Our study was unique in examining a large number of combinations of plant species, AMF species and N sources, but provides little evidence for N partitioning or a role for AMF in N acquisition of old-field plant species. Rather, AMF were at best neutral and most often negative in their effect on plant growth and N acquisition under conditions of low N supply. Although AMF did not promote plant growth in this study, we detected specificity in whether plants experienced AMF as neutral or detrimental to growth as well as in plant species responses to N sources.
Plant species responses to AMF
Mycorrhizal fungi are known to act as facultative parasites at relatively high phosphorus availability (Buwalda & Goh, 1982), especially when photosynthate is limiting, such as may occur when young plants are establishing (Smith, 1980; Hayman, 1983; Brundrett, 1991) or when light levels are low (Smith, 1980; Smith et al., 1986; Brundrett, 1991). We initiated our plants as newly germinated seedlings, so it is possible that AMF drains on limited photosynthate during early establishment contributed to the growth depressions observed at final harvest. However, early establishment drains typically disappear after 1–2 wk (Hayman, 1983). Because our plants grew for four months, it is likely that some other factor contributed to the observed growth depressions. Light availability is a possible factor, although the day length and light intensity provided in our experiment were sufficient to promote vigorous growth of our S. bicolor cultures and were above those found to depress or reverse positive growth responses to mycorrhizal colonization in other studies (onion, Hayman, 1974; maize, Daft & El-Giahmi, 1978).
Photosynthate production is also strongly limited by N availability; plants require substantial amounts of N (2–5% N content by dry weight, compared with 0.3–0.5% phosphorus content, Marschner, 2002), and well over 50% of leaf N is devoted to photosynthesis (Field & Mooney, 1986). In order to benefit plant growth under N-limiting conditions, AMF must enhance plant access to N to an extent sufficient to allow photosynthate production in excess of AMF demand for photosynthate. Given the relatively high N demands of plants, the expectation that net benefits of AMF exceed net costs in unfertilized soil and fall below net costs in fertilized soil (Johnson et al., 1997) may not apply when N is the more limiting nutrient. In our study, percentage N in shoots increased for those plant–AMF combinations that typically resulted in lower overall plant growth and total N acquisition. Also, percentage N in roots tended to increase, and percentage C in shoots tended to decrease, with inoculation by any of the three AMF species (A. trappei, G. decipiens, G. gigaspora) that caused growth declines in most plant species. When inoculated with these same AMF species, percentage C in roots declined for the two plant species that showed the strongest growth depressions. These results suggest that AMF often imposed a C drain on plants, preventing plants from using acquired N for increased growth.
A recent study involving a greater range of old-field plants and AMF, grown in low-nutrient soil with supplemental phosphorus, found growth-depressing effects of AMF in half of the plant–AMF combinations (Klironomos, 2003). Yet, other studies, including a number conducted under low N conditions, have used 15N labeling to show that AMF can transport N to the plant host, increasing host acquisition of 15NH4+ (celery, Ames et al., 1983; cucumber, Johansen et al., 1992), 15NO3− (ryegrass, Cliquet et al., 1997; lettuce, Azcón et al., 2001), 15N-labelled amino acids (ryegrass, Cliquet et al., 1997; wheat, transformed Queen Ann's Lace roots, Hawkins et al., 2000, experiments i, iv), or 15N derived from plant litter (plantain, Hodge et al., 2001). However, 15N-labelling studies must be interpreted with care because of artifacts related to differences in size of nonmycorrhizal vs mycorrhizal plants (Högberg et al., 1994). Furthermore, the amount of 15N transferred in tracer studies is small compared with total plant needs, and in four of the six studies no increases in plant growth or total N were detected in mycorrhizal vs nonmycorrhizal treatments, despite the evidence for N transfer. Cliquet et al. (1997) and Azcón et al. (2001) did find significantly enhanced plant growth and N content for mycorrhizal treatments, although in the latter study plants were fertilized with a phosphorus-free fertilizer, so mycorrhizal benefit might best be explained by phosphorus-limiting conditions.
Mechanisms of AMF-mediated N acquisition include increased surface area for uptake or increased mineralization of N from organic forms. The dominant perspective has been that AMF are not saprotrophic (Brundrett, 1991), although recent work has shown that AMF may stimulate mineralization of organic patches in soil (Hodge, 2001; Hodge et al., 2001; but see Hodge et al., 2000). Our results do not support a significant role for either mechanism of AMF-mediated N acquisition, at least not in individual pot culture with homogeneous soil (but see Cliquet et al., 1997). Perhaps AMF-mediated N acquisition manifests itself only under more realistic field conditions, where plants are competing for limited N in spatially unbounded and heterogenous soil. Then, one plant's gain in N is another's loss, and AMF may enhance N acquisition by virtue of access to a larger soil volume than roots and/or via exploitation of nutrient-rich hot spots (Hawkins et al., 2000). Few such studies yet exist, although Šmilauerová & Šmilauer (2002) found that foraging responses to N and phosphorus patches were unrelated to AM mycorrhizal dependency under competitive field conditions. Similarly, Hodge (2003) found no effect of AM mycorrhizal inoculum on plant N capture from a dispersed vs patchy organic N source, although competing plant species did obtain more N from either source with AMF inoculation. More generally, competitive environment might also affect whether AMF act as parasites or mutualists, altering our understanding of the parasitism to mutualism continuum emerging from single plant pot culture studies (e.g. Klironomos, 2003).
Although both native and exotic plant species experienced growth-depressing effects of AMF in this study, the two native species were affected most. Yet the natives were also the smallest species on average, and we found that mean plant size was inversely correlated with MSS. We found no relationship between other measured plant traits (percentage N and C and root length, surface area or diameter) and MSS, although percentage N in shoots was negatively correlated with mycorrhizal dependency. The latter relationship was due to the fact that A. odoratum had the lowest percentage N in shoots (highest N use efficiency; Chapin & Van Cleve, 1989) of all our plant species, and was the one species whose growth and N acquisition was unaffected by colonization with any AMF species – all other species had negative values of mycorrhizal dependency. Other work has suggested that root morphology (Hetrick et al., 1990; Brundrett, 2002), perhaps in combination with phenology (Hetrick et al., 1988) is important in determining plant responses to AMF. Also, although not addressed here, greater allocation to nonstructural carbohydrates, especially in roots, has been associated with greater mycorrhizal dependency (Graham, 2000), and could thus also influence vulnerability to parasitic effects of AMF. Traits of AMF species may be of equal importance in understanding why AMF are beneficial or parasitic under any given resource conditions or for any particular plant species. Aggressiveness in rate of root colonization, for example, can predict nonbeneficial AMF effects on plants at high phosphorus supply (Graham, 2000; Graham & Abbott, 2000). C and N stoichiometry (especially relative to that of plant species), size and growth rate, and nativity and phenology of AMF are also logical traits to explore. Such information may have helped to understand why Glomus D1 was most frequently neutral and the other AMF species most frequently nonbeneficial in effect on the plant species in our experiment.
Plant species responses to N
Even if AMF play such a little role in N partitioning, plant species might partition forms or sources of N by virtue of root characteristics or associations with other microbes (Reynolds et al., 2003). Our plants were exposed to other indigenous soil microbes in addition to the AMF treatments, yet still we found little evidence that different plant species grew better on different forms of N. Ability to use multiple forms and sources of N may be a generally advantageous strategy for a sessile organism in a temporally and spatially heterogenous world. It is also possible that partitioning manifests itself only under competitive conditions. We grew our plants singly in pots, and so perhaps we saw only the ‘fundamental N niche’ of these species, and the ‘realized N niches’ when grown in mixture would have been different. Recent work has shown, for example, that alpine plant species exhibit preferences for different forms of N in the presence vs absence of neighbors (A. Miller et al., National Park Service, unpublished data).
Although we did not detect N partitioning, we found that natives and exotics again behaved differently in response to treatments. The exotic species experienced some release from N limitation under all N treatments (although less so with chitin, the most recalcitrant N source), but the two native species did not. Yet tissue N data show that the natives were able to acquire more N from N treatments. These results are consistent with the greater growth reductions due to AMF for natives. That is, in S. lyrata and P. sphaerocarpon, gains in photosynthate from added N may have more often gone to AMF rather than to growth.