AOBPreview originally published online on June 12, 2006
Annals of Botany 2006 98(4):693-713; doi:10.1093/aob/mcl114
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INVITED REVIEW |
Root Structure and Functioning for Efficient Acquisition of Phosphorus: Matching Morphological and Physiological Traits
1 School of Plant Biology, Faculty of Natural and Agricultural Sciences, The University of Western Australia, 35 Stirling Highway, WA 6009, Australia and 2 Department of Botany, University of Cape Town, Private Bag, Rondebosch, 7701, South Africa
* For correspondence. E-mail hans.lambers{at}uwa.edu.au
Received: 11 February 2006 Returned for revision: 14 March 2006 Accepted: 27 March 2006 Published electronically: 12 June 2006
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ABSTRACT |
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TOP
ABSTRACT
INTRODUCTION• Background Global phosphorus (P) reserves are being depleted, with half-depletion predicted to occur between 2040 and 2060. Most of the P applied in fertilizers may be sorbed by soil, and not be available for plants lacking specific adaptations. On the severely P-impoverished soils of south-western Australia and the Cape region in South Africa, non-mycorrhizal species exhibit highly effective adaptations to acquire P. A wide range of these non-mycorrhizal species, belonging to two monocotyledonous and eight dicotyledonous families, produce root clusters. Non-mycorrhizal species with root clusters appear to be particularly effective at accessing P when its availability is extremely low.
• Scope There is a need to develop crops that are highly effective at acquiring inorganic P (Pi) from P-sorbing soils. Traits such as those found in non-mycorrhizal root-cluster-bearing species in Australia, South Africa and other P-impoverished environments are highly desirable for future crops. Root clusters combine a specialized structure with a specialized metabolism. Native species with such traits could be domesticated or crossed with existing crop species. An alternative approach would be to develop future crops with root clusters based on knowledge of the genes involved in development and functioning of root clusters.
• Conclusions Root clusters offer enormous potential for future research of both a fundamental and a strategic nature. New discoveries of the development and functioning of root clusters in both monocotyledonous and dicotyledonous families are essential to produce new crops with superior P-acquisition traits.
Key words: Actinorhizal, capillaroid roots, carboxylates, Casuarinaceae, cluster roots, Cyperaceae, dauciform roots, exudation, Fabaceae, Proteaceae, proteoid roots, Restionaceae
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INTRODUCTION |
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Phosphorus (P) is an essential inorganic nutrient for all living organisms. It is required as a structural component in nucleic acids and phospholipids, as an element in intermediates in carbon metabolism, and to allow (in)activation of a wide range of enzymes. After nitrogen (N), P is quantitatively the most important inorganic nutrient for plant growth, and often limits primary productivity in natural systems as well as cropping systems, unless supplied as fertilizer (Vance et al., 2003
). P is a non-renewable resource, unlike N, which can be assimilated from N2 into NH3 by free-living and symbiotic N2-fixing micro-organisms, or converted into NH3, 
or urea industrially. Moreover, global P reserves are rapidly being depleted; depending on the assumed scenario, current P reserves will be halved (relative to the reserves at the turn of the twentieth century) by 2040 or, more likely, by 2060 (Steen, 1998). Whilst our global P reserves are being depleted, P levels in many agricultural soils are building up, because 80–90 % of P applied as fertilizer is sorbed by soil particles, rendering it unavailable for plants that lack specific adaptation to access sorbed P (Gerke et al., 1994; Jones, 1998a). With decreasing global P reserves, P-fertilizer prices are bound to increase. There is an urgent need to develop crops that are more efficient at acquiring inorganic P (Pi) from soil and/or at using P more efficiently. Equally, it is becoming increasingly important to use crops that reduce the off-site effects of P fertilization, thus reducing the risks of pollution of streams and rivers.
Unlike nitrate, which readily moves in soil towards the roots via both mass flow and diffusion, phosphate (Pi) is highly immobile. Mass flow typically delivers as little as 1–5 % of a plant's P demand, and the amount intercepted by growing roots is only half of that (Lambers et al., 1998). The rest of all required Pi must reach the root surface via diffusion; diffusion coefficients for phosphate in soil are typically very low compared with those for other nutrients: 0·3–3·3 x 10–13 m2 s–1 (Clarkson, 1981). Diffusion is particularly slow in dry soil (e.g. Turner and Gilliam, 1976; Bhadoria et al., 1991). Increasing Pi delivery to roots via mass flow can be achieved by enhanced transpiration rates, but this cannot have a major effect, and would be at the expense of a plant's water-use efficiency. Root interception of Pi can be increased by root proliferation, increased frequency and length of root hairs, a modified root architecture that enhances allocation to shallow soil horizons, and mycorrhizal symbioses. Diffusion of Pi toward the root can be increased by increasing the moisture content of dry soil, or by increasing the Pi concentrations in the soil solution through release of Pi from complexed, sorbed or organic forms of P. This review focuses on structural and functional root traits that enhance Pi acquisition from soil with a low availability of Pi, i.e. soils with a reasonable amount of total P, but where diffusion of Pi towards the root limits plant growth. In particular, it deals with traits of native species naturally occurring on soils with a low Pi availability, to explore the potential of these traits for future crop plants. The focus will be on species native to south-western Australia and the Cape region in South Africa, two of the world's 25 biodiversity hotspots. Both regions were once part of the southern-hemisphere super-continent Gondwanaland, and their soils are ancient and deeply weathered, especially those in Western Australia, some of which are estimated to over be 3 billion years old (White, 1986). As soils weather over thousands to millions of years, both the total P levels and the availability of Pi decline (Walker and Syers, 1976; Crews et al., 1995; Richardson et al., 2004). The decline in availability results from the main initial soil P-containing mineral, calcium apatite, being utilized by organisms to form organic P, and by sorption of P onto the surfaces of other minerals. This mineral-sorbed P is labile and can be desorbed in response to diffusion gradients as a result of Pi uptake by plant roots, or can be chemically displaced by root exudates. Over (geological) time, mineral-sorbed P can also be surrounded (occluded) by Fe and Al, rendering it essentially unavailable to plants. The evolutionary consequence of this decline in P levels and Pi availability is an incredibly diverse array of plant species with present-day root adaptations with remarkable ability to acquire sparingly available soil P, and to use internal P efficiently, and that could be explored for future use in crops.
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COMMON ROOT TRAITS TO ENHANCE Pi ACQUISITION |
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Root architecture
This denotes the spatial configuration of roots of different order and age, with the implication that the overall configuration has some functional significance (Lynch, 1995
). In terms of P nutrition, the root architecture of Phaseolus vulgaris when grown at low Pi supply is significant for immobile nutrients such as P because more laterals are produced in shallow soil horizons, where most of the P is located (Lynch and Brown, 2001). Fitter et al. (2002) showed that root architecture had a major effect in Arabidopsis thaliana, both in a pot experiment and in a field experiment conducted under natural conditions, on the fitness of a mutant with a reduced number of lateral roots relative to its isogenic wild-type when P was the limiting nutrient. As expected, when the more mobile nitrate was the limiting nutrient for plant growth, there was no difference in fitness between mutant and wild-type.
Root biomass
Most species allocate more biomass to roots when Pi is limiting for their growth (Brouwer, 1963, 1983). Some of the observed difference in biomass allocation pattern between plants grown with a high vs. a low supply of Pi may be ontogenetic, owing to comparisons of plants at different sizes, rather than a truly plastic response (Kemp and Blair, 1994; Niklas, 1994). However, there is also clear evidence that Pi supply has a direct effect on biomass partitioning, independent of ontogeny (Ryser et al., 1997; De Groot et al., 2001). Interestingly, many Lupinus species, some known to be highly P-efficient, show little change in biomass partitioning to roots as dependent on Pi supply (Keerthisinghe et al., 1998; Pearse et al., 2006; S. J. Pearse, unpubl. data). This low plasticity has been found in both cluster-root-bearing and non-cluster-root-forming Lupinus species (S. J. Pearse, unpubl. data). Effects of Pi supply on biomass partitioning between roots and shoots are thought to involve a decreased production in and export of cytokinins from roots at a low Pi supply, possibly associated with a decreased rate of uptake and metabolism of nitrogen (Kuiper et al., 1989). In the case of Lupinus species, would that mean that they show no change in cytokinin production and export as dependent on their P status? Or might they have a limited response to cytokinins? Do they respond to variation in N supply? Given that we do know that L. albus and L. mutabilis do increase their biomass partitioning to roots (increased root mass ratio) under water stress (Carvalho et al., 2004), a low plasticity in root mass ratio supply is either not typical for all species in this genus or is restricted to effects of P. Is the lack of response to Pi as found for some Lupinus species linked to the capacity of some species in this genus to produce cluster roots? Interestingly, cytokinins play a role in both biomass partitioning (Kuiper et al., 1989) and cluster-root formation, as antagonists of auxins (Neumann et al., 2002). Would that mean that the Lupinus species lacking the capacity to produce root clusters can relatively easily be modified into cluster-forming plants? So far, there are no data in the literature to provide satisfactory answers to many of the questions raised here.
Root length
In field plots of Beta vulgaris, total root-length production over the entire growing season was 3–4 times the size of the living root system at harvest (120 km m–2) in high-P plots, and five times (200 km m–2) in low-P plots (Steingrobe, 2001). The author calculated a 25 % increase in Pi uptake at low P supply as a result of this enhanced root-length production compared with that at the root production of high-P plants. A similarly enhanced root-length production at a low P supply has been observed for Hordeum vulgare (Steingrobe et al., 2001). Increased root production, without a proportional increase in living-root biomass, i.e. enhanced root turnover, allows greater amounts of uptake of immobile soil resources, such as P. Fast root turnover is a very important trait of cluster-root-producing species, as discussed below (Shane and Lambers, 2005a).
Specific root length
In the cases where plants were found to respond to Pi supply with a change in specific root length (SRL), their SRL increased with decreasing Pi supply (Powell, 1974; Christy and Moorby, 1975; Schroeder and Janos, 2005). The increase in SRL is associated with a decrease in root diameter (Powell, 1974), especially for the apical regions of the root system (Mollier and Pellerin, 1999). However, a decrease in root diameter is by no means a universal response to a low Pi supply (e.g. Borch et al., 1999; Schroeder and Janos, 2005).
Root hairs
Root hairs are a fairly common root structure, and increased root-hair length and numbers are considered to be an adaptation that enhances Pi acquisition and a plant's competitive advantage when soil Pi is limiting for growth (Bates and Lynch, 2001). Species that develop more and/or longer root hairs, e.g. Lolium perenne, are far more efficient at accessing Pi from soils, and thus show less of a growth response in P-fertilized soils than do species that lack these traits, e.g. Podocarpus totara (Clarkson, 1981). This point was elegantly demonstrated in a comparison of genotypes of Hordeum vulgare; genotypes with the capacity to form longer root hairs (about 1 mm) took up more P, and tended to yield better when Pi was limiting crop growth compared with genotypes having roots hairs half the length (about 0·5 mm) (Gahoonia and Nielsen, 2004). Root-hair abundance and length is enhanced by P deficiency (Schmidt, 2001). The increased growth of root hairs observed for plants grown at low Pi availability can be mimicked in plants grown at high Pi supplies by adding an ethylene precursor to ‘high-P’ roots. Similarly, root-hair growth can be inhibited by adding the ethylene inhibitor 1-amino-cyclopropane-1-carboxylate (ACC) to the medium of ‘low-P’ roots (Zhang et al., 2003). This suggests that ethylene plays a major role in modulating the growth of root hairs in response to plant P nutrition. A recent detailed anatomical analysis in A. thaliana has shown that the effects of low Pi availability and ethylene on root-hair development differ, with low P status leading to a decreased size and increased number of cortical cells, and increased numbers (approximately double) of root-hair-bearing epidermal cell files, whereas ethylene does not (Zhang et al., 2003). Split-root experiments have established that a shoot-derived signal is required for root hairs to increase in length; the signal is translocated to the roots only when the shoot senses a low P status, and root-hair length is even further enhanced by a low P status in the roots (Jungk, 2001). The signal is unknown but given that auxin is also involved in root-hair formation (Schmidt, 2001), the shoot-derived signal might be an auxin. Increased length and abundance of root hairs is one of the typical adaptive P-starvation-induced plant responses (Fig. 1).
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Mycorrhizal associations
The vast majority (82 %) of all higher plant species have the capacity to form a symbiotic association with a mycorrhizal fungus (Brundrett, 2002
). It is widely accepted that the ancestral mode of Pi acquisition in higher terrestrial plants was through an association with arbuscular mycorrhizal (AM) fungi; more recent mycorrhizal associations include ectomycorrhizas and orchidaceous mycorrhizas (Brundrett, 2002). At low Pi availability, the mycorrhizal symbiosis often enhances plant Pi uptake and growth, especially when the species has a root system that is relatively coarse with few root hairs, e.g. in Citrus species (Graham and Eissenstat, 1994). However, even when there is no effect on net Pi uptake or growth, there can be a major down-regulation of the roots' high-affinity Pi transporters (Smith et al., 2003), possibly in response to an improved plant P status, owing to rapid Pi uptake by the external mycelium. Further research is required to confirm this contention. For further information on the role of mycorrhizas for plant Pi uptake, the reader is referred to recent reviews (Brundrett, 2002; Söderström, 2002). Present-day non-mycorrhizal species must have lost their ability to form an association with AM fungi. Of the minority of species that do not form a mycorrhizal symbiosis, some have specialized root clusters such as proteoid and dauciform root clusters, and specialized physiology associated with rapid rates of carboxylate exudation, as discussed below. However, it should be noted that these specialized adaptations are not restricted to non-mycorrhizal species. There are, indeed, several mycorrhizal species that also have the capacity to produce root clusters, e.g. Casuarina species (Casuarinaceae), Myrica species (Myricaceae) and Viminaria viminalis (Fabaceae) (Neumann and Martinoia, 2002; Shane and Lambers, 2005a). Mycorrhizal associations can even be formed with the root clusters of Hakea verrucosa, a species naturally occurring on soils containing high levels of nickel (Boulet and Lambers, 2005). Furthermore, some non-mycorrhizal species (e.g. Brassicaceae) lack morphological adaptations like root clusters, but show rapid rates of carboxylate exudation (Hoffland et al., 1989). Finally, some species that lack mycorrhizal associations, e.g. Chenopodiaceae and Urticaceae (Lambers et al., 1998), would appear to have no specialized morphological adaptations, and these species tend to be restricted to relatively nutrient-rich sites and habitats with low competitive pressure (Olsson and Tyler, 2004).
High-affinity Pi transporters
Much remains to be discovered about the expression of high-affinity Pi transporters as dependent on plant P status. Recent discoveries have revealed that sugars are integrally related to P-deficiency-induced expression of one of these transporters in L. albus (Liu et al., 2005). Exogenous sugars stimulate accumulation of transcripts of a high-affinity transporter in dark-grown, P-sufficient seedlings. Conversely, in intact P-deficient plants, expression of this transporter in cluster roots was reduced in girdled plants, and in dark-grown plants in which expression was rapidly restored upon re-exposure to light. Similar results were also obtained for a gene encoding acid phosphatase and a third gene, both being P-deficiency induced. There is obviously cross-talk between phosphorus acquisition and carbon metabolism, similar to that between nitrogen and sulfur uptake and carbon metabolism (Lejay et al., 2003). The promoters of the genes encoding the high-affinity transporter and the acid phosphatase contain a short sequence that is identical to the binding site for a transcription activator for P-deficiency-induced genes in A. thaliana (Rubio et al., 2001).
Enhanced expression of high-affinity, plasma-membrane-bound Pi transporters in roots, and a concomitantly increased P-uptake capacity, is a typical P-starvation response (Burleigh and Harrison, 1999; Dong et al., 1999) (Fig. 1). This response is usually interpreted as an acclimation to a low availability of Pi in soil. However, diffusion of Pi in soil is the key limiting factor for Pi uptake, and changes in kinetic parameters of the roots' P-uptake system, including an increase in Imax (maximum Pi inflow rate), have little effect on a plant's capacity to acquire Pi from soil (Silberbush and Barber, 1983; Raghothama and Karthikeyan, 2005). This is not to say that plants do not need a high-affinity system for Pi uptake; rather, it shows that enhancing the expression of this Pi transport system does not have a proportional effect on Pi uptake, and may have no effect at all. This may explain, partly, why over-expression of a high-affinity phosphate transporter in transgenic Hordeum vulgare had no effect on Pi uptake from soil (Rae et al., 2004). What might be the adaptive significance of differential expression of high-affinity Pi transporters? Species that have a very low capacity to adjust their Pi uptake capacity in response to changes in Pi supply in the root environment show signs of P toxicity at elevated Pi supply (Shane et al., 2004a, b; Shane and Lambers, 2006). Therefore, we suggest that the capacity to down-regulate Pi transporters at high Pi supply is the trait that has adaptive significance, rather than the capacity to up-regulate Pi transporters at a low Pi supply. Although over-expression of high-affinity Pi transporters may not enhance Pi acquisition from soil, we envisage that it might improve internal P utilization. P-starved A. thaliana plants have been found to express high-affinity Pi transporters in roots as well as in developing flowers and fruits (Karthikeyan et al., 2002). If expression of high-affinity Pi transporters can be reduced in reproductive organs of grain crops, accumulation of P in seeds may be decreased, allowing P to be utilized in photosynthetic tissues, which could lead to increased grain production, and greater P return to the soil in organic form.
Effects of phosphite on P-starvation responses
A low plant P status induces the P-starvation responses as discussed above. Many of these responses are suppressed by phosphite, which acts as an analogue of phosphate. Typically suppressed P-starvation responses include increased allocation to root biomass (Varadarajan et al., 2002), enhanced root-hair formation (Ticconi et al., 2001), up-regulation of the high-affinity phosphate transporters and acid phosphatases (Varadarajan et al., 2002), and cluster-root formation (Gilbert et al., 2000). Phosphite inhibits mycorrhiza formation in Zea mays (Seymour et al., 1994), but not in Allium cepa (Sukarno et al., 1998), Eucalyptus marginata, E. globulus or Agonis flexuosa (Howard et al., 2000). The contrasting results for mycorrhization might be due to the fact that phosphite inhibits the expression of high-affinity phosphate transporters (Varadarajan et al., 2002), which would lower the plant P status, and thus indirectly enhance mycorrhization. Phosphite is also used as a fungicide, e.g. to combat the soil-borne plant pathogen Phytophthora cinnamomi in natural ecosystems (Hardy et al., 2001). Considering the effect of phosphite on P-starvation responses, the use of phosphite as a fungicide in pristine ecosystems clearly needs further scrutiny.
Rhizosphere alteration
As discussed above, enhanced root production is an adaptive response to acquire poorly mobile soil resources. An alternative strategy is to enhance the availability of Pi in soil. There are two fundamentally different mechanisms to enhance Pi availability. First, in superficial, dry soil horizons, where most of the P will be located, the mobility of Pi can be enhanced by the release of water into that dry soil. The released water would originate from moister regions in the soil, and be transported inside the root system in a process termed ‘hydraulic redistribution’ (Burgess et al., 1998, 2000). When this process was first described for desert plants that took up water from deep soil layers and released it, at night, into superficial layers, it was termed hydraulic lift. However, it has since been established that water can flow downwards as well as upwards, and it is envisaged that it will also move horizontally, from roots in moist superficial patches via the stem to the roots in drier soil, where the water can be released. Secondly, the concentration of Pi, the only form of P that is taken up by roots, can be enhanced by the release of root exudates, particularly carboxylates and phosphatases (Raghothama and Karthikeyan, 2005). These two strategies to enhance Pi availability are discussed in the next two sections.
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ENHANCED Pi UPTAKE ASSOCIATED WITH HYDRAULIC REDISTRIBUTION? |
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When some roots are in contact with moist soil, while others on the same plant are in dry soil, water may move from moist to dry patches. This was first discovered in desert shrubs, where water can move from moist deep soil layers into shallower dry soil, and was termed ‘hydraulic lift’ (Caldwell and Richards, 1989
). The water that is transported from deeper and moist layers into shallower dry soil is available for the species that exhibited hydraulic lift as well as for neighbouring plants (Caldwell and Richards, 1989). Hydraulic lift usually occurs at night, in C3 species, but can also occur during the day in CAM species (Yoder and Novak, 1999). However, water can also move from roots in shallow layers, moistened after rain, to roots deep in the profile (Burgess et al., 1998). Equally, water can move horizontally, via the stem, depending on soil water content (Hultine et al., 2003), and hence the term ‘hydraulic redistribution’ is more appropriate (Burgess et al., 1998, 2000). Hydraulic redistribution is not restricted to woody species, but has also been demonstrated in the crop species Pennisetum americanum (Vetterlein and Marschner, 1993). Water uptake by roots requires the expression of water-channel proteins (aquaporins); these proteins are expressed in a diurnal pattern, with low expression levels at night and increasing expression early in the day (Vandeleur et al., 2005). One would expect that water release also requires expression of water-channel proteins, but this has not yet been investigated.
Because diffusion of Pi in dry soil is very slow (Amijee et al., 1991; Bhadoria et al., 1991), Pi uptake declines with decreasing soil moisture content (Turner and Gilliam, 1976; Vig and Singh, 1983; Mouat and Nes, 1986). Therefore, nutrient uptake from dry shallow patches in soil is expected to increase when the soil is moistened due to hydraulic redistribution (Vetterlein and Marschner, 1993; Horton and Hart, 1998; Huang, 1999), and this could be especially significant for poorly mobile nutrients such as P. Equally, when deeper soil layers contain abundant P reserves, hydraulic redistribution down the profile might enhance the uptake of Pi (McCulley et al., 2004). This interesting concept of enhancing Pi availability by hydraulic redistribution is, however, particularly difficult to approach experimentally, and hence there is little convincing evidence to support it. Valizadeh et al. (2003) found that P banded in dry topsoil was accessed by Triticum aestivum with access to moist subsoil, as a result of to the release of hydraulically lifted water.
In summary, there is a wealth of information on the occurrence of hydraulic redistribution and the use of hydraulically lifted water by neighbouring plants. It is highly likely that hydraulic redistribution enhances Pi acquisition from P-enriched, dry soil patches. However, further research is required to establish the extent to which hydraulic redistribution may favour Pi acquisition.
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ENHANCED Pi UPTAKE ASSOCIATED WITH THE RELEASE OF ROOT EXUDATES |
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In addition to increasing the diffusion coefficient of Pi in soil by hydraulic redistribution as discussed above, root activity can also enhance the concentration of Pi in soil, owing to the release of exudates.
Carboxylates
Carboxylates (e.g. citrate, malate) can be major components of exudates released by roots, especially under P deficiency (Gardner et al., 1983; Hoffland et al., 1989; Keerthisinghe et al., 1998). However, some high-exuding plant species, e.g. Cicer arietinum, appear to release carboxylates (mainly malonate) constitutively (Wouterlood et al., 2004). Carboxylates mobilize both inorganic P and organic P (Po), because they complex metal cations that bind phosphate and displace phosphate from the soil matrix by ligand exchange (Fig. 2) (Gerke et al., 2000; Hayes et al., 2000; Jones et al., 2003). The cations excreted together with the carboxylates to maintain charge balance may be protons, leading to rhizosphere acidification (Hinsinger, 2001; Hinsinger et al., 2003). However, other cations, especially K+, are at least as important (Y Zhu et al., 2005), and carboxylate exudation is not invariably associated with acidification (Roelofs et al., 2001). Transport of carboxylates in the anionic form from the cytosol (pH 
7·2–7·5) into a more acidic rhizosphere is likely to result in protonation of the carboxylates in the rhizosphere. This is likely to contribute to scavenging of H+ from the rhizosphere, and hence increase the rhizosphere pH. In fact, unless the soil pH is initially alkaline, acidification does not enhance Pi availability; rather, acidification immobilizes Pi at low pH due to the formation of Fe and Al complexes (Lambers et al., 1998). In addition to P immobilization, acidification can influence the extent of ionization of carboxylates (Jones, 1998b; Hinsinger et al., 2003), which can reduce their chelating ability, potentially rendering them ineffective in acidified soil (Pearse et al., 2006)
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Phenolics and mucilage
Exudation of phenolics may also increase under P deficiency (Neumann and Römheld, 2001
; Juszczuk et al., 2004), but this has received less attention in the literature. Similarly, release of mucilage can be enhanced under P deficiency, and this can also enhance Pi availability in soil (Nagarajah et al., 1970; Gaume et al., 2000; Grimal et al., 2001). Phenolics and mucilage act in the same way as carboxylates (Guppy et al., 2005), but tend to be less effective than carboxylates (Neumann and Römheld, 2001). In addition, release of phenolics may serve a fungistatic role (Weisskopf et al., 2006), as further discussed in the section below dealing with root clusters.
Phosphatases
Organic P typically accounts for 30–80 % of total P in soil (Pederson, 1953; Tarafdar and Claassen, 1988; Adams, 1992). Soil organic P compounds (mainly phosphate mono- and di-esters, Sumann et al., 1998), after having been mobilized by carboxylates, must first be hydrolysed, to release Pi for plant uptake (George et al., 2002) (Fig. 2). Acid phosphatases can hydrolyse a range of organic P compounds (Tarafdar and Claassen, 2001), and both expressed sequence tags for phosphatase (Uhde-Stone et al., 2003) and these enzymes are more abundant in the rhizosphere when plants are P starved (e.g. Li et al., 1997; Gilbert et al., 1999; Yun and Kaeppler 2001; Wasaki et al., 2003). Phytases are required to hydrolyse phytate (= myo-inositol penta- and hexa-phosphates), which is fairly resistant to other phosphatases (Hayes et al., 2000). Phytate can be a major component of the soil organic P pool (Pederson, 1953; McKercher and Anderson, 1968). Phosphatases and phytases in soil may be of microbial origin (Tarafdar and Claassen, 2001), but roots also exude phosphatases (Tarafdar and Claassen, 2001, 2005), and roots of some species also release significant amounts of phytases (Li et al., 1997). Most plants have a very limited capacity to access phytate in the rhizosphere, except in the presence of micro-organisms that can dephosphorylate phytate (Richardson et al., 2001). Transgenic plants of A. thaliana, exhibiting enhanced exudation of extracellular phytase (derived from Medicago truncatula) from their roots, have greater access to phytate than their wild-type (Xiao et al., 2005). Similarly, transgenic plants of Trifolium subterraneum, exhibiting enhanced, constitutive expression and exudation of a phytase derived from Aspergillus niger, had better access to phytate than wild-type plants (George et al., 2004). However, this effect was only pronounced when plants were grown in non-sorbing, sterile laboratory media, and much less so when plants were grown in soil where phytase is rapidly immobilized, limiting its ability to interact with phytate (George et al., 2005). This suggests that phytate can only be dephosphorylated by phytase after it has been mobilized into the soil solution by, for example, carboxylates (Fig. 2).
Exudation as dependent on soil moisture
Phosphate-starvation responses are controlled systemically, via signals originating in the shoot (Abel et al., 2002; Fig. 1). This explains why a low soil moisture content, which reduces the mobility of Pi in soil (e.g. Turner and Gilliam, 1976; Bhadoria et al., 1991), and hence tends to lower the plant's P status, enhances root exudation (Liebersbach et al., 2004). As a consequence, Pi uptake is affected much less by water shortage than expected on the basis of the effect of soil moisture on Pi mobility in soil.
In summary, roots of many species release an array of exudates (e.g. carboxylates, phenolics, protons and other cations, phosphatases, water, mucilage), and thus enhance the availability of Pi in the rhizosphere. The nature and effectiveness of the exudates depends on species as well as environmental conditions. A low plant P status tends to enhance exudation.
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SPECIALIZED ROOT STRUCTURES: ROOT CLUSTERS |
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The specialized roots discussed here, collectively called ‘root clusters’, combine a specialized structure and specialized physiology (see below) to maximize Pi acquisition from soils of low fertility, especially when P is present in ‘sorbed’ or insoluble sources (e.g. rock phosphate and iron phosphate). Proteoid (e.g. Keerthisinghe et al., 1998
) and dauciform (Shane et al., 2005; Playsted et al., 2006) root clusters are induced by P deficiency and occasionally by Fe deficiency (reviewed in Shane and Lambers, 2005a). P deficiency induces a wide range of genes in cluster roots of L. albus, including genes involved in carbon metabolism, secondary metabolism, P scavenging and remobilization, plant hormone metabolism, and signal transduction, when compared with P-sufficient and P-deficient non-cluster roots (Uhde-Stone et al., 2003).
There are several ‘types’ of root clusters, occurring in both monocotyledonous and in dicotyledonous species (Fig. 3). The best known examples are the ‘bottlebrush-like’ proteoid roots (Fig. 3A, B) described by Purnell (1960) for woody species of Proteaceae. Proteacean taxa are distributed primarily in Australia and South Africa, but proteoid-like roots have also been described in a range of other species from several families, e.g. in Fabaceae [Lupinus albus (white lupin) and Aspalathus linearis (rooibos)] (Fig. 3E, F) (Dinkelaker et al., 1995; Shane and Lambers, 2005a). Monocotyledonous families containing rushes (Restionaceae from the southern hemisphere) and sedges (Cyperaceae with a worldwide distribution) form root clusters termed ‘dauciform’ roots (in sedges; Fig. 3C, D) and ‘capillaroid’ roots (in rushes; Fig. 3G–I).
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1 µM [P] (except for the roots of a plant shown in E, which were collected in soil containing Root cluster morphologies involve formation of compact clusters of (determinate) branch roots (rootlets), or root hairs, in a small soil volume which markedly increases the surface area of the root system (Fig. 3). Moreover, root clusters are ephemeral; even in the woody species that develop them, rootlets remain in the primary state of growth until they senesce, and although we know little about their turnover, it is becoming apparent that these fine roots are physiologically active for little more than a few weeks (Shane et al., 2004c
, 2005b; Playsted et al., 2006). The root architecture of field-grown, root-cluster-forming species is patterned toward strongly soil-binding root-cluster development in upper soil layers where levels of nutrients are often enriched. Briefly, the basic root-cluster structures are as follows (the reader is referred to Lamont, 2003, for more details). Proteoid roots formed by species in Proteaceae are either ‘simple’ or ‘compound’, but occasionally both types are found on single root systems (e.g. in South African genera of Leucadendron and Protea; Lamont, 1983). Only a few genera within Proteaceae (e.g. Australian Banksia and Dryandra and South African Orothamnus; Lamont 1982, 1983) produce ‘compound’ proteoid root clusters (e.g. Banksia grandis; Fig. 3A). The ‘compound’ proteoid roots are essentially multiples of the ‘simple’ root clusters, but there are likely to be distinct ecophysiological reasons for the differences between the simple cluster roots and the compound clusters (Fig. 3), which tend to form dense root-mats in natural systems. Many proteacean genera (e.g. Australian Hakea and South African Serruria), and genera of other families [e.g. Fabaceae including the South African Aspalathus (rooibos) and the crop species Lupinus albus] produce ‘simple’ proteoid roots (Fig. 3B, E, F) that include numerous short, determinate rootlets, and each simple proteoid root is separated by unbranched regions (Fig. 3B). The main difference between the simple root clusters of Proteaceae (e.g. Hakea; Fig. 3B) and some species in the Fabaceae (e.g. Aspalathus linearis; Fig. 3F) is most notably the density of rootlets produced per unit root axis, which is far greater in the Proteaceae. Within the genus Lupinus, some species, e.g. L. albus, produce easily recognizable root clusters, whereas others form structures that are distinct from non-cluster roots and have been termed ‘cluster-like’ roots by Hocking and Jeffery (2004), who showed that their physiology (below) is rather similar to that of the root clusters in L. albus.
Dauciform root clusters were first described by Russian plant scientists for Cyperaceae (sedges) (Selivanov and Utemova, 1969, and references cited therein). They were subsequently found in cyperacean species in Great Britain (Davies et al., 1973; Ballard, 2001), continental Europe (Bakker et al., 2005; Güsewell, 2005), and in many parts of Australia (Lamont, 1974; Phillips and Weste, 1984; Shane et al., 2005a; Playsted et al., 2006) and New Zealand (Powell, 1973). Lamont (1974) named them ‘dauciform’ roots, because of the carrot-shape of the dauciform root axis (Fig. 3C, D). Dauciform roots often occur in groupings of 20–30 individuals (Lamont, 1974) and each dauciform root may be as short as 2 mm, e.g. in Carex (cosmopolitan) species, or much longer, e.g. up to 12 mm in Lepidosperma (Western Australian) and Tetraria (South African) species (Fig. 3C, D, respectively) (Lamont, 1974; Shane et al., 2005a; Neumann and Römheld, 2006). Instead of the usual formation of dense clusters of short rootlets, the mature axis of a dauciform root is covered with dense clusters of long (approx. 2 mm; Fig. 3C, D) root hairs. The tips of the dauciform root axis are either indeterminate (and may form additional dauciform roots in sequence along the main axis; see Fig. 3C), or the dauciform root tip is determinate, and the entire dauciform root senesces (cf. figure 1 in Shane et al., 2005b).
The monocotyledonous family of the Restionaceae has a Gondwanan distribution (Pate and Meney, 1999). Approximately 486 species are located mainly in Africa (over 300 species in South Africa) and in mainland Australia and Tasmania (approx. 150 species), and a few species are found in New Zealand, South America (Chile) and South East Asia (Indochina). Approximately half of the Australian taxa develop root clusters, especially species adapted to arid environments, where their development begins only after the onset of seasonal rains (figure 1·3 in Meney and Pate, 1999). These ‘capillaroid’ roots were discovered and named by Lamont (figure 4 in Lamont, 1982), and are characterized by clumps of roots or rootlets, densely covered with exceptionally long root hairs (Fig. 3G–I). Their name (capillaroid) stems from the sponge-like properties on holding soil water (Lamont, 1980, 1982). Little is known about their structure and development in species of Restionaceae and how these specialized roots contribute to plant nutrition and water balance. We have recently found root clusters in South African Restionaceae that are remarkably ‘proteoid-like’ in their morphology (Fig. 3H), and produce distinct (ephemeral) clusters separated by unbranched main root axis. However, most species observed thus far have the morphology typical of that shown in Fig. 3G, I. We hypothesize that the physiology and functioning of capillaroid roots is similar to that of proteoid roots.
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Most physiological information about root-cluster functioning has been derived from studies of L. albus, but much has also been discovered about root-cluster functioning in native plants adapted to soils of extremely low Pi concentration. One of the most important aspects is the importance of the influence of the stage of development on root-cluster functioning. The finding that carboxylate (e.g. citrate) release in L. albus (Watt and Evans, 1999
) and in proteacean species such as Hakea (H. prostrata, Shane et al., 2004c; H. undulata, Dinkelaker et al., 1997) occurs in an exudative burst strongly supports the view that root development and physiological activity are closely linked as the components of root systems grow and mature (McCully, 1999). Studies of the ‘compound’ (mat-forming) proteoid roots of Banksia integrifolia have also shown that carboxylates, such as citrate, are released into the rhizosphere (Grierson, 1992; Roelofs et al., 2001), but there are no reports on the time course of carboxylate exudation in taxa that produce compound roots. It has now become apparent that dauciform roots in Cyperaceae, although morphologically and anatomically very distinct, also release carboxylates (citrate) in large quantities during a developmentally programmed exudative burst, thus functioning in a way very similar to proteoid roots (Shane et al., 2005b, 2006; Playsted et al., 2006). Another parallel between dauciform and proteoid roots is that both are suppressed when plants have a high P status. Like proteoid roots, dauciform roots release a variety of other compounds, as further discussed below. There is no physiological information on capillaroid roots. In summary, root clusters differ greatly in their anatomy and morphology, but are rather similar with respect to their physiology. Our knowledge on capillaroid roots is restricted to their anatomy and morphology; their physiology remains to be investigated. The release of carboxylates from root clusters of Proteaceae and Fabaceae, and dauciform roots of Cyperaceae in an exudative burst is bound to be vital for their function, as further discussed in the section ‘Root clusters: combining structure and functioning’.
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PHYLOGENY OF ROOT-CLUSTER-FORMING SPECIES |
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Proteoid root clusters were perhaps once considered as a curiosity associated with many proteacean species (Purnell, 1960
) and L. albus (Gardner et al., 1983). However, proteoid roots have since been described for a wide range of species in families that are not at all closely related to Proteaceae (Shane and Lambers, 2005a). Although very few species in the genus Lupinus produce the kind of clusters that are found in L. albus (Clements et al., 1993; Skene and James, 2000), many others produce ‘cluster-like roots’, which function in a similar way to the ‘true cluster roots’ (Hocking and Jeffery, 2004). Within the Cyperaceae (sedges), dauciform roots are restricted to two tribes: Cariceae and Rhynchosporeae (Lamont, 1981). Outside the Cyperaceae, dauciform roots have only been reported for Juncus pauciflorus and J. squarrosus (Juncaceae, reeds) (Powell, 1973). Capillaroid roots have been reported exclusively in the Restionaceae (rush) (Lamont, 1982). Viewed in this way, it is obvious that root clusters (a term we use here to refer to all types of clusters: simple and compound proteoid or cluster roots, dauciform roots, capillaroid roots, cluster-like roots; Fig. 3) are actually more widespread in the plant kingdom than considered before.
Root clusters are found in two large monocotyledonous families: Cyperaceae (dauciform roots) and Restionaceae (capillaroid roots) (Fig. 4). Root clusters also occur in several dicotyledonous families. Proteoid roots were first discovered in the Proteaceae (Purnell, 1960), which belong to the Proteales (Eudicots). This accounts for the name ‘proteoid’ roots. There are no records of root clusters in other families within the Proteales; while growing over a year in low-P nutrient solution in the glasshouse, Platanus hybrida (Platanaceae, Proteales) never produced any root clusters (H. M. Stace and H. Lambers, unpubl. data). Apart from the Proteales, within the eudicots, root clusters have been described in Core Eudicots (Rosids) only; that is, they occur in several families that are phylogenetically very distantly related to the Proteaceae (Fig. 4) (Skene, 2000; Shane and Lambers, 2005a). Within the Rosids, root clusters occur in four orders belonging to the eurosids I (fabids): Fagales (Betulaceae, Casuarinaceae, Myricaceae), Cucurbitales (Cucurbitaceae), Rosales (Elaeagnaceae, Moraceae) and Fabales (Fabaceae). Root clusters have obviously evolved several times. What is very striking is that actinorhizal species belonging to different orders and families, namely Betulaceae (Fagales), Casuarinaceae (Fagales), Eleagnaceae (Rosales) and Myricaceae (Fagales), all have the capacity to produce root clusters. Why is there an association between root-cluster-bearing habit and being actinorhizal? Does a plant's capacity to recognize and form an association with Frankia species have something in common with its capacity to develop clusters? Further investigations of the signal-transduction pathways involved in the actinorhizal symbioses and root-cluster formation may lead to fascinating new discoveries. In addition, a careful study of species belonging to the four other actinorhizal families, Rosaceae, Rhamnaceae (both Rosales), Coriariaceae and Datiscaceae (both Cucurbitales) (Swensen, 1996; Vessey et al., 2005), might well reveal more records of cluster-root-bearing species.
Root clusters are no longer the curiosity restricted to plants from ‘down under’, but occur in many distantly related families throughout the plant kingdom. Many species are used as crops, for nuts (Macadamia species), a source of protein (Lupinus species), tea (Aspalathus linearis), timber and pulpwood (Grevillea species) (Shane and Lambers, 2005a). Others may be used in pastures; for example, in eastern Canada and western North America some sedges (Carex sp.) are recognized for their potential for use as forage for grazing (Uresk, 1986; Catling et al., 1994), and Kennedia species for introduction as food (Rivett et al., 1983) or pasture plants (Cocks, 2001). Considering our dwindling P reserves, these cluster-bearing species need to receive greater emphasis in future research.
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RELATIONSHIPS BETWEEN P-ACQUISITION STRATEGIES AND SOIL TYPES |
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Having discussed the structure and functioning of root clusters in different plant families, we now explore where root-cluster-bearing species fit in the landscape. The Western Australian flora offers a unique opportunity to explore that question. Both non-mycorrhizal, cluster-bearing species belonging to the Cyperaceae and Proteaceae, and mycorrhizal species without root clusters are common in south-western Australia, a global biodiversity hotspot (Myers et al., 2000
). In addition, there are several species that are both mycorrhizal and cluster-bearing (e.g. Casuarinaceae and Fabaceae). Are the different nutrient acquisition adaptations distributed randomly or linked to certain habitat factors? We can address this question in some detail using McArthur's (1981) detailed descriptions of soil and vegetation for 150 reference sites in south-western Western Australia. Non-mycorrhizal, cluster-bearing Proteaceae predominate on the most P-impoverished soils in the region, whereas mycorrhizal Myrtaceae without root clusters predominate on soils that have somewhat higher P levels (Fig. 5). Whereas Myrtaceae in Western Australia generally dominate forests and tall woodlands, and Proteaceae generally dominate shrublands/heaths and low woodlands, the occurrences of species of the two families are by no means mutually exclusive. Proteaceae understorey sp