Introduction

Soil respiration plays a critical role in controlling carbon (C) balance in terrestrial ecosystems (Raich and Schlesinger 1992). A small change in soil respiration has the potential to alter the buildup of CO2 in the atmosphere, providing feedbacks to climate change (Davidson and Janssens 2006). Consequently, it is imperative to accurately estimate soil respiration as affected by environmental changes.

Soil respiration (R S) includes autotrophic respiration generated by living roots and that by the associated rhizosphere microbes (R A) and heterotrophic respiration produced by microbial decomposition of soil organic matter (R H) (Subke et al. 2006). However, the relative contribution of these two processes is poorly known, despite its importance to evaluate C sequestration in forest ecosystems, especially under climate change, and calculate forest net ecosystem productivity (NEP) by the difference between net primary productivity and R H (Chapin et al. 2002; Bond-Lamberty et al. 2004). In addition, R A mainly depends on photosynthesis, root biomass, and activity (Högberg et al. 2001; Tang et al. 2009), while R H depends on soil properties and microbial activity (Whitaker et al. 2014; Wei et al. 2015). For these reasons, R A and R H may respond differently to environmental variables, vegetation types, and climate change (Wang et al. 2014; Yan et al. 2015).

Reforestation is a key land management practice across the world, and how this land use change affects the C budget is still unclear (Arora and Boer 2010). This understanding can provide insight into the C balance in soil. The vegetation changes may alter the aboveground photosynthetic supply to belowground and soil physicochemical and microbiological properties, which inevitably affect microbial activity and soil respiration (Lü et al. 2015; Chodak et al. 2016; Guo et al. 2016). Indeed, soil respiration was lower in woodlands than in grasslands due to the lower photosynthate input to belowground (Raich and Tufekcioglu 2000) or to the vegetation-mediated reduction in soil temperature under the canopy (Smith and Johnson 2004). Grasslands showed lower soil CO2 efflux than the woodlands due to plant-mediated changes in soil chemical properties (Jenkins and Adams 2010; Wang et al. 2013).

Due to intensive anthropogenic disturbances, most of forests in southern China were degraded into grassland or pastures (Ren et al. 2007b). This promoted reforestation of several areas in the past 30 years to increase forest cover and improve ecosystem functions and services (SFA 2005). Changes in plant species, soil nutrient conditions, and microbial properties during reforestation have been studied (Ren et al. 2007a; Wu et al. 2013), and these changes are expected to influence soil respiration and the relative sources. The influence of land use changes on C dynamics in tropical ecosystems is expected to be equal or more than that due to climate change in the coming decades (Espírito-Santo et al. 2014; Cavaleri et al. 2015). Therefore, investigations of soil respiration and the relative sources in response to reforestation in subtropics may be important for better predicting the response to future climate changes.

Here, the trenching method was used to partition soil respiration into R A and R H in a natural shrubland and three common plantations in southern China. Soil respiration is affected by soil temperature and moisture, and at the regional scale, it also depends on soil pH, substrate availability (soil C, nitrogen (N), and phosphorus (P)), biomass, and composition of soil microbial communities and root (Shi and Jin 2016). For example, the extreme soil pH values can stress and kill some bacterial species (Wang et al. 2015), and the soil C/N ratio can affect the microbial decomposition of plant residues (Shi and Jin 2016). The increase in soil P availability can also stimulate soil respiration in subtropics (Liu et al. 2012). Thus, biotic and abiotic factors, such as soil temperature and moisture; soil pH; contents of total C, N, and P and available P; biomass and composition of microbial community; and fine root biomass, were also determined in order to get insights in the variations of soil respiration and the relative sources among the four stands. The aims of this study were to (1) study the effects of reforestation on both R A and R H and (2) identify the abiotic and biotic factors responsible for the variations in soil respiration types after reforestation.

Materials and methods

Site description

This study was carried out at the Heshan National Field Research Station of Forest Ecosystem, Chinese Academy of Sciences (112° 54′ E, 22° 41′ N), which is located at Heshan City, Guangdong Province. This station has a typical subtropical monsoon humid climate with a wet season (from March to September) and a dry season (from October to February). The mean annual temperature is 21.7 °C. The mean annual precipitation ranges from 1460 to 1892 mm. Soils are classified as acrisol (IUSS Working Group WRB 2015).

In 1984, experimental plantations were established at this station on a homogenous degraded hilly land in order to restore the ecosystem services and functions. The topography, initial soil properties, and initial vegetation composition were similar across this hilly land (Wang et al. 2010). The main initial soil properties at 0–10 cm were 4.03 for soil pH, 15.3 g kg−1 for soil organic C, 1.23 g kg−1 for total N, 0.55 g kg−1 for total P, and 10 mg kg−1 for available P (Li et al. 2002). The dominant species before plantation establishment were grass, including Ischaemum indicum, Eriachne pallescens, and Baeckea frutescens. A degraded land with an area of 3.5 ha was left unplanted so that it naturally succeeded to the shrubland (SL) stage. The plantations selected in this study included a 3.17-ha mixed-conifer plantation (CP), a 3.99-ha mixed-legume plantation (LP), and a 2.68-ha mixed-native plantation (NP) species. All the trees in the three plantations were planted with a spacing of 2.5 m × 2.5 m. All the plantations and the shrubland were not subjected to any anthropogenic disturbance since 1984 (Wang et al. 2010). Further information of the selected shrubland and plantations is reported in Table 1.

Table 1 Characteristics of the four vegetation types in subtropical China (data from Wang et al. 2010, 2011a)
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Root exclusion treatments and soil CO2 efflux

Six trenched plots and six control plots were randomly laid out at each site. The size of each plot was 0.6 m × 0.6 m (Lee et al. 2003). For each trenched plot, we dug a trench (0.2 m wide and 0.6 m deep) belowground (deeper than the bottom of the root zone) using a steel knife and shovel in May 2011. The trench walls were lined with four sheets of polyethylene nets with a mesh size of 37 μm to prevent root entrance and allow soil water flowing laterally. The trenches were then refilled with the soil according to its original soil profiles to minimize disturbance. All the trenched plots were kept free of vegetation during the experiment.

One PVC soil collar with 20 cm diameter and 15 cm height was inserted into the soil 3 cm of depth in each plot and remained fixed throughout the experiment. Soil CO2 efflux in all plots at the four stands was measured twice a month in the wet season (from March to September) and monthly in the dry season (from October to February) using a Li-8100 Automated Soil CO2 Flux System (Li-Cor, Lincoln, NE, USA) from November 2011 to October 2012. The soil CO2 efflux of the trenched plots was assumed to be heterotrophic respiration (R H) because living roots in the trenched plots were assumed to be absent 6 months after trenching (Wang et al. 2011b). The autotrophic respiration (R A) was quantified by the difference between soil CO2 efflux from the control minus that of the trenched plots. The soil CO2 efflux was measured between 9:00 a.m. and noon because the obtained values approximate to the daily mean of the region (Tang et al. 2006). Soil temperature and moisture were recorded simultaneously when measuring soil CO2 efflux. Soil temperature (°C) at 5 cm depth was measured by a digital thermometer (CEM DT-131). The volumetric soil moisture (cm3 H2O cm−3 soil, %) at 5 cm depth was recorded by a MPKit including three amplitude domain reflectometry (ADR) moisture probes (MP406) and a data logger (MPM160 meter).

Field sampling and laboratory analysis

Soil samples were collected from 0 to 10 cm depth (A horizon) in the trenched plots and the controls in August 2011. Three soil cores (2.5 cm diameter) were randomly taken and then combined into one sample. Soil samples were sieved (<2 mm) and stored at 4 °C before microbial biomass C (MBC) determination. A subsample of soils was air-dried before being analyzed for pH, soil organic C, total N, total P, and available P content.

Soil pH was measured in a soil suspension with deionized water at a ratio of 25 mL to 10 g soil. Soil organic C was determined by the dichromate oxidation method (Nelson and Sommers 1996). Total N and total P were analyzed by the micro-Kjeldahl digestion procedure (Bremmer and Mulvaney 1982). Available P was extracted with 0.03 mol L−1 NH4F–0.025 mol L−1 HCl and determined colorimetrically (Bray and Kurtz 1945).

Soil microbial biomass C was analyzed by the fumigation-extraction method (Vance et al. 1987). The composition of soil microbial community was determined by phospholipid fatty acid (PLFA) analysis as described by Bossio and Scow (1998). The bacterial PLFA ratios were 14:0, 15:0, and 17:0; those of gram-negative bacteria (G− bacteria) were 16:1ω7c, 15:0 3OH, cy17:0, 16:1 2OH, 18:1ω7c, and cy19:0ω8c; and those of gram-positive bacteria (G+ bacteria) were i14:0, i15:0, a15:0, i16:0, i17:0, and a17:0 (Frostegård and Bååth 1996). The fungal PLFA biomarkers were represented by 18:2ω6,9 and 18:1ω9c (Kaiser et al. 2010; Frostegård et al. 2011).

The fine root biomass was determined in October 2011 using a sequential soil coring method (Persson 1978). Five soil cores were taken from each of eight randomly selected subplots at each site using a steel corer (10 cm in diameter, 20 cm in depth). All sample cores of each subplot were combined and carefully sieved (<2 mm) to separate the fine roots (<2 mm). The root samples were oven-dried at 60 °C for 72 h and weighed.

Data analysis

Repeated measures ANOVA was used to examine the differences in soil CO2 efflux, soil temperature, and soil moisture between trenched and the respective control and among the four stands through the experimental period. Two-way ANOVA was used to determine the difference in soil chemical properties and in biomass and composition of microbial community using four stands and two treatments (trenched and the control) as the main factors. The difference in root biomass among the four stands was tested by one-way ANOVA. Tukey’s multiple comparison test was further conducted to separate differences among means. The relationship between soil CO2 efflux and temperature was fitted by an exponential function, and the relationship between soil CO2 efflux and soil moisture followed a linear regression model (Tang et al. 2006). Pearson correlation coefficients were calculated to examine the relationship between the R S (or R H) and the relevant soil variables. Differences were considered significant at P < 0.05. All data analyses were carried out using the statistical package R (R Core Team 2014).

Results

Soil respiration and its sources

The R S at the four stands was relatively higher from April to October than that in the other months (Fig. 1). The R S of soils under the mixed-legume and the mixed-native species was significantly greater than that of the mixed-conifer and the shrubland soils (P < 0.001). The annual R S was 727 g C m−2 year−1 in the shrubland soil, 666 g C m−2 year−1 in the mixed-conifer soil, 1086 g C m−2 year−1 in the mixed-legume soil, and 1075 g C m−2 year−1 in the mixed-native species soil.

Fig. 1
figure 1

a–d Monthly mean soil total respiration (R S), heterotrophic respiration (R H), and autotrophic respiration (R A) in the four vegetation types in subtropics from November 2011 to October 2012. Error bars indicate standard errors (n = 6)

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Soil respiration was significantly decreased by trenching throughout the experiment period (P = 0.002). The temporal pattern of R H over the year was similar to that of R S (Fig. 1). The mean R H of soils under the mixed-legume (2.24 μmol CO2 m−2 s−1) and the mixed-native (2.04 μmol CO2 m−2 s−1) species was significantly higher than those of the shrubland (1.54 μmol CO2 m−2 s−1) and the mixed-conifer (1.41 μmol CO2 m−2 s−1) soils (P < 0.001 for all).

The soil R A peaked in May or June in the shrubland, the mixed-conifer, and the mixed-legume soils (Fig. 1). In the mixed-native species, soil R A was relatively higher in August and September than that in the other months. The soil under the mixed-native species showed significantly higher R A than that in the shrubland (P = 0.007) and the mixed-conifer (P = 0.004) soils. There was not a clear seasonal pattern of the percentage of R A/R S of each stand (Fig. 2). This percentage was significantly greater in soil under the mixed-native species than that in the shrubland soil (P = 0.046).

Fig. 2
figure 2

Monthly mean percentage of total respiration respired as autotrophic respiration (R A/R S) in soils under the four vegetation types in subtropics. SL shrubland, CP mixed-conifer, LP mixed-legume, NP mixed-native species

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Soil temperature and moisture

Soil temperature was relatively higher from April to November than that in the other months. Reforestation did not significantly influence soil temperature. Soil moisture was relatively greater from March to August than that in the other months (Fig. 3). The mixed-native species and the shrubland exhibited significantly higher soil moisture than the other two plantations (P < 0.010). There were no significant differences in soil temperature between the trenching and controls at the four stands.

Fig. 3
figure 3

Monthly mean soil temperature and moisture (both at 5 cm depth) in the controls (a, c) and trenched plots (b, d) in the four vegetation types in subtropics from November 2011 to October 2012. Error bars indicate standard errors (n = 6). SL shrubland, CP mixed-conifer, LP mixed-legume, NP mixed-native species

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Soil chemical properties

Soil pH was significantly higher in the mixed-conifer than in the mixed-legume (P < 0.01) and in the shrubland. Total N, total P, and available P contents of the shrubland soil were the lowest among the four stand soils (Table 2). The soil under the mixed-legume showed significantly higher soil organic C (SOC) (P < 0.001), total N (P < 0.001), and total P (P < 0.001) contents than the shrubland soil. The C/N ratio of soil under the three plantations was significantly lower than that in the shrubland soil. Total P and available P contents of soil under mixed-native species were also significantly greater than those in the shrubland soil (P < 0.01).

Table 2 Soil chemical properties of controls and trenched soil (0–10 cm depth) under the four vegetation types after 27-year plantations
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Soil microbes and fine root biomass

Soil microbial biomass C was the highest under the mixed-native species and the lowest under the mixed-conifer species (Fig. 4a). The value in the soil under the mixed-native species was significantly greater than that in soils under the shrubland (P = 0.002) and the mixed-conifer (P < 0.001) species. The significant difference was also observed between values of the mixed-legume and the mixed-conifer soils (P = 0.011). The trenching affected significantly the microbial biomass C of the shrubland soil (P = 0.009) (Fig. 4a).

Fig. 4
figure 4

a–d Soil microbial biomass C and composition of microbial community at 0–10 cm depth in the controls and trenched plots in the four vegetation types in subtropics. Error bars indicate standard errors (n = 6). The lowercase letters indicate significant differences between the treatments at each site. The uppercase letters indicate significant differences in the average of the controls and the trenching plots among the four sites. SL shrubland, CP mixed-conifer, LP mixed-legume, NP mixed-native species

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The percentage of total PLFAs present as bacterial PLFAs was significantly lower in soil under the mixed-conifer than in the shrubland soil (P = 0.030) (Fig. 4b), while the percentage of total PLFAs present as fungal PLFAs was significantly greater in soil under the mixed-conifer than in the shrubland soil (P = 0.014) and in the soil under the mixed-legume (P = 0.038) (Fig. 4c). Correspondingly, the ratio of fungal to bacterial PLFAs was significantly greater in the soil under mixed-conifer than in the shrubland soil (P = 0.016) (Fig. 4d). The bacterial PLFAs, fungal PLFAs, and their ratio were not significantly influenced by the trenching in the soil under each stand due to the high variability of the measured values.

The fine root biomass was 129 g m−2 in the shrubland, 127 g m−2 in the mixed-conifer, 113 g m−2 in the mixed-legume, and 197 g m−2 in the mixed-native species. The mean fine root biomass of the mixed-native species was significantly higher than that of the other three stands (P = 0.018 for the shrubland, P = 0.034 for the mixed-conifer, and P = 0.003 for the mixed-legume).

Relationships of soil CO2 efflux with abiotic and biotic factors

Soil CO2 efflux at each stand was significantly correlated with soil temperature (Table 3), and this correlation was not significantly affected by trenching. The linear relationship between soil CO2 efflux and soil moisture was only observed in the control of the shrubland and in the control and trenched plots of the mixed-native species.

Table 3 Relationship between soil CO2 efflux and soil temperature and moisture in the four vegetation types
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R S was significantly and positively correlated with soil chemical properties (except for pH, C/N ratio, and available P content) and microbial biomass C. Significant positive correlations were observed between R H and SOC, total N and P contents, and microbial biomass C (Table 4).

Table 4 Pearson correlation coefficients between soil CO2 efflux and soil chemical and microbiological properties in the four vegetation types
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Discussion

Trenching effects on soil CO2 efflux

Soil CO2 efflux was significantly decreased by trenching, thus confirming what already reported (Sayer and Tanner 2010; Bond-Lamberty et al. 2011). Soil microclimate (soil temperature or moisture) was not significantly influenced by trenching, and thus, the difference in soil CO2 efflux between the controls and trenched plots was not ascribed to the trenching effect on soil microclimate. Drake et al. (2012) found that the trenching reduced the R H following the removal of live-root-C inputs and the consequent shifts in the microbial composition and activity. In our study, the composition of soil microbial community was not significantly affected by the root exclusion, which agreed with the report by Wu et al. (2011b) in the same region.

Effects of reforestation on soil respiration and its sources

The annual R S of our sites ranged from 666 to 1086 g C m−2 year−1, values similar to those of adjacent plantations in this region (Wu et al. 2011a; Yu et al. 2015). The mixed-legume and the mixed-native species significantly increased both R S and R H compared with the values of the shrubland. The spatial variation of R S was quite similar to that of R H, which contradicts the hypothesis that the variation in R S is mainly driven by changes in R A due to the significant correlation found between R A and SOC (Wang et al. 2013). We did not observe an increasing trend in R A by increasing the SOC content, and this may explain this discrepancy. Therefore, our findings indicate that R H may be the major driver of changes in R S with reforestation.

The percentage of R S respired as R A was 20–28%, confirming that R H dominates the total soil respiration in soils under the plantations of southern China (Yu et al. 2015). Our percentages, however, were lower than those of nearby natural forests in subtropics (~36% on average) (Huang et al. 2016); probably, this is due to the lower root biomass at our sites because our plants were younger (~30 years old) than those of the older nature forests (>50 years old) (Zeng et al. 2008; Yan et al. 2009). Indeed, root biomass often increases with forest age (Yan et al. 2009). The significantly increased percentage of R S respired as R A in the soil under mixed-native species (28%) compared to the shrubland (22%) soil indicated more belowground investment in establishing root systems (Yan et al. 2015), despite showing high soil respiration.

Factors driving the changes in soil respiration and its sources after reforestation

Previous studies reported that plant-mediated effects on soil microclimate were responsible for changes in CO2 efflux from soils under different vegetation types (Vesterdal et al. 2012). Although soil respiration was correlated with soil temperature, soil temperature was not significantly affected by reforestation. The change in soil moisture among the four stands did not parallel changes in R H, which depended on soil chemical properties and microbial biomass. It is well established that organic C and nutrient contents of soil control the supply of substrates for microbial respiration (Allen and Schlesinger 2004; Zhang et al. 2015). The increased organic C, total N, and total P contents and the decreased C/N ratio of soil under the mixed-legume compared to values of the shrubland soil indicate that more energy and nutrients were available for microbes, giving a higher R H in the former than the latter soil. Moreover, the P content is a limiting factor for soil respiration in this region (Liu et al. 2012). The higher P content of the mixed-native species soil than that of the shrubland soil probably contributed to the higher microbial biomass and thus greater R H. In addition, the composition of soil microbial community was altered by the mixed-conifer, with a higher ratio of fungal-bacterial PLFAs than that in the shrubland soil. Fungi generally store more C than bacteria, and their cell walls are resistant to decomposition (Singh et al. 2010). This may explain that R H in the mixed-conifer soil was not higher than that in the shrubland soil since there were no increases in soil C, nutrients, or microbial biomass in soil under the mixed-conifer. Our results suggested that soil chemical and biological properties are important in controlling the spatial variations of microbial decomposition of plant residues after reforestation.

Root respiration depends on plant productivity and root biomass (Saiz et al. 2006; Tang et al. 2009). Previous studies have shown that the net primary productivity (NPP) was the lowest in the shrubland but the highest in the mixed-legume (Ren et al. 2007a; Zeng et al. 2008). The pattern of NPP across the four stands was apparently inconsistent with the R A pattern. However, the mixed-native species had the highest R A and the significantly greatest fine root biomass and probably the high metabolic activity was responsible for the highest R A (Pregitzer 2002; Tomotsune et al. 2013). Hence, we speculated that R A was probably controlled by fine root biomass than plant productivity after reforestation. Also in poplar plantation, the fine root biomass is a key driver of R A (Yan et al. 2015). Moreover, the increased percentage of R S respired as R A in the mixed-native species after reforestation also corresponded with its higher fine root biomass.

Conclusions

There were temporal variations of R S and R H of soils at the four stands, with higher values in the wet than dry season. The higher value of R A was also observed in the wet season. After reforestation, R S increased in soils under the mixed-legume and the mixed-native species but not in the mixed-conifer soil. R H of the four stands exhibited the same pattern of R S. The changes in R H after reforestation were mainly determined by changes in some soil chemical properties and microbial biomass. The R A and the percentage of R S respired as R A were the highest in the soil under mixed-native species after reforestation, which was related to the greater fine root biomass. Our findings highlight the importance of vegetation types in influencing different sources of soil respiration and indicate the necessity to consider the changes in some soil chemical and microbiological properties of the different plantations when evaluating soil CO2 efflux. The difference in the percentage of R S respired as R A after reforestation suggests that the mixed-native species may be more efficient in allocating C to belowground than the other two plantations, and this suggests to diversify plantations for forest management of degraded areas.