Soil phosphorus depletion capacity of arbuscular mycorrhizae formed by maize hybrids

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1 Soil phosphorus depletion capacity of arbuscular mycorrhizae formed by maize hybrids A. Liu, C. Hamel 1, S. H. Begna 2, B. L. Ma 3, and D. L. Smith 2 1 Department of Natural Resource Sciences and 2 Department of Plant Science, Macdonald Campus, McGill University, Lakeshore Road, Ste-Anne-de-Bellevue, Quebec, Canada H9X 3V9 ( liu_aiguo@hotmail.com); 3 Central Experimental Farm, Agriculture Canada, Ottawa, Ontario, Canada K1A 0C6. Received 22 June 2002, accepted 20 March Liu, A., Hamel, C., Begna, S. H., Ma, B. L. and Smith, D. L Soil phosphorus depletion capacity of arbuscular mycorrhizae formed by maize hybrids. Can. J. Soil Sci. 83: The ability of arbuscular mycorrhizal (AM) fungi to help their host plant absorb soil P is well known, but little attention has been paid to the effect of AM fungi on soil P depletion capacity. A greenhouse experiment was conducted to assess, under different P levels, the effects of mycorrhizae on extractable soil P and P uptake by maize hybrids with contrasting phenotypes. The experiment had three factors, including two mycorrhizal treatments (mycorrhizal and non-mycorrhizal), three P fertilizer rates (0, 40, and 80 mg kg 1 ) and three maize hybrids [leafy normal stature (LNS), leafy reduced stature (LRS) and a conventional hybrid, Pioneer 3979 (P3979)]. Extractable soil P was determined after 3, 6 and 9 wk of maize growth. Plant biomass, P concentration and total P content were also determined after 9 wk of growth. Fertilization increased soil extractable P, plant biomass, P concentration in plants and total P uptake. In contrast to P3979, the LNS and LRS hybrids had higher biomass and total P content when mycorrhizal. Mycorrhizae had less influence on soil extractable P than on total P uptake by plants. The absence of P fertilization increased the importance of AM fungi for P uptake, which markedly reduced soil extractable P under AM plants during growth. This effect was strongest for LNS, the most mycorrhizae-dependent hybrid, intermediate for LRS, and not significant for the commercial hybrid P3979, which did not respond to AM inoculation. Key words: Arbuscular mycorrhizal fungi, extraradical hyphae, maize hybrid, plant biomass, P uptake, soil extractable P Liu, A., Hamel, C., Begna, S. H., Ma, B. L. et Smith, D. L Capacité d hybrides de maïs mycorhizés à reduire le niveau de phosphore du sol. Can. J. Soil Sci. 83: Il est bien connu que les champignons mycorhiziens à arbuscules (CMA) peuvent aider leur plante-hôte à absorber le P du sol. Par contre, peu d attention a été porté sur la capacité de ces champignons à réduire la quantité de P du sol. Un essai a été mené en serre sous différents niveaux de P du sol, pour déterminer l effet des mycorhizes formées par des hybrids de maïs de génotypes contrastants sur la capacité d absorption du P par ces hybrids et sur le niveau de P extractible du sol. L essai avait trois facteurs: deux niveaux de mycorhization (mycorhizé et non-mycorhizé), trois taux de fertilisation en P (0, 40 et 80 kg 1 ) et trois hybrids de maïs [leafy normal stature (LNS), leafy reduced stature (LRS) et un hybride conventionel, le Pioneer 3979 (3979)]. Le P extractible du sol a été mesuré après 3, 6 et 9 semaines de croissance du maïs. La biomasse végétale, la concentration en P et le contenu en P des plants furent déterminées après 9 semaines. La fertilisation a augmenté le P extractible du sol, la biomasse végétale, la concentration des plants en P et l absorption du P. Les hybrids LNS et LRS, contrairement au P3979, avaient une biomasse et un contenu en P plus important lorsqu ils étaient mycorhizés. Les mycorhizes eurent moins d influence sur le P extractible du sol que sur l absorption du P. L absence de fertilisation phosphatée a augmenté la contribution des CMA à l absorption du P, ce qui a réduit de façon marquée le niveau de P extractible du sol durant la croissance du maïs. Cet effet fut particulièrement important pour LNS, l hybride le plus dépendant des mycorhizes, moyen pour LRS et nonsignificatif pour P3979, l hybride commercial, qui n a pas répondu à l inoculation mycorhizienne. Mots clés: Champignons mycorhiziens à arbuscules, hyphes extraradiculaire, hybride de maïs, biomasse végétale, absorption du P, P extractible du sol Leafy and reduced-stature hybrids, including LRS and LNS, were recently developed at the Central Experimental Farm, Agriculture and Agri-Food Canada, Ottawa, ON. The yields, shoot architectures and other traits of these leafy hybrids were reported by Modarres et al. (1997) and Begna et al. (1999). However, the nutrient uptake ability of these new hybrids is not known. The effects of P fertilization on soil available P and other forms of P have been studied in agricultural fields under both short- and long-term cropping systems (McKenzie et al. 1992a, b; Richards et al. 1995). The amount of P required by plants does not depend only on 337 plant species, but also on plant genotype. The P requirement of a given genotype depends on its biomass and tissue P concentration (Koide 1991). The ability of plants to extract P is largely driven by their root characteristics, including the ability to form mycorrhizae (Plenchette et al. 1983; Daft 1991; Khalil et al. 1994). There can be considerable genotype-dependent effects on AM root colonization and response to P application, and efficiency of P extraction (Baon et al. 1993). A previous experiment reported that, of a set of three contrasting hybrids, LNS genotype was the hybrid most responsive to AM fungi (Liu et al. 2000a).

2 338 CANADIAN JOURNAL OF SOIL SCIENCE Usually, mycorrhizal plants not only have a greater biomass as compared to non-mycorrhizal controls, but also a higher concentration of P (Barea 1991). While the impact of AM fungi on plant P uptake has been intensively studied, very little effort has been devoted to understanding the effect of these fungi on soil available P and, to date, it is not known whether the better P uptake by mycorrhizal plants can also decrease soil P level. After decades of fertilization, P has accumulated in agricultural soils to become a major threat to surface water quality (Sharpley et al. 1994; Sharpley 1995; Sims 1997). High P load in runoff results in eutrophication of rivers and lakes (Schindler 1977; Sharpley et al. 1994; Carpenter et al. 1998). Regulations are being implemented to prohibit P fertilization in soils with excessive P levels. This means effective approaches need to be found to achieve high agricultural production with low fertilizer input. Mycorrhizal fungi may help to maintain yields for non-p fertilized crops and reduce excessive soil P levels (Bethlenfalvay 1992; Schreiner and Bethlenfalvay 1995). Plant genotypes that develop particularly extensive and effective symbioses would seem particularly appropriate in such sustainable agricultural systems. Although several extractants may be used to assess plant available P in soil, none of them are universally effective (Troeh and Thompson 1993). The Mehlich-3 (Mehlich 1984) solution is the preferred extractant for available P determination in the neutral to acid soils prevailing in the province of Quebec, Canada. Because this study was conducted in the province of Quebec, Mehlich-3 extractable P was used as an indicator of soil available P in this study. The objective of this study was to define the effects of mycorrhizae on extractable soil P and plant P uptake, using maize hybrids with contrasting genotypes. MATERIALS AND METHODS Experimental Design and Plant Growth Conditions A factorial experiment was laid out in a completely randomized design with four replicates. The three factors included three rates of P fertilizer (0, 40, and 80 mg P kg 1 soil), three maize hybrids (P3979, LRS and LNS) and two mycorrhizal treatments (mycorrhizal and non-mycorrhizal). Maize was grown in 10-L pots (25 cm diameter 22 cm depth) filled with 10 kg of a pasteurized growth medium (soil) consisting of two parts of sand and one part of a light field soil (coarse -silty, mixed, nonacid, frigid Humaquept). Prior to seeding, it was determined that the soil had a ph of 6.5, and KCl extractable NH + 4 and NO 3 and Mehlich-3 (Mehlich 1984) extractable P in the soil were 3.22, 9.30 and 42 mg kg 1, respectively. Potassium dihydrogen phosphate (KH 2 PO 4 ) was applied to establish target P levels and ammonium nitrate (NH 4 NO 3 ) was applied to all pots at the same level (20 mg N kg 1 soil) to prevent N deficiency. Designated pots were inoculated with 30 g of commercial inoculant containing Glomus intraradices Schenck & Smith, DAOM , (Mycorizes, Premier Tech. Inc, Rivière-du-Loup, QC). Control pots received the same amount of sterilized inoculum (30 min at 121 C). Maize seeds were surface sterilized with 30% H 2 O 2 for 10 min and washed several times with distilled water. They were then germinated in Petri dishes on moist filter papers for 3 d, and germinated seeds were sown in the pots. Plants were grown under greenhouse conditions with a 16/8 h day/night regime, 300 µmol m 2 s 1 photon flux density, 75% relative humidity and at 26/22 C. Pots were watered to field capacity three times per week. Sampling and Data Collection In order to monitor changes in soil extractable P, soils were sampled two times during plant growth and again at harvest (at 3, 6 and 9 wk). Three soil cores (0 15 cm) per pot were taken 5 cm from the plant (forming a triangle) and pooled to make a composite sample. Five grams of each sample were used for P extraction (Mehlich 1984) and colorimetric determination (LACHAT AELABCHEM autoanalyzer, Milwaukee), and the rest of the soil was returned to the holes of each respective pot, to reduce the effect of soil sampling on plant growth. Whole plants were harvested after 9 wk of growth. All roots were collected and washed with tap water. A 3-g root sample was taken randomly from each pot. The root samples were cleared with 10% KOH (121 C for 30 min) and stained with 0.02% acid fuchsin. The percentage of root length colonized by AM fungi was estimated under a dissecting microscope using the grid line intersect method (Giovannetti and Mosse 1980) by counting out of 100 root sections intersecting the grid lines, the number of these rootline intersections that bear mycorrhizal colonization. Total hyphal length was measured using modifications of the procedures of Miller and Jastrow (1992). After harvest, all the soil in each pot was taken out, mixed and randomly sampled. Moist 20-g soil samples were placed in 50-mL centrifugation tubes with 40 ml of 2 M sucrose solution containing 2% sodium hexametaphosphate. Tubes were shaken for 30 min on an end-to-end shaker and centrifuged for 15 min at 450 g. The upper part of the supernatant was collected with a pipette after each centrifugation. Soil samples were thoroughly extracted by repeating the extraction procedure three consecutive times. Hyphae were recovered on a filter paper and stained with a 0.02% acid fuchsin in lactoglycercol solution for 12 h. A line intersect method (Tennant 1975) was used to determine the length of hyphae recovered on the filter. Hyphal length was expressed on a dry soil basis. Plant dry mass (roots and shoots together) was determined after drying at 70 C for 48 h. Dried plants were ground and digested in concentrated H 2 SO 4 and 30% H 2 O 2 (Thomas et al. 1967). Concentrations of P from both the digested plants and soil samples extracted by Mehlich-3 were analyzed colorimetrically using a flow injection autoanalyzer (LACHAT AELABCHEM, Milwaukee). Statistical Analyses Data were subjected to analysis of variance (ANOVA), using the ANOVA procedure of the SAS program (SAS Institute, Inc. 1990). Statistical significance was determined at the 5%

3 LIU ET AL. SOIL P DEPLETION CAPACITY OF ARBUSCULAR MYCORRHIZAE 339 Table 1. Summary of F significance from analysis of variance Soil extractable P Degree of Root Extraradical Plant Plant P Plant total Sampling time Sources freedom colonization hyphal length biomass concentration P content 3 wk 6 wk 9 wk Hybrid (H) 2 * ** ** * ** NS * ** Phosphorus (P) 2 ** ** ** ** ** ** ** ** Mycorrhizal (M) 1 ** ** ** * ** NS * * H P 4 NS z NS NS NS NS NS NS NS H M 2 NS NS * * ** NS * * P M 2 NS NS NS NS NS NS * * H P M 4 NS NS NS NS NS NS * * *, ** P < 0.05 and P < 0.01, respectively; NS, not significant. probability level. Means of main effects were compared using the least significant difference test following a significant ANOVA test. When there was an interaction between two factors, the means of the combinations of each level of the factors were compared using the LSD test. When there was a three-factor interaction, the data were sorted by the factor that was most significant (with the highest F value), and then the means of the combination of each level of the other two factors were compared. RESULTS Root Colonization and Extraradical Hyphal Development The significance of the F values from the ANOVA analysis is summarized in Table 1. In non-mycorrhizal pots, there was no mycorrhizal colonization of plants roots (Table 2). The amount of extraradical hyphae found in non-mycorrhizal soils was only 8% of the amount found in mycorrhizal soils (Table 2). Phosphorus fertilization reduced percentage root colonization and extraradical hyphal length. The extraradical hyphal length was greater at low P than at high P fertilization when averaged over hybrids (Table 2). Averaged over P levels, LNS had higher percentage root colonization than the other two hybrids and more extraradical hyphae than P3979 (Table 2). Plant Biomass, P Concentration and Total P Content There was an interaction between mycorrhizae and maize hybrid for plant biomass and P concentration (Table 1). Mycorrhizal inoculation increased the biomass of LNS and LRS, but did not affect the biomass of P3979 (Table 3). LNS plants inoculated with AM fungi had a higher average P concentration than non-mycorrhizal LNS plants (Table 3). Mycorrhizal inoculation did not influence total P content of hybrid P3979, but the total P content of LNS and LRS hybrids was greater when these hybrids were mycorrhizal (Table 3). LNS and P3979 corn hybrids had higher biomass and total P content than the LRS hybrid (Table 3). Averaged over hybrids, P fertilization increased plant total P content (Table 4). Plant biomass increased with increasing application of P fertilizer (Table 4). Plants grown under high P fertilization had a higher P concentration than those grown in the soil without P fertilization when averaged over the three hybrids (Table 4). Table 2. Root colonization and extraradical hyphal length as influenced by mycorrhizal status, P application rate and hybrid Root Extraradical Factor Level colonization (%) hyphal length (m g 1 ) Mycorrhizal Myc+ z 38.8 ± 1.23a 1.30 ± 0.041a status Myc 0.00 ± 0.00b 0.10 ± 0.003b P P ± 0.75a 0.86 ± 0.029a P ± 0.62 b 0.73 ± 0.020b P ± 0.65b 0.44 ± 0.015c Hybrid P ± 0.60b 0.59 ± 0.026b LNS 22.7 ± 0.82a 0.79 ± 0.061a LRS 18.5 ± 0.58b 0.65 ± 0.066ab z Myc+ and Myc represent mycorhizal and non-mycorrhizal treatments; P 0, P 40 and P 80 represent P application levels (0 mg kg 1, 40 mg kg 1, and 80 mg kg 1 soil); P3979, LNS and LRS represent Pioneer 3979, leafy normal stature and leafy reduced stature hybrids, respectively. a,b Means ± SE followed by the same letter within the same factor and same measurement are not significantly different at P < 0.05 according to the LSD test. Soil Extractable P Mycorrhizal treatments did not influence soil extractable P after 3 wk (Table 5). However, after 6 wk, there was less soil extractable P under LNS grown without P fertilizer in mycorrhizal pots than in non-mycorrhizal pots. There was no difference in soil extractable P between the mycorrhizal and non-mycorrhizal treatments for P3979 at any sampling time (Table 5). After 9 wk, soil extractable P was lower under mycorrhizal than under non-mycorrhizal LNS plants under low P (40 mg P kg 1 soil) or no P (0 mg P kg 1 soil) fertilization. When no P fertilizer was added, soil extractable P was lower under mycorrhizal than under nonmycorrhizal LRS. DISCUSSION The enhanced growth, higher P concentration and greater total P uptake (15 mg) for LNS hybrids, when mycorrhizal, can be attributed to the more extensive internal and external mycelial systems that develop on this hybrid. The magnitude of a plant response to mycorrhizal formation depends on the extent of mycorrhizal development (Smith and Read 1997). A more extensive external mycelial network may be better able to exploit the soil P resource (Jakobsen et al. 1992; Liu et al. 2000b). In addition, the P requirement of the host plant and the level of soil available P will also influence

4 340 CANADIAN JOURNAL OF SOIL SCIENCE Table 3. Plant biomass, P concentration and total plant P content as influenced by mycorrhizal status and hybrid P3979 z LNS LRS Measurements Myc+ Myc Myc+ Myc Myc+ Myc Plant biomass (g) 27.5 ± 1.12a 26.4 ± 1.01ab 27.8 ± 1.10a 24.9 ± 0.80b 22.6 ± 0.80c 19.4 ± 0.7d P concentration (g kg 1 ) 3.05 ± 0.12bc 2.92 ± 0.08c 3.36 ± 0.14a 3.16 ± 0.10b 3.07 ± 0.10bc 2.99 ± 0.06c Total P content (mg) 83.7 ± 3.34b 77.2 ± 3.95bc 93.4 ± 3.72a 78.4 ± 4.47bc 69.5 ± 4.48c 58.0 ± 2.4d z P3979, LNS and LRS represent Pioneer 3979, Leafy normal stature and Leafy reduced stature hybrids, Myc+ and Myc represent mycorhizal and non-mycorrhizal treatments, respectively. a d Means ± SE followed by the same letter within the same row are not significantly different at P < 0.05 according to the LSD test. the extent of plant response to mycorrhizae (Abbott and Robson 1984). The larger the plant P requirement and the smaller the available P pool, the larger the benefit of mycorrhizal formation to plant growth. The LNS hybrids seem to rely highly on the AM symbiosis. It has a higher shoot:root ratio than the other hybrids (Liu et al. 2000a). This may create a condition of high P demand by the shoot and low P supply by the root when there is no symbiotic mycelial network to extend the root system. It is likely that the response to AM inoculation was a result of increased uptake from sources of available phosphate through mycorrhizae. It is true that early experiments with rock phosphate have yielded conflicting results (Mosse et al. 1976; Barea et al. 1980; Islam et al. 1980; Powell et al. 1980) leading to the view that AM fungi could provide access to P sources otherwise unavailable. However, later experiments clearly demonstrated that mycorrhizal infection did not significantly improve the availability of P from insoluble rock phosphate or calcium phosphate sources (Morel and Plenchette 1994; Nurlaeny et al. 1996). Whether AM fungi influenced soil extractable P level depends on the following parameters: soil extractable P level, plant development and plant mycorrhizal dependency. First, at a low level of soil extractable P, in this experiment, mycorrhizae in LNS decreased soil extractable P compared with non-mycorrhizal control. At the highest level of P fertilization (80 mg P kg 1 soil), mycorrhizae did not reduce extractable P, while without P fertilization, P availability was reduced under mycorrhizal conditions, when both the LNS and LRS hybrids were grown. It appears therefore, that mycorrhizae can reduce soil extractable P more effectively where the total capacity of the extractable P pools is not very high. This may be explain by the fact that soil extractable P level is a determining factor controlling the development of mycorrhizae and thus plant P uptake capacity (Abbott and Robson 1984; Koide and Li 1990; Lambert and Weidensaul 1991). Root colonization, extraradical hyphal length and mycorrhizae-mediated plant P uptake are stimulated by low soil P levels (Koide and Li 1990; Liu et al. 2000b). Extraradical hyphae extend several centimeters from the root to absorb available P, in this way increasing the thickness of the P depletion zone surrounding roots in mycorrhizal soil (Li et al., 1991a, b). Second, the impact of mycorrhizae on soil extractable P developed with plant development. After 3-wk of plant growth, although AM LNS and LRS hybrids had absorbed more P from the soil than their non-mycorrhizal counterparts, soil extractable P was not significantly reduced. Table 4. Plant biomass, P concentration and total P content per plant as influenced by P application rate Applied P rate (mg kg 1 ) Measurements P z 0 P 40 P 80 Plant biomass (g) 18.7 ± 0.96c 25.8 ± 1.04b 29.8 ± 1.16a P concentration (g kg 1 ) 2.84 ± 0.156b 3.15 ± 0.168ab 3.39 ± 0.173a Total P content (mg) 52.9 ± 2.21c 81.8 ± 3.08b 96.1 ± 3.35a z P 0, P 40 and P 80 represent P application levels (0 mg kg 1, 40 mg kg 1, and 80 mg kg 1 soil). y Means ± SE followed by the same letter within the same row are not significantly different at P < 0.05 according to the LSD test. However, with increasing plant size and P content, the difference in soil P availability became measurable. Third, the impact of mycorrhizae on soil extractable P was greater for hybrids with greater mycorrhizal dependency. The most mycorrhizae-dependent hybrid, LNS, reduced soil extractable P after 6 and 9 wk of plant growth in mycorrhizal pots without P fertilization, compared to non-mycorrhizal pots, and after 9 wk of growth in mycorrhizal pots with low P fertilization (40 mg P kg 1 soil). In contrast, soil extractable P levels were similar in mycorrhizal pots and non-mycorrhizal pots containing the least mycorrhizaedependent hybrid, P3979. This indicates that the more responsive to AM the hybrid, the more likely the soil extractable P level reduction. Therefore, decreased soil extractable P under mycorrhizal plants was found when all the following conditions were present: (1) soil extractable P level is not high (allowing for better development of AM fungi), (2) highly mycorrhizae-dependent hybrids, and (3) large plant size (leading to large total P uptake). This suggests that soil extractable P can be significantly reduced by the AM fungal effects only when the mycorrhizae-mediated P uptake is large enough to exceed inputs of extractable P from other P pools in soil such as organic P and precipitated inorganic P. The extent of mycorrhizae-mediated P uptake by a hybrid depends on the reliance of this hybrid on AM fungi for P uptake. The impacts of these findings on agricultural production are likely to be large in areas where P fertilization in amounts exceeding crop exportation has led to P build-up in soils. In the province of Quebec, Canada, for example, the very high P level of most agricultural soils leads to excessive P loads in runoff, causing eutrofication of fresh water bodies. As a consequence, it has become urgent to find sustainable alternatives to reduce excessive soil P

5 LIU ET AL. SOIL P DEPLETION CAPACITY OF ARBUSCULAR MYCORRHIZAE 341 Table 5. Soil extractable P as influenced by mycorrhizal status, P application rate and maize hybrid Soil extractable P (mg kg 1 ) Sampling Applied P rate P3979 z LNS LRS time (mg kg 1 ) Myc+ Myc Myc+ Myc Myc+ Myc 3 wk P ± 1.72 a y 36.7 ± 2.13a 36.7 ± 1.85a 35.7 ± 1.58a 37.6 ± 1.92a 38.2 ± 2.06a P ± 3.17a 64.5 ± 2.84a 61.7 ± 3.15a 62.4 ± 2.76a 66.2 ± 3.52a 65.3 ± 3.23a P ± 3.43a 77.5 ± 3.75a 80.2 ± 3.18a 75.2 ± 3.82a 77.3 ± 4.21a 79.8 ± 2.91a 6 wk P ± 1.41b 30.2 ± 1.62b 27.2 ± 1.32c 30.5 ± 1.47b 34.8 ± 1.36a 35.7 ± 1.54a P ± 2.38b 56.5 ± 1.37b 55.3 ± 1.66b 54.7 ± 2.67b 60.5 ± 4.85ab 62.5 ± 3.53 a P ± 2.27b 70.5 ± 1.95ab 69.3 ± 3.25ab 71.2 ± 2.38ab 73.5 ± 2.27ab 75.2 ± 2.85 a 9 wk P ± 0.91 c 23.7 ± 0.88c 19.5 ± 0.78d 22.8 ± 0.84c 29.7 ± 1.12b 33.2 ± 1.31a P ± 1.73b 44.7 ± 1.72b 38.9 ± 1.63c 43.7 ± 1.65b 51.8 ± 1.88a 53.6 ± 2.04a P ± 2.23b 51.2 ± 1.95b 51.2 ± 2.28b 52.1 ± 2.36b 58.6 ± 2.84a 60.7 ± 2.38a z P3979, LNS and LRS represent Pioneer 3979, Leafy normal stature and Leafy reduced stature hybrids; Myc+ and Myc represent mycorhizal and nonmycorrhizal treatments; P 0, P 40 and P 80 represent P application levels (0 mg kg 1, 40 mg kg 1, and 80 mg kg 1 soil), respectively. y Means ± SE followed by the same letter in the same row are not significantly different at P < 0.05 according to the LSD test. without decreasing crop yield. Our results show that the use of maize hybrids with superior ability to develop mycorhizae could help maintain crop yield while P fertilizer is reduced or even eliminated in high-p soils. The reduction of soil P levels, in turn, could benefit AM fungal development. Field experiments should be conducted to verify these conclusions. In conclusion, this study revealed a relationship between P fertilization, mycorrhizae development, P uptake, mycorrhizal dependency of hybrids and soil extractable P. The absence of P fertilization increased the importance of mycorrhizae for P uptake, which could lead to decreased soil extractable P during later stages of plant development. This effect was large in highly mycorrhizae-dependent hybrids such as LNS, and non-significant in the commercial hybrid P3979, which did not respond to AM inoculation. ACKNOWLEDGMENTS The authors thank Dr. X. M. Zhou and Mr. T. Spedding for their assistance. 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