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Plant communities, and the ecosystems of which they are a part, face multiple global changes, including elevated atmospheric CO2 concentrations ([CO2]) and nitrogen (N) enrichment. An increasing number of multiple-factor global change experiments have been established to test whether community and ecosystem responses to one global change factor are modified by a second (Reich et al., 2006b; Templer & Reinmann, 2011). These multifactor experiments reveal that the effects of one global change on community and ecosystem processes, such as plant abundance and productivity, are often contingent (sensu Schmitz, 2010) on a second global change (Dukes et al., 2005; Reich et al., 2006a; Kardol et al., 2010; Morgan et al., 2011). Given the logistical and economic constraints, multifactor experiments are usually performed at two levels of a specific factor, such as at ambient vs elevated levels. However, many of these factors vary across time and space in their magnitude, and this variation can alter their effects on community and ecosystem processes (Phoenix et al., 2012). To investigate the effects of these varying magnitudes, single-factor but multilevel experiments have been established; these experiments reveal nonlinearities in community and ecosystem responses across increasing availabilities of global change factors, such as [CO2] and N enrichment (Gill et al., 2002; Granados & Körner, 2002; Bradford et al., 2008; Fay et al., 2009). Furthermore, meta-analyses suggest that the combined effects of these factors vary nonadditively across levels. For example, ecosystem responses to elevated [CO2] are highest in studies in which N addition rates are greatest (van Groenigen et al., 2006). To determine definitively whether these nonadditive interactions are level dependent requires multifactor, multilevel global change experiments.
Managed grasslands comprise 70% of agricultural lands, and global changes that influence their productivity by changing forage amount and quality will influence human food production (Soussana & Lüscher, 2007). Elevated [CO2] is expected to increase forage amount where N is not limited (e.g. through fertilization), but impacts on forage quality are more uncertain as they are dependent on foliar trait plasticity (e.g. N concentration), as well as plant community composition (e.g. legumes vs grasses; Soussana & Lüscher, 2007). Many of the C3 species from managed grasslands were introduced by European settlers to the New World and so are common on multiple continents, across different fertilizer managements and under widely varying levels of N deposition (Hunt et al., 1991, 1993). Although it is recognized that acute (e.g. fertilizer) vs chronic (e.g. atmospheric deposition) N enrichment is likely to influence community and ecosystem processes differently (Bernot & Dodds, 2005; Phoenix et al., 2012), direct experimental tests appear to be lacking. Further, in the context of multifactor global change experiments, it appears unknown whether the regime (i.e. acute vs chronic) of N amendment will modify interactive effects of N with elevated [CO2].
We assessed community (species dominance) and ecosystem (aboveground net primary productivity, ANPP) responses to multiple global change factors using model grassland ecosystems. We applied six levels of [CO2] across six levels of N enrichment, with the latter added following either a chronic or acute regime to simulate deposition or fertilization, respectively. Although chronic and acute regimes varied in the [N] per addition, we made 56 chronic vs four acute additions across the 28 wk of the experiment, ensuring that chronic and acute treatments of the same N enrichment level (e.g. 70 kg N ha−1 yr−1) received the same total amount of N. We expected a three-way interaction between [CO2] × N enrichment × N regime, where N enrichment would reduce N limitation and hence yield greater ANPP gains with increasing [CO2], which, in turn, would alleviate moisture limitations through improved water use efficiency and so give greater ANPP responses to N enrichment. We expected these gains to be greater with acute vs chronic N addition, because we reasoned that the higher concentrations at which the acute additions were added would permit the plants to acquire more of the added N for growth. This expectation was based on the fact that plants switch from high- to low-affinity N uptake kinetics at higher N concentrations, and low-affinity systems have much higher maximal rates (Kraiser et al., 2011). We also expected community composition to be determined by a three-way interaction, with productivity gains being mirrored by abundance increases in those species with higher relative growth rates (RGRs; representative of improved pastures and higher competitive ability) and decreases in those species with lower RGRs. We examined additional response variables (e.g. specific leaf area (SLA), foliar N concentration, root biomass) across a subset of the experimental ecosystems to attempt to explain the composition and productivity responses across the full experiment.
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For cumulative ANPP across the seven harvests, the best-fit model retained significant interaction terms for [CO2] × N rate (P = 0.0000) and N rate × N regime (P = 0.0014). The [CO2] × N rate interaction appeared to be a product of the fact that responses to increasing levels of [CO2] and N were generally greater when the other resource was in higher supply. For example, ANPP was c. 2.3 times greater at 700 ppm [CO2] when the N rate was 240 as opposed to 10 kg N ha−1 yr−1, but it less than doubled between the same N rates at 280 ppm [CO2] (Fig. 1). Equally, at 240 kg N ha−1 yr−1, ANPP increased by c. 1.4 times across 280–700 ppm [CO2], but by only c. 1.2 times across the same [CO2] levels for 10 kg N ha−1 yr−1 (Fig. 1). Also evident is that the shape of the ANPP response across [CO2] appeared to be strongly dependent on the N rate. For example, with 0 kg N ha−1 yr−1, maximum ANPP was recorded across 375–450 ppm, but, at 240 kg N ha−1 yr−1, the maximum occurred across 450–550 ppm, and, at 70 kg N ha−1 yr−1, the maximum ANPP occurred at 700 ppm (Fig. 1).
Figure 1. Interactive effects of atmospheric CO2 concentration and nitrogen (N) enrichment rate on aboveground net primary productivity (ANPP). Shown are the means ± 1SE (n = 10, data pooled across N regime: Table 1). Note that n = 5 for the 0 kg N ha−1 yr−1 values. To aid visual interpretation, mean values within an N enrichment level are connected by solid or dashed lines.
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The N rate × N regime interaction arose because ANPP under acute N amendment did not appear to differ from ANPP under chronic N amendment at the lowest N rates, but became progressively greater with increasing N rate (Fig. 2). At 240 kg N ha−1 yr−1, acute additions resulted in c. 1.15 times greater ANPP than chronic additions, giving a mean difference of 126 g m−2 (Fig. 2).
Figure 2. Interactive effects of nitrogen (N) enrichment rate and regime (chronic, closed circles; acute, open circles) on aboveground net primary productivity (ANPP). ANPP for plants watered but without N amendment is shown by the closed triangle. Shown are means ± 1SE (n = 30, data pooled across CO2 concentration: Table 1).
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In contrast with the ANPP responses, there were no interactions between the CO2 and N treatments on community dominance. However, as for ANPP, the community response was dependent on the N enrichment rate. The best-fit LMM only retained this factor and its positive coefficient (P = 0.0001) indicated the shifting biomass dominance from H. lanatus to A. odoratum with increasing N rate (Fig. 3).
Figure 3. Effect of nitrogen (N) enrichment rate on plant community composition, represented as the proportional biomass contribution to aboveground net primary productivity (ANPP) of the co-dominant species Anthoxanthum odoratum (closed circles) and Holcus lanatus (open circles). Shown are means ± 1SE (n = 60, data pooled across CO2 concentration and N regime: Table 1). Note that n = 30 for the 0 kg N ha−1 yr−1 values because data could only be pooled across CO2 concentration.
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We evaluated a suite of additional response variables on a subset of 60 microcosms to help to explain the ANPP and community composition responses across the full experiment. Before using these variables, we checked that ANPP (estimated for the subset as the standing aboveground biomass at the final harvest, permitting direct comparison with the other plant biomass variables measured for the subset) and community dominance responded similarly to the full experiment. This was true for ANPP, where there was a [CO2] × N rate interaction (P = 0.07), with ANPP responses to increasing [CO2] or N rate being greater when the level of the other resource was greater (Table 2). This same response appeared to be mirrored in the whole-community shoot and root C biomass values (Table 2). Perhaps facilitating these synergistic effects between [CO2] and N rate, [CO2] was a significant predictor of soil moisture (P = 0.0001), which, for example, increased steadily from 16% to 24% at the highest N rate as [CO2] increased from 280 to 700 ppm (Table 2). Equally, acidification of mineral soil pH with increasing N appeared to be muted by higher [CO2] ([CO2] × N rate interaction: P = 0.062). Other response variables appeared to be less directly tied to the [CO2] × N rate interaction on ANPP, but were typical for single factor [CO2] and N rate effects, such as increased (P = 0.0001) and reduced (P = 0.0001) shoot C : N ratios with increasing [CO2] and N, respectively (Table 2). Further, shoot N (P = 0.0001) and the proportion of 15N-labeled amendment acquired by the shoots (P = 0.0001) generally increased with N rate (Table 2). Root responses were more idiosyncratic: root C : N ratios appeared to be relatively unresponsive to [CO2] and N rate (P > 0.01), the proportion of 15N-labeled amendment acquired by roots was lower (P = 0.0001) at 240 vs 70 kg N ha−1 yr−1, and root mass N was dependent on [CO2] and N rate (interaction: P = 0.02). This interaction occurred because root mass N was unaffected by N rate at certain [CO2] (e.g. 375 ppm), but at others (e.g. 550 and 700 ppm) responded positively to at least the highest N level (Table 2).
Table 2. Atmospheric CO2 concentration and nitrogen (N) enrichment rate effects on soil and community variables for the subset of 60 microcosmsa
|[CO2] (ppm)||N rate (kg N ha−1 yr−1)||pHb||Moisture (%)c||ANPP (g m−2)d||Shoot mass C (g m−2)||Root mass C (g m−2)||Shoot mass N (g m−2)||Root mass N (g m−2)||Shoot C : N||Root C : N||Shoot 15N (%)e||Root 15N (%)|
|280|| 0||4.66 + 0.02||20 ± 0.3||339 ± 26||372 ± 15||820 ± 89||5.1 ± 0.2||15 ± 1.5||74 ± 3.2||56 ± 0.8||na||na|
| 70||4.55 + 0.01||20 ± 0.3||488 ± 46||434 ± 39||859 ± 33||5.7 ± 0.6||18 ± 1.6||77 ± 3.7||50 ± 4.1||39 ± 4.4||29 ± 3.3|
|240||4.40 + 0.03||16 ± 0.8||673 ± 32||632 ± 43||1104 ± 69||10.0 ± 0.6||22 ± 2.2||63 ± 3.0||51 ± 3.7||48 ± 6.4||27 ± 1.3|
|375|| 0||4.66 + 0.05||23 ± 1.4||445 ± 42||337 ± 16||719 ± 66||3.7 ± 0.3||15 ± 2.7||92 ± 2.8||49 ± 8.3||na||na|
| 70||4.57 + 0.03||22 ± 0.4||579 ± 32||434 ± 24||869 ± 41||5.2 ± 0.4||15 ± 0.9||84 ± 3.3||60 ± 2.9||36 ± 7.5||26 ± 1.1|
|240||4.48 + 0.04||21 ± 0.8||883 ± 87||692 ± 45||988 ± 66||10.9 ± 0.6||16 ± 1.4||64 ± 3.8||61 ± 1.5||46 ± 5.3||21 ± 2.1|
|550|| 0||4.75 + 0.02||22 ± 0.4||411 ± 6||401 ± 53||737 ± 111||3.7 ± 0.5||12 ± 2.7||109 ± 6.0||67 ± 5.3||na||na|
| 70||4.69 + 0.04||24 ± 0.4||654 ± 64||443 ± 13||871 ± 66||5.0 ± 0.1||17 ± 2.5||89 ± 3.2||54 ± 4.5||36 ± 4.9||28 ± 2.1|
|240||4.55 + 0.05||22 ± 0.9||1119 ± 50||675 ± 60||1187 ± 81||9.5 ± 0.5||24 ± 1.6||71 ± 4.9||51 ± 3.4||49 ± 6.6||25 ± 1.7|
|700|| 0||4.63 + 0.04||26 ± 3.7||342 ± 28||377 ± 26||893 ± 215||4.0 ± 0.2||18 ± 4.7||94 ± 4.1||52 ± 7.2||na||na|
| 70||4.66 + 0.04||24 ± 0.7||742 ± 44||504 ± 35||1008 ± 82||5.8 ± 0.6||19 ± 1.4||90 ± 7.7||52 ± 2.7||39 ± 5.6||30 ± 1.0|
|240||4.54 + 0.03||24 ± 0.5||947 ± 46||705 ± 49||1467 ± 67||9.2 ± 0.4||31 ± 2.8||76 ± 3.0||49 ± 4.0||41 ± 6.1||28 ± 0.9|
Unlike the [CO2] × N rate interaction, the N rate × N regime interaction for the subset did not mirror the full experiment as it was not significant (P = 0.13) and, indeed, there was little difference in standing ANPP between chronic and acute treatments (Table 3). The absence of a chronic vs acute difference was reflected in the majority of the other variables, with the only obvious effects being of N rate itself, which decreased pH (P = 0.0001), soil moisture (P = 0.03) and shoot C : N ratios (P = 0.0001), and increased masses of shoot and root C and N (P = 0.0001 in all four instances; Table 3). The only variable for which chronic vs acute treatments differed was the proportion of the 15N-labeled amendment recovered in the shoots; it was c. 1.7 times greater when added acutely (P = 0.0001), and this acute vs chronic difference was slightly larger at 240 vs 70 kg N ha−1 yr−1 (N rate × N regime interaction: P = 0.0001; Table 3).
Table 3. Nitrogen (N) enrichment rate and N regime effects on soil and community variables for the subset of 60 microcosmsa
|N regime||N rate (kg N ha−1 yr−1)||pHb||Moisture (%)c||ANPP (g m−2)d||Shoot mass C (g m−2)||Root mass C (g m−2)||Shoot mass N (g m−2)||Root mass N (g m−2)||Shoot C : N||Root C : N||Shoot 15N (%)e||Root 15N (%)|
|na|| 0||4.67 + 0.02||23 ± 1.0||384 ± 18||372 ± 15||792 ± 61||4.1 ± 0.2||15 ± 1.5||92 ± 4.2||56 ± 3.2||na||na|
|Chronic|| 70||4.59 + 0.03||22 ± 0.8||604 ± 29||489 ± 34||964 ± 66||5.9 ± 0.6||19 ± 1.5||86 ± 3.7||51 ± 2.2||29 ± 3.4||29 ± 1.6|
|Acute|| 70||4.63 + 0.03||23 ± 0.5||657 ± 48||433 ± 16||890 ± 32||5.3 ± 0.3||16 ± 1.6||83 ± 3.7||58 ± 2.4||48 ± 2.4||27 ± 1.2|
|Chronic||240||4.48 + 0.03||21 ± 1.1||904 ± 69||696 ± 35||1135 ± 64||9.6 ± 0.4||24 ± 2.4||73 ± 2.7||50 ± 3.0||34 ± 2.4||26 ± 1.5|
|Acute||240||4.50 + 0.04||20 ± 1.0||908 ± 53||657 ± 33||1238 ± 75||10.3 ± 0.4||23 ± 1.6||64 ± 2.6||56 ± 1.8||58 ± 1.5||25 ± 1.2|
For community dominance, the subset of 60 microcosms behaved almost identically to the full experiment, with increasing N rate shifting biomass dominance from H. lanatus to A. odoratum (P = 0.0002, Table 4). At higher N rates, both species had higher foliar %N (P = 0.0001 for both species), lower foliar C : N ratios (A. odoratum, P = 0.0001; H. lanatus, P = 0.03) and higher SLA (Table 4), although the increase was only significant for A. odoratum (A. odoratum, P = 0.06; H. lanatus, P = 0.22). A significant N rate × species interaction for foliar %N (P = 0.0032) and C : N (P = 0.0028) revealed that A. odoratum showed greater relative increases in these leaf traits: for example, its foliar %N increased c. 1.34 times from 0 to 240 kg N ha−1 yr−1, but only c. 1.18 times for H. lanatus (Table 4). For both species, SLA responded most between 0 and 70 kg N ha−1 yr−1, and foliar N (and C : N) most between 70 and 240 kg N ha−1 yr−1, revealing marked differences and nonlinearities in the response size of different leaf traits across N rate. In addition, although, on average, H. lanatus was still more abundant than A. odoratum at 70 kg N ha−1 yr−1, the latter acquired almost three times as much N from the 15N-labeled amendments, and even greater amounts at 240 kg N ha−1 yr−1 (N rate × species interaction: P < 0.05; Table 4).
Table 4. Effect of nitrogen (N) enrichment rate on soil, community and leaf variables of the co-dominant species Anthoxanthum odoratum (Ant) and Holcus lanatus (Hol) for the subset of 60 microcosmsa
|N rate (kg N ha−1 yr−1)||Spp.||pHb,c||Moisture (%)d||ANPP (g m−2)e||Comp. (%)f||Foliar [N] (%)||Foliar C : N||SLA (cm−2 g−1)||Foliar 15N (%)g|
|0||Ant||4.46 + 0.02||23 ± 1.0||384 ± 18||37 ± 3.2||0.96 ± 0.06||46 ± 2.2||79 ± 8.0|| na|
|70||Ant||4.34 + 0.02||23 ± 0.4||616 ± 30||44 ± 3.3||1.06 ± 0.03||42 ± 1.3||106 ± 8.1||5.8 ± 0.6|
|240||Ant||4.21 + 0.02||21 ± 0.7||906 ± 43||59 ± 3.1||1.29 ± 0.05||35 ± 1.4||109 ± 5.7||8.6 ± 0.9|
|0||Hol|| na|| na|| na||57 ± 3.2||0.97 ± 0.06||44 ± 2.3||117 ± 17.1|| na|
|70||Hol|| na|| na|| na||51 ± 3.1||1.01 ± 0.04||43 ± 1.9||134 ± 9.1||2.0 ± 0.4|
|240||Hol|| na|| na|| na||38 ± 3.1||1.14 ± 0.07||40 ± 2.4||139 ± 7.7||1.9 ± 0.5|
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Our hypothesis was that we would see a three-way interaction between [CO2], N rate and N regime, with greatest ANPP and shifting composition to dominance by the species with higher RGR (e.g. H. lanatus) at the highest [CO2] and N rate, when N was applied acutely. Our rationale for this hypothesis was that elevated [CO2] responses would be N limited, as is often observed with natural soils (Reich et al., 2006a,b; Leakey et al., 2009; Drake et al., 2011; Luo et al., 2011). Hence, we expected N enrichment to remove this limitation, favoring species of improved pastures, and that higher soil solution N concentrations from acute additions would make N most available to the plants (Kraiser et al., 2011). Greater ANPP with higher N rates, especially when applied acutely (Fig. 2), and greater relative increases in productivity with increasing [CO2] at higher N rates (Fig. 1), seemed to confirm these expected mechanisms. However, ecosystem responses to the global change factors were simpler than projected, with only two-way interactions between [CO2] and N rate, and N rate and N regime (Figs 1, 2). Further, the community dominance only responded to N rate and, in contrast with our expectations, by shifting dominance to a species with lower RGR and more typical of less improved pasture (Fig. 3).
Multifactor global change experiments are still rare, especially longer term experiments (Dukes et al., 2005; Reich et al., 2006b; Dawes et al., 2011; Luo et al., 2011), and multifactor, multilevel experiments are rarer still (Lewis et al., 2010). Given the cost and logistics of such experiments, one proposed solution is to use mechanistic insights from single-factor experiments to construct models that project ecosystem response and feedback to global environmental change (Ostle et al., 2009). Our observations contribute empirical data highlighting the limitation of this approach, where combinations of factors yield ecological responses that may not be predicted from single-factor responses (Luo et al., 2011). In our case, the surprise was less contingency than anticipated, where, although we observed N limitation of the [CO2] response, and higher productivity with acute vs chronic N, we did not observe a three-way interaction across our factors on the ecosystem or community variables. Perhaps, most importantly, our data support the idea that both the level and regime of applied N matters in determining the system response (Phoenix et al., 2012). Meta-analyses reveal that the effects of elevated [CO2] on productivity may be greater under higher vs lower N addition (van Groenigen et al., 2006) and we provide direct empirical support for this (Fig. 1). Further, many experimental studies add N at concentrations and frequencies not representative of deposition (Phoenix et al., 2012), and we show that productivity responses differ with acute vs chronic N addition, especially at the higher addition rates typical of many experiments (Neff et al., 2002; Mack et al., 2004; Bradford et al., 2008). Lastly, low rates of N addition, whether chronic or acute, are rarely investigated (Phoenix et al., 2012), but our data suggest that low rates still drive community change (Fig. 3).
We find general support for the idea that ANPP responses to [CO2] are N limited, whether by initial N availability or progressive N limitation (PNL; Reich et al., 2006b; Luo et al., 2011). The phenomenon of PNL might explain why productivity under 0 kg N ha−1 yr−1 peaked at ambient [CO2] (Fig. 1), especially given the pronounced increase in shoot C : N at higher [CO2] (Table 2). Such an increase might feed back to bind N in organic detrital forms that are less available for plant growth. Higher soil moistures with higher [CO2] (Table 2) are consistent with the expectation that C3 plants have higher water use efficiencies under elevated [CO2] (Morgan et al., 2011); this reduced moisture limitation (despite higher ANPP), combined with reduced N limitation through N addition, probably explained why the highest productivity was observed with higher [CO2] and N rate (Fig. 1). This interaction may also have been facilitated by the fact that N addition was less acidifying at higher vs lower [CO2] concentrations (Table 2), where acidification can decrease plant performance given the increased mobility of toxic ions in soils (Phoenix et al., 2012). We cannot uncouple the relative contributions of moisture, N and H+ availabilities in determining ANPP responses to [CO2] × N rate, but our data do suggest that the interaction is probably driven by more than just N limitation.
Our data confirm the expectation that the productivity of fertilized grasslands will increase with rising atmospheric [CO2] (van Kessel et al., 2000; Soussana & Lüscher, 2007). However, such an increase is accompanied by declines in forage quality (Table 2; shoot data), and these declines in quality are expected to alter biotic interactions and foodweb structure aboveground (Reich et al., 2006a; Tylianakis et al., 2008; Antoninka et al., 2009). Notably, root responses to [CO2] × N rate were idiosyncratic across traits and factor levels, with changes in C : N ratios and total N amount at some [CO2] but not others, and lower acquisition of the 15N tracer under higher vs lower N rates (Table 2). These differences between the foliar and root responses to our global change treatments question whether aboveground responses to global changes are a suitable proxy for belowground responses. If they are not a suitable proxy, then to understand the consequences of multiple global changes for root–herbivore-based foodwebs requires that we redress the paucity of investigations into root mass and stoichiometric responses to [CO2] and N enrichment (Antoninka et al., 2009; Anderson et al., 2010).
Data from the subset of 60 microcosms added little to our understanding of what drives the N rate × N regime interaction on ANPP (Fig. 2). The N rate showed large and expected consequences on variables such as ANPP, shoot C : N and pH. However, unlike ANPP in the full experiment, N regime had little influence on ANPP in the subset and, indeed, little influence on shoot and root N dynamics, pH and moisture (Table 3). The lack of effect on ANPP may have been a result of reduced statistical power (as our n dropped from five to three per treatment) and/or because ANPP for the subset of microcosms was based on the standing plant biomass at the final experimental harvest, but calculated across harvests 2–8 for the full experiment from the biomass clipped 6 cm above the soil height. The only variable influenced by the N regime was the amount of fertilizer acquired by the shoots, estimated through the percentage recovery of the 15N tracer, which was higher under acute N (Table 3). This greater acquisition under the acute regime might explain why ANPP responses to acute additions in the full experiment increasingly diverged from chronic additions as the N rate increased (Fig. 2). The most plausible mechanism to explain this divergence is that plants switch from high- to low-affinity uptake kinetics as soil solution and concentrations increase (Kraiser et al., 2011). Low-affinity kinetics are associated with first-order and much higher rates of N uptake, meaning that plants should acquire N for growth more rapidly when soil solution N concentrations are higher. However, other than ANPP, our data suggest that many soil and plant variables are relatively insensitive to the N regime (Table 3). Therefore, although the single to few large doses of N applied in many field studies may poorly represent N deposition to ecosystems (Phoenix et al., 2012), they may still permit valid inferences about N deposition effects on a range of response variables. To test this possibility robustly will require large-scale field manipulations that investigate directly N regime effects.
Lawton (1999) infamously argued that community ecology is a mess, opining that contingency in local processes makes community responses to perturbations overwhelmingly complicated, and hence unpredictable. In reply to Lawton, emerging syntheses in community ecology (Vellend, 2010) outline frameworks for making general statements about fundamental processes that underlie community dynamics at local scales, but remain pessimistic about our ability to predict how particular processes shape a specific community. However, we observed less ‘mess’ in our community than in ecosystem data, with the community response only driven by N rate and not contingent on [CO2] or N regime (Fig. 3). The greater phenotypic plasticity across N rate in A. odoratum leaf traits, associated with light interception (SLA) and photosynthetic rates (%N), provides the most parsimonious explanation for why it increased in relative abundance at the expense of H. lanatus (Fig. 3 and Table 4). Such a clear mechanistic interpretation was not obvious for the ecosystem data, but, as in the ANPP response, the importance of the magnitude of a global change factor in determining its relative effect size is evident. For example, the most abrupt shift in community dominance was between 10 and 35 kg N ha−1 yr−1 (Fig. 3), showing not only that low additional N loadings associated with N deposition can influence community processes (Phoenix et al., 2012), but also that they can have the greatest relative effects, making even ‘clean’ systems vulnerable to N deposition. Leaf traits also exhibited different sensitivities across different N levels, with SLA increasing most from 0 to 70 kg N ha−1 yr−1, and foliar %N from 70 to 240 kg N ha−1 yr−1 (Table 4). These results demonstrate that interpretations of functional leaf trait responses to a global change factor might need to carefully consider the level at which the factor is applied.
We used a multispecies plant community and field soils, which presumably included beneficial and pathogenic soil organisms, but our work was still shorter term (6 months) and applied factors in a stepwise increase. In longer term studies, shifts in plant community composition can eventually negate positive effects of a factor, such as [CO2], on productivity (Langley & Megonigal, 2010), and incremental or continuous increases in [CO2] have the potential to yield very different productivity and community consequences compared with stepwise increases (Luo & Reynolds, 1999; Klironomos et al., 2005). Nevertheless, none of the longer term field studies have assessed grassland community responses to [CO2] × N rate at more than two levels, nor have they looked at the N regime. Yet, we demonstrate that level and regime can alter ecosystem and community responses, making the magnitude and shape of field responses uncertain across space and time as the amount and nature of the factors vary.
Notably, as observed in [CO2] gradient studies (Gill et al., 2002; Fay et al., 2009), we saw some of the strongest responses to [CO2] between sub- to ambient concentrations and, at some N rates, a decline in ANPP between 550 and 700 ppm [CO2] (Fig. 1). Such nonlinearities and even declines at [CO2] around 700 ppm have been highlighted (Berntson & Bazzaz, 1998; Granados & Körner, 2002), but are not represented in the modeling efforts to project vegetation responses to rising atmospheric [CO2] (Ostle et al., 2009). There is a clear need for multifactor, multilevel field studies to permit us to assess uncertainty in model projections, but the required number of experimental treatments and replicates (66 and 330, respectively, in our study) may preclude their establishment given the logistics and cost to set up and maintain so many plots. If their establishment is unlikely, data from laboratory experiments, such as ours, should be used in efforts to quantify certainty bounds for model projections of global change effects on communities and ecosystems.
The paucity of two-level (ambient and elevated), multifactor experiments makes the prediction of future responses to global environmental change highly uncertain (Tylianakis et al., 2008; Ostle et al., 2009). This uncertainty is magnified further by the lack of multifactor, multilevel experiments, because the outcome of interacting global changes on communities and ecosystem processes may be dependent on the amount of the factor (van Groenigen et al., 2006). We find direct experimental support for this possibility, showing that amount matters in determining interactive and main effects of factors, and that nonadditivity and nonlinearity in ecosystem responses are likely to be the norm rather than the exception. The call has been made for more long-term, multifactor field experiments to clarify how global environmental changes will influence terrestrial systems (Luo et al., 2011). Our results suggest that this clarity will only be achieved if these long-term studies include multiple levels of the global change factors.