Mycorrhizal fungi improve the yield and balance of grass–legume mixtures by increasing nutrient access and reducing competition

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Cobb, Gaowen Yang, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2674682/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 05 Sep, 2023 Read the published version in Plant and Soil → Version 1 posted 5 You are reading this latest preprint version Abstract Aims Plant species mixtures with different functional traits or microbial associations can more fully utilize soil nutrient pools. However, there is a gap in our understanding of the synergistic influences of arbuscular mycorrhizal (AM) fungi in nutrient capture and resource partitioning across different soil depths in mixed plant communities, as well as their possible effects on plant coexistence. To address these knowledge gaps, we assessed the effect of AM fungi on nutrient uptake and competition between shallow- and deep-rooted pairs of grass and legume species. Methods To quantify the mycorrhizal mediation of N uptake at different rooting depths, 15 N tracer was injected into soil at 3- and 25-cm depths of monoculture or bi-species mixtures, with or without AM fungi. Results AM symbioses reduced plant competition by equalizing access to N and subsequently increasing shoot N and P concentrations of co-cultivated plant species. Niche differentiation, based on rooting depth, had minimal effects on grass and legume competitive interactions. Both deep-rooted plant species, i.e., Medicago sativa and Dactylis glomerata , and shallow-rooted species, i.e., Lolium perenne and Trifolium repens , primarily acquired nutrients from shallow soil layers. However, AM fungi significantly increased the rooting depth of both shallow- and deep-rooted plant species, enhancing nutrient uptake and N 2 fixation from depths of both 3 and 25 cm. Conclusion Our results suggest a strong influence of AM fungi on host-plant competitiveness and vertical nutrient capture in mixed plant communities, underscoring that microbial mutualist-mediated nutrient acquisition is a key driver of gains in productivity of grass–legume mixtures. Arbuscular mycorrhizal (AM) fungi Competitive balance Grass–legume mixtures Rooting depth Vertical nutrient capture Tracer injection Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Introduction Increasing grassland productivity while reducing environmental impacts remains a substantial agricultural challenge (Chang et al., 2021 ; Nyfeler et al. 2011 ; Reiss and Drinkwater 2018 ). Grass–legume mixtures are widely cultivated to increase agroecosystem productivity and stability while decreasing costly and environmentally damaging nitrogen (N) fertilizer use (Reiss and Drinkwater 2018 ). The benefits of grass–legume mixtures are primarily attributed to complementary partitioning and utilization of soil resources (Thilakarathna et al. 2016 ). This enhanced nutrient uptake efficiency is driven by plant species functional trait diversity (Barry et al. 2019 ; Husse et al. 2016 ; Husse et al. 2017 ; Nyfeler et al. 2011 ). The benefits of grassland mixtures are maximized when constituent species have different N 2 -fixing abilities as well as vertically and temporally differentiated strategies of nutrient acquisition (Hoekstra et al. 2015 ; Husse et al. 2017 ). Vertical niche complementarity between plant species with different canopy and/or rooting depths may also increase the utilization of available light and soil resources (Hoekstra et al. 2015 ; Husse et al. 2016 ; Husse et al. 2017 ; Roscher et al. 2016 ). While previous studies have demonstrated that diverse plant communities can more fully utilize the vertical aerial space (Niklaus et al. 2017 ; Pennekamp et al. 2018 ; Reiss and Drinkwater 2018 ), relatively few studies have assessed belowground spatial resource use (Barry et al. 2019 ). There are some reports that plant communities are more likely to invest into root biomass in deeper soil layers as plant community richness increases, and belowground complementarity effects are linked to the abundance of deep-rooted plant species (Mueller et al. 2013 ; Oram et al. 2018 ). Additionally, more plant roots may aggregate in topsoil as species richness increases (Mommer et al. 2010 ; Ravenek et al. 2014 ). Recent studies suggest that species coexistence and ecosystem productivity are enhanced by positive biotic feedbacks between plants and microbial mutualists, such as arbuscular mycorrhizal (AM) fungi (Barry et al. 2019 ; Bever et al. 2010 ; Coban et al. 2022 ; Yang et al. 2018 ; Zhou et al. 2022 ). AM fungi can enhance nutrient acquisition of plants and may even transfer nutrients among neighboring plants (Barry et al. 2019 ). However, increased plant diversity can alter the outcomes of interactions of plants between microbial mutualists (Eisenhauer 2012 ), thus potentially enhancing compensatory responses among plant species and thereby ultimately increasing yield and plant density (van der Heijden et al., 1998 ; Yang et al. 2018 ). However, more empirical evidence is needed to understand the functioning of roots and their associated mycorrhizal fungi in nutrient acquisition in mixtures of forage plant species. Mycorrhizal fungi generally influence root characteristics (Chen et al. 2017 ), such as total root and taproot lengths and root volume (Shao et al. 2018 ). Kong et al. ( 2014 ) suggested mycorrhizal colonization should be integrated into models linking root formation, maintenance, and persistence based on the finding that thicker roots are more heavily colonized. AM fungi are more strongly linked with belowground than aboveground plant richness (Hiiesalu et al. 2014 ). Although the literature reports that mycorrhizal associations impact root system architecture (Wu et al. 2013 ), there is a knowledge gap regarding AM fungal influence on nutrient uptake of individual plant species in mixed grass–legume communities. To address this gap, we examined the role of AM fungi in plant N nutrition via (i) symbiotic N 2 fixation and (ii) uptake from different soil depths. We hypothesized that N uptake by mycorrhizal plant species mixtures occurs from all soil depths, resulting in increased overall nutrient uptake and biomass yield compared to monocultures. We further hypothesized that AM fungi enhance soil nutrient uptake and N acquisition from the atmosphere via N 2 fixation in both shallow and deep soil layers, leading to greater plant nutrient access throughout the soil profile and consequently increased total N acquisition. Materials And Methods Experimental design Microcosm experiments were conducted in a glasshouse facility maintained at 20–25°C and 50–70% relative humidity. Four species were selected based on their differential symbiotic N 2 -fixation ability (N 2 -fixing legumes versus non-N 2 -fixing grasses) and differential rooting depth (deep-rooted species versus shallow-rooted species): (1) Medicago sativa (cv. ‘Western Star’), a perennial N 2 -fixing legume with a deep taproot and a well-developed fibrous root system; (2) Trifolium repens (cv. ‘Haifa’), a perennial N 2 -fixing legume with a relatively small and shallow root system; (3) Dactylis glomerata (cv. ‘Baoxing’), a perennial non-N 2 -fixing, C 3 tussock grass with deep roots and highly branched rhizomes; (4) Lolium perenne (cv. ‘Kaili’), a perennial non-N 2 -fixing, vigorously tillering C 3 grass with relatively shallow and fine roots (see Fig. 1 a). M. sativa and D. glomerata are reported to access nutrients from deeper soil depths, whereas T. repens and L. perenne typically exploit soil down to an approximately 10-cm depth (Hoekstra et al. 2015 ; Husse et al. 2017 ). These four species were planted in monocultures, for comparison with four bi-species mixtures ( M. sativa + D. glomerate ; M. sativa + L. perenne; T. repens + D. glomerata ; and T. repens + L. perenne ), resulting in eight types of cultures. All plant combinations were grown in sterilized soil or sterilized soil with local AM fungal inoculum. Each culture and soil treatment combination was replicated eight times, resulting in a total of 128 microcosm. To assess the influence of AM fungi on nutrient capture between deep-rooted and shallow-rooted plant species, 15 N was used as a tracer to assess the nutrient uptake from shallow (3 cm) and deep (25 cm) soil depths, following the recommendations of Hoekstra et al ( 2015 and 2017). Four replicate microcosms each were 15 N labelled at 3 cm and at 25 cm soil depths in week 12 (see Fig. 1 b). Growth Conditions Experimental soil was collected from the top 10 cm of an established local grass–legume system at the Modern Agricultural Science and Technology Station of Sichuan Agricultural University, Ya'an, Sichuan, China (103°14′ E, 30°08′ N). Soil was sieved to particles ≤ 2-mm to remove large roots and stones. Each microcosm (21 cm diameter, 30 cm height) was filled with 3:1 ( v : v ) soil:sand mixture with a final pH of 6.2, soil organic matter content of 14.6 g kg − 1 , total N of 1.21 g kg − 1 , total P of 0.61 g kg − 1 , plant-available N of 20.6 mg kg − 1 , and plant-available P of 4.73 mg kg − 1 . Seeds of M. sativa , T. repens , and L. perenne were obtained from the Evergreen International Grass Co., Ltd. (Beijing, China). Seeds of D. glomerata were obtained from the Sichuan Agricultural University (Ya'an, China). The seeds were surface sterilized with 75% alcohol for 2 min and then germinated in vermiculite for 4 weeks. Six seedlings of each species were transplanted into one microcosm for monocultures, and three seedlings of each grass and legume species in the bi-species mixtures were transplanted in an alternating pattern, such that directly neighboring plants were heterospecific. We replaced any seedlings that did not survive within two weeks of transplantation with similar-age seedlings. Fresh soil was used for mycorrhizal inoculations, and sterilized soil (autoclaved at 121°C for 120 minutes and allowed to cool for 72 h) was used for non-mycorrhizal controls. A 100-ml microbial wash, composed of non-AM microbes, was uniformly amended to all microcosms. The microbial wash was prepared by blending soil and water in a 1:2 ratio ( v : v ) for 10 seconds and passing the slurry through a 25-µm filter to exclude the relatively large mycorrhizal spores (Johnson et al. 2010 ). M. sativa was inoculated with rhizobia strain ACCC17676, and T. repens was inoculated with rhizobia strain ACCC18007 (Zhou et al. 2017 ). Both rhizobial inoculants were obtained from the Agricultural Culture Collection of China (ACCC). Two milliliters of a liquid culture of rhizobia strains (grown in yeast extract mannitol medium to exponential phase, > 1×10 9 CUF rhizobia per ml) was applied to seedling roots at transplantation (Larimer et al. 2014 ). All microcosms were arranged randomly in a glasshouse with a 14-h photoperiod per day, a temperature cycle of 25°C day/20°C night, and 50–60% relative humidity. Each microcosm was irrigated daily and amended with 100 ml of modified Hoagland nutrient solution (N- and P-free) biweekly throughout the 16-week experiment following Scheublin and Van Der Heijden ( 2006 ). Labeling With 15 N Tracer Following Hoekstra et al. ( 2014 ), Hoekstra et al. ( 2015 ), and Husse et al. ( 2017 ), 15 N (double labelled ammonium-nitrate, 99 atom%) was employed as a tracer to determine the effect of AM fungi on nutrient uptake from shallow (3 cm) and deep (25 cm) soil depths. Four of the eight treatment replicates were labelled either at a 3- or 25-cm soil depth through six injection holes opened with a 0.5-cm-diameter screwdriver. One milliliter of 15 N was then injected into the corresponding 3- or 25-cm-deep holes, using a 5-mL multi-pipette attached to a hollow steel needle via a silicon tube (Hoekstra et al. 2014 a) during week 12. This resulted in a tracer application rate of 0.08 g N m − 2 (2.512 mg of N) per microcosm. Tools used for 15 N labeling were sterilized to minimize cross-contamination. Although the surface 15 N tracers move vertically downward in response to daily irrigation (He, Zhang, Cao, & Ke, 2022 ; Hoekstra, Suter, Finn, Husse, & Lüscher, 2015 ; Zhang, 2020 ), shallow 15 N tracers can only descend a finite distance, even under conditions of high precipitation. Both He et al. ( 2022 ) and Hoekstra et al. ( 2015 ) indicated that tracers move downward an average distance of 5–10 cm. The majority of tracers remain stationary (Hoekstra et al., 2015 ), facilitating effective interpretation of tracer data when injected at the depths we selected. Sampling Plant shoots and roots were harvested and sorted by species 16 weeks after transplantation and weighed after oven-drying at 60°C for 48 h. A subsample of shoot tissue of each species was ground and analyzed spectrophotometrically for P concentration after digestion in nitric acid (Leonardi et al. 1999 ). Another subsample of shoot tissue was ground in a bead mill and assessed for 15 N and total N by continuous-flow isotope ratio mass spectrometry at the Stable Isotope Laboratory at the Chinese Academy of Agricultural Sciences (Beijing, China). Roots were washed, and then, nodules were counted. Subsamples of roots were clipped into 1-cm segments, cleared in 10% KOH, and stained with trypan blue in lacto-glycerol (modified from Phillips and Hayman 1970 ) to determine the fractional root length colonized by AM fungal structures, intra-radical hyphae, arbuscules, vesicles, or coils at 30 positions per root subsample, using the magnified gridline intersect method at 40× magnification. The remaining roots were then dried at 60°C and weighed. Data analysis Competitive interactions between plant species were calculated as the relative yield per individual (RYind) following the equation RYind = ( O ij / M ij ), where O ij is the shoot biomass of an individual plant species i grown in a bi-species mixture j , and M ij is the mean shoot biomass per individual of species i in the same mycorrhizal treatment j (De Wit 1960 ; Wagg et al. 2011 ) across the biomass data of all bi-species mixtures. A graph was plotted according to Williams and McCarthy ( 2001 ) representing all possible outcomes of competitive interactions between the two gramineous species and two leguminous species (Williams and McCarthy 2001 ). The RY values of the legume and grass components of the bi-species mixtures were plotted against each other to determine the relative prevalence of and underlying cause of yield advantages. The relative yield totals (RYTs) were calculated by summing up the relative yields of the mixture components per treatment (Wagg et al. 2011 ). RYTs are used to check for overyielding in cultivation mixtures, where values greater than 1 indicate a greater biomass production in mixture than expected based on the average of the monocultures of the mixture components (Ren et al. 2017 ). Mycorrhizal growth response (MGR) was calculated as MGR = ln(Myc/Nonmyc), where Myc is the total plant dry matter of mycorrhizal plants and Nonmyc is the mean value of the total dry matter of corresponding non-mycorrhizal plants (Johnson 2010 ; Liu et al. 2021 ). Tracer concentrations (A) were corrected for background concentration (A 0 ) to determine the concentration of excess 15 N using the following formula: atom% 15 N excess = A – A 0 . Tracer uptake (TU) in aboveground biomass of individual species was calculated as TU = atom% 15 N excess × Biomass, where Biomass is aboveground biomass (g plant − 1 ). To calculate N resource utilization from shallow or deep soil, the total uptake of 15 N tracer (TU tot ) was calculated as the sum of TU of both mixture component species. The proportional contribution of uptake from the 25-cm soil depth to total tracer uptake (TU_P 25 ) was calculated as TU_ P 25 = TU 25 / (TU 3 + TU 25 ), where TU 3 is tracer shoot uptake from the 3-cm soil depth and TU 25 is the tracer shoot uptake from the 25-cm soil depth (Hoekstra et al. 2015 ). Percent N derived from the atmosphere via symbiotic N 2 fixation (NDFA%) was calculated using the formula %NDFA = (1 - atom% 15 N excess mix / atom% 15 N excess mon ) × 100, where atom% 15 N excess mix is the 15 N abundance of the legume in each mixture and atom% 15 N excess mon is the 15 N abundance in monoculture conditions. Generalized linear mixed models were used as implemented in SAS version 9.2 (SAS Institute, Cary, NC, USA) to test whether AM fungi interacted with plant community effects to influence response variables, with AM fungi and plant community as fixed factors and block as a random factor. To further test the effect of soil depth on N acquisition, tracer depth was included as a further fixed effect. Tukey’s HSD was utilized for post-hoc mean comparisons at p ≤ 0.05 . Redundancy analysis (RDA) in CANOCO version 5.2 was used to visualize the correlational relationships and effects of treatments by defining the latter as environmental dummy variables (Lai, 2013 ). Log-transformations were used to standardize variables with different scales. Monte Carlo tests were run with restricted random permutations of samples reflecting the experimental design. Results Plant productivity M. sativa produced less above- and belowground biomass in the M. sativa + L. perenne bi-species mixture compared to both M. sativa monoculture and the M. sativa + D. glomerata bi-species mixture. Regardless of mycorrhizal inoculation, the above- and belowground biomass of L. perenne in both the M. sativa + L. perenne and T. repens + L. perenne mixtures was greater than that in L. perenne monoculture, while D. glomerata produced more above- and belowground biomass in the T. repens + D. glomerata mixture than in both D. glomerata monoculture and the M. sativa + D. glomerata mixture (Table 1 and Fig. 2 ). Table 1 Effects of culture system (CS; monoculture or mixture), arbuscular mycorrhizal fungi (AMF) and their interactions on biomass production, nodulation of legume roots, shoot N concentration, and shoot P concentration of model plant species. Significant effects of treatments are indicated in bold. CS AMF CS × AMF df (factor) 2 1 2 df (error) 42 Aboveground biomass Plant community F -value 10.962 14.652 3.207 P -value < 0.001 < 0.001 0.004 Medicago sativa F -value 13.317 8.549 11.652 P -value < 0.001 0.006 < 0.001 Trifolium repens F -value 2.67 26.762 2.913 P -value 0.081 < 0.001 0.065 Lolium perenne F -value 34.403 1.854 0.196 P -value < 0.001 0.181 0.823 Dactylis glomerata F -value 15.551 12.235 1.488 P -value < 0.001 < 0.001 0.237 Belowground biomass Plant community F -value 28.143 12.175 2.316 P -value < 0.001 < 0.001 0.030 Medicago sativa F -value 4.201 17.492 13.191 P -value 0.022 < 0.001 < 0.001 Trifolium repens F -value 8.772 1.406 3.205 P -value 0.001 0.242 0.051 Lolium perenne F -value 22.535 2.583 0.9 P -value < 0.001 0.115 0.414 Dactylis glomerata F -value 10.54 0.978 0.506 P -value < 0.001 0.328 0.607 Total biomass Medicago sativa F -value 6.624 14.437 12.189 P -value 0.003 < 0.001 < 0.001 Trifolium repens F -value 0.987 18.443 3.387 P -value 0.381 < 0.001 0.043 Lolium perenne F -value 30.439 2.663 0.66 P -value < 0.001 0.110 0.522 Dactylis glomerata F -value 14.774 5.661 0.855 P -value < 0.001 0.022 0.433 Nodule number Medicago sativa F -value 18.18 6.244 3.063 P -value < 0.001 0.016 0.057 Trifolium repens F -value 2.742 35.937 4.384 P -value 0.076 < 0.001 0.019 Nodule weight Medicago sativa F -value 0.829 5.686 0.776 P -value 0.444 0.022 0.467 Trifolium repens F -value 5.469 25.981 4.286 P -value 0.008 < 0.001 0.020 Shoot N concentration Medicago sativa F -value 0.69 14.488 2.573 P -value 0.509 0.001 0.093 Trifolium repens F -value 1.386 59.134 0.037 P -value 0.266 < 0.001 0.964 Lolium perenne F -value 5.853 1.594 0.424 P -value < 0.007 0.216 0.659 Dactylis glomerata F -value 0.293 1.777 7.925 P -value 0.748 0.193 0.002 Shoot P concentration Medicago sativa F -value 14.338 55.353 10.045 P -value < 0.001 < 0.001 < 0.001 Trifolium repens F -value 22.993 52.881 0.413 P -value < 0.001 < 0.001 0.665 Lolium perenne F -value 7.601 13.346 1.658 P -value 0.002 0.001 0.208 Dactylis glomerata F -value 4.12 1.027 2.247 P -value 0.026 0.319 0.123 Relative yield total (RYT) F -value 15.705 6.377 0.281 P -value < 0.001 0.014 0.839 In the M. sativa + D. glomerate and T. repens + D. glomerata mixtures, the above- and belowground biomass of M. sativa and T. repens were also significantly greater with mycorrhizal inoculation, whereas aboveground biomass of D. glomerata was significantly reduced for mycorrhizal plants, compared to non-mycorrhizal controls (Table 1 and Fig. 2 ). There were no significant effects of mycorrhizal association on either the above- or belowground biomass of L. perenne. Competitive Relations In Grass–legume Mixtures The relative yield of the mixture components (RYcomp) of the four forage species were strongly influenced by mycorrhizal association and cultivation conditions (i.e., monoculture or bi-species mixture) (Table 1 and Fig. 3 a). For the mixtures of T. repens + D. glomerata and M. sativa + D. glomerata , the biomass of both T. repens or M. sativa were reduced by D. glomerata when AM fungi were not present. However, the competitive pressure of D. glomerata was reduced when AM fungi were present, resulting in enhanced growth of legume partners in mixed microcosms. T. repens and L. perenne were more productive in mixtures than in either monoculture, regardless of the presence of AM fungi; L. perenne showed a competitive advantage over M. sativa in mixtures regardless of mycorrhizal status. AM fungi significantly increased the relative yield total (RYT) of all four grass–legume mixtures, compared to corresponding non-mycorrhizal microcosms (Table 1 ). The mixtures of T. repens + D. glomerata and T. repens + L. perenne in the presence of AM fungi had greater RYTs than the other two mixtures (Fig. 3 b). Rooting Depth Addition of AM fungi significantly increased the rooting depth of all four plant species (Fig. 4 ) compared to corresponding non-mycorrhizal microcosms. The rooting depth of T. repens in non-mycorrhizal T. repens + D. glomerata mixtures was significantly lower than that of non-inoculated T. repens monoculture and the non-inoculated T. repens + L. perenne mixture. Meanwhile, the rooting depth of T. repens increased when AM fungi were present, resulting in no significant differences between monoculture and bi-species mixtures. In the presence of AM fungi, the rooting depth of L. perenne in the M. sativa + L. perenne and T. repens + L. perenne mixtures was greater than that in L. perenne monoculture (Fig. 4 ). Mycorrhizal Growth Response The mycorrhizal growth response (MGR) of M. sativa was greatest in the M. sativa + D. glomerata mixture, and the MGR of T. repens was greatest in the T. repens + D. glomerata mixture (Fig. 5 ). However, mycorrhizal association had no significant effects on L. perenne and D. glomerata productivity when comparing these species between monoculture and mixed cultivation conditions. Am Fungi Root Colonization And Nodulation In the presence of AM fungi, root colonization of the grasses, L. perenne and D. glomerata , was significantly less than root colonization of the legumes, i.e., M. sativa and T. repens (Fig S1 ). Mycorrhizal root colonization was significantly greater for M. sativa in monoculture conditions than in mixtures, whereas T. repens showed significantly greater root colonization in the T. repens + L. perenne mixture than in the T. repens + D. glomerata mixture or monoculture. No significant differences in mycorrhizal root colonization of both L. perenne and D. glomerata were detected between monocultures and mixtures. Both legumes produced more nodules when inoculated with AM fungi (Table 1 ). When AM fungi were absent, the nodule numbers of M. sativa in monoculture were greater than in mixed cultivation. Arbuscular mycorrhizal symbiosis promoted nodulation of M. sativa in the M. sativa + D. glomerata mixture, resulting in similar nodule numbers in the M. sativa + D. glomerata mixture compared to monoculture conditions (Fig. S2 a). Mycorrhizal T. repens produced more nodules in mixed cultivation compared to monoculture conditions (Fig. S2 b). Shoot Nitrogen And Phosphorus Concentrations Compared to non-mycorrhizal controls, AM associations significantly increased the shoot N and phosphorous (P) concentrations of M. sativa in monoculture (Fig. 6 a, b), as well as the shoot P concentration of M. sativa in the M. sativa + D. glomerata mixture (Fig. 6 b). The shoot N and P concentrations of T. repens in mycorrhizal monoculture and mixture cultivation were greater than those of the corresponding non-mycorrhizal microcosms (Table 1 and Fig. 6 c, d). However, AM fungi were associated with reduced shoot P concentration of L. perenne in both M. sativa + L. perenne and T. repens + L. perenne mixtures relative to non-mycorrhizal controls (Fig. 6 f). Shoot N concentration of D. glomerata in the M. sativa + D. glomerata mixture and shoot P concentration of D. glomerata in the T. repens + D. glomerata mixture were significantly greater in mycorrhizal plants (Table 1 and Fig. 6 g, h). N Acquisition And Soil Depth Of N Uptake Tracer injection at a depth of 3 cm resulted in a significantly greater shoot atom% 15 N excess for all four plant species than injection at a depth of 25 cm, regardless of the presence of AM fungi (Table 2 ). Shoot atom% 15 N excess values of M. sativa and T. repens were substantially lower in the legume–grass mixtures than in monocultures. Compared to non-mycorrhizal controls, mycorrhizal M. sativa and T. repens in monocultures showed a lower shoot atom% 15 N excess at a depth of 3 cm. AM fungi reduced shoot atom% 15 N excess of T. repens in the T. repens + D. glomerata mixture at 25 cm. In contrast, a significantly greater shoot atom% 15 N excess was observed for D. glomerata in all mycorrhizal monocultures and mixtures compared to the non-mycorrhizal treatments at a depth of 3 cm (Fig. S3 ). Tracer uptake by M. sativa and T. repens was generally greater in monocultures than legume-grass mixtures, whereas tracer uptake by L. perenne and D. glomerata was significantly reduced in monocultures compared to legume-grass mixtures. Tracer uptake at depths of both 3 and 25 cm were greater for M. sativa in M. sativa + D. glomerata mixtures inoculated with AM fungi than that in non-mycorrhizal microcosms (Fig. 7 a). Conversely, AM fungi were associated with reduced tracer uptake of D. glomerata in M. sativa + D. glomerata mixtures at depths of both 3 and 25 cm (Fig. 7 d). AM fungi increased tracer uptake by T. repens and L. perenne from both soil depths (Table 2 ). Table 2 Effects of culture system (CS; monoculture or mixture), arbuscular mycorrhizal fungi (AMF), tracer depth (TD), and their interactions on atom% 15 N excess, tracer uptake of plant species ( Medicago sativa , Trifolium repens , Lolium perenne , and Dactylis glomerata ), and N-fixation rate of legumes ( M. sativa and T. repens ). Significant effects of treatments are indicated in bold. CS AMF TD CS× AMF CS× TD AMF×TD CS×AMF×TD d.f. (factor) 2 1 1 2 2 1 2 d.f. (error) 24 atom% 15 N excess M. sativa F -value 190.457 0.1 16.351 1.871 15.643 8.013 2.93 P -value < 0.001 0.987 < 0.001 0.176 < 0.001 0.009 0.073 T. repens F -value 218.139 8.581 6.14 3.24 11.692 0.995 4.626 P -value < 0.001 0.007 0.021 0.057 < 0.001 0.329 0.020 L. perenne F -value 0.413 1.203 5.424 0.949 3.984 1.833 0.092 P -value 0.666 0.284 0.029 0.401 0.032 0.188 0.912 D. glomerata F -value 16.704 12.984 10.741 0.024 0.652 16.642 1.137 P -value < 0.001 < 0.001 0.003 0.977 0.530 < 0.001 0.338 Tracer uptake M. sativa F -value 266.632 2.716 4.501 5.543 2.819 0.894 0.687 P -value < 0.001 0.112 0.044 0.010 0.080 0.354 0.513 T. repens F -value 591.523 8.662 69.584 3.434 59.854 1.064 3.814 P -value < 0.001 0.007 < 0.001 0.049 0.000 0.313 0.036 L. perenne F -value 39.802 4.25 2.916 0.877 0.073 0.945 0.171 P -value < 0.001 0.050 0.101 0.429 0.930 0.341 0.844 D. glomerata F -value 29.413 13.495 1.604 12.338 2.003 1.526 0.67 P -value < 0.001 0.001 0.218 0.000 0.157 0.229 0.521 N-fixation rate M. sativa F -value 168.829 1.775 13.696 0.627 8.668 20.235 4.128 P -value < 0.001 0.195 0.001 0.543 0.001 < 0.001 0.029 T. repens F -value 196.45 10.905 1.825 4.085 6.409 2.442 5.938 P -value < 0.001 0.003 0.189 0.030 0.006 0.131 0.008 The percentage of N derived from the atmosphere via symbiotic N 2 fixation (%NDFA) was greater for M. sativa and T. repens in legume–grass mixtures than in monocultures, regardless of mycorrhizal status (Table 2 ). In the non-inoculated M. sativa monoculture as well as the non-inoculated M. sativa + D. glomerata mixture, M. sativa had greater %NDFA when 15 N was added at a depth of 25 cm rather than at 3 cm; however, % NDFA did not differ for mycorrhizal plants between 3- and 25-cm depths (Fig. 7 e). In mycorrhizal T. repens monoculture, %NDFA was greater when 15 N was applied at a depth of 3 cm (Fig. 7 f). Co-variation Of Traits Of The Four Forage Plant Species Redundancy analysis revealed that measured variation of traits of the four forage plant species strongly co-varied with AM fungal root colonization (Fig. 8 ). The first axis explained 76.1%, 85.8%, 70%, and 65.7% of the total variance for M. sativa , T. repens , L. perenne , and D. glomerata , respectively. The shoot P, shoot N, aboveground biomass, nodule number, and nodule weight of M. sativa and T. repens were strongly correlated with mycorrhizal root colonization. Discussion Our study revealed how AM fungi shape competitive interactions in legume–grass mixtures by mediating nutrient uptake from different soil layers. The presence of AM fungi dramatically shifted the competitive balance between grasses and legumes, decreasing resource competition. Niche differentiation based on differences in rooting depth had relatively minimal effects on the competitive interactions in the legume–grass mixtures. Although the influence of vertical root distribution and soil mutualists on belowground resource partitioning have been examined separately in grassland ecosystems (Barry et al. 2019 ; Husse et al. 2017 ; Liu et al. 2021 ), our study is the first to assess synergistic effects. In addition to enhancing the complementarity between N 2 -fixing legumes and non-N 2 -fixing grasses via stimulation of symbiotic N 2 fixation, AM fungi significantly influenced overall nutrient capture and the absolute and relative growth of the grass and legume species in bi-species mixtures, generally increasing overall yield and equalizing production of both plant species in the mixtures. Previous research on the effects of microbial meditated processes in grass–legume mixtures revealed the integral role of mycorrhizal fungi in mixture dynamics (Bashan et al. 2014 ; Stevens et al. 2020 ; Wang et al. 2021 ), in relation to both P acquisition (Hinsinger et al. 2011 ) as well as symbiotic N 2 fixation and transfer between legumes and grasses (Isaac and Borden 2019 ; Jalonen et al. 2009 ; Thilakarathna et al. 2016 ). In the absence of AM fungi, D. glomerata and L. perenne monocultures were less productive than grass–legume mixtures, whereas M. sativa and T. repens produced greater biomass in monocultures than mixtures. AM fungi enhanced the competitiveness of M. sativa and T. repens , particularly compared to D. glomerata in mixtures, and thus increased the relative yields of both the grass and legume species. Our results concur with claims that plant species coexistence is promoted by more equal soil resource uptake in mycorrhizal plant communities (Hiiesalu et al. 2014 ). We also observed that AM fungi facilitated shoot P and N concentrations in both M. sativa and T. repens with neighboring D. glomerata . The greater shoot N and P contents of D. glomerata were likely related to increased soil P availability and fixed N transfer, as N fixation was greater for legumes associated with AM fungi. Additionally, AM fungi reduced shoot atom% 15 N excess in both M. sativa and T. repens and increased shoot atom% 15 N excess in D. glomerata , suggesting plants contended with fungal competition for mineral N (Hodge & Fitter, 2010 ; Püschel et al., 2016 ). Greater reliance of the legumes on symbiotic N 2 fixation can leave more plant-available N for neighboring grasses, effectively enabling a N transfer from legumes to grasses (Hodge, Robinson, & Fitter, 2000 ); this effect is like more relevant in a short-term experiment that has less root and nodule turnover. We also observed that M. sativa , and T. repens exhibited greater root colonization of and positive responsiveness to AM fungi than either D. glomerata or L. perenne . Previous studies reported that AM fungi promote legume growth more than cool-season grass growth under competition (Bahadur et al. 2019 ; Zhou et al. 2022 ), balancing grass–legume mixtures (Klabi et al. 2014 ). AM fungi assist legumes in acquisition of P and other growth-limiting nutrients, ultimately stimulating rhizobial symbioses (Larimer et al. 2014 ; Liu et al. 2020 ). In our study, AM symbiosis had positive effects on nodulation and N 2 -fixation of M. sativa and T. repens . However, AM fungi had a minimal influence on the growth and competitiveness of L. perenne . Indeed, AM fungi reduced the shoot P concentration of L. perenne , potentially attenuating the competitiveness of L. perenne in grass–legume mixtures. Plant species differ in AM fungus-mediated nutrient acquisition (Bahadur et al. 2019 ; Ren et al. 2017 ), providing highly mycotrophic plants a growth advantage and less mycotrophic plants a growth disadvantage when in competition (Wang, Li, Li, & Rosendahl, 2021 ; Liu et al. 2021 ). Our results also support ‘positive mixture effects,’ suggesting greater plant richness leads to greater complementarity in nutrient capture in plant communities (Barry et al. 2019 ; Husse et al. 2017 ). Except for the M. sativa + D. glomerata mixtures, the relative yield totals (RYTs) of grass–legume mixtures were, on average, greater than 1, indicating overyielding relative to yields of monocultures. Our results demonstrate differences in above- and belowground vertical growth strategies among species facilitate coexistence and greater yields, driven by greater soil nutrient use relative to monocultures (Barry et al. 2019 ; Hoekstra et al. 2015 ). We applied 15 N tracer at depths of 3 cm and 25 cm to detect differential N uptake by deep- and shallow-rooted species as well as the potential of AM fungal association to modify nutrient capture. In monocultures, the mean shoot atom% 15 N excess from a 3-cm soil depth was significantly greater for both deep- and shallow-rooted mycorrhizal species, whereas there were no AM fungi-mediated differences in tracer uptake from either soil depth in grass–legume mixtures. This indicates that deep-rooted M. sativa and D. glomerata as well as shallow-rooted L. perenne and T. repens primarily take up nutrients from shallow soil layers. Similar differences in nutrient capture were reported by Hoekstra et al. ( 2014 ). However, some studies have shown significant differences in nutrient uptake between deep- and shallow-rooted species (Berendse 1982 ; Pirhofer-Walzl et al. 2013 ), especially when nutrients were supplied to shallow soil layers (Hoekstra et al. 2015 ). Notably, species co-existence is generally attributed to niche differentiation owing to differences in root architecture (Erktan et al. 2018 ). However, Kraft et al. ( 2015 ) reported that niche differences are likely explained by multiple plant traits, such as rooting depth, phenology, etc. and that individual functional traits may not explain species coexistence or complementarity. Roots tend to proliferate in fertile patches, and thus, plant species with different overall growth strategies may be equally capable of acquiring soil resources from different soil depths. Complementarity belowground is shaped by interactions with symbiotic partners such as AM fungi and rhizobia, markedly different mechanisms than those driving aboveground dynamics (Hiiesalu et al. 2014 ). In the present study, AM fungi significantly increased the rooting depth of all four species and decreased differences in the vertical root distribution between deep- and shallow-rooted species. Accordingly, belowground niche overlap associated with similar root morphologies may have less influence on species coexistence. Plants with deep root systems and extensive horizontal spread are strong competitors, while species with deep but less branched root systems may be less tolerant of direct competition (Erktan et al. 2018 ; Oram et al. 2018 ). Moreover, Semchenko et al. ( 2018 ) demonstrate that direct competitive and facilitative interactions are more influential than differences in the vertical root distribution (Semchenko et al. 2018 ). Both D. glomerata and L. perenne in mixture with either legume species were superior in acquiring 15 N from both shallow and deep soil layers, whereas both M. sativa and T. repens accessed less overall 15 N in mixtures. M. sativa and T. repens acquired more N from symbiotic N 2 fixation than soil sources when grown in mixtures. N derived from the atmosphere can be shared with non-leguminous plants growing in mixtures, promoting grass–legume coexistence when plant-available N is limiting (Pirhofer-Walzl et al. 2013 ). Consistent with our hypothesis, mycorrhizal fungi improved N uptake from shallow and deep soil layers, as evident from the significant shoot atom% 15 N excess in D. glomerata and uptake of the 15 N tracer by mycorrhizal M. sativa, T. repens , and L. perenne . Symbiosis with AM fungi stimulated N 2 fixation in both leguminous species examined, i.e., deep-rooting M . sativa and shallow-rooting T . repens . This is consistent with results reported by Lin et al. ( 2015 ), as AM fungi can modify root structure and function, as well as competitive interactions between plant species. Indeed, the influence of AM fungi on N acquisition was more pronounced in shallow soil layers, likely owing to greater root and AM hyphal density (Morikawa et al. 2022 ; Zhang et al. 2021 ). Our study demonstrates the critical role of AM fungi in vertical nutrient acquisition and stimulation of N 2 fixation, ultimately enhancing productivity of grass–legume plant communities. AM fungi promoted competition–cooperation balance as well as competition–production balance between grasses and legumes, increasing N 2 fixation in legumes and equalizing the distribution of soil nutrients between legumes and grasses in mixtures. Grass–legume mixtures showed overyielding associated with complementarity between grasses and legumes. Both deep and shallow-rooting species relied on the shallow soil layer for most N acquisition, and AM fungi altered plant root depth as well as enhanced N uptake and symbiotic N 2 fixation. Our study clearly demonstrates multiple AM fungus-mediated effects on grass–legume competition and growth, suggesting strategies for resource-efficient forage production based on functional root–microbe interactions. Abbreviations AM: Arbuscular mycorrhizal N: Nitrogen C: Carbon P: Phosphorus TU: Tracer uptake MGR: Mycorrhizal growth response RYT: Relative yield total Declarations Authors’ contributions JZ, JG, SL, XZ and LL conceived the study and selected the methodology; JZ, JG, SL and XL collected the data; JZ, SL, FS and YP analyzed the data; JZ led the writing of the manuscript; AC, GY, YY and LL revised the manuscript; all authors contributed to the drafts and gave final approval for publication. Acknowledgements This study was supported by the National Natural Science Foundation of China (32271776, 31901379), a fellowship from the China Postdoctoral Science Foundation (2022M712289), and a special Sichuan Postdoctoral Research Projects fellowship. Competing interest The authors have no relevant financial or non-financial interests to disclose. 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Mycorrhiza. https://doi.org/10.1007/s00572-021-01061-2 Supplementary Files zjqeditssupportinginfo.docx Cite Share Download PDF Status: Published Journal Publication published 05 Sep, 2023 Read the published version in Plant and Soil → Version 1 posted Reviewers invited by journal 14 Apr, 2023 Reviewers agreed at journal 03 Apr, 2023 Editor assigned by journal 26 Mar, 2023 First submitted to journal 23 Mar, 2023 Editorial decision: Major revisions 17 Mar, 2023 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2674682","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":187198507,"identity":"4735368e-e9ba-49a1-bb01-d54c39db052b","order_by":0,"name":"JIQIONG ZHOU","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA6ElEQVRIiWNgGAWjYBACxmYGhgNglgQQfwBiPgYGZuK1MM4AUmyEtCAAUAszDzFamNuZNx74uKOWQX528zFpm4rDdWzszYcNGGpsonE7jK3g4MwzxxkY5xxLk845c1iCjedYcgLDsbTcBpxaeAwO87YdY2CWyDGTzm0DapHIMT7A2HCYsBY2kBZLErTUMPCAtDBCtSTg1wLyS9sBBgmJtGTLnjPpkm1Avxgk4PGLYf/hzR8+ttUxyM9IPnjjR4U1Pz8wxCQ+1Njg1tLAYACkDtejKkjAoRwE5BnAWurwKBkFo2AUjIIRDwBnEVHWGsYrqgAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0002-0171-9047","institution":"China Agricultural University","correspondingAuthor":true,"prefix":"","firstName":"JIQIONG","middleName":"","lastName":"ZHOU","suffix":""},{"id":187198508,"identity":"bea48561-60f2-444e-9046-49788fe40569","order_by":1,"name":"Jinchao Gong","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Jinchao","middleName":"","lastName":"Gong","suffix":""},{"id":187198509,"identity":"2ee646e6-639a-4670-810f-632eaf9980d5","order_by":2,"name":"Shan Liu","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Shan","middleName":"","lastName":"Liu","suffix":""},{"id":187198510,"identity":"42f6f8da-8194-4939-ac30-2e241790c3bb","order_by":3,"name":"Adam B. Cobb","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Adam","middleName":"B.","lastName":"Cobb","suffix":""},{"id":187198511,"identity":"e3652378-8af2-46b4-9585-65872e71c5a4","order_by":4,"name":"Gaowen Yang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Gaowen","middleName":"","lastName":"Yang","suffix":""},{"id":187198512,"identity":"9649466c-27f2-4c4c-bad3-e984c765a048","order_by":5,"name":"Xiangjun Li","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Xiangjun","middleName":"","lastName":"Li","suffix":""},{"id":187198513,"identity":"86ba3696-2dc2-4805-9676-74897aa876c7","order_by":6,"name":"Feida Sun","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Feida","middleName":"","lastName":"Sun","suffix":""},{"id":187198514,"identity":"0e1e0466-78d4-4d12-9f4f-5dc7e30e1e66","order_by":7,"name":"Yan Pen","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Yan","middleName":"","lastName":"Pen","suffix":""},{"id":187198515,"identity":"b3a8afa5-6917-40a3-95a7-85fe8e194e2b","order_by":8,"name":"Yanhong Yan","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Yanhong","middleName":"","lastName":"Yan","suffix":""},{"id":187198516,"identity":"8a548752-629d-458a-afe9-f71d27302b11","order_by":9,"name":"Lin Liu","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Lin","middleName":"","lastName":"Liu","suffix":""},{"id":187198517,"identity":"dff83b77-4ed9-4584-ac40-1f1cd6d7adb6","order_by":10,"name":"Xinquan Zhang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Xinquan","middleName":"","lastName":"Zhang","suffix":""}],"badges":[],"createdAt":"2023-03-09 16:19:32","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2674682/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2674682/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s11104-023-06261-7","type":"published","date":"2023-09-05T15:01:28+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":35065120,"identity":"81e5481c-0959-48f4-a725-ab26c1f3f2ac","added_by":"auto","created_at":"2023-03-30 22:14:08","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":3157177,"visible":true,"origin":"","legend":"\u003cp\u003eExperimental design\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/d5f80aae5f95d47a511fe207.png"},{"id":35064357,"identity":"fdd2a676-9fed-4ba9-98c7-a2dfccdfc05f","added_by":"auto","created_at":"2023-03-30 22:06:07","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":2180937,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of arbuscular mycorrhizal fungi (AMF) on above- and belowground biomass of \u003cem\u003eMedicago sativa\u003c/em\u003e(a, b), \u003cem\u003eTrifolium repens\u003c/em\u003e (c, d), \u003cem\u003eLolium perenne\u003c/em\u003e (e, f), and \u003cem\u003eDactylis glomerata\u003c/em\u003e (g, h) in monoculture and grass–legume mixture microcosms. The means of treatments labelled with the same letter(s) were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e). The tops and bottoms of boxes represent the upper (75th percentile) and lower (25th percentile) quartiles, respectively; the Mean is shown as a square in a box; the median is shown as a horizontal line inside each box (middle quartile); whiskers show the 1.5-fold interquartile range added to and subtracted from the upper and lower quartiles, respectively\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/c80df2bcb33a31e5ddac8c6f.png"},{"id":35064361,"identity":"b5418798-5ca3-40c9-84c7-d32941b0bd81","added_by":"auto","created_at":"2023-03-30 22:06:08","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":901510,"visible":true,"origin":"","legend":"\u003cp\u003e(a) Graphic representation of all possible outcomes of competitive interactions between two leguminous species (RY\u003csub\u003eL\u003c/sub\u003e, relative yield of legumes; Ms, \u003cem\u003eMedicago sativa\u003c/em\u003e; Tr, \u003cem\u003eTrifolium repens\u003c/em\u003e) and the two gramineous species (RY\u003csub\u003eG\u003c/sub\u003e, relative yield of grasses; Lp, \u003cem\u003eLolium perenne\u003c/em\u003e; Dg, \u003cem\u003eDactylis glomerata\u003c/em\u003e). The diagonal reference line indicates the boundary between the areas of the graph in which leguminous species showed a competitive advantage over gramineous species (area above) and in which gramineous species showed a competitive advantage over leguminous species (area below). Area Ⅰ is where leguminous plant growth is enhanced in grass–legume mixture while gramineous growth is suppressed by leguminous species. Area Ⅱ indicates where gramineous species and leguminous species are both facilitated in grass–legume mixtures compared to corresponding monocultures. Area Ⅲ indicates the conditions under which both gramineous species and leguminous species are suppressed in grass–legume mixtures. Area Ⅳ indicates conditions in which gramineous species had a clear competitive advantage over leguminous species in grass–legume mixtures. Yellow points indicate grass–legume mixtures without arbuscular mycorrhizal fungi (AMF); blue points represent grass–legume mixtures inoculated with AMF. (b) Effects of AMF on the relative yield totals in monoculture and grass–legume mixture microcosms. The means of treatments labelled with the same letter(s) were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e). The tops and bottoms of boxes represent the upper (75th percentile) and lower (25th percentile) quartiles, respectively; the Mean is shown as a square in a box; the median is shown as a horizontal line inside each box (middle quartile); whiskers show the 1.5-fold interquartile range added to and subtracted from the upper and lower quartiles, respectively\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/da3474fdb3522e0d1499edb1.png"},{"id":35064358,"identity":"a9948120-574c-4a6f-8250-0dd0f3e4152c","added_by":"auto","created_at":"2023-03-30 22:06:08","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":80997,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of arbuscular mycorrhizal fungi (AMF) on rooting deepth of \u003cem\u003eMedicago sativa\u003c/em\u003e (a), \u003cem\u003eTrifolium repens\u003c/em\u003e (b), \u003cem\u003eLolium perenne\u003c/em\u003e (c), and \u003cem\u003eDactylis glomerata\u003c/em\u003e (d) in monoculture and grass–legume mixture microcosms. The means of treatments labelled with the same letter(s) were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e). The tops and bottoms of boxes represent the upper (75\u003csup\u003eth\u003c/sup\u003e percentile) and lower (25\u003csup\u003eth\u003c/sup\u003e percentile) quartiles, respectively; the Mean is shown as a square in a box; the median is shown as a horizontal line inside each box (middle quartile); whiskers show the 1.5-fold interquartile range added to and subtracted from the upper and lower quartiles, respectively\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/5c1df33c3fb695283a41fe44.png"},{"id":35066009,"identity":"fd0f2170-b622-4dbb-9bb4-d6046d2dd655","added_by":"auto","created_at":"2023-03-30 22:22:08","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":231630,"visible":true,"origin":"","legend":"\u003cp\u003eThe influence of mycorrhizal growth responses (MGR) of \u003cem\u003eMedicago sativa\u003c/em\u003e, \u003cem\u003eTrifolium repens\u003c/em\u003e, \u003cem\u003eLolium perenne\u003c/em\u003e, and \u003cem\u003eDactylis glomerata\u003c/em\u003e in monoculture and grass–legume mixture microcosms. The means of treatments labelled with the same letter(s) were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e). The tops and bottoms of boxes represent the upper (75\u003csup\u003eth\u003c/sup\u003e percentile) and lower (25\u003csup\u003eth\u003c/sup\u003e percentile) quartiles; the Mean is shown as a square in a box; the median is shown as a horizontal line inside each box (middle quartile); whiskers show the 1.5-fold interquartile range added to and subtracted from the upper and lower quartiles, respectively\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/98a3aa388c99a2953f535979.png"},{"id":35065122,"identity":"0b9a18cf-ff52-40e8-be96-9b8fe28455eb","added_by":"auto","created_at":"2023-03-30 22:14:08","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":1844544,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of arbuscular mycorrhizal fungi (AMF) on shoot N concentration and shoot P concentration of \u003cem\u003eMedicago sativa\u003c/em\u003e (a, b), \u003cem\u003eTrifolium repens\u003c/em\u003e (c, d), \u003cem\u003eLolium perenne\u003c/em\u003e (e, f), and \u003cem\u003eDactylis glomerata\u003c/em\u003e (g, h) in monoculture and grass–legume mixture microcosms. The means of treatments labelled with the same letter(s) were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e). The tops and bottoms of boxes represent the upper (75th percentile) and lower (25th percentile) quartiles, respectively; the Mean is shown as a square in a box; the median is shown as a horizontal line inside each box (middle quartile); whiskers show the 1.5-fold interquartile range added to and subtracted from the upper and lower quartiles, respectively\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/30a716c8910fc4ea9d3ed0ef.png"},{"id":35064365,"identity":"18f00e9c-5553-43e8-8655-c01a7eae0583","added_by":"auto","created_at":"2023-03-30 22:06:08","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":2003780,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of arbuscular mycorrhizal fungi (AMF) on tracer uptake of \u003cem\u003eMedicago sativa\u003c/em\u003e (a), \u003cem\u003eTrifolium repens\u003c/em\u003e (b), \u003cem\u003eLolium perenne\u003c/em\u003e (c), and \u003cem\u003eDactylis glomerata\u003c/em\u003e (d), and the percent N derived from the atmosphere fixed by legumes (% NDFA) for \u003cem\u003eMedicago sativa\u003c/em\u003e (e), and \u003cem\u003eTrifolium repens\u003c/em\u003e (f) injected at a 3- or 25-cm soil depth in monoculture and grass–legume mixture microcosms. The plots show means +/- SE; the means of treatments labelled with the same letter were not significantly different based on Tukey’s HSD test (\u003cem\u003ep ≤ 0.05\u003c/em\u003e)\u003c/p\u003e","description":"","filename":"floatimage7.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/317d7551c3b33bfeeafd6f97.png"},{"id":35064364,"identity":"5827ccf2-7843-4543-9440-080a39d069a3","added_by":"auto","created_at":"2023-03-30 22:06:08","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":335777,"visible":true,"origin":"","legend":"\u003cp\u003eRedundancy analysis (RDA) of plant variables in axis 1 × axis 2 ordination planes constrained by the presence of arbuscular mycorrhizal fungi (+AMF or -AMF) and \u003csup\u003e15\u003c/sup\u003eN tracer depths (3 cm or 25 cm)\u003c/p\u003e","description":"","filename":"floatimage8.png","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/269016cd7e562c8bf5a190bb.png"},{"id":35066010,"identity":"ad41d359-0662-4cc2-abe2-f226925ff823","added_by":"auto","created_at":"2023-03-30 22:22:08","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":3183217,"visible":true,"origin":"","legend":"","description":"","filename":"zjqeditssupportinginfo.docx","url":"https://assets-eu.researchsquare.com/files/rs-2674682/v1/00ddecefa95c5102e1ebf7be.docx"}],"financialInterests":"","formattedTitle":"Mycorrhizal fungi improve the yield and balance of grass–legume mixtures by increasing nutrient access and reducing competition","fulltext":[{"header":"Introduction","content":"\u003cp\u003eIncreasing grassland productivity while reducing environmental impacts remains a substantial agricultural challenge (Chang et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Nyfeler et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Reiss and Drinkwater \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Grass\u0026ndash;legume mixtures are widely cultivated to increase agroecosystem productivity and stability while decreasing costly and environmentally damaging nitrogen (N) fertilizer use (Reiss and Drinkwater \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). The benefits of grass\u0026ndash;legume mixtures are primarily attributed to complementary partitioning and utilization of soil resources (Thilakarathna et al. \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). This enhanced nutrient uptake efficiency is driven by plant species functional trait diversity (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Nyfeler et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). The benefits of grassland mixtures are maximized when constituent species have different N\u003csub\u003e2\u003c/sub\u003e-fixing abilities as well as vertically and temporally differentiated strategies of nutrient acquisition (Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eVertical niche complementarity between plant species with different canopy and/or rooting depths may also increase the utilization of available light and soil resources (Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Roscher et al. \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). While previous studies have demonstrated that diverse plant communities can more fully utilize the vertical aerial space (Niklaus et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Pennekamp et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Reiss and Drinkwater \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2018\u003c/span\u003e), relatively few studies have assessed belowground spatial resource use (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). There are some reports that plant communities are more likely to invest into root biomass in deeper soil layers as plant community richness increases, and belowground complementarity effects are linked to the abundance of deep-rooted plant species (Mueller et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Oram et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Additionally, more plant roots may aggregate in topsoil as species richness increases (Mommer et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Ravenek et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2014\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eRecent studies suggest that species coexistence and ecosystem productivity are enhanced by positive biotic feedbacks between plants and microbial mutualists, such as arbuscular mycorrhizal (AM) fungi (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Bever et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Coban et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Yang et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Zhou et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). AM fungi can enhance nutrient acquisition of plants and may even transfer nutrients among neighboring plants (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). However, increased plant diversity can alter the outcomes of interactions of plants between microbial mutualists (Eisenhauer \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2012\u003c/span\u003e), thus potentially enhancing compensatory responses among plant species and thereby ultimately increasing yield and plant density (van der Heijden et al., \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e1998\u003c/span\u003e; Yang et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, more empirical evidence is needed to understand the functioning of roots and their associated mycorrhizal fungi in nutrient acquisition in mixtures of forage plant species.\u003c/p\u003e \u003cp\u003eMycorrhizal fungi generally influence root characteristics (Chen et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), such as total root and taproot lengths and root volume (Shao et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Kong et al. (\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) suggested mycorrhizal colonization should be integrated into models linking root formation, maintenance, and persistence based on the finding that thicker roots are more heavily colonized. AM fungi are more strongly linked with belowground than aboveground plant richness (Hiiesalu et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Although the literature reports that mycorrhizal associations impact root system architecture (Wu et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), there is a knowledge gap regarding AM fungal influence on nutrient uptake of individual plant species in mixed grass\u0026ndash;legume communities. To address this gap, we examined the role of AM fungi in plant N nutrition via (i) symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation and (ii) uptake from different soil depths. We hypothesized that N uptake by mycorrhizal plant species mixtures occurs from all soil depths, resulting in increased overall nutrient uptake and biomass yield compared to monocultures. We further hypothesized that AM fungi enhance soil nutrient uptake and N acquisition from the atmosphere via N\u003csub\u003e2\u003c/sub\u003e fixation in both shallow and deep soil layers, leading to greater plant nutrient access throughout the soil profile and consequently increased total N acquisition.\u003c/p\u003e"},{"header":"Materials And Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eExperimental design\u003c/h2\u003e \u003cp\u003eMicrocosm experiments were conducted in a glasshouse facility maintained at 20\u0026ndash;25\u0026deg;C and 50\u0026ndash;70% relative humidity. Four species were selected based on their differential symbiotic N\u003csub\u003e2\u003c/sub\u003e-fixation ability (N\u003csub\u003e2\u003c/sub\u003e-fixing legumes versus non-N\u003csub\u003e2\u003c/sub\u003e-fixing grasses) and differential rooting depth (deep-rooted species versus shallow-rooted species): (1) \u003cem\u003eMedicago sativa\u003c/em\u003e (cv. \u0026lsquo;Western Star\u0026rsquo;), a perennial N\u003csub\u003e2\u003c/sub\u003e-fixing legume with a deep taproot and a well-developed fibrous root system; (2) \u003cem\u003eTrifolium repens\u003c/em\u003e (cv. \u0026lsquo;Haifa\u0026rsquo;), a perennial N\u003csub\u003e2\u003c/sub\u003e-fixing legume with a relatively small and shallow root system; (3) \u003cem\u003eDactylis glomerata\u003c/em\u003e (cv. \u0026lsquo;Baoxing\u0026rsquo;), a perennial non-N\u003csub\u003e2\u003c/sub\u003e-fixing, C\u003csub\u003e3\u003c/sub\u003e tussock grass with deep roots and highly branched rhizomes; (4) \u003cem\u003eLolium perenne\u003c/em\u003e (cv. \u0026lsquo;Kaili\u0026rsquo;), a perennial non-N\u003csub\u003e2\u003c/sub\u003e-fixing, vigorously tillering C\u003csub\u003e3\u003c/sub\u003e grass with relatively shallow and fine roots (see Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea). \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e are reported to access nutrients from deeper soil depths, whereas \u003cem\u003eT. repens\u003c/em\u003e and \u003cem\u003eL. perenne\u003c/em\u003e typically exploit soil down to an approximately 10-cm depth (Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). These four species were planted in monocultures, for comparison with four bi-species mixtures (\u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerate\u003c/em\u003e; \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne; T. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e; and \u003cem\u003eT. repens\u0026thinsp;+\u0026thinsp;L. perenne\u003c/em\u003e), resulting in eight types of cultures. All plant combinations were grown in sterilized soil or sterilized soil with local AM fungal inoculum. Each culture and soil treatment combination was replicated eight times, resulting in a total of 128 microcosm. To assess the influence of AM fungi on nutrient capture between deep-rooted and shallow-rooted plant species, \u003csup\u003e15\u003c/sup\u003eN was used as a tracer to assess the nutrient uptake from shallow (3 cm) and deep (25 cm) soil depths, following the recommendations of Hoekstra et al (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e and 2017). Four replicate microcosms each were \u003csup\u003e15\u003c/sup\u003eN labelled at 3 cm and at 25 cm soil depths in week 12 (see Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eGrowth Conditions\u003c/h3\u003e\n\u003cp\u003eExperimental soil was collected from the top 10 cm of an established local grass\u0026ndash;legume system at the Modern Agricultural Science and Technology Station of Sichuan Agricultural University, Ya'an, Sichuan, China (103\u0026deg;14\u0026prime; E, 30\u0026deg;08\u0026prime; N). Soil was sieved to particles\u0026thinsp;\u0026le;\u0026thinsp;2-mm to remove large roots and stones. Each microcosm (21 cm diameter, 30 cm height) was filled with 3:1 (\u003cem\u003ev\u003c/em\u003e:\u003cem\u003ev\u003c/em\u003e) soil:sand mixture with a final pH of 6.2, soil organic matter content of 14.6 g kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, total N of 1.21 g kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, total P of 0.61 g kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, plant-available N of 20.6 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, and plant-available P of 4.73 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eSeeds of \u003cem\u003eM. sativa\u003c/em\u003e, \u003cem\u003eT. repens\u003c/em\u003e, and \u003cem\u003eL. perenne\u003c/em\u003e were obtained from the Evergreen International Grass Co., Ltd. (Beijing, China). Seeds of \u003cem\u003eD. glomerata\u003c/em\u003e were obtained from the Sichuan Agricultural University (Ya'an, China). The seeds were surface sterilized with 75% alcohol for 2 min and then germinated in vermiculite for 4 weeks. Six seedlings of each species were transplanted into one microcosm for monocultures, and three seedlings of each grass and legume species in the bi-species mixtures were transplanted in an alternating pattern, such that directly neighboring plants were heterospecific. We replaced any seedlings that did not survive within two weeks of transplantation with similar-age seedlings.\u003c/p\u003e \u003cp\u003eFresh soil was used for mycorrhizal inoculations, and sterilized soil (autoclaved at 121\u0026deg;C for 120 minutes and allowed to cool for 72 h) was used for non-mycorrhizal controls. A 100-ml microbial wash, composed of non-AM microbes, was uniformly amended to all microcosms. The microbial wash was prepared by blending soil and water in a 1:2 ratio (\u003cem\u003ev\u003c/em\u003e:\u003cem\u003ev\u003c/em\u003e) for 10 seconds and passing the slurry through a 25-\u0026micro;m filter to exclude the relatively large mycorrhizal spores (Johnson et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). \u003cem\u003eM. sativa\u003c/em\u003e was inoculated with rhizobia strain ACCC17676, and \u003cem\u003eT. repens\u003c/em\u003e was inoculated with rhizobia strain ACCC18007 (Zhou et al. \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Both rhizobial inoculants were obtained from the Agricultural Culture Collection of China (ACCC). Two milliliters of a liquid culture of rhizobia strains (grown in yeast extract mannitol medium to exponential phase, \u0026gt;\u0026thinsp;1\u0026times;10\u003csup\u003e9\u003c/sup\u003e CUF rhizobia per ml) was applied to seedling roots at transplantation (Larimer et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). All microcosms were arranged randomly in a glasshouse with a 14-h photoperiod per day, a temperature cycle of 25\u0026deg;C day/20\u0026deg;C night, and 50\u0026ndash;60% relative humidity. Each microcosm was irrigated daily and amended with 100 ml of modified Hoagland nutrient solution (N- and P-free) biweekly throughout the 16-week experiment following Scheublin and Van Der Heijden (\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2006\u003c/span\u003e).\u003c/p\u003e\n\u003ch3\u003eLabeling With \u003csup\u003e15\u003c/sup\u003eN Tracer\u003c/h3\u003e\n\u003cp\u003eFollowing Hoekstra et al. (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), Hoekstra et al. (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), and Husse et al. (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), \u003csup\u003e15\u003c/sup\u003eN (double labelled ammonium-nitrate, 99 atom%) was employed as a tracer to determine the effect of AM fungi on nutrient uptake from shallow (3 cm) and deep (25 cm) soil depths. Four of the eight treatment replicates were labelled either at a 3- or 25-cm soil depth through six injection holes opened with a 0.5-cm-diameter screwdriver. One milliliter of \u003csup\u003e15\u003c/sup\u003eN was then injected into the corresponding 3- or 25-cm-deep holes, using a 5-mL multi-pipette attached to a hollow steel needle via a silicon tube (Hoekstra et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2014\u003c/span\u003ea) during week 12. This resulted in a tracer application rate of 0.08 g N m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e (2.512 mg of N) per microcosm. Tools used for \u003csup\u003e15\u003c/sup\u003eN labeling were sterilized to minimize cross-contamination. Although the surface \u003csup\u003e15\u003c/sup\u003eN tracers move vertically downward in response to daily irrigation (He, Zhang, Cao, \u0026amp; Ke, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Hoekstra, Suter, Finn, Husse, \u0026amp; L\u0026uuml;scher, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Zhang, \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), shallow \u003csup\u003e15\u003c/sup\u003eN tracers can only descend a finite distance, even under conditions of high precipitation. Both He et al. (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) and Hoekstra et al. (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e) indicated that tracers move downward an average distance of 5\u0026ndash;10 cm. The majority of tracers remain stationary (Hoekstra et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), facilitating effective interpretation of tracer data when injected at the depths we selected.\u003c/p\u003e\n\u003ch3\u003eSampling\u003c/h3\u003e\n\u003cp\u003ePlant shoots and roots were harvested and sorted by species 16 weeks after transplantation and weighed after oven-drying at 60\u0026deg;C for 48 h. A subsample of shoot tissue of each species was ground and analyzed spectrophotometrically for P concentration after digestion in nitric acid (Leonardi et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e1999\u003c/span\u003e). Another subsample of shoot tissue was ground in a bead mill and assessed for \u003csup\u003e15\u003c/sup\u003eN and total N by continuous-flow isotope ratio mass spectrometry at the Stable Isotope Laboratory at the Chinese Academy of Agricultural Sciences (Beijing, China).\u003c/p\u003e \u003cp\u003eRoots were washed, and then, nodules were counted. Subsamples of roots were clipped into 1-cm segments, cleared in 10% KOH, and stained with trypan blue in lacto-glycerol (modified from Phillips and Hayman \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e1970\u003c/span\u003e) to determine the fractional root length colonized by AM fungal structures, intra-radical hyphae, arbuscules, vesicles, or coils at 30 positions per root subsample, using the magnified gridline intersect method at 40\u0026times; magnification. The remaining roots were then dried at 60\u0026deg;C and weighed.\u003c/p\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eData analysis\u003c/h2\u003e \u003cp\u003eCompetitive interactions between plant species were calculated as the relative yield per individual (RYind) following the equation RYind = (\u003cem\u003eO\u003c/em\u003e\u003csub\u003e\u003cem\u003eij\u003c/em\u003e\u003c/sub\u003e / \u003cem\u003eM\u003c/em\u003e\u003csub\u003e\u003cem\u003eij\u003c/em\u003e\u003c/sub\u003e), where \u003cem\u003eO\u003c/em\u003e\u003csub\u003e\u003cem\u003eij\u003c/em\u003e\u003c/sub\u003e is the shoot biomass of an individual plant species \u003cem\u003ei\u003c/em\u003e grown in a bi-species mixture \u003cem\u003ej\u003c/em\u003e, and \u003cem\u003eM\u003c/em\u003e\u003csub\u003e\u003cem\u003eij\u003c/em\u003e\u003c/sub\u003e is the mean shoot biomass per individual of species \u003cem\u003ei\u003c/em\u003e in the same mycorrhizal treatment \u003cem\u003ej\u003c/em\u003e (De Wit \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1960\u003c/span\u003e; Wagg et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) across the biomass data of all bi-species mixtures. A graph was plotted according to Williams and McCarthy (\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2001\u003c/span\u003e) representing all possible outcomes of competitive interactions between the two gramineous species and two leguminous species (Williams and McCarthy \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). The RY values of the legume and grass components of the bi-species mixtures were plotted against each other to determine the relative prevalence of and underlying cause of yield advantages.\u003c/p\u003e \u003cp\u003eThe relative yield totals (RYTs) were calculated by summing up the relative yields of the mixture components per treatment (Wagg et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). RYTs are used to check for overyielding in cultivation mixtures, where values greater than 1 indicate a greater biomass production in mixture than expected based on the average of the monocultures of the mixture components (Ren et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eMycorrhizal growth response (MGR) was calculated as MGR\u0026thinsp;=\u0026thinsp;ln(Myc/Nonmyc), where Myc is the total plant dry matter of mycorrhizal plants and Nonmyc is the mean value of the total dry matter of corresponding non-mycorrhizal plants (Johnson \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Liu et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTracer concentrations (A) were corrected for background concentration (A\u003csub\u003e0\u003c/sub\u003e) to determine the concentration of excess \u003csup\u003e15\u003c/sup\u003eN using the following formula: atom% \u003csup\u003e15\u003c/sup\u003eN excess\u0026thinsp;=\u0026thinsp;A \u0026ndash; A\u003csub\u003e0\u003c/sub\u003e.\u003c/p\u003e \u003cp\u003eTracer uptake (TU) in aboveground biomass of individual species was calculated as TU\u0026thinsp;=\u0026thinsp;atom% \u003csup\u003e15\u003c/sup\u003eN excess \u0026times; Biomass, where Biomass is aboveground biomass (g plant\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e).\u003c/p\u003e \u003cp\u003eTo calculate N resource utilization from shallow or deep soil, the total uptake of \u003csup\u003e15\u003c/sup\u003eN tracer (TU\u003csub\u003etot\u003c/sub\u003e) was calculated as the sum of TU of both mixture component species.\u003c/p\u003e \u003cp\u003eThe proportional contribution of uptake from the 25-cm soil depth to total tracer uptake (TU_P\u003csub\u003e25\u003c/sub\u003e) was calculated as TU_ P\u003csub\u003e25\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;TU\u003csub\u003e25\u003c/sub\u003e / (TU\u003csub\u003e3\u003c/sub\u003e\u0026thinsp;+\u0026thinsp;TU\u003csub\u003e25\u003c/sub\u003e), where TU\u003csub\u003e3\u003c/sub\u003e is tracer shoot uptake from the 3-cm soil depth and TU\u003csub\u003e25\u003c/sub\u003e is the tracer shoot uptake from the 25-cm soil depth (Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e).\u003c/p\u003e \u003cp\u003ePercent N derived from the atmosphere via symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation (NDFA%) was calculated using the formula %NDFA = (1 - atom% \u003csup\u003e15\u003c/sup\u003eN excess\u003csub\u003emix\u003c/sub\u003e / atom% \u003csup\u003e15\u003c/sup\u003eN excess\u003csub\u003emon\u003c/sub\u003e) \u0026times; 100, where atom% \u003csup\u003e15\u003c/sup\u003eN excess\u003csub\u003emix\u003c/sub\u003e is the \u003csup\u003e15\u003c/sup\u003eN abundance of the legume in each mixture and atom% \u003csup\u003e15\u003c/sup\u003eN excess\u003csub\u003emon\u003c/sub\u003e is the \u003csup\u003e15\u003c/sup\u003eN abundance in monoculture conditions.\u003c/p\u003e \u003cp\u003eGeneralized linear mixed models were used as implemented in SAS version 9.2 (SAS Institute, Cary, NC, USA) to test whether AM fungi interacted with plant community effects to influence response variables, with AM fungi and plant community as fixed factors and block as a random factor. To further test the effect of soil depth on N acquisition, tracer depth was included as a further fixed effect. Tukey\u0026rsquo;s HSD was utilized for post-hoc mean comparisons at \u003cem\u003ep\u0026thinsp;\u0026le;\u0026thinsp;0.05\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eRedundancy analysis (RDA) in CANOCO version 5.2 was used to visualize the correlational relationships and effects of treatments by defining the latter as environmental dummy variables (Lai, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Log-transformations were used to standardize variables with different scales. Monte Carlo tests were run with restricted random permutations of samples reflecting the experimental design.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003ePlant productivity\u003c/h2\u003e \u003cp\u003e \u003cem\u003eM. sativa\u003c/em\u003e produced less above- and belowground biomass in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e bi-species mixture compared to both \u003cem\u003eM. sativa\u003c/em\u003e monoculture and the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e bi-species mixture. Regardless of mycorrhizal inoculation, the above- and belowground biomass of \u003cem\u003eL. perenne\u003c/em\u003e in both the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e mixtures was greater than that in \u003cem\u003eL. perenne\u003c/em\u003e monoculture, while \u003cem\u003eD. glomerata\u003c/em\u003e produced more above- and belowground biomass in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture than in both \u003cem\u003eD. glomerata\u003c/em\u003e monoculture and the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eEffects of culture system (CS; monoculture or mixture), arbuscular mycorrhizal fungi (AMF) and their interactions on biomass production, nodulation of legume roots, shoot N concentration, and shoot P concentration of model plant species. Significant effects of treatments are indicated in bold.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAMF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c6\" namest=\"c5\"\u003e \u003cp\u003eCS \u0026times; AMF\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003edf (factor)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003edf (error)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c6\" namest=\"c3\"\u003e \u003cp\u003e42\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAboveground biomass\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePlant community\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e10.962\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e14.652\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e3.207\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.004\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e13.317\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e8.549\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e11.652\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.006\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.67\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e26.762\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.913\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.081\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.065\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eLolium perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e34.403\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1.854\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.196\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.181\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.823\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eDactylis glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e15.551\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e12.235\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.488\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.237\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eBelowground biomass\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePlant community\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e28.143\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e12.175\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e2.316\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.030\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e4.201\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e17.492\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e13.191\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.022\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e8.772\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1.406\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.205\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.242\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.051\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eLolium perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e22.535\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e2.583\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.115\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.414\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eDactylis glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e10.54\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.978\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.506\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.328\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.607\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTotal biomass\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e6.624\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e14.437\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e12.189\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.003\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.987\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e18.443\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e3.387\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.381\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.043\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eLolium perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e30.439\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e2.663\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.66\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.110\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.522\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eDactylis glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e14.774\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e5.661\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.855\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.022\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.433\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eNodule number\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e18.18\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e6.244\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.063\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.016\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.057\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.742\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e35.937\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e4.384\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.076\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.019\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eNodule weight\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.829\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e5.686\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.776\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.444\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.022\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.467\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e5.469\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e25.981\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e4.286\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.008\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.020\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eShoot N concentration\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e14.488\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.573\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.509\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.093\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.386\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e59.134\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.037\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.266\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.964\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eLolium perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e5.853\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1.594\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.424\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.007\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.216\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.659\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eDactylis glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.293\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1.777\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e7.925\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.748\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.193\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.002\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eShoot P concentration\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eMedicago sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e14.338\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e55.353\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e10.045\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eTrifolium repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e22.993\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e52.881\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.413\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.665\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eLolium perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e7.601\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e13.346\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.658\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.002\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.208\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eDactylis glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e4.12\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e1.027\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.247\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e0.026\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e0.319\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.123\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eRelative yield total (RYT)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e15.705\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e6.377\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.281\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c5\" namest=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.014\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.839\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerate\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixtures, the above- and belowground biomass of \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e were also significantly greater with mycorrhizal inoculation, whereas aboveground biomass of \u003cem\u003eD. glomerata\u003c/em\u003e was significantly reduced for mycorrhizal plants, compared to non-mycorrhizal controls (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). There were no significant effects of mycorrhizal association on either the above- or belowground biomass of \u003cem\u003eL. perenne.\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eCompetitive Relations In Grass–legume Mixtures\u003c/h3\u003e\n\u003cp\u003eThe relative yield of the mixture components (RYcomp) of the four forage species were strongly influenced by mycorrhizal association and cultivation conditions (i.e., monoculture or bi-species mixture) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea). For the mixtures of \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e and \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e, the biomass of both \u003cem\u003eT. repens\u003c/em\u003e or \u003cem\u003eM. sativa\u003c/em\u003e were reduced by \u003cem\u003eD. glomerata\u003c/em\u003e when AM fungi were not present. However, the competitive pressure of \u003cem\u003eD. glomerata\u003c/em\u003e was reduced when AM fungi were present, resulting in enhanced growth of legume partners in mixed microcosms. \u003cem\u003eT. repens\u003c/em\u003e and \u003cem\u003eL. perenne\u003c/em\u003e were more productive in mixtures than in either monoculture, regardless of the presence of AM fungi; \u003cem\u003eL. perenne\u003c/em\u003e showed a competitive advantage over \u003cem\u003eM. sativa\u003c/em\u003e in mixtures regardless of mycorrhizal status.\u003c/p\u003e \u003cp\u003eAM fungi significantly increased the relative yield total (RYT) of all four grass\u0026ndash;legume mixtures, compared to corresponding non-mycorrhizal microcosms (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The mixtures of \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e in the presence of AM fungi had greater RYTs than the other two mixtures (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eRooting Depth\u003c/h3\u003e\n\u003cp\u003eAddition of AM fungi significantly increased the rooting depth of all four plant species (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e) compared to corresponding non-mycorrhizal microcosms. The rooting depth of \u003cem\u003eT. repens\u003c/em\u003e in non-mycorrhizal \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixtures was significantly lower than that of non-inoculated \u003cem\u003eT. repens\u003c/em\u003e monoculture and the non-inoculated \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e mixture. Meanwhile, the rooting depth of \u003cem\u003eT. repens\u003c/em\u003e increased when AM fungi were present, resulting in no significant differences between monoculture and bi-species mixtures. In the presence of AM fungi, the rooting depth of \u003cem\u003eL. perenne\u003c/em\u003e in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e mixtures was greater than that in \u003cem\u003eL. perenne\u003c/em\u003e monoculture (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eMycorrhizal Growth Response\u003c/h3\u003e\n\u003cp\u003eThe mycorrhizal growth response (MGR) of \u003cem\u003eM. sativa\u003c/em\u003e was greatest in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture, and the MGR of \u003cem\u003eT. repens\u003c/em\u003e was greatest in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). However, mycorrhizal association had no significant effects on \u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e productivity when comparing these species between monoculture and mixed cultivation conditions.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eAm Fungi Root Colonization And Nodulation\u003c/h3\u003e\n\u003cp\u003eIn the presence of AM fungi, root colonization of the grasses, \u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e, was significantly less than root colonization of the legumes, i.e., \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e (Fig \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). Mycorrhizal root colonization was significantly greater for \u003cem\u003eM. sativa\u003c/em\u003e in monoculture conditions than in mixtures, whereas \u003cem\u003eT. repens\u003c/em\u003e showed significantly greater root colonization in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e mixture than in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture or monoculture. No significant differences in mycorrhizal root colonization of both \u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e were detected between monocultures and mixtures.\u003c/p\u003e \u003cp\u003eBoth legumes produced more nodules when inoculated with AM fungi (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). When AM fungi were absent, the nodule numbers of \u003cem\u003eM. sativa\u003c/em\u003e in monoculture were greater than in mixed cultivation. Arbuscular mycorrhizal symbiosis promoted nodulation of \u003cem\u003eM. sativa\u003c/em\u003e in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture, resulting in similar nodule numbers in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture compared to monoculture conditions (Fig. \u003cspan refid=\"MOESM2\" class=\"InternalRef\"\u003eS2\u003c/span\u003e a). Mycorrhizal \u003cem\u003eT. repens\u003c/em\u003e produced more nodules in mixed cultivation compared to monoculture conditions (Fig. \u003cspan refid=\"MOESM2\" class=\"InternalRef\"\u003eS2\u003c/span\u003e b).\u003c/p\u003e\n\u003ch3\u003eShoot Nitrogen And Phosphorus Concentrations\u003c/h3\u003e\n\u003cp\u003eCompared to non-mycorrhizal controls, AM associations significantly increased the shoot N and phosphorous (P) concentrations of \u003cem\u003eM. sativa\u003c/em\u003e in monoculture (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ea, b), as well as the shoot P concentration of \u003cem\u003eM. sativa\u003c/em\u003e in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eb). The shoot N and P concentrations of \u003cem\u003eT. repens\u003c/em\u003e in mycorrhizal monoculture and mixture cultivation were greater than those of the corresponding non-mycorrhizal microcosms (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ec, d). However, AM fungi were associated with reduced shoot P concentration of \u003cem\u003eL. perenne\u003c/em\u003e in both \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eL. perenne\u003c/em\u003e mixtures relative to non-mycorrhizal controls (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ef). Shoot N concentration of \u003cem\u003eD. glomerata\u003c/em\u003e in the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture and shoot P concentration of \u003cem\u003eD. glomerata\u003c/em\u003e in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture were significantly greater in mycorrhizal plants (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eg, h).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eN Acquisition And Soil Depth Of N Uptake\u003c/h3\u003e\n\u003cp\u003eTracer injection at a depth of 3 cm resulted in a significantly greater shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess for all four plant species than injection at a depth of 25 cm, regardless of the presence of AM fungi (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess values of \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e were substantially lower in the legume\u0026ndash;grass mixtures than in monocultures. Compared to non-mycorrhizal controls, mycorrhizal \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e in monocultures showed a lower shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess at a depth of 3 cm. AM fungi reduced shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess of \u003cem\u003eT. repens\u003c/em\u003e in the \u003cem\u003eT. repens\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture at 25 cm. In contrast, a significantly greater shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess was observed for \u003cem\u003eD. glomerata\u003c/em\u003e in all mycorrhizal monocultures and mixtures compared to the non-mycorrhizal treatments at a depth of 3 cm (Fig. \u003cspan refid=\"MOESM3\" class=\"InternalRef\"\u003eS3\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTracer uptake by \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e was generally greater in monocultures than legume-grass mixtures, whereas tracer uptake by \u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e was significantly reduced in monocultures compared to legume-grass mixtures. Tracer uptake at depths of both 3 and 25 cm were greater for \u003cem\u003eM. sativa\u003c/em\u003e in \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixtures inoculated with AM fungi than that in non-mycorrhizal microcosms (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ea). Conversely, AM fungi were associated with reduced tracer uptake of \u003cem\u003eD. glomerata\u003c/em\u003e in \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixtures at depths of both 3 and 25 cm (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ed). AM fungi increased tracer uptake by \u003cem\u003eT. repens\u003c/em\u003e and \u003cem\u003eL. perenne\u003c/em\u003e from both soil depths (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eEffects of culture system (CS; monoculture or mixture), arbuscular mycorrhizal fungi (AMF), tracer depth (TD), and their interactions on atom% \u003csup\u003e15\u003c/sup\u003eN excess, tracer uptake of plant species (\u003cem\u003eMedicago sativa\u003c/em\u003e, \u003cem\u003eTrifolium repens\u003c/em\u003e, \u003cem\u003eLolium perenne\u003c/em\u003e, and \u003cem\u003eDactylis glomerata\u003c/em\u003e), and N-fixation rate of legumes (\u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e). Significant effects of treatments are indicated in bold.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"9\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCS\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAMF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eTD\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCS\u0026times;\u003c/p\u003e \u003cp\u003eAMF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eCS\u0026times;\u003c/p\u003e \u003cp\u003eTD\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eAMF\u0026times;TD\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eCS\u0026times;AMF\u0026times;TD\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ed.f. (factor)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ed.f. (error)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"7\" nameend=\"c9\" namest=\"c3\"\u003e \u003cp\u003e24\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eatom%\u003c/b\u003e\u003csup\u003e\u003cb\u003e15\u003c/b\u003e\u003c/sup\u003e\u003cb\u003eN excess\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eM. sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e190.457\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e16.351\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.871\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e15.643\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e8.013\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e2.93\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.987\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.176\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e0.009\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.073\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eT. repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e218.139\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e8.581\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e6.14\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.24\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e11.692\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.995\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e4.626\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.007\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.021\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.057\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.329\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.020\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eL. perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.413\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.203\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e5.424\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.949\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e3.984\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e1.833\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.092\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.666\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.284\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.029\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.401\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.032\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.188\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.912\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eD. glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e16.704\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e12.984\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e10.741\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.024\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.652\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e16.642\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.003\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.977\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.530\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.338\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTracer uptake\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eM. sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e266.632\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2.716\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e4.501\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e5.543\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e2.819\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.894\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.687\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.112\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.044\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.010\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.080\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.354\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.513\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eT. repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e591.523\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e8.662\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e69.584\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e3.434\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e59.854\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e1.064\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e3.814\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.007\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.049\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.000\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.313\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.036\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eL. perenne\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e39.802\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e4.25\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.916\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.877\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.073\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.945\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.171\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.050\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.101\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.429\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.930\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.341\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.844\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eD. glomerata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e29.413\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e13.495\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.604\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e12.338\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e2.003\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e1.526\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.67\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.218\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.157\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.229\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.521\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eN-fixation rate\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eM. sativa\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e168.829\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.775\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e13.696\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.627\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e8.668\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e20.235\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e4.128\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.195\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.543\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.029\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eT. repens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eF\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e196.45\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e10.905\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.825\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e4.085\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e6.409\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e2.442\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e5.938\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eP\u003c/em\u003e-value\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e\u0026lt;\u0026thinsp;0.001\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.003\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.189\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003e0.030\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u003cb\u003e0.006\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e0.131\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e\u003cb\u003e0.008\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eThe percentage of N derived from the atmosphere via symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation (%NDFA) was greater for \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e in legume\u0026ndash;grass mixtures than in monocultures, regardless of mycorrhizal status (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). In the non-inoculated \u003cem\u003eM. sativa\u003c/em\u003e monoculture as well as the non-inoculated \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixture, \u003cem\u003eM. sativa\u003c/em\u003e had greater %NDFA when \u003csup\u003e15\u003c/sup\u003eN was added at a depth of 25 cm rather than at 3 cm; however, % NDFA did not differ for mycorrhizal plants between 3- and 25-cm depths (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ee). In mycorrhizal \u003cem\u003eT. repens\u003c/em\u003e monoculture, %NDFA was greater when \u003csup\u003e15\u003c/sup\u003eN was applied at a depth of 3 cm (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ef).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eCo-variation Of Traits Of The Four Forage Plant Species\u003c/h3\u003e\n\u003cp\u003eRedundancy analysis revealed that measured variation of traits of the four forage plant species strongly co-varied with AM fungal root colonization (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e). The first axis explained 76.1%, 85.8%, 70%, and 65.7% of the total variance for \u003cem\u003eM. sativa\u003c/em\u003e, \u003cem\u003eT. repens\u003c/em\u003e, \u003cem\u003eL. perenne\u003c/em\u003e, and \u003cem\u003eD. glomerata\u003c/em\u003e, respectively. The shoot P, shoot N, aboveground biomass, nodule number, and nodule weight of \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e were strongly correlated with mycorrhizal root colonization.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eOur study revealed how AM fungi shape competitive interactions in legume\u0026ndash;grass mixtures by mediating nutrient uptake from different soil layers. The presence of AM fungi dramatically shifted the competitive balance between grasses and legumes, decreasing resource competition. Niche differentiation based on differences in rooting depth had relatively minimal effects on the competitive interactions in the legume\u0026ndash;grass mixtures. Although the influence of vertical root distribution and soil mutualists on belowground resource partitioning have been examined separately in grassland ecosystems (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Liu et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), our study is the first to assess synergistic effects. In addition to enhancing the complementarity between N\u003csub\u003e2\u003c/sub\u003e-fixing legumes and non-N\u003csub\u003e2\u003c/sub\u003e-fixing grasses via stimulation of symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation, AM fungi significantly influenced overall nutrient capture and the absolute and relative growth of the grass and legume species in bi-species mixtures, generally increasing overall yield and equalizing production of both plant species in the mixtures.\u003c/p\u003e \u003cp\u003ePrevious research on the effects of microbial meditated processes in grass\u0026ndash;legume mixtures revealed the integral role of mycorrhizal fungi in mixture dynamics (Bashan et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Stevens et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Wang et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), in relation to both P acquisition (Hinsinger et al. \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) as well as symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation and transfer between legumes and grasses (Isaac and Borden \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Jalonen et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Thilakarathna et al. \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). In the absence of AM fungi, \u003cem\u003eD. glomerata\u003c/em\u003e and \u003cem\u003eL. perenne\u003c/em\u003e monocultures were less productive than grass\u0026ndash;legume mixtures, whereas \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e produced greater biomass in monocultures than mixtures. AM fungi enhanced the competitiveness of \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e, particularly compared to \u003cem\u003eD. glomerata\u003c/em\u003e in mixtures, and thus increased the relative yields of both the grass and legume species. Our results concur with claims that plant species coexistence is promoted by more equal soil resource uptake in mycorrhizal plant communities (Hiiesalu et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). We also observed that AM fungi facilitated shoot P and N concentrations in both \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e with neighboring \u003cem\u003eD. glomerata\u003c/em\u003e. The greater shoot N and P contents of \u003cem\u003eD. glomerata\u003c/em\u003e were likely related to increased soil P availability and fixed N transfer, as N fixation was greater for legumes associated with AM fungi. Additionally, AM fungi reduced shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess in both \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e and increased shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess in \u003cem\u003eD. glomerata\u003c/em\u003e, suggesting plants contended with fungal competition for mineral N (Hodge \u0026amp; Fitter, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; P\u0026uuml;schel et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Greater reliance of the legumes on symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation can leave more plant-available N for neighboring grasses, effectively enabling a N transfer from legumes to grasses (Hodge, Robinson, \u0026amp; Fitter, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2000\u003c/span\u003e); this effect is like more relevant in a short-term experiment that has less root and nodule turnover.\u003c/p\u003e \u003cp\u003eWe also observed that \u003cem\u003eM. sativa\u003c/em\u003e, and \u003cem\u003eT. repens\u003c/em\u003e exhibited greater root colonization of and positive responsiveness to AM fungi than either \u003cem\u003eD. glomerata\u003c/em\u003e or \u003cem\u003eL. perenne\u003c/em\u003e. Previous studies reported that AM fungi promote legume growth more than cool-season grass growth under competition (Bahadur et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Zhou et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), balancing grass\u0026ndash;legume mixtures (Klabi et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). AM fungi assist legumes in acquisition of P and other growth-limiting nutrients, ultimately stimulating rhizobial symbioses (Larimer et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Liu et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). In our study, AM symbiosis had positive effects on nodulation and N\u003csub\u003e2\u003c/sub\u003e-fixation of \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e. However, AM fungi had a minimal influence on the growth and competitiveness of \u003cem\u003eL. perenne\u003c/em\u003e. Indeed, AM fungi reduced the shoot P concentration of \u003cem\u003eL. perenne\u003c/em\u003e, potentially attenuating the competitiveness of \u003cem\u003eL. perenne\u003c/em\u003e in grass\u0026ndash;legume mixtures. Plant species differ in AM fungus-mediated nutrient acquisition (Bahadur et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Ren et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), providing highly mycotrophic plants a growth advantage and less mycotrophic plants a growth disadvantage when in competition (Wang, Li, Li, \u0026amp; Rosendahl, \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Liu et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Our results also support \u0026lsquo;positive mixture effects,\u0026rsquo; suggesting greater plant richness leads to greater complementarity in nutrient capture in plant communities (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Husse et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Except for the \u003cem\u003eM. sativa\u003c/em\u003e\u0026thinsp;+\u0026thinsp;\u003cem\u003eD. glomerata\u003c/em\u003e mixtures, the relative yield totals (RYTs) of grass\u0026ndash;legume mixtures were, on average, greater than 1, indicating overyielding relative to yields of monocultures. Our results demonstrate differences in above- and belowground vertical growth strategies among species facilitate coexistence and greater yields, driven by greater soil nutrient use relative to monocultures (Barry et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eWe applied \u003csup\u003e15\u003c/sup\u003eN tracer at depths of 3 cm and 25 cm to detect differential N uptake by deep- and shallow-rooted species as well as the potential of AM fungal association to modify nutrient capture. In monocultures, the mean shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess from a 3-cm soil depth was significantly greater for both deep- and shallow-rooted mycorrhizal species, whereas there were no AM fungi-mediated differences in tracer uptake from either soil depth in grass\u0026ndash;legume mixtures. This indicates that deep-rooted \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eD. glomerata\u003c/em\u003e as well as shallow-rooted \u003cem\u003eL. perenne\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e primarily take up nutrients from shallow soil layers. Similar differences in nutrient capture were reported by Hoekstra et al. (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). However, some studies have shown significant differences in nutrient uptake between deep- and shallow-rooted species (Berendse \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Pirhofer-Walzl et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), especially when nutrients were supplied to shallow soil layers (Hoekstra et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Notably, species co-existence is generally attributed to niche differentiation owing to differences in root architecture (Erktan et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, Kraft et al. (\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2015\u003c/span\u003e) reported that niche differences are likely explained by multiple plant traits, such as rooting depth, phenology, etc. and that individual functional traits may not explain species coexistence or complementarity. Roots tend to proliferate in fertile patches, and thus, plant species with different overall growth strategies may be equally capable of acquiring soil resources from different soil depths. Complementarity belowground is shaped by interactions with symbiotic partners such as AM fungi and rhizobia, markedly different mechanisms than those driving aboveground dynamics (Hiiesalu et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). In the present study, AM fungi significantly increased the rooting depth of all four species and decreased differences in the vertical root distribution between deep- and shallow-rooted species. Accordingly, belowground niche overlap associated with similar root morphologies may have less influence on species coexistence. Plants with deep root systems and extensive horizontal spread are strong competitors, while species with deep but less branched root systems may be less tolerant of direct competition (Erktan et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Oram et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Moreover, Semchenko et al. (\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2018\u003c/span\u003e) demonstrate that direct competitive and facilitative interactions are more influential than differences in the vertical root distribution (Semchenko et al. \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Both \u003cem\u003eD. glomerata\u003c/em\u003e and \u003cem\u003eL. perenne\u003c/em\u003e in mixture with either legume species were superior in acquiring \u003csup\u003e15\u003c/sup\u003eN from both shallow and deep soil layers, whereas both \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e accessed less overall \u003csup\u003e15\u003c/sup\u003eN in mixtures. \u003cem\u003eM. sativa\u003c/em\u003e and \u003cem\u003eT. repens\u003c/em\u003e acquired more N from symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation than soil sources when grown in mixtures. N derived from the atmosphere can be shared with non-leguminous plants growing in mixtures, promoting grass\u0026ndash;legume coexistence when plant-available N is limiting (Pirhofer-Walzl et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eConsistent with our hypothesis, mycorrhizal fungi improved N uptake from shallow and deep soil layers, as evident from the significant shoot atom% \u003csup\u003e15\u003c/sup\u003eN excess in \u003cem\u003eD. glomerata\u003c/em\u003e and uptake of the \u003csup\u003e15\u003c/sup\u003eN tracer by mycorrhizal \u003cem\u003eM. sativa, T. repens\u003c/em\u003e, and \u003cem\u003eL. perenne\u003c/em\u003e. Symbiosis with AM fungi stimulated N\u003csub\u003e2\u003c/sub\u003e fixation in both leguminous species examined, i.e., deep-rooting \u003cem\u003eM\u003c/em\u003e. \u003cem\u003esativa\u003c/em\u003e and shallow-rooting \u003cem\u003eT\u003c/em\u003e. \u003cem\u003erepens\u003c/em\u003e. This is consistent with results reported by Lin et al. (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), as AM fungi can modify root structure and function, as well as competitive interactions between plant species. Indeed, the influence of AM fungi on N acquisition was more pronounced in shallow soil layers, likely owing to greater root and AM hyphal density (Morikawa et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Zhang et al. \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOur study demonstrates the critical role of AM fungi in vertical nutrient acquisition and stimulation of N\u003csub\u003e2\u003c/sub\u003e fixation, ultimately enhancing productivity of grass\u0026ndash;legume plant communities. AM fungi promoted competition\u0026ndash;cooperation balance as well as competition\u0026ndash;production balance between grasses and legumes, increasing N\u003csub\u003e2\u003c/sub\u003e fixation in legumes and equalizing the distribution of soil nutrients between legumes and grasses in mixtures. Grass\u0026ndash;legume mixtures showed overyielding associated with complementarity between grasses and legumes. Both deep and shallow-rooting species relied on the shallow soil layer for most N acquisition, and AM fungi altered plant root depth as well as enhanced N uptake and symbiotic N\u003csub\u003e2\u003c/sub\u003e fixation. Our study clearly demonstrates multiple AM fungus-mediated effects on grass\u0026ndash;legume competition and growth, suggesting strategies for resource-efficient forage production based on functional root\u0026ndash;microbe interactions.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eAM: Arbuscular mycorrhizal\u003c/p\u003e\n\u003cp\u003eN: Nitrogen\u003c/p\u003e\n\u003cp\u003eC: Carbon\u003c/p\u003e\n\u003cp\u003eP: Phosphorus\u003c/p\u003e\n\u003cp\u003eTU: Tracer uptake\u003c/p\u003e\n\u003cp\u003eMGR: Mycorrhizal growth response\u003c/p\u003e\n\u003cp\u003eRYT: Relative yield total\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eJZ, JG, SL, XZ and LL conceived the study and selected the methodology; JZ, JG, SL and XL collected the data; JZ, SL, FS and YP analyzed the data; JZ led the writing of the manuscript; AC, GY, YY and LL revised the manuscript; all authors contributed to the drafts and gave final approval for publication.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by the National Natural Science Foundation of China (32271776, 31901379), a fellowship from the China Postdoctoral Science Foundation (2022M712289),\u0026nbsp;and a special Sichuan Postdoctoral Research Projects fellowship.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no relevant financial or non-financial interests to disclose.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBahadur A, Jin Z, Long X, Jiang S, Zhang Q, Pan J, Liu Y, Feng H (2019) Arbuscular mycorrhizal fungi alter plant interspecific interaction under nitrogen fertilization. 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Mycorrhiza. https://doi.org/10.1007/s00572-021-01061-2\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Arbuscular mycorrhizal (AM) fungi, Competitive balance, Grass–legume mixtures, Rooting depth, Vertical nutrient capture, Tracer injection","lastPublishedDoi":"10.21203/rs.3.rs-2674682/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2674682/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eAims\u003c/h2\u003e \u003cp\u003ePlant species mixtures with different functional traits or microbial associations can more fully utilize soil nutrient pools. However, there is a gap in our understanding of the synergistic influences of arbuscular mycorrhizal (AM) fungi in nutrient capture and resource partitioning across different soil depths in mixed plant communities, as well as their possible effects on plant coexistence. To address these knowledge gaps, we assessed the effect of AM fungi on nutrient uptake and competition between shallow- and deep-rooted pairs of grass and legume species.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eTo quantify the mycorrhizal mediation of N uptake at different rooting depths, \u003csup\u003e15\u003c/sup\u003eN tracer was injected into soil at 3- and 25-cm depths of monoculture or bi-species mixtures, with or without AM fungi.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eAM symbioses reduced plant competition by equalizing access to N and subsequently increasing shoot N and P concentrations of co-cultivated plant species. Niche differentiation, based on rooting depth, had minimal effects on grass and legume competitive interactions. Both deep-rooted plant species, i.e., \u003cem\u003eMedicago sativa\u003c/em\u003e and \u003cem\u003eDactylis glomerata\u003c/em\u003e, and shallow-rooted species, i.e., \u003cem\u003eLolium perenne\u003c/em\u003e and \u003cem\u003eTrifolium repens\u003c/em\u003e, primarily acquired nutrients from shallow soil layers. However, AM fungi significantly increased the rooting depth of both shallow- and deep-rooted plant species, enhancing nutrient uptake and N\u003csub\u003e2\u003c/sub\u003e fixation from depths of both 3 and 25 cm.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eOur results suggest a strong influence of AM fungi on host-plant competitiveness and vertical nutrient capture in mixed plant communities, underscoring that microbial mutualist-mediated nutrient acquisition is a key driver of gains in productivity of grass\u0026ndash;legume mixtures.\u003c/p\u003e","manuscriptTitle":"Mycorrhizal fungi improve the yield and balance of grass–legume mixtures by increasing nutrient access and reducing competition","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-03-30 22:06:03","doi":"10.21203/rs.3.rs-2674682/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewersInvited","content":"","date":"2023-04-14T09:03:32+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2023-04-03T10:29:27+00:00","index":0,"fulltext":""},{"type":"editorAssigned","content":"","date":"2023-03-27T02:27:51+00:00","index":"","fulltext":""},{"type":"submitted","content":"Plant and Soil","date":"2023-03-23T12:35:10+00:00","index":"","fulltext":""},{"type":"decision","content":"Major revisions","date":"2023-03-17T07:55:22+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"1eb36b44-1278-4786-bb16-2a6b455659bc","owner":[],"postedDate":"March 30th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2023-09-11T15:08:01+00:00","versionOfRecord":{"articleIdentity":"rs-2674682","link":"https://doi.org/10.1007/s11104-023-06261-7","journal":{"identity":"plant-and-soil","isVorOnly":false,"title":"Plant and Soil"},"publishedOn":"2023-09-05 15:01:28","publishedOnDateReadable":"September 5th, 2023"},"versionCreatedAt":"2023-03-30 22:06:03","video":"","vorDoi":"10.1007/s11104-023-06261-7","vorDoiUrl":"https://doi.org/10.1007/s11104-023-06261-7","workflowStages":[]},"version":"v1","identity":"rs-2674682","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2674682","identity":"rs-2674682","version":["v1"]},"buildId":"_2-kVJe1T_tPrBINL-cwx","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}