Global variation of soil microbial carbon-use efficiency in relation to growth temperature and substrate supply

Soil microbial carbon-use efficiency (CUE), which is defined as the ratio of growth over C uptake, is commonly assumed as a constant or estimated by a temperature-dependent function in current microbial-explicit soil carbon (C) models. The temperature-dependent function (i.e., CUE = CUE0 + m × (T − 20)) simulates the dynamic CUE based on the specific CUE at a given reference temperature (i.e., CUE0) and a temperature response coefficient (i.e., m). Here, based on 780 observations from 98 sites, we showed a divergent spatial distribution of the soil microbial CUE (0.5 ± 0.25; mean ± SD) at the global scale. Then, the key parameters CUE0 and m in the above equation were estimated as 0.475 and −0.016, respectively, based on the observations with the Markov chain Monte Carlo technique. We also found a strong dependence of microbial CUE on the type of C substrate. The multiple regression analysis showed that glucose influences the variation of measured CUE associated with the environmental factors. Overall, this study confirms the global divergence of soil microbial CUE and calls for the incorporation of C substrate beside temperature in estimating the microbial CUE in different biomes.

Soil microbial dynamic is a key regulator of ecosystem carbon (C) cycling because of its important role in decomposition and stabilization of soil organic carbon (SOC) [1][2][3] . The mechanistic understanding of soil microbial dynamics has been rapidly improved in recent decades 4 , but their numeric representation in the Earth system models (ESMs) has lagged behind 5 . Recent studies have reported that it might be reasonable to incorporate microbial processes into global C model [5][6][7][8][9] . For example, the simulated global SOC by the Community Land Model (CLM) has been more accurate with the implementation of microbial dynamics [7][8][9] . In those microbial-explicit SOC models, the soil microbial carbon-use efficiency (CUE; i.e., the ratio of growth over C uptake) is a key parameter. However, there is a lack of consensus on the CUE estimate among different models 10 . Synthesis based on observations has shown that the natural variation in soil microbial CUE is larger than aquatic, coastal and estuarine ecosystems 11 . Thus, a better understanding of the microbial CUE variability in the soil is critical for improving the simulation accuracy of the microbial-explicit SOC models.
Because CUE represents the ratio of growth to assimilation rates, differences in the temperature sensitivity of these two components causes the variations in CUE as a function of temperature change. Generally, respiration increases more than growth as a function of temperature, so CUE tends to decrease with temperature in both soil and aquatic systems 6,[12][13][14][15][16][17] . This pattern has already been represented in many microbial-explicit SOC models as a linear temperature sensitivity function 5,9,10,[18][19][20] : where CUE 0 is the CUE at reference temperature, m is the temperature response coefficient (i.e., the change in CUE per °C temperature change) and T 0 is the reference temperature (usually set as 20 °C). However, the In contrast to temperature, substrate quality and accessibility are other key factors which affect the soil microbial CUE 11,21 . As summarized by Manzoni et al. 11 , substrate quality regulates CUE variation through two different approaches. First, substrates with different chemical structures have to undergo different metabolic pathways to be completely decomposed. For example, substrates consist of more degradable compounds such as carbohydrates and protein could result in higher CUE than those contains more recalcitrant compounds, e.g., aliphatic, aromatic and lignin 11,[22][23][24][25] . Second, microbial CUE is higher for the C substrate with a high degree of reduction (γ S ), i.e., a measure of the chemical energy per unit mole of C 11,22,26,27 . The value of γ S ranges from 1 (e.g., oxalate) to 8 (e.g., methane) and is approximately equal to 4.2 in the microbial biomass 11,27 . Hence, the microbial CUE is mainly limited by energy when the γ S of substrate is <4.2, but reaches the maximum for substrates with γ S > 4.2 11,27 . Both of these two approaches highlight that substrate quality is critical in affecting the microbial CUE variability in the soil. Besides the substrate quality, microbial growth is strongly related to the substrate accessibility which is collectively affected by environmental and soil conditions such as water availability 28 and aggregates 29 .
The objective of this study is to examine the impacts of temperature and substrate type on soil microbial CUE based on observations at the global scale. A data-assimilation approach based on the Markov chain Monte Carlo (MCMC) technique is applied to constrain the parameters in the Eq. (1). Specifically, the aims of this study are to: ① constrain the key parameters in the Eq. (1) based on observations; and ② explore the dependence of soil microbial CUE upon substrate type.

Results
Global divergence of soil microbial CUE. Overall, 780 observations from 98 sites across the globe were collected in our analysis (Fig. 1, Tables S1, S2), with the measurements between the year of 1973 to 2017. The latitude of the sites ranged from 71°S to 78°N, and the longitude from 147°W to 174°E. Our data covered most terrestrial biomes, including forests, shrublands, grasslands, croplands and tundra (Fig. 2a). Globally, the mean value of CUE is 0.5 ± 0.25 (Mean ± SD) (Fig. 2b). The frequency distribution of CUE values suggests a distinct divergent distribution of CUE at the global scale. The observed CUE were highest in shrublands (0.73 ± 0.04) and grasslands (0.65 ± 0.22) but lowest in forests (0.41 ± 0.22).
Temperature dependence of soil microbial CUE. The linear regression analysis detected no correlation of CUE with latitude, longitude, mean annual precipitation, mean annual temperature, soil pH or carbon to nitrogen ratio across the given sites (Fig. S1). However, a significant negative relationship was found between the CUE and the corresponding incubated temperature based on 718 values from our database (Fig. 3a). The range of the incubation temperature in this synthetic database was from 2 °C to 28 °C. The regression of CUE with temperature was matched well by Eq. (1), in which the maximum likelihood estimator of the CUE 0 and m was 0.475 and −0.016, respectively (Fig. 3). Therefore, the temperature dependence of CUE could be better expressed as the following formula: www.nature.com/scientificreports www.nature.com/scientificreports/ Dependence of CUE upon substrate type and degree of reduction. Soil microbes using different types of substrate as C sources have different CUE (Fig. 4). The microbial CUE was highest in the soils which were incubated with glucose plus organic carbon addition (0.75 ± 0.05), while was lowest in that with the residue plus N addition (0.15 ± 0.15). The mean soil microbial CUE under the addition of amino acid (0.51 ± 0.20) was comparable with that under the addition high-molecular compounds (0.51 ± 0.19). Soil microbial CUE was similar under the additions of other organic acid (0.33 ± 0.21) and plant residues (0.32 ± 0.24). Nutrient availability  www.nature.com/scientificreports www.nature.com/scientificreports/ plays an important role in regulating microbial CUE. For example, the microbial CUE was high in soils which were incubated with glucose plus N solution (0.60 ± 0.16) or mixed inorganic solution (0.41 ± 0.17). However, for those soils with glucose addition, no positive impact of N addition on microbial CUE was detected. On the contrary, adding N lead to 51% and 53% declines of CUE under the incubation of high-molecular compounds and plant residue, respectively (Fig. 4). Based on the 462 observations of CUE with according substrate type, soil microbial CUE also varied with the γ S of different substrate (Turkey test, P < 0.05; Fig. 5).

Discussion
Soil microbial CUE is a commonly used parameter to quantify how soil carbon (C) is partitioned between microbial growth and respiration. The global mean CUE in our analysis (0.5) is comparable to the result of a former data-analysis across the world, where soil CUE was about 0.55 11 . These values are close to the thermodynamic  www.nature.com/scientificreports www.nature.com/scientificreports/ maximum of metabolic efficiency (0.6) 26,27 , but is much higher than the CUE value of 0.3, which has been recommended for large-scale models by Sinsabaugh et al. 30 . Furthermore, the global divergence of microbial CUE with a wide range (i.e., 0.002 to 0.993) in our analysis implies that a constant CUE in biogeochemical models is inappropriate. For example, the mean CUE of soil microbes in the shrublands (0.73) is much higher than that in grasslands (0.65) or forests (0.41). The great variability of CUE at global and ecosystem-level scale emphasizes the importance of understanding the mechanisms underlying the microbial CUE variation.
Several recent modelling studies have used the Eq. (1) to simulate the variation of microbial CUE 5,31 . Among these models, however, the CUE 0 is not identical and the parameter m ranges largely from -0.016 to 0 10,18 . In this study, we recommend an equation of CUE = 0.475 − 0.016 × (T − 20) by assimilating the observations to the Eq. (1) with the Markov chain Monte Carlo (MCMC) technique. The temperature response coefficient m in this formula is not 0 as in some previous studies 10,18 . Thus, the CUE is not a constant value but must changes with temperature in biogeochemical models according to this amended function. Because the rate of climate warming is faster at high than low latitudes 32 , the CUE could reduce quicker in cold than warm regions. Thus, it remains unclear how future soil C cycling in global land models will be affected if they incorporate the temperature dependent of CUE. However, it should be noted that numerous studies have shown a lack of temperature effect on CUE in the short term 33  The selection of glucose rather than other types of C substrate suggests that glucose has a large impact on the global variation of measured CUE. We further compared the two fitted equations between data of two substrate categories (i.e., glucose and others) (Table S4). We found that the CUE 0 was higher with adding glucose (0.491) than other types of substrate (0.319). This study also found that latitude is another important factor influencing microbial CUE. The percentage contribution of latitude reaches 22.31% when all factors (Latitude, Longitude, Temperature, Substrate type, MAP, pH) are included in the multiple stepwise regression analysis for explaining the CUE variation (Fig. 6), though there is no significant relationship between latitude and CUE in the univariate regression analysis (Fig. S1(a)). We also assessed all the environmental factors (i.e., Latitude, Longitude, Temperature, MAP, and pH) with a Principal Component Analysis (PCA) (Fig. S2).
Numerous laboratory studies and field investigations have found strong dependence of CUE upon substrate type and nutrient condition [35][36][37] . In this study, the soil microbes have higher CUE with the easily degradable (such as glucose, amino acid and high-molecular organic matter) than the recalcitrant (plant residues) substrates (Fig. 4). This could be due to the greater activation energies are consumed for enzyme production and excretion during the decomposition of the recalcitrant than easily degradable compounds 11 . Another potential mechanism is that the substrate accessibility of plant residues is lower than others, e.g., glucose, because of the occlusion of organic matters by aggregation 38 . Increasing nutrient availability is assumed to enhance microbial CUE mainly because of its negative effect on microbial respiration [39][40][41] . This assumption is partially supported by our analysis, which shows greater CUE after adding inorganic N or mixed inorganic solution than only adding water (Fig. 4). However, adding N leads to no changes and even reductions of microbial CUE in the soils which were incubated with glucose and high-molecular compounds or plant residue, respectively. The variation of microbial CUE also depends on the γ S of different substrate. Our study also suggests that more degradable compounds (such as glucose) with high γ S owns higher microbial CUE than the recalcitrant compounds (such as oxalate) with low γ S (Figs 4 and 5). Also, because the chemical compound of litter differs greatly among plant species 42 , the substrate www.nature.com/scientificreports www.nature.com/scientificreports/ type must be considered to estimate the soil microbial CUE in different regions of the globe. Thus, more research efforts are still needed to explore the differences in substrate type among vegetation types and their roles in regulating the dynamics of soil microbial CUE.
In summary, this study used a data assimilation approach to constrain the temperature dependence of soil microbial CUE based on the existing observations at the global scale. The observed CUE shows a divergent distribution globally, suggesting that the parameterization of CUE is still one key challenge for the microbial-explicit soil C models. We recommend to use the constraint parameters in the Eq. (2) (i.e., CUE = 0.475 − 0.016 × (T − 20)) if soil microbial CUE is used in future global land models. However, it should be noted that tremendous additional efforts are still needed before we can correctly simulate the variations of microbial CUE in the soil. For example, as shown in this study, it is clear that the type of substrate (especially glucose) affects the simulation of soil microbial CUE. Besides, the random sampling of data has been one major challenge for almost all meta-analysis studies, because the observations of almost all variables are distributed in three hotspot regions (i.e., North America, West Europe, and China). In our study, most observations were from forest, grassland and cropland. So the data set cannot perfectly represent the distributions of global biome type and soil community in the real Earth system. We also call for more observations in other biomes such as shrubland, tundra and desert. Overall, this study highlights that the predictive ability of microbial-explicit soil C models can be enhanced by an improved understanding on the impacts of temperature and substrate type on microbial CUE.

Methods
Data collection. Peer-reviewed literatures related to soil microbial CUE published before July 2017 were searched using the Web of Science, according to the PRISMA (preferred reporting items for systematic reviews and meta-analyses) guidelines 43,44 (Fig. S3). There are different methods to quantify the soil microbial CUE, so the selected studies must directly have reported the CUE or provided at least one of the following information: (1) microbial C uptake and substrate C consumption; (2) microbial C uptake and cumulative C respiration; and (3) microbial C uptake, respiration rate and the length of experiment time. The following Eqs (3)(4)(5) in this study has shown the methods for CUE calculation based on these data. Ancillary information such as latitude, longitude, mean annual temperature, mean annual precipitation, pH, C:N ratios, and substrate type were also extracted from the papers or cited papers or, in the case that it was not reported, extracted from the database at http://www. worldclim.org/ using the location information (e.g., latitude and longitude).
Methods for data conversion. Soil microbial CUE was not directly reported but can be calculated based on its definition in many literatures. If one study reported the microbial C accumulation (ΔMBC) and the C substrate consumption (ΔC substrate ), then the CUE can be calculated as: substrate If both the microbial C accumulation (ΔMBC) and cumulative microbial respiration (R cumulative ) were reported, we calculated the CUE by using the following formula: In some studies, the cumulative microbial respiration can be obtained from the rate of respiration (R) over a given incubation time (t), so the CUE was calculated as: Bayesian framework to determine CUE dependence on temperature. The Eq. (1) commonly represents the temperature dependence of CUE in most microbial-explicit SOC models 5,9,18 . The parameterization of the Eq. (1) (i.e., CUE 0 and m) is usually arbitrary in soil enzyme driven models. For example, the CUE 0 has been fixed as 0.31, 0.5 or 0.63 and parameter m ranges from −0.016 to 0 in previous studies 10,18 . This study employed the Bayesian probability inversion and a Markov chain Monte Carlo (MCMC) technique to evaluate the CUE 0 and m. To get the maximum likelihood estimators of the CUE 0 and m, we specified them as the uniform distribution over a set of intervals, which were set as (0, 1) and (−0.1, 0), respectively. The collected CUE values and the corresponding incubation temperatures were used as inversing data to constrain the two parameters. We formally made five parallel runs using the Metropolis-Hastings (M-H) algorithm 45,46 as the MCMC sampler with 100,000 simulations for each run. Each run started from a random initial point in their respective parameter intervals to eliminate the effect of the initial condition on stochastic sampling. The acceptance rates for the five runs ranged from 5% to 10% which tested by the Gelman-Rubin (G-R) diagnostic method. The initial samples (approximately 1,000 for each run) were discarded after the running means and standard deviations (SDs) were stabilized (regarded as the burn-in period). All the accepted samples from five runs after the burn-in periods (approximately 20,000 samples in total) were used to construct maximum likelihood estimators of both the CUE 0 and m (inset of Fig. 3). The evaluation of CUE 0 and m were conducted with MATLAB 2016b (The Mathworks, Natick, MA, USA).