Utilization of AIPO4 as a phosphorus source by ectomycorrhizal Pinus rigida Mill, seedlings

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1 7-2 New Phytol. (1990), 116, Utilization of AIPO4 as a phosphorus source by ectomycorrhizal Pinus rigida Mill, seedlings BY JONATHAN R. CUMMING* AND LEONARD H. WEINSTEIN The Boyce Thompson Institute for Plant Research and the Department of Natural Resources, Cornell University, Ithaca, NY 148, USA (Received 15 November 1989; accepted 2 April 1990) SUMMARY Mycorrhizal infection of roots of Pinus rigida Mill, seedlings by Pisolithus tinctorius Coker & Couch maintained normal foliar P relations when seedlings were exposed to Al in solution. To investigate how this symbiosis aids in P acquisition in the presence of Al, seedlings were grown in sand culture with solutions containing between 1 and /im NaHjPO^ (Pj) with or without insoluble AIPO4 in the substrate. Mycorrhizal seedlings had a higher affinity for P^ than non-mycorrhizal seedlings as judged by superior growth and foliar P contents at all P, levels tested. In addition, mycorrhizal seedlings, hut not nonmycorrhizal seedlings, effectively extracted P from AlPO^ through the dissolution of this insoluble salt. Measurement of matrix solution ph indicated that increased proton production, in association with high proliferation of mycorrhizal roots under P-limiting conditions, led to increased AlPO^ solubilization. Key words: Ectomycorrhizas, aluminium, phosphorus, Pinus rigida, Pisolithus tinctorius. INTRODUCTION Aluminium is phytotoxic to many species at levels commonly found in moderately to highly acidic soils. Although the mechanisms of Al toxicity are not yet understood, experimental evidence suggests that interactions between Al and P^ and enzyme transformations involving P assimilation play a role in toxicity response (Clarkson, 1966, 1967; Loughman, 1969; Foy, Chaney & White, 1978; Haug, 1984; Cumming, Eckert & Evans, 1986; Pfeffer et al., 1986; Pettersson, Hallbom & Bergman, 1988). The matrix of the root apoplast is negatively charged and will bind the highly electropositive AP^ ion. From this position, Al complexes P,, reducing its availability for plant uptake (Rorison, 1964; Clarkson, 1966, 1967; McCormick & Borden, 1974; Cumming et al., 1986). Plants exposed to Al often exhibit reduced foliar P concentrations (Foy et al., 1978; Cumming & Weinstein, 1990). This occurs even in cases where reduced shoot growth has offset reductions in P uptake, indicating that P availability is severely limited and that the specific uptake capacity of the roots has been altered. * Present address: Department of Botany, University of Alberta, Edmonton, Alberta T6G 2E9, Canada; to whom correspondence should be addressed. Cumming & Weinstein (1990) noted that the association of the ectomycorrhizal symbiont Pisolithus tinctorius with the roots of Pinus rigida seedlings grown in sand culture led to the maintenance of normal foliar P concentrations under Al exposure whereas uninoculated seedlings exhibited altered patterns of foliar P accumulation. Mycorrhizal infection may alter P relations in several ways. Changes in root morphology and extramatrical hyphal growth lead to increased root surface area, decreased Pj diffusive resistance in the rhizosphere, and enhanced plant P uptake (Bowen, 1973; Harley & Smith, 1983; Reid, 1984). In addition, two potential mechanisms can be envisaged whereby mycorrhizal infection mediates changes in inorganic Al-P interactions in the rhizosphere and root free space. First, acidification of the rhizosphere by the dissociation of excreted organic acids or through elevated H^ production associated with preferential NH^"^ use by ectomycorrhizas (France & Reid, 1983 ; Rygiewicz, Bledsoe & Zasoski, 1984 a, b) or altered H'^/NH^^ stoichiometry during uptake (Bledsoe & Rygiewicz, 1986) may increase the availability of P through the solubilization of inorganic metalphosphate salts (Snoeyink & Jenkins, 1980). Second, excretion of organic compounds by fungal symbionts may dissolve metal-phosphate precipitates through

2 100 J. R. Gumming and L. H. Weinstein the formation of soluble organo-aluminum complexes (Jurinak et al., 1986; Lapeyrie, 1988). This would both increase P availability and chelate and detoxify metals in the rhizosphere (Harley & Smith, 1983; Reid, 1984; Cumming & Weinstein, 1990). The aim of this research was to distinguish between possible miechanisms involved in ectomycorrhizal amelioration of Al-induced P deficiency in Pinus rigida seedlings. Experiments were undertaken to determine differences in the ability of mycorrhizal and nonmycorrhizal seedlings to obtain P from solutions containing a range of low concentrations of Pj and the role of the mycorrhizal association in obtaining P from insoluble AlPO^ in the rhizosphere. Seedling growth, foliar P concentrations, root surface acid phosphatase activity, and rhizosphere chemistry were assessed. MATERIALS AND METHODS Pinus rigida seeds (collected from single trees on Cape Cod, Barnstable County, Massachusetts, USA) were surface sterilized by soaking in 30 % HgOg for 60 min followed by rinsing in sterile distilled HgO. Seeds were aseptically transferred to sterile perlite and stratified for four weeks at 4 C. Seedlings were grown under controlled environmental conditions and were watered twice daily with a nutrient solution consisting of 1-0 mm KNOg, 05 mm Ca(NO3)2, 1-0 mm NH4NO3, 0-5 mm NaHgPO^, 0-25 mm 4, /tm KCI, IS iim H3BO4, liim ZnClg, 0-5/fM CUSO4, 0-5//M MOO3, 20 Fe,Na-EDTA. All solutions were adjusted to ph 4-0 ±0-1 with H2SO4 and autoclaved prior to use. Within the growth chamber, photoperiod was 14/10 h light/dark, with a photosynthetic photon flux density of 5 ± 10 /^mol m~^ s~^ from combined high pressure sodium and metal halide lamps. Air temperature was 24/19 ± 1 C light/dark and relative humidity was %. Seedlings from one open-pollinated (half-sib) family were transferred to sand culture in 163 cm* plastic columns (Ray Leach Conetainer Nursery, Canby, Oregon, USA). One-half of the columns contained sand to which solid AIPO^ powder (Aldrich Chemical Company, Milwaukee, Wisonsin, USA) was added to a level of 10/^gg"^ sand, and one-half of the seedlings were inoculated with P. tinctorius as previously reported (Cumming & Weinstein, 1990) in a crossed statistical design with AIPO4 treatments. Nutrient solutions were delivered to seedlings via individual tubes from a manifoldpump-reservoir system which automatically delivered between 20 and 24 ml of solution in the morning and between 10 and 12 ml of solution in the afternoon. A subset of columns acted as flow-through lysimeters, and solution samples were collected and assessed for volume, ph, and monomeric Al concentration by the eriochrome cyanine R method (Anon, 1985). The concentration of Al in solution collected from columns containing AIPO4 was assumed to be equivalent to the concentration of P in solution due to the dissolution of AlPO^. Solution [Al] in a set of columns containing AIPO4 without seedlings was found to range between 1-2 and 13-1yWM which represented passive dissolution of AIPO4 in the sand columns. A baseline value of lo/fm was adopted for subsequent analyses of seedling responses. The volume and calculated [H^ of solutions added and subsequently collected from lysimeters were used to calculate net proton budgets. Dissolved organic carbon was not measured and, as dissolved organic carbon buffers solution ph, proton budgets are underestimated, especially for mycorrhizal seedlings. Three weeks following transfer to sand columns, seedlings were watered with 1-0, 5-0, 10-0 or -0 [lu NaHgPO^ (designated Pj) solutions for an additional 28 d. Na2SO4 was added to maintain constant Na concentrations across all solution Pj treatments. Pj treatments were applied in a crossed design with AIPO4 and mycorrhizal treatments. Seedling height was recorded at weekly intervals during the experiment. Seedling root surface acid phosphatase activity, and root and shoot growth were assessed at the end of the experiment. Dry needles from three seedlings within each treatment combination were pooled equally by weight, ground with a mortar and pestle, and analysed for P and Al concentration by inductively coupled plasma emission spectrophotometry (ICP). Seedling root systems from inoculation treatments were highly infected with P. tinctorius, although quantification was not undertaken in this study. In a previous study under similar conditions, > 90 % of short roots were mycorrhizal and were characterized by well-developed mantles and extramatrical hyphae (Cumming and Weinstein, 1990). Acid phosphatase (NPPase) activity was determined on entire seedling root systems as follows. Excised root systems were gently separated from sand and transferred to 20 ml of standard nutrient solution without Pj at ph 4-0 to which disodium p- nitrophenyl phosphate (NPP) had been added to a concentration of 1 mm. Extramatrical hyphae remained attached to roots. systems in assay solution were placed in a bell jar and evacuated for 5 min to infiltrate the free space of the root with substrate. Aliquots of assay solution were taken at 15 and 75 min and assessed for ^-nitrophenol (NP) colorimetrically by reaction with 0-5 M NaOH (Tabatabai & Bremner, 1969; Ho, 1987). The amount of NP produced during the intervening 60 min incubation period was corrected for background colour by reacting similar aliquots with distilled HgO. This was an important step, since mycorrhizal roots released pigments during the assay which interfered with absorbance readings. No

3 Utilization ofalpo, by ectomycorrhizas NPPase activity was detected in 12 random samples of rhizosphere sand, indicating that acid phosphatase activity was due to root or mycorrhizal NPPase activity and not to microbial activity in the sand column. Treatments were arranged in a 4 x 2 x 2 factorial design. Treatment factors were solution P concentration (4 levels), mycorrhizal inoculation (2 classes), and solid AIPO4 addition (2 classes). Linear regression analyses were made across solution P^ concentration and regression parameters obtained for responses to solution P^ were assessed for treatment differences by two-way analyses of variance due to AIPO4 and mycorrhizal inoculation. Least square means and estimates for the intercept and linear responses to solution P^ concentration are presented for each variable assessed. For seedling heights, regression coefficients obtained by orthogonal polynomial regression analysis over time (Snedecor & Cochran, 1980) were assessed for treatment differences by two-way analysis of variance in which responses to solution Pj were analysed as dose responses also using orthogonal polynomial coefficients. Multivariate analysis of variance was undertaken to investigate partial correlations between variates accounting for variation association with treatments. All analyses were executed using SAS procedures (SAS Institute, 1985). RESULTS Overall height growth rate was a good predictor of final shoot weight (r = 0-848, «= 96, P<0-01), except in inoculated seedlings grown with AIPO4 (r = 0-210, n = 24), and was used to determine the time-course of seedling response to treatments. Differences in seedling height due to mycorrhizal treatment were noted as early as three weeks after transfer to sand columns, when mean mycorrhizal seedling height was 19% greater than comparable height of nonmycorrhizal seedlings. Mycorrhizal seedling height growth over time across all Pj treatments was 35 and 99% greater than nonmycorrhizal rates without and with AIPO4, respectively (Table 1). The rate of increase in seedling height over time among AIPO4 4 by y inoculation treatments in response to the Pj gradient were linear and exhibited positive slopes (Table 1). However, mycorrhizal seedlings grown with AIPO4 did not respond to increasing Pj, indicating that Pj was not limiting in this treatment. Mycorrhizal infection enhanced shoot and root weights across the full range of Pj applied (Table 2). Slopes and intercepts for shoot and root weight as functions of solution Pj between mycorrhizal treatments within AIPO4 treatments were significantly different, with mycorrhizal plants more effectively adding biomass for each additional increment of P, (Table 2). The significantly greater intercepts for 101 Table 1. Height growth rate* 0/Pinus rigida seedlings in response to [PJ as affected by AlPO^ and mycorrhizal inoculation AlPO, Inoculation + Parameter estimate f Meant (0-014) (0-014) (0-024) (0-024) Slope ( ) ( ) ( ) n.s." * Seedling height measured from the cotyledonary node to the tip of the longest primary needle. t Parameter estimates presented with standard errors in parentheses, w = 24 seedlings for each regression. X Mean = mm d^^ over all [PJ within each AlPO^ by inoculation treatment. Slope = mm d^^ fiw'^ P,. " Not significant. Table 2. and root weights of Pinus rigida seedlings in response to [PJ as affected by AlPO^ and mycorrhizal inoculation AlPO, - + Inoculation Parameter estimate* Meant (4-45) (4-40) (4-52) (4-46) (10-66) (6-15) 335- (10-66) (6-15) Slope 1-60 (0-24) 1-27 (0-25) 3-04 (0-24) 1-81 (0-26) 1-49 (0-72) 0-89 (0-40) 2-04 (0-72) n.s. * Parameter estimates presented with standard errors in parentheses, n = 24 seedlings for each regression. t Mean = mg dry weight over all [PJ within each AIPO4 by inoculation treatment. X Slope = mg d. wt /tm"^ P^. Not significant. both mycorrhizal and non-mycorrhizal seedlings grown with AIPO4 in contrast to those grown without AIPO4 suggests that both obtained P from this source (Table 2). However, the larger increase in growth of mycorrhizal seedlings in response to AIPO4 addition indicates that they were more effective in obtaining P from this source than the non-mycorrhizal seedlings (Table 2). Foliar P concentrations increased linearly with

4 102 y.r. Cumming and L. H. Weinstein Table 3. Foliar P concentration of Pinus rigida seedlings in response to [P^] as affected by AlPO^ and mycorrhizal inoculation ^* (mg P g-i d. wt) 4t (mg P g-i d. wt) [PJ (/IM) Non- inoculated Inoculated Non-inoculated Inoculated Slope (0-0013) (0-0013) t (0-0020) (0-0020) * Standard error of least square mean = t Standard error of least square mean = 0-04 and 0-07 for 10 and /im, respectively. t Equilibrium [PJ in solution with AlPO^ in the rhizosphere equalled approximately \0 fim. Slope = mg P g"^ d. wt ftm~^ P, (standard errors in parentheses). Table 4. Foliar Al concentration of Pinus rigida seedlings in response to [PJ as affected by AlPO^ and mycorrhizal inoculation -AlPO,* (/*g Al g-i d. wt) Al g-i d. wt) [PJ (/*M) Non-inoculated Inoculated Non-inoculated Inoculated Slope (0-161) t n.s n.s. * Standard error of least square mean = 6-4. t Standard error of least square mean = 11-5 and 19-5 for 10 and /im, respectively. X Equilibrium [PJ with AlPO^ equalled approximately 10 fim. Slope = /ig Al g"' d. wt. fim~^ Pj (standard errors in parentheses). " Not significant. 200 I I lings were able to utilize AIPO4 as a P source and, as a consequence, did not respond markedly to increasing soluble Pj (Table 3). AIPO4 treatments led to elevated tissue Al in both mycorrhizal treatments (Table 4). In both mycorrhizal and non-mycorrhizal seedlings, NPPase activity decreased with increasing [Pj] in solution (Fig. 1). However, root NPPase activity was depressed by mycorrhizal infection (Fig. 1). NPPase activity of non-mycorrhizal seedlings was unaffected by AIPO4 in the root zone, indicating that the presence of AIPO4 in the rhizosphere did not effectively reduce P deficiency in these seedlings (Fig. 1). Mean root NPPase activity of mycorrhizal Figure 1. surface acid phosphatase activity of nonmycorrhizal and mycorrhizal Pinus rigida seedlings as affected by nutrient solution P, concentration. O ^ D, +AIPO4; open symbols, non-inoculated; filled symbols, inoculated. solution [PJ in all treatments (Table 3). The smallest response was shown by mycorrhizal seedlings grown with AIPO4, again indicating that mycorrhizal seed- seedlings was lower and did not respond to [PJ when grown with AIPO4, further reflecting the acquisition of P from AIPO4 by roots infected with P. tinctorius (Fig. 1). Table 5 presents partial correlation coeflicients for growth and P relations separated by mycorrhizal treatment. Differences in correlation coefficients reflect different patterns of P acquisition and growth between the two classes of seedlings. There was a significant correlation between foliar P concentration and P uptake efficiency (mg P in foliage g~^ root weight) in mycorrhizal seedlings that was not evident

5 Utilization of AlPO^ by ectomycorrhizas 103 Table 5. Partial correlation coefficients and probabilities {in parentheses) for measures of growth and phosphorus nutrition of Pinus rigida seedlings separated by inoculation treatment Non-inoculated Inoculated weight weight Foliar P concn P uptake efficiency NPPase activity weight weight Foliar P concn P uptake efficiency NPPase activity weight (0-0004) (0-9963) (0-0080) (0-0187) ( (0-0001) (0-0003) (0-0554) (0-16) weight (0-6092) (0-9859) (0-0001) (0-0008) (0-0004) (0-0261) Foliar P concn (0-1011) (0-6901) (0-0005) (0-4897) P uptake efficiency (0-6724) (0-1849) NPPase activity P sources and the putative action of this enzyme system. Measurement of leachate ph indicated that mycorrhizal seedlings produced more H^ ions than nonmycorrhizal seedlings which was accompanied by elevated AIPO4 solubilization (Fig. 2). Proton production in both mycorrhizal and non-mycorrhizal seedlings increased with increasing solution Pj concentration (Table 6), more so in mycorrhizal seedlings, reflecting the degree of proliferation of these root systems under P limitation. Mean proton production and linear estimates for proton production in response to solution [PJ were significantly different between the two classes of seedlings (Table 6). Table 6. Solution acidification and ^ solubilization by Pinus rigida seedlings in response to mycorrhizal inoculation and solution [PJ* -20 Inoculation Time (days) Figure 2. Time-course for matrix solution proton (top) and Al (bottom) concentration for non-mycorrhizal and mycorrhizal Pinus rigida seedlings receiving /im P, solutions. O, -AlPO^; D, +AIPO4; open symbols, non-inoculated; filled symbols, inoculated. in their non-mycorrhizal counterparts. In nonmycorrhizal seedlings, shoot and root weights were not correlated with foliar P concentration or P uptake efficiency. In contrast, growth was negatively correlated with foliar P concentration and P uptake efficiency in mycorrhizal seedlings. For both groups of seedlings, growth was negatively correlated with NPPase activity whereas foliar P concentration and P uptake efficiency were uncorrelated with NPPase activity, as would be expected given the nature of the Slope " Slope -6-5t (0-43) (0-43) [Aline. (fimol 20-4J n.s.? (0-049) * Least square means for [PJ pooled over time (omitting day = 0 sampling time). t Standard error of least square mean = X Standard error of least square mean = 6-9. nmol H^ r* /im~^ P, (standard errors in parentheses). ' /imol AP^ 1"^ /im~^ P, (standard errors in parentheses). 5 Not significant.

6 104. R. Cumming and L. H. Weinstein DISCUSSION Seedlings of P. rigida inoculated with the ectomycorrhizal fungus P. tinctorius grew better than non-mycorrhizal seedlings under conditions of P limitation. Greater increases in height growth, shoot and root biomass, and foliar P concentrations in mycorrhizal seedlings, in response to increasing solution [PJ, indicate that mycorrhizal roots have a higher affinity for P, than nonmycorrhizal roots (Bowen, 1973; Cress, Throneberry & Lindsey, 1979). In addition, mycorrhizal seedlings effectively acquired P from insoluble AlPO^ when soluble P was both limiting and nonlimiting, suggesting that the mechanism involved is not induced by P limitation, but is the product of normal physiological function of mycorrhizal roots. Nonmycorrhizal P. rigida seedlings were unable to significantly alter the solubility of AlPO^ above baseline levels, hence could not exploit AIPO4 as an alternative P source. As solution Al leads to the precipitation of complex aluminium phosphates in the soil, rhizosphere, and root free space (Clarkson, 1966, 1967; McCormick & Borden, 1974; Cumming et al, 1986), soluble Pj present at the plasma tnembrane will be limited for plant uptake. Given a higher carrier affinity and greater instantaneous uptake rate for the phosphate ion (Cress et al., 1979; Dighton & Harrison, 1983), mycorrhizal roots will be able to maintain P uptake and normal physiological function where P deficiency will lead to reductions in seedling vigour in non-mycorrhizal seedlings. This may represent one mechanism whereby the symbiosis between P. rigida and P. tinctorius prevents Al toxicity development in mycorrhizal seedlings (Cumming & Weinstein, 1990). Mycorrhizal roots also apparently alter the reactions between Al and Pj, preventing their precipitation and subsequent P limitation. One mechanism whereby the fungus enhanced the solubilization of AIPO4, and possibly more complex amorphous aluminium phosphates, is through greater rhizosphere acidification. Inorganic phosphate salts exhibit ph dependent solubilities (Snoeyink & Jenkins, 1980). For example, AIPO4 solubility increases by approximately 170/^moir^ for each ph unit increase in acidity, and it is evident from the results presented here (Fig. 2) that elevated rhizosphere acidification by mycorrhizal seedlings enhanced AlPO^ dissolution. Non-mycorrhizal seedlings did not produce this effect, possibly reflecting loss of proton extrusion capacity of the root system with increasing P limitation (Fig. 2 and Table 6). An additional mechanism whereby mycorrhizal infection with P. tinctorius alters the solubilization of AIPO4 may be through the production of organic acids or other organic ligands which chelate Al in solution and may lead to changes in the chemical equilibria between AIPO4, Al^*, and H^PO^" in solution. The chelation of Al removes this species from equilibrium reactions between the solid and solution phases and leads to enhanced AIPO4 solubilization (Struthers & Sieling, 19; Agnihotri, 1970; Smith, 1976; Jurinak et al., 1986; Lipton, Blanchar & Blevins, 1987; Lape>rie, 1988). The production of Al-chelating substances by mycorrhizal roots may, in addition to increasing P availability, reduces Al toxicity by reducing the concentration of phytotoxic Al species in solution (Bartlett & Riego, 1972; Hue, Craddock & Adams, 1986). In a previous study employing similar methods we found that mycorrhizal infection greatly reduced the accumulation of Al in the foliage of P. rigida seedlings when seedlings were exposed to Al in solution, potentially by the above chelation mechanism (Cumming & Weinstein, 1990). This pattern was not evident in the present study, as both nonmycorrhizal and mycorrhizal seedlings exhibited similar foliar [Al]. The reason for this is not known. Seedlings inoculated with P. tinctorius exhibited a high degree of short root infection, although infection may not have been as complete as the previous study. However, the strong responses of seedlings inoculated with P. tinctorius in the present study to solution [PJ and AIPO4 in comparison to noninoculated seedlings suggests that the symbiosis was indeed vigorous. surface acid phosphatase (NPPase) activity increased under P limitation in both nonmycorrhizal and mycorrhizal seedlings in the present study (Fig. 1), suggesting that this enzyme system may be de-repressed by low soluble P conditions (Woolhouse, 1969; Bowen, 1973; Alexander & Hardy, 1981) or that stressed root cells exhibited elevated NPPase activity associated with senescence (Raudaskoski, 1976; Alexander & Hardy, 1981). The exception to this pattern was that NPPase activity of mycorrhizal seedlings grown with AIPO4 in the rhizosphere did not respond to changes in soluble Pj concentrations (Fig. 1). This pattern, along with patterns of foliar P nutrition and growth, further supports the concept that these seedlings obtained sufficient P from AIPO4. Antibus et al. (1981) and others have reported elevated acid phosphatase levels in mycorrhizal compared to nonmycorrhizal roots and have postulated that this enzyme system is responsible for obtaining P from organic forms (Bartlett & Lewis, 1973 ; Williamson & Alexander, 1975) and represents a major advantage to mycorrhizal plants in forest soils where a large proportion of soil P is bound in organic form (Sutton & Gunary, 1969; Dalai, 1977; Harrison, 1983). It is difficult to reconcile patterns of NPPase activity noted here with these reports. The effect of mycorrhizal infection on root acid phosphatase activity will depend on the inherent activity

7 Utilization of AlPO^ by ectomycorrhizas 105 of the fungal symbiont and potential interactions between the symbiont and the host resulting from the infection process. Ho (1987) reported in vitro acid phosphatase rates for eight isolates of P. tinctorius ranging between 1-0 and 9-2 mmol NPP g~^ d. wt. h-\ Six other species screened by Ho & Zak (1979) were characterized by rates ranging between 16-2 and /tmol NPP g-^ d. wt. h'^ It is evident that, of these species, P. tinctorius exhibits a superior acid phosphatase activity, although it is not known whether these authors accounted for optical interference resulting from pigments released from P. tinctorius cultures (see Materials and Methods). How these rates relate to mycorrhizas formed by these ectomycorrhizal symbionts cannot be readily ascertained, given the limitations of in vitro/in vivo comparisons. The overall depression of NPPase activity by mycorrhizal infection with P. tinctorius noted in the present study may reflect greater Pj absorbing power of mycorrhizal roots (Bowen, 1973; Cress et al., 1979; Dighton & Harrison, 1983; Harley & Smith, 1983), hence their roots are less P stressed. In addition, since rates of root cell senescence and root turnover are lower in mycorrhizal in comparison to non-mycorrhizal roots, acid phosphatase activity associated with cell senescence would also be lower (Raudaskoski, 1976; Alexander & Hardy, 1981). Relationships among growth and P nutrition variates (Table 5) of non-mycorrhizal and mycorrhizal seedlings suggest that P acquisition and growth processes were related differently in the two classes of seedlings. Partial correlation coefficients reflect relationships among variates which exist after relationships resulting from treatment factors have been taken into account. In the present study, partial correlation analyses account for variation associated with both Pj concentration and AIPO4 treatments. In non-mycorrhizal seedlings, shoot and root weights were uncorrelated with foliar P concentration, suggesting that Pj concentration was the primary factor controlling both growth and P translocation to foliage. In contrast, the growth of mycorrhizal seedlings was negatively correlated to foliar P concentration suggesting that mycorrhizal seedlings added disproportionately more biomass for any increment of available P^. The ability of mycorrhizal seedlings to more efficiently add biomass for a given increment of a nutrient has been previously reported (Bowen, 1973 ; Harley & Smith, 1983) and appears to hold in the present study. In summary, mycorrhizal infection of P. rigida seedlings increased the seedlings' capacity to grow and accumulate P under P-limiting conditions. Association of the symbiont P. tinctorius with seedling roots conferred a higher uptake affinity for Pj, as evidenced by elevated growth and foliar P concentrations. Enhanced root proliferation and associated proton extrusion by roots of mycorrhizal seedlings under P deprivation led to superior AIPO4 dissolution. Production of organic chelating compounds by the mycorrhizal symbiont may additionally alter AIPO4 solubility equilibria and may represent another P-scavenging mechanism. ACKNOWLEDGEMENTS We would like to thank Anne Buckelew and Paul Wetzel for assistance in the laboratory, Gail Rubin for invaluable statistical guidance, and Mike Rutzke of the Department of Pomology, Cornell University for ICP analyses. This research was funded in part by a grant from the Andrew W. Mellon Foundation to J. R. Cumming from Cornell University, and grant from the United States EPA- USDA-Forest Service Forest Response Program. REFERENCES AGNIHOTRI, V. P. (1970). 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