Summary The capacity for nitrate reduction, as measured by nitrate reductase activity (NRA), was generally low for a range of plant communities in Australia (coastal heathland, rainfor-est, savanna woodland, monsoon forrainfor-est, mangrove, open Euca-lyptus forest, coral cay open forest) and only a loose relationship existed between NRA and leaf nitrogen concen-tration. This suggests that nitrate ions are not the sole nitrogen source in these communities. Based on 15N labeling experi-ments, we found a range of tree species exhibiting a pro-nounced preference for uptake of ammonium over nitrate. Analysis of soil solutions from several forest and heathland communities indicated that ammonium ions were more preva-lent than nitrate ions and that soluble forms of organic nitrogen (amino acids and protein) were present in concentrations simi-lar to those of mineral nitrogen. To determine the extent to which root adaptations and associations might broaden nitro-gen source utilization to include organic nitronitro-gen, we assessed the effects of various nitrogen sources on seedling growth in sterile culture. Non-mycorrhizal seedlings of Eucalyptus gran-dis W. Hill ex Maiden. and Eucalyptus maculata Hook. grew well on mineral sources of nitrogen, but did not grow on organic sources of nitrogen other than glutamine. Mycorrhizal seed-lings grew well on a range of organic nitrogen sources. When offered a mixture of inorganic and organic nitrogen sources at low concentrations, mycorrhizal seedlings derived a significant proportion of their nitrogen budget from organic sources. We also demonstrated that a species of the obligately non-mycor-rhizal genus Hakea, a heathland proteaceous shrub possessing cluster roots, had the ability to incorporate 15N-labeled organic sources (e.g., glycine). We conclude that mycorrhizal associa-tions and root adaptaassocia-tions confer the ability to substantially broaden the nitrogen source base on some plant species. Keywords: ammonium, cluster root, mycorrhiza, nitrate, ni-trate reductase.
Introduction
Several root characteristics are reported to enhance mineral nutrition, including various kinds of mycorrhizal roots (Harley and Smith 1983), cluster roots (Lamont 1982) and N2-fixing
nodulated roots (Sprent and Sprent 1990). With the exception of nodulated roots, the significance of these root specializa-tions for nitrogen nutrition of plants in natural ecosystems is less clear than for phosphorus nutrition (Alexander 1983, Dinkelaker et al. 1995). Nitrogen availability may be a key factor determining photosynthetic capacity of natural plant communities (Field and Mooney 1986), especially in plant communities for which acute deficiencies of nitrogen are com-mon.
Vesicular-arbuscular, ericoid, orchid and ecto-mycorrhizae all play a role in plant nitrogen acquisition (Read 1991). Alexander (1983) showed that ectomycorrhizae enhance ni-trate and ammonium uptake by plants, because the fungal hyphae increase the effective volume of soil exploited by the roots. Mycorrhizal fungi can also utilize organic forms of nitrogen (Abuzinadah and Read 1986, Finlay etal. 1992), thus making forms of nitrogen available to the host that would otherwise be unavailable to it. This capacity may be of particu-lar importance for woody plants growing in nutrient-poor soils (Read 1991).
Root clusters are aggregations of hairy rootlets that are produced on the root systems of many plants (Lamont 1982). Species with cluster roots are able to grow on poor soils, especially those low in phosphorus and nitrogen (Lamont 1993). Cluster roots improve the nutrition of P, Fe, and Mn by altering rhizosphere conditions through excretion of organic acids and phenolics (Dinkelaker et al. 1995). Cluster roots are induced in species of the genus Hakea under N-limiting con-ditions and are mainly produced in soil horizons rich in organic matter (Lamont 1973). The possible significance of proteoid roots for N nutrition was noted by Pate and Jeschke (1993) who found higher concentrations of amino acids in the xylem sap of proteoid roots than of non-proteoid roots of Banksia prionotes Lindley.
Three distinct groups of N2-fixing root associations can be identified: the rhizobial associations of leguminous species, the actinorrhizal associations of the genus Casuarina and the coralloid roots of cycads (Lamont 1984). The haustoria of root hemiparasites constitute another potentially important root specialization implicated in the uptake of organic nitrogen
Root adaptation and nitrogen source acquisition in natural ecosystems
MATTHEW H. TURNBULL,
1SUSANNE SCHMIDT,
2PETER D. ERSKINE,
2SUANNE RICHARDS
2and GEORGE R. STEWART
2,31 Department of Plant and Microbial Sciences, University of Canterbury, Christchurch, New Zealand 2
Department of Botany, The University of Queensland, Brisbane, Queensland 4072, Australia 3 Author to whom correspondence should be addressed
Received October 25, 1995
(e.g., in the genera Exocaropus, Anthobolus, Santalum and Striga) (Lamont 1984).
To date, the importance of mycorrhizal associations and other root adaptations to the nitrogen nutrition of species from forest systems in sub-tropical and tropical regions has received little attention. Because the supply of nitrogen in these systems may be limiting to plant growth (Bowen 1981), we have explored the impact of root specializations on the potential to utilize various nitrogen sources and their subsequent metabo-lism in several subtropical species. We have analyzed enzy-matic activities and metabolism in plants from a range of sub-tropical and tropical communities, and demonstrated that different plant species utilize different nitrogen sources and that the type of root specialization strongly influences the characteristics of nitrogen assimilation.
Materials and methods
Study sites and plant material
Measurements were made in nine plant communities from sub-tropical and tropical northern Australia: (1) sub-tropical coastal heathland (Beerwah State Forest, 60 km north of Bris-bane, Queensland); (2) semi-arid ‘‘mulga’’ woodland (Curraw-inya National Park, 1000 km west of Brisbane, Queensland); (3) sub-tropical rainforest (Lamington National Park, Green Mountains, 120 km south of Brisbane, Queensland); (4) sa-vanna woodland (Kakadu National Park, 200 km east of Dar-win, Northern Territory); (5) monsoon open forest (Kakadu National Park, 200 km east of Darwin, Northern Territory); (6) eucalypt open forest (Brisbane Forest Park, Brisbane, Queens-land); (7) tropical mangrove forest (Kakadu National Park, 200 km east of Darwin, Northern Territory); (8) coral cay forest (Heron Island, Great Barrier Reef Marine Park, Queensland); and (9) tropical rainforest (Kakadu National Park, 200 km east of Darwin, Northern Territory). Data for other plant communi-ties have been included where appropriate and are sourced from previous works as indicated in the text.
Nitrate reductase assays
Terminal leafy material from the principal overstory and un-derstory species was collected from each community. The suite of species studied generally represented over 80% of the com-munity biomass--ground cover at the respective sites. Nitrate reductase activity (NRA) was determined on freshly harvested leaf material by an in vivo assay (Stewart et al. 1986). Leaf nitrogen concentration was determined in oven-dried and finely ground material by automated combustion.
15N Labeling experiments
The pathway of incorporation and metabolism of 15N-labeled substrates (NH4+, NO3−, glycine) was determined after the seed-lings had been incubated for 8 h in a fresh aerated liquid medium containing 1 mol m−3 nitrogen (99 or 65 atom% excess 15N). Experiments were conducted on
Eucalyptus sp. seedlings grown in sterile culture and on Hakea seedlings (Hakea sp. Mt. Coolum P.R. Sharpe 3338; formerly H.
gib-bosa)excavated from the field and transferred to liquid culture. Hakea sp. seedlings were fed either a single 15N-labeled nitro-gen source (NH4+, NO3− or glycine) or were offered the three nitrogen sources simultaneously, with one being 15N labeled and the other two at natural abundance. At the end of the incubation period, leaves and roots were extracted in methanol and analyzed as described below.
Gas chromatography--mass spectrometry
Methanolic extracts from labeling experiments were prepared and derivatized as described by Kershaw and Stewart (1992). The amount of 15N incorporated into each amino acid (includ-ing glutamine and asparagine) was determined by GC--MS analysis as described by Turnbull et al. (1995). The presence of free amino acids in the methanol-soluble fraction of fungal mycelium was determined by a post-column ninhydrin derivi-tization, HPLC-based amino acid analyzer (Model 6300, Beckman Instruments, Palo Alto, CA).
Soil solution analysis
Soils from the coastal heathland (Site 1) and eucalypt open forest (Site 6) were sampled. At Site 6, soils from two distinct open-forest types were analyzed, a moist fertile forest domi-nated by Eucalyptus grandis W. Hill ex Maiden. and a ridge top dominated by Eucalyptus maculata Hook.Soluble compo-nents were extracted from the soil samples (0--5 cm depth) in distilled water or in 1 mol m−3 KCl, and nitrate, ammonium, amino acids and soluble protein were determined as described by Turnbull et al.(1995). At the coastal heathland site (Site 1), we compared the immediate availability of ammonium, nitrate and amino acids in the soil solution of the rooting zone by means of mixed-bed ion exchange resin bags (Stewart et al. 1993). Ammonium and nitrate in the eluate were assayed as described by Stewart et al. (1993). Amino acid concentration was determined by HPLC.
fine powder in a vibratory ball mill (Retsch MM-2, Haan, Germany) and nitrogen concentration of the leaf material was determined by automated combustion. Duplicate samples of approximately 120 µg nitrogen were analyzed for 15N with a continuous flow isotope ratio mass spectrometer (CF-IRMS, Tracer Mass, Europa Scientific, Crewe, U.K.) set to the single nitrogen mode. Precision of the instrument, based on multiple analysis (n = 133) of a laboratory standard (Eucalyptus crebra F.J. Muell. leaves) was 0.21 SD. The percentage of plant nitrogen derived from the unlabeled organic source (Forg) was calculated as:
Forg = 1 −
Atom% 15N excess in plant Atom% 15N excess in labeled source
100.
Results and discussion
Nitrate reduction
Figure 1a describes the relationship between community-aver-aged leaf NRA and leaf nitrogen concentration for nine distinct plant communities from northern Australia. The capacity for reduction of nitrate ions was generally low compared with values reported for plants for which nitrate ions are an impor-tant source of nitrogen (Smirnoff et al.1984). Average NRA ranged from 22 pkat gfw−1 for plants in the coastal heathland (Site 1) to 117 pkat gfw−1 for plants in the semi-arid ‘‘mulga’’ woodland (Site 2). For eight of the nine communities, average NRA was below 100 pkat gfw−1. A few species appeared to be predominantly nitrate utilizers based on their relatively high NR activities and high concentrations of nitrate in leaf extracts and xylem sap (data not shown). Although NRA was generally low in the ecosystems studied, it was not always associated with low leaf nitrogen concentration. There was a weak rela-tionship between community-averaged NRA and leaf nitrogen concentration (r2 = 0.38, Figure 1a). This was also the case within a given community (Figure 1b). For example, in the tropical monsoon forest (Site 5), there was no relationship between NRA and leaf nitrogen concentration, despite the presence of species with a moderately high capacity for nitrate reduction (about 400 pkat gfw−1).
The low rates of nitrate reduction observed in the leaves of species in these systems could be explained if these species are predominantly root nitrate assimilators. However, root NRA in these types of species is also generally low (e.g., average root NRA in mature phase rainforest trees is 39 pkat gfw−1 (Stewart et al.1988), 37 pkat gfw−1 in a Banksia woodland (Stewart et al.1993), 18 pkat gfw−1 on the coral cay and 24 pkat gfw−1 in the coastal heathland). These results are consistent with the con-clusion that the majority of species in these ecosystems utilize sources of nitrogen other than nitrate ions.
Using data for a range of plant groups in tropical, sub-tropi-cal and temperate ecosystems, it is possible to identify patterns of nitrate utilization that appear to be based on the presence or absence of mycorrhizal associations. Putatively non-mycorrhi-zal families (e.g., the Urticaceae, Chenopodiaceae, Amaran-thaceae and Polygonaceae) had average NRA values in the
range of 1200--2400 pkat gfw−1 (Figure 2), indicating that nitrate ions are the primary source of nitrogen. In contrast, in a range of largely vesicular-arbuscular mycorrhizal (VAM) and vesicular-arbuscular and ecto-mycorrhizal (VAM-ECM) her-baceous and woody species, the potential for nitrate reduction was much lower, with average values in the range of 50--350 pkat gfw−1. Of the species sampled from families with very specialized mycorrhizal associations (the Ericaceae and Epac-ridaceae), all displayed low capacities for nitrate reduction.
Contribution of nitrogen fixation
Putative N2-fixing species often displayed the highest nitrogen concentrations, indicative of the importance of nitrogen fixa-tion to the nitrogen status of species in nutrient-poor ecosys-tems. For example, in the tropical savanna woodland (Site 4), nitrogen-fixing species had an average nitrogen concentration
of 2.6% (± 0.43 SD) compared with a community average of 1.3% (± 0.61 SD). A similar trend was observed in the sub-tropical eucalypt open forest (Site 6), where nitrogen fixing species had average nitrogen concentrations of 2.92% (± 1.36 SD) compared with 1.52% (± 0.60 SD) for non-fixing species. At the most nitrogen-limited site (Site 1), the average nitrogen concentration of nitrogen-fixing species was signifi-cantly higher than that of non-fixing species (1.51 ± 0.35 versus 1.0 ± 0.19%). However, the relative importance of nitrogen fixation as a strategy and the extent to which N2-fixing species might contribute to ecosystem nitrogen turnover may be small. Thus, at the heathland site (Site 1), putative N2-fixing species represent 10% of the total species, but contribute only 1.5% to the total ecosystem biomass (Bolton 1986). In a similar community type (mixed Banksia woodland--heath) on Stradbroke Island in southeastern Queensland, 10% of the species are potential nitrogen fixers (Clifford and Specht 1979), as are 10% of the species in a Banksia woodland in Western Australia (Bennett 1988) and 16% of the species in a typical Eucalyptus open forest (Clifford and Specht 1979). Of the greater than 400 species represented in the semi-arid Aca-cia aneura F.J. Muell.dominated ‘‘mulga’’ woodland (repre-sented by Site 2), only 13% are potential nitrogen fixers (Nelder 1984).
Because of the potentially low rates of nitrogen fixation under natural conditions (e.g., Hansen et al.1987), the contri-bution of symbiotic N2-fixation to total ecosystem nitrogen may be relatively small. However, assessments of the
contribu-tion of nitrogen-fixers to ecosystem nitrogen status are compli-cated by the fact that published values for rates of N2-fixation in the field vary widely (0.10--1.6 kg N ha−1 year−1 for four common shrub legumes of jarrah forest (Hansen et al.1987); 2.2 kg N ha−1 year−1 for Acacia pulchella R. Br. (Monk et al. 1981); 18.6 kg N ha−1 year−1 for Macrozamia riedlei (Fisch. ex Gaud.--Beaup) C. Gardn., an understory component of mixed Banksia woodland in southwest Western Australia (Hal-liday and Pate 1976)). Pate et al.(1993) cite δ15N values for a range of species in a Banksia woodland that suggest that the woody species were either assimilating a common nitrogen source and that nitrogen fixation was of minor importance to the nitrogen-fixing species, or that shoot nitrogen was derived from mobilized reserves rather than current assimilation. Het-erogeneity in 15N discrimination of soil precludes the use of the 15N natural abundance technique for assessing legume N
2 fixa-tion in a jarrah forest (Hansen and Pate 1987).
Nitrogen source availability in soils
Analysis of water extracts of soil yielded similar nitrogen profiles for the E. grandis and E. maculata forest sites (Ta-ble 1). Ammonium and nitrate were present in similar concen-trations; however, the water extracts tended to underestimate the presence of ammonium. Organic forms of nitrogen contrib-uted significantly to total soluble soil nitrogen, and the soluble soil components accounted for approximately 0.1% of the total soil nitrogen in both the E. grandis and E. maculata forest soils. In the wet season, soluble protein concentrations in soils of the E. grandis and E. maculata forests were 32.9 and 37.0 nmol N gdw−1 soil, respectively. The concentration of soluble proteins was lower during the drier months (6.9 and 8.1 nmol N gdw−1 soil in July in the E. grandis and E. maculata forests, respectively) than the wet months of the year. The highest concentrations of amino acids were found in the E. grandis forest soil (4.05 nmol N gdw−1) and this pool decreased dra-matically in July. The major amino acids in the total pool were serine, alanine, glycine, aspartate and leucine (data not shown).
In the subtropical wet heathland site, ammonium, amino acids and nitrate were present in the soil solution (Table 2). The absolute quantities of each component varied because of envi-ronmental conditions, but ammonium dominated at all times, and amino acids were abundant during waterlogging or imme-diately after fire as a result of increased rates of mobilization of organic sources. Total available nitrogen in the soil solution also varied considerably, and increased 100-fold in the week following fire. Nitrate availability increased immediately after fire, whereas the amino acid concentration in the soil solution was significantly reduced, presumably because of increased ammonification and nitrification.
Recent studies have stressed the importance of organic forms of nitrogen, such as amino acids, for plant nutrition in temperate and arctic regions (Abuarghub and Read 1988, Kiel-land 1994). Plants can absorb amino acids from the soil solu-tion by an active transport mechanism (Jones and Darrah 1994), and species such as the non-mycorrhizal arctic sedge Eriophorum vaginatum L. display a preference for amino acids Figure 2. Average nitrate reductase activities (± SEM) for species from
over inorganic forms of nitrogen (Chapin et al. 1993). The relative importance of mineral and organic forms of nitrogen for plant nutrition likely depends on their relative availability in the organic and mineral horizons of the soil. The soil nitrogen profiles for both the open Eucalyptus forests and the coastal heathland indicate that multiple sources of nitrogen may be available for plants in natural ecosystems and that organic forms may contribute significantly to total soil nitro-gen. Because the pool of amino acid nitrogen is highly labile (Schmidt et al. 1960), its presence in soil solution extracts is highly variable (Read et al. 1989). The availability of proteins in the soil solution is also likely to be subject to significant seasonal release and immobilization (Abuzinadah et al. 1986).
Incorporation of nitrate versus ammonium
Although it is known that the availability of nitrate and ammo-nium varies with the soil type, it is not known how much plant species differ in their ability to assimilate these sources. We found a preference for uptake and assimilation of ammonium ions over nitrate ions in a range of species (Table 3). The rate of incorporation of ammonium ions was nearly 3 times that of nitrate in Eucalyptus grandis, 5--6 times that of nitrate in the mangroves Avicennia marina (Forssk.) Vierh and Rhizophora mangle L., 10 times that of nitrate in Hakea sp. and 25 times that of nitrate in Leptospermum sp. In the coral cay forest species, the preference for uptake of ammonium over nitrate was 15 times in Pandanus sp. aff. heronensis H. St. John and 25 times in Argusia argentea (L.f.) Heine.
The nitrogen concentration and the generally low nitrate reductase activity in the communities studied suggest that many of the species utilize nitrogen sources other than nitrate
(Tables 1--3). This is consistent with previous findings for a range of soil types and successional stages (Smirnoff and Stewart 1985, Stewart et al.1993).
Utilization of organic sources of nitrogen
Although soluble organic nitrogen represents a significant pool in soils of natural ecosystems, few studies have been undertaken to determine the extent to which these pools can be accessed by plants. Non-mycorrhizal seedlings of both E. grandis and E. maculata grew well on nitrate and ammo-nium, but showed little capacity to utilize organic nitrogen (Table 4). Neither species showed growth beyond that sup-ported by seed reserves on asparagine, glycine, histidine or BSA. In both species, mycorrhizal infection conferred on seedlings the ability to grow on organic sources of nitrogen (Table 4). Both E. grandis and E. maculata seedlings infected with Elaphomyces sp. displayed significantly greater growth than non-mycorrhizal seedlings on asparagine, glycine, histid-ine and BSA (P < 0.01). The potential to utilize a broad spectrum of organic sources has important implications for the nutrition of Eucalyptus. In field conditions, a complex mixture of nitrogen sources, all at potentially low concentrations, may be available. Thus, the ability to access a diversity of nitrogen sources may confer distinct nutritional advantages on mycor-rhizal plants in forest ecosystems. When mycormycor-rhizal seedlings of Eucalyptus grandis were offered a mixed source of nitrogen in the growth medium, they displayed a strong ability to utilize organic nitrogen. Seedlings derived 48 and 55% of their nitro-gen from asparagine when it was offered with nitrate and ammonium, respectively (Table 5). The BSA protein source provided 42 and 33% of plant nitrogen when it competed with Table 1. Mean nitrogen concentrations for inorganic and organic components of soil solution (nmol N gdw−1 soil) from forest sites dominated by Eucalyptus grandis and E. maculata for the months of December (wet season) and July (dry season). Each value represents the mean (± SD) for replicate soil samples at each site (n = 3 for nitrate, ammonium and amino acid determinations; n = 6 for protein determinations); nd = not detected.
Nitrogen source E. grandis forest site E. maculata forest site
December July December July
Ammonium 26.2 (24.1) 21.9 (5.7) 37.1 (23.7) 26.8 (4.3)
Nitrate 2.8 (1.7) 46.3 (30.6) 54.4 (23.0) 18.9 (11.4)
Amino acids (total) 4.1 (1.25) 0.01 1.1 (0.50) nd
Soluble protein 32.9 (7.2) 6.9 (1.9) 37.0 (3.55) 8.1 (2.5)
Table 2. Mean nitrogen concentrations for inorganic and organic components of soil solution (nmol N g−1 ion exchange resin) from a wet coastal heathland in Beerwah State Forest, southeastern Queensland. Each value represents the mean for replicate samples at points during a burning cycle and during waterlogging. The proportion of each component as a percentage of the total is indicated in parentheses.
NH4+ NO3− Amino acids Total
Before burn 2.28 (46) 0.2 (4) 2.5 (50) 4.98
After burn -- 1 week 578 (94) 28 (4.5) 12 (1.5) 618
After burn -- 3 weeks 157 (91) 8 (5) 6.7 (4) 172
nitrate and ammonium, respectively. Thus, organic sources of nitrogen may provide a significant nitrogen source even in the presence of readily assimilable inorganic ions, especially in conditions where plants must scavenge for available resources (Table 5).
Although it has been established that mycorrhizal plant species in heathland ecosystems can utilize complex organic nitrogen through ericoid or ecto-mycorrhizae (Bajwa et al. 1985) and have access to NH4+ through VAM (Marschner and Dell 1994), the question arises whether predominantly non-mycorrhizal and non N2-fixing plants, such as members of the Proteaceae, Cyperaceae and Restionaceae, also have special means of nitrogen nutrition. In Hakea sp. seedlings offered single nitrogen sources, a preference in the order of ammo-nium, glycine, nitrate was displayed in both roots and proteoid roots (Figure 3a). Rates of incorporation of labeled glycine were in the order of 40--60% those of ammonium and 4--5 times those of nitrate. A similar trend in preference was dis-played in both root types when all three nitrogen sources were offered simultaneously (Figure 3b). Uptake of ammonium was reduced by 35% in proteoid roots and by 20% in roots under
the competitive influence of the other nitrogen sources in the triple feeding experiment. The uptake of nitrate and glycine was not significantly influenced by the presence of other nitro-gen sources in the medium. Although incorporation of 15 N-la-beled substrates was consistently higher in roots than in proteoid roots on a fresh weight basis, direct comparisons of rates of uptake between the two root types are difficult because of the retention of small amounts of sand in cluster roots.
Because proteoid roots are mainly found in the upper or-ganic rich soil layer, Dinkelaker et al. (1995) have suggested that proteoid roots have a special function in the utilization of organic nitrogen, such as preferential uptake of amino acids. We have demonstrated that root systems of the genus Hakea can take up and assimilate exogenous sources of amino acids from the soil solution, but we found no qualitative difference between roots and proteoid roots with respect to uptake of labeled glycine (Figure 3). We conclude, therefore, that the Table 3. Rates of incorporation of nitrate and ammonium (nmol 15N
gdw−1 h−1) in a range of plant species from diverse plant communities. Each value represents the mean of replicate determinations.
Species Nitrate Ammonium
Eucalypt openforest
Eucalyptus grandis 246 714
Coastal heathland
Hakea sp. 180 1715
Leptospermum sp. 29 724
Mangrove forest1
Avicennia marina 360 1818
Rhizophora mangle 66 408
Coral cay open forest
Argusia argentea 29 724
Pandanus sp. aff. heronensis 98 1450 1 Data taken from Stewart et al. 1991.
Table 4. Dry weight (mg) of seedlings of Eucalyptus grandis and Eucalyptus maculata on a range of inorganic and organic sole nitrogen sources in buffered agar culture (pH 5.5). Seedlings were grown in non-mycorrhizal form or in association with Elaphomyces sp. (isolate NQ732). Each value represents the mean (± SEM) for replicate growth assays for seedlings (n = 12).
Nitrogen source Eucalyptus grandis Eucalyptus maculata
Non-mycorrhizal Mycorrhizal Non-mycorrhizal Mycorrhizal
N free 3 (0.3) 25 (0.5)
Nitrate 111 (12.5) 65 (2.0) 144 (35) 156 (5.0)
Ammonium 85 (8.0) 65 (5.0) 125 (13) 70 (2.5)
Asparagine 13 (0.5) 29 (0.5) 26 (4.5) 82 (2.0)
Glycine 4 (0.5) 18 (1.0) 19 (0.5) 46 (4.0)
Histidine 3 (0.5) 35 (0.5) 16 (1.5) 64 (2.0)
Protein (BSA) 3 (0.5) 45 (0.5) 14 (3.0) 162 (10)
1 Adapted from Turnbull et al.(1995).
Table 5. Atom% excess and calculated percentage utilization of inor-ganic and orinor-ganic sources of nitrogen in mycorrhizal seedlings of Eucalyptus grandis grown in sterile medium at pH 5.5. Plants were grown in medium to which was added either a single labeled source of 15N-labeled ammonium or nitrate (10 atom% excess) or a mixture of these labeled inorganic sources with either asparagine or BSA in the combinations shown. Values are the means of duplicate bulked sam-ples (n = 10--12) (± range) except for 15NO3− / Asparagine and 15NO3−/ BSA which are single bulked samples (n = 10). The percentage of plant nitrogen derived from the unlabeled source was calculated ac-cording to the formula given in Materials and methods.
Source Atom% excess Calculated %
leaf material utilization of unlabeled N source 15NO
3
− 9.89 (0.47)
15NO 3 − / NH
4
+ 2.28 (0.15) 77
15NO 3
− / Asparagine 5.10 48
15NO 3
− / BSA 5.79 42
15NH 4
+ 9.33 (0.11)
15NH 4
+ / Asparagine 4.20 (0.10) 55 15NH
4
ability to utilize organic nitrogen is common to both proteoid and normal roots in Hakea. The structure of proteoid roots and their placement in the organic soil horizons suggests that their functional role is intensive exploration of small soil volumes (cf. Dinkelaker et al. 1995).
The haustoria of root hemiparasites constitute another po-tentially important root specialization related to the uptake of organic nitrogen. Such specializations are found in a range of genera (e.g., Exocarpus, Anthobolus, Santalum and Striga) that are common in nutrient-poor ecosystems (Lamont 1984). Not only do these structures enable the hemiparasite to tap into host nutrient reserves, but studies with Olax phyllanthi L. have shown that these species have the ability to transform host organic solutes before they enter the xylem (Pate et al.1994), which enables this species to parasitize hosts with a wide variety of organic transport compounds. This, in addition to the ability to induce the production of nitrate reductase in hosts containing high concentrations of nitrate in xylem sap, has the potential to confer on species with such haustorial
connec-tions, the ability to tap a broad spectrum of nitrogen sources that have been acquired and concentrated by the host.
Conclusion
Our results confirm those obtained in previous studies (e.g., Stewart et al. 1993, Pate et al. 1993) suggesting complex patterns of utilization of ecosystem nitrogen sources. We con-clude that species that are predominantly nitrate assimilating are likely to be non-mycorrhizal or weakly VAM. In contrast, species that have specific associations with ectomycorrhizal or ericoid mycorrhizal fungi assimilate ammonium, amino acids or even more complex forms of nitrogen such as protein. Many of these species have a relatively low potential to assimilate nitrate. This is likely to have important consequences to spe-cies composition and community structure in natural ecosys-tems that are not highly nitrifying, and which may become exposed to anthropogenic inputs of nitrogen. In addition, we have demonstrated that obligately non-mycorrhizal species may have the ability to compete with mycorrhizal species for organic pools of nitrogen that are present in many natural ecosystems. Other root specializations (nitrogen-fixing sym-bioses, hemiparasitic haustoria) extend further the spectrum of nitrogen source utilization in natural ecosystems. There is, thus, the possibility of both competitive and complementary patterns of exploitation of different nitrogen sources within the soil profile in relation to plant life from physiology, rooting morphology and the presence of specialized feeding roots or mycorrhizal associations (Pate et al.1993).
Acknowledgments
This study was supported by the Australian Research Council (Grant Nos. A19230676 and A19332711) and the Australian Flora Founda-tion. Research activities in Kakadu National Park were supported both financially and logistically by the Environmental Research Institute of the Supervising Scientist, Jabiru. The assistance of Dr N. Ashwath during field work in the Northern Territory is gratefully acknow-ledged. We thank Dr Paul Reddell (CSIRO Division of Soils, Towns-ville) for the provision of fungal cultures.
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