH for root growth of Bromus benekenii was between 3.8 and 4.0. In the Al experiments, root growth started to decrease at 20 M of quickly reacting Al and almost ceased at 70 M Al. This characterizes Bromus benekenii as an Al sensitive species. In the pH experiment, shoot concentrations of Ca, Mg, K and P decreased with decreasing pH, but root concentrations were not affected. In the Al experiments, the Al concentrations of both shoots and roots increased with Al in the nutrient solution. At treatments of 70 M Al or higher, Ca, Mg, K and P concentrations in the shoots were reduced. The critical concentrations of H- and Al ions in the experiments were similar to the highest concentrations found at field sites of Bromus benekenii, analysed in soil solutions obtained by centrifugation technique. Both Al and H toxicity were considered to be of importance as limiting factors for the distribution of Bromus benekenii in south Sweden. Probably, Al toxicity starts to limit growth when also pH itself influences growth negatively. The importance of simulating natural soil solutions in experiments is emphazised, in order to obtain information on the importance of chemical soil factors to the distribution of plants.     

Aluminium resistance requires resistance to acid stress: a case study with spinach that exudes oxalate rapidly when exposed to Al stress

Spinach is a vegetable with a high oxalate concentration in its tissues. Oxalate efflux from spinach (Spinacia oleracea L. cv. Quanneng) roots was rapidly stimulated (within 30 min) by aluminium (Al) treatment. The efflux was constant within 6 h, but increased with increasing Al concentration. The efflux was confined to the root tip (0-5 mm), which showed a 5-fold greater efflux than the root zone distal to the tip (5-10 mm). Oxalate efflux could not be triggered by treatment with the trivalent cation lanthanum or by phosphorus deficiency, indicating that the efflux was specific to the Al treatment. All this evidence suggested that spinach possesses Al-resistance mechanisms. However, spinach was found to be as sensitive to Al toxicity as the Al-sensitive wheat line ES8, which had no Al-dependent organic acids efflux. The Al accumulated in the apical 5 mm of the roots of spinach which was also similar to that in the Al-sensitive wheat after 24 h treatment with 50 microM AlCl(3), indicating a non-exclusion mechanism. In addition, root elongation in spinach was significantly inhibited at pH 4.5, compared with that at pH 6.5. Based on this evidence, it is concluded that the sensitivity to acid stress in spinach could mask the potential role for oxalate to protect the plant roots from Al toxicity.     

Altered root growth and plant chemistry ofPinus sylvestris seedlings subjected to aluminum in nutrient solution

One-year-old Scots pine (Pinus sylvestris L.) seedlings were grown for 9 weeks in nutrient solutions containing 0, 0.5, 1, 2 and 4 mM aluminum nitrate (Al(NO3)3) at pH 4.2. Nine weeks exposure to Al significantly reduced total plant, shoot and root mass and caused a linear decline in proportional allocation of biomass to roots. Relative growth rate of roots declined to as low as zero. Aluminum treatment decreased calcium and magnesium uptake and increased Al content in roots and needles. After 3 weeks of exposure a 10–60% increase in total phenols in roots and a 20–40% increase in o-diphenols in roots and needles were noted. Roots affected by Al showed degeneration of meristematic cells, fewer cell divisions, deformation in cell walls and higher lignification and suberization. The majority of root apices were structurally similar to dormant roots, and a premature senescence of the entire root system was observed. Net photosynthetic rate after 6 weeks of treatment was negatively correlated with needle Al content and Al/Ca ratio (r < -0.9, P < 0.1). The results suggest that Scots pine may be more susceptible to Al than was expected based on previous experiments.     

Colonization of wheat by Azospirillum brasilense Cd is impaired by saline stress

The effect of saline stress on the colonization of wheat was analyzed by using Azospirillum brasilense Cd carrying the fusion of the reporter gene lacZ (β-galactosidase) with the N₂ fixation gene promoter nifA. Colonization was also studied by inducing para-nodules on wheat roots using 2,4-D, establishing that these structures acted as bacterium protected niches. Bacteria grown under standard conditions were distributed along the whole root system, except the elongation zone, and colonized the para-nodules. Bacteria experiencing saline stress were mainly localized at the root tips and the lateral roots. In 2,4-D treated plants, most of the bacteria were present around the basal surface of the modified lateral root structures. Using the MPN method, there were not statistical differences between the numbers of control and stressed bacteria. As this method estimates endophytic colonization in contrast with the one using X-gal, which emphasizes colonization on the root surface, both procedures demonstrated to be necessary, concluding that salt treatment reduced surface colonization (X-gal) but not colonization inside the root. The bacterial counts made on inoculated wheat roots indicated higher numbers of both control and stressed bacteria in roots treated with 2,4-D compared with untreated roots. This revised version was published online in June 2006 with corrections to the Cover Date.     

Aluminum Partitioning in Intact Roots of Aluminum-Tolerant and Aluminum-Sensitive Wheat (Triticum aestivum L.) Cultivars

Aluminum (Al) partitioning in intact roots of wheat (Triticum aestivum L.) cultivars that differ in sensitivity to Al was investigated. Roots of intact seedlings were exposed to Al for up to 24 hours and distribution of Al was assessed visually by hematoxylin staining or by direct measurement of concentration of Al by atomic absorption spectrophotometry or ion chromatography. Major differences in Al accumulation between Al-tolerant (Atlas 66) and Al-sensitive (Tam 105) cultivars were found in the growing regions 0 to 2 and 2 to 5 millimeters from the root apex. Al content was 9 to 13 times greater in the 0 to 2 millimeters root tips of cv Tam 105 than in the tips of cv Atlas 66 when exposed to 50 micromolar Al for 19 to 24 hours. The oxidative phosphorylation inhibitor carbonyl cyanide m-chlorophenylhydrazone and the protein synthesis inhibitor cycloheximide increased Al uptake by intact root tips of cv Atlas 66. Also, loss of Al from the roots of both cultivars was measured after the roots were “pulsed” with 50 micromolar Al for 2 hours and then placed in an Al-free nutrient solution for 6 hours. The 0 to 2 millimeter root tips of cv Tam 105 lost 30% of the absorbed Al, whereas the tips of cv Atlas 66 lost 60%. In light of these results, we conclude that the differential Al sensitivity in wheat correlates with the concentration of Al in the root meristems. The data support the hypothesis that part of the mechanism for Al tolerance in wheat is based on a metabolism-dependent exclusion of Al from the sensitive meristems.     

Improved correlation between soil exchangeable K and plant K contents on a tissue water basis

In acid volcanic soils, plant roots are thought to be injured by acidity (low pH) and/or solubilized aluminium (Al) ions. An attempt was made to separate the effects of low pH from those of Al on the elongation and viability of alfalfa (Medicago sativa L.) radicles in water culture. Root elongation was irreversively curtailed by 20 hours treatment at pH 4.0 without Al or 20 mmol m^-3 Al at pH 5.0. Viability of surface cells of root tips was detected as a degrading activity of fluorescein diacetate (FDA) by cellular esterases and subsequent accumulation of derived fluorescein within cells. Large numbers of the surface cells lost their viability after four hours exposure at the low pH. In contrast, surface cells maintained both FDA degrading activity and ability to accumulate fluorescein 20 h after initial exposure to the Al solution (20 mmol Al m^-3, pH 5.0). These results suggest that there are some significant differences in the mechanisms of phytotoxicity to alfalfa root between the two stress factors.