Increases in nitrogen (N) deposition can greatly stimulate
ecosystem net carbon (C) sequestration through positive N-induced effects on
plant productivity. However, how net ecosystem CO
Anthropogenic reactive nitrogen (N) inputs to the terrestrial biosphere has increased more than 3-fold over the past century and is predicted to increase further (Lamarque et al., 2005; Galloway et al., 2008). Because of the strong coupling of ecosystem carbon (C) and N cycles, excess N deposition could have significant impacts on the ecosystem C cycle (LeBauer and Treseder, 2008; Liu and Greaver, 2010; Lu et al., 2011). Ecosystem net C sequestration is predicted to increase or have no significant change under rising N deposition (Nadelhoffer et al., 1999; Magnani et al., 2007; Reay et al., 2008; Niu et al., 2010; Lu et al., 2011; Fernandez-Martinez et al., 2014). However, we have a limited understanding of the dynamic N responses of C sequestration in terrestrial ecosystems, which is crucial for model projection of the future terrestrial C cycle under rising N deposition (Reay et al., 2008).
Although N addition generally enhances plant growth and ecosystem net
primary productivity (NPP) based on global syntheses of N addition
experiments (LeBauer and Treseder, 2008; Xia and Wan, 2008; Lu et al.,
2011), the responses of ecosystem C fluxes vary with N loading rates (Liu
and Greaver, 2010; Lu et al., 2011). According to the N saturation
hypothesis, NPP is assumed to slowly increase with N addition rates first,
then reach its maximum value at the N saturation point, and finally decline with
further increase in N input (Aber et al., 1989; Lovett and Goodale, 2011).
During this process, NPP shifts from an N-limited, an N intermediate, to an N
saturation stage as N deposition increases. Similarly, net ecosystem
CO
Moreover, ER can be divided into aboveground plant respiration, belowground plant respiration (root respiration), and soil microbial respiration. These components of ER could be affected by plant aboveground biomass, root biomass, soil organic matter, and microbial biomass C, which may respond variously to N addition (Phillips and Fahey, 2007; Hasselquist et al., 2012). For example, root respiration would be enhanced or not significantly changed under N addition, while soil microbial respiration may be suppressed by N addition (Zhou et al., 2014). The different responses of various components of ER to N addition will also consequently change the response of NEE. Nevertheless, there is limited knowledge on how various components of NEE respond differentially to N addition gradient. In addition, the N responses of ecosystem C fluxes may shift over time because of changes in plant community structure and other limiting factors (Niu et al., 2010). It is not clear when and how ecosystem C fluxes get N saturated under increasing N input. The mechanisms underlying the saturation response of C fluxes are even far from clear, which prevent us from accurately predicting the C cycle in response to rising N deposition.
In this study, we explored the responses of various ecosystem C cycle
processes to an N addition gradient in an alpine meadow on the Qinghai–Tibetan
Plateau. The Qinghai–Tibetan Plateau has an area of 2.5 million km
The study site is located in an alpine meadow in Hongyuan County, Sichuan
Province, China, which is on the eastern Qinghai–Tibetan Plateau
(32
We conducted an N addition experiment with six levels of N addition rate (0,
2, 4, 8, 16, 32 gN m
Ecosystem C fluxes were measured using a transparent static chamber
(0.5
Soil respiration (SR) was assessed following the measurement of NEE and ER.
It was also measured with LI-6400XT attaching a soil CO
Soil samples were collected from the topsoil (0–10 cm) of the 30 plots on
15 August 2014 and 14 August 2015. Two soil cores (8 cm in diameter and
10 cm in depth) were taken at least 1 m from the edge in each plot, and
then completely mixed to get a composite sample. The soil samples were sieved
by a 2 mm mesh and then were air-dried for chemical analysis. Soil pH was
determined with a glass electrode in a
Repeated-measures ANOVA was used to examine N addition effects on
each ecosystem C flux over the growing season in 2014 and 2015. When we
evaluate N addition effects on the different components of ER and their
proportions, we averaged their values across the year and then used one-way
ANOVA to test the differences among treatments. To test the response pattern
of ecosystem C cycle properties to the N addition gradient, we fitted the
response parameter to linear or quadratic functions which had higher
Seasonal dynamics of net ecosystem CO
Relationships between N addition rate and net ecosystem CO
Results (
NEE varied throughout the growing seasons
in both 2014 and 2015. The maximum rates of net CO
Comparisons of the Akaike information criterion (AIC) among functions
describing the relationships between NEE, ER, GEP, SR, and
The N addition gradient had significant effects on ER (
Relationships between N addition rate and soil respiration (SR)
Plant respiration and its components in response to the N addition
gradient in 2014 and 2015 (mean
We divided ER into
In addition, the proportions of different efflux components to ER differed in
response to the N addition gradient between years (Fig. 5). The proportions
of
In order to examine the causes of the N saturation responses of NEE and ER
in 2015, we examined the relationship between ER and its various components
and also NEE. The results showed that ER had a significantly positive
correlation with
Our results showed that initial ecosystem C fluxes (NEE, ER, and GEP) in 2014 suggested ecosystem N limitation, whereas in 2015 these C fluxes clearly suggested N saturation under high N addition rates. These findings not only extend the N saturation hypothesis for the response of NPP to N addition (Aber et al., 1998, 1989; Lovett and Goodale, 2011) but also provide comprehensive evidence of potential relationships between various ecosystem C fluxes and ecosystem N dynamics. Previous N addition studies used only one level of N addition and found that NEE showed a positive (Niu et al., 2010; Huff et al., 2015) or no significant response (Harpole et al., 2007; Bubier et al., 2007) to N addition. Using one level of N addition only might not be enough to capture or quantify complex ecosystem responses to N addition. By using an N addition gradient experiment, this study comprehensively showed the saturation responses of NEE and its components to different N loading rates.
The contributions of different source components to ecosystem
respiration (ER) in response to the N addition gradient in 2014 and 2015
(mean
The N saturation response of NEE in 2015 was mainly attributed to the saturation responses of ER and GEP (Fig. 2), while the N saturation response of ER was likely caused by the saturation response of aboveground plant respiration and decreasing soil microbial respiration along the N addition gradient. The decrease in aboveground plant respiration under N32 treatment was primarily due to N addition stimulating plant growth and thus standing litter accumulation after plant senescence (Supplement Figs. S1–S2). In 2014, plant aboveground biomass (AGB) was stimulated under the high N addition treatment, especially AGB of grasses (Fig. S2). In this grassland, grasses usually have higher height than other plants. The accumulation of grasses' standing litter under the N32 treatment limited light conditions for other plants and negatively influenced plant growth in the early growing season in 2015. Therefore, GEP and NEE did not keep increasing at the highest N addition rate, leading to N saturation response. The N-induced light limitation for plant growth was also observed in other ecosystems, like temperate grassland (Niu et al., 2010; Kim and Henry, 2013). Moreover, our results showed that most components of ER had a similar response patterns between the 2 years except for soil microbial respiration, which increased in 2014 but decreased in 2015 along with N addition rates. Thus, we propose that soil microbial respiration might play a key role in mediating the N saturation effects for ER and thus NEE, which is not reported in previous studies. The decline in microbial respiration under high N addition conditions was primarily due to the N-induced reduction in soil pH (Fig. 7). Although many factors can influence soil microbial respiration, such as soil N availability and microbial community structure (Janssens et al., 2010), previous studies with a similar N addition gradient suggested that soil pH was the most important driver for responses of microbes under high N addition rates (Liu et al., 2014; Song et al., 2014; Chen et al., 2016). N addition can lead to soil acidification and have negative impacts on soil microbial growth and activities (Liu et al., 2014; Tian et al., 2016). In this study, the decreased soil pH may cause toxicity effects on microbial activity (Treseder, 2008; Zhou et al., 2012) and thus reduces microbial respiration after 2 years of N addition.
Relationships between aboveground plant respiration
(
N-induced changes in soil pH (pH)
Our findings demonstrate that N responses of ecosystem C fluxes shifted from a linear response to a saturation response over the 2 years of treatments (Fig. 2). A recent study revealed that ecosystem C fluxes exhibited saturating responses to N addition during two consecutive measurement years in a temperate grassland (Tian et al., 2016). However, their measurement was conducted after 10 years of N addition treatments (similar N addition rates to our study), so it did not capture the early response signals of ecosystem C exchange. Results of another N addition gradient experiment carried out in three marsh ecosystems showed that aboveground plant biomass increased linearly with N addition rates after 7 months of treatment but showed saturating responses after 14 months of N addition (Vivanco et al., 2015). Taken together with our results, this suggests that N saturation of ecosystem C fluxes might happen within a couple years of N input. The different responses between years in this study are not likely due to climate differences because temperature and precipitation were not significantly different between 2014 and 2015. We acknowledge that our findings are only based on the short-term study, while a long-time experiment may capture more robust patterns in N saturation and the underlying mechanisms, but the findings of the initial shift in N responses are helpful to better understand the dynamics of the ecosystem in response to external N input.
The N saturation threshold for ecosystem C fluxes of this alpine meadow is
approximately 8 gN m
The components of ER showed diverse responses to the N addition gradient (Figs. 4, 5). For example, in 2014, aboveground plant respiration and its proportion to ER increased, but belowground plant respiration and its proportion to ER decreased with N addition amounts (Figs. 4b, c, 5a, b). To our knowledge, no previous study examined the different components of ER in response to the N addition gradient. Some studies conducted in alpine grassland demonstrated that N addition had no significant effects on ER (Jiang et al., 2013; Gong et al., 2014), since aboveground biomass did not respond to N addition in their studies. In this study, compared to the control treatment (without N addition), greater plant growth and aboveground biomass under N addition enhanced aboveground plant respiration and thus stimulated ER. The lack of N effect on SR in 2014 may be attributed to the counteractive responses of soil microbial respiration and root respiration to N addition. In the first year, N addition ameliorated the nutrient limitation for microbes; thus, soil microbial activity and biomass increased in the short term (Treseder, 2008) and subsequently stimulated microbial respiration (Peng et al., 2011). On the other hand, N addition could reduce belowground biomass allocation (Haynes and Gower, 1995), leading to a decrease in root respiration. The increase in soil microbial respiration partly offsets the decrease in root respiration. As a result, SR had no significant difference among N treatments in the first year. However, in the second year, soil microbial respiration declined under high N addition levels, in combination with the low root respiration, resulting in decreases in SR under N16 and N32 treatments. This decrease in SR was also observed in other ecosystems under long-term or high levels of N addition (Yan et al., 2010; Zhou and Zhang, 2014; Maaroufi et al., 2015). We are fully aware that there are some limitations for the partitioning technique, in which we used deep versus shallow collars to partition root from microbial respiration. This approach cuts roots and excludes the effects of changes in plant C allocation on microbial respiration. Soil moisture content may also change in the deep collars, which likely affects microbial respiration. However, this method has the advantage of exploring mechanisms of microbial responses in the absence of plant effects; it is a common and useful technique to partition the components of ER and is widely used in previous studies (Wan et al., 2005; Zhou et al., 2007).
Based on a field N addition gradient experiment, this study tested N saturation theory against multiple C cycle processes. We found that the ecosystem C fluxes of NEE, GEP, and ER shifted from linear responses to saturation responses over 2 years of N addition. The saturation responses of NEE and ER were mainly caused by the N-induced saturation response of aboveground plant respiration and decreasing soil microbial respiration along the N addition gradient. N-induced reduction in soil pH was the main mechanism underlying declines in microbial respiration under high N addition. We also revealed that various components of ER, including aboveground plant respiration, soil respiration, root respiration, and microbial respiration, responded differentially to the N addition gradient. The findings suggest that the C cycle processes have differential responses to N addition between aboveground and belowground plant parts and between plants and microbes. Our findings provide experimental evidence for the dynamic N responses of the ecosystem C cycle, which is helpful for parameterizing biogeochemical models and guiding ecosystem management in light of future increasing N deposition.
No data sets were used in this article.
The authors declare that they have no conflict of interest.
The authors thank Xiaojing Qin, Yanfang Li, Fangyue Zhang, Quan Quan, Zheng Fu, Qingxiao Yang, and Xiaoqiong Huang for their help in field measurement. We thank the staff of Institute of Qinghai–Tibetan Plateau at Southwest University for Nationalities. This study was financially supported by the National Natural Science Foundation of China (31625006, 31470528), the Ministry of Science and Technology of China (2016YFC0501803, 2013CB956300), the “Thousand Youth Talents Plan”, and the West Light Foundation of the Chinese Academy of Sciences. Edited by: Michael Bahn Reviewed by: three anonymous referees