Environmental Microbiology Reports (2022) 14(1), 85–95
13033
Brief Report
Bacterial rather than fungal diversity and community
assembly drive soil multifunctionality in a subtropical
forest ecosystem
Shun Han,1,2 Shuang Tan,1 Achen Wang,1
Wenli Chen 1* and Qiaoyun Huang 1,3
1
State Key Laboratory of Agricultural Microbiology,
College of Life Science and Technology, Huazhong
Agricultural University, Wuhan, 430070, China.
2
Institute for Environmental Genomics, Department of
Microbiology and Plant Biology, University of Oklahoma,
Norman, OK, 73019, USA.
3
Key Laboratory of Arable Land Conservation (Middle
and Lower Reaches of Yangtze River), Ministry of
Agriculture, College of Resources and Environment,
Huazhong Agricultural University, Wuhan, 430070,
China.
Summary
Microbial diversities are key drivers of soil multi-
functionality in terrestrial ecosystems and are impor-
tant for stability and productivity of ecosystems.
However, the relationships among microbial diver-
sity, community assembly and soil multifunctionality
in forest ecosystems remained unclear. Here, soil
samples were collected from a subtropical forest
ecosystem, Lushan Mountain, China. High-
throughput sequencing was employed to reveal the
bacterial/fungal community assembly and biodiver-
sity, as well as 10 enzyme activities were measured
to assess soil multifunctionality. We found that soil
multifunctionality was negatively regulated by bacte-
rial and fungal alpha diversity, implying a higher
potential functional redundancy in this forest soil.
The null model indicated that deterministic processes
(variable selection) and stochastic processes (dis-
persal limitation) govern bacterial and fungal phylo-
genetic turnover, respectively. Correlation analysis
revealed that bacterial rather than fungal community
assembly processes have a significant linkage to soil
Received 4 May, 2021; accepted 29 November, 2021. *For corre-
spondence. E-mail wlchen@mail.hzau.edu.cn; Tel: +86-27-
87280670; Fax: +86-27-87280670
© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd.
doi:10.1111/1758-2229.
multifunctionality. These observations projected that
soil variables could regulate multifunctionality by
shaping the phylogenetic and taxonomic turnover of
bacteria rather than fungi. In summary, our study
highlighted that soil multifunctionality is mainly
driven by bacterial diversity and community assem-
bly processes while not fungal, presenting different
views and knowledge of microbial diversity and com-
munity assembly processes in ecosystem
functioning.
Introduction
Soil-dwelling microorganisms are highly diverse and gen-
erally play an important role in governing and regulating
various ecosystem functions (Zavaleta et al., 2010;
Byrnes et al., 2014; Jing et al., 2015; Lefcheck
et al., 2015; Delgado-Baquerizo et al., 2017; Han
et al., 2018; Han et al., 2021a), such as climate change,
carbon and nitrogen cycling, biomass production and
plant residue decomposition. Traditionally, many studies
have focused mainly on the impact of diversity on a sin-
gle ecosystem function (e.g., plant productivity, biomass,
soil extracellular enzyme activity and/or nutrition cycling)
(Cardinale et al., 2011; Isbell et al., 2011; Bardgett and
Van der Putten, 2014; Wagg et al., 2014; Han
et al., 2020). However, the lack of a comprehensive
investigation on the function of the overall ecosystem
may lead to a limited understanding of how soil microbial
diversity influences ecosystem functions (Soliveres
et al., 2016). Thus, comprehensive knowledge on the
linkage between biodiversity and ecosystem functions is
essential for the maintenance of ecosystem health and
sustainability.
In recent decades, ecologists have explored the links
between microbial diversity (or community diversity) and
multiple simultaneous ecosystem processes and services
(multifunctionality) (Hector and Bagchi, 2007; Bardgett
and Van der Putten, 2014; Jing et al., 2015; Delgado-
Baquerizo et al., 2017; Manning et al., 2018; Wagg
et al., 2019; Han et al., 2021b). Growing evidence from

86 S. Han et al.
many studies has demonstrated that the linkage between
microbial diversity and ecosystem multifunctionality is
more frequently positive and linear than saturating across
a broad range of ecosystems – e.g., grassland (Byrnes
et al., 2014; Jing et al., 2015; Wagg et al., 2019), farm-
land (Luo et al., 2019; Chen et al., 2020), and boreal for-
est soils (Li et al., 2019; Giguere-Tremblay et al., 2020),
implying that microbial diversity could enhance soil multi-
functionality. Meanwhile, observational experiments
showed that losses in microbial functional diversity could
result in the lowering (reduction) of several specific soil
processes (e.g., emission of methane and nitrous oxide)
(Trivedi et al., 2019). Investigations of grassland ecosys-
tems have shown enhanced ecosystem functioning by
microbial biodiversity, and their interaction complexity
promotes ecosystem functioning (Wagg et al., 2019). As
such, higher microbial diversity improved the
multifunctional resistance to drought and climate change
in agricultural and grassland ecosystems (Jing
et al., 2015; Luo et al., 2019). Environmental factors can
also affect ecosystem processes and functions (Bastida
et al., 2016; Delgado-Baquerizo et al., 2016a; Chen
et al., 2020; Hu et al., 2020). For example, soil pH affects
ecosystem multifunctionality via the influence of soil
microbial activity and community structure (Delgado-
Baquerizo et al., 2017). Organic carbon impacts the activ-
ities of bacterial and fungal populations and further deter-
mines the division of functional niches in semiarid soils
(Bastida et al., 2016). Additionally, climate changes
(e.g., mean annual temperature and precipitation) and
plant types are also critical factors influencing soil eco-
system multifunctionality (Jing et al., 2015; Delgado-
Baquerizo et al., 2016b; Zhou et al., 2020). The under-
standing of microbial biodiversity and abiotic factors
affecting soil multifunctionality has advanced dramatically
in recent years, but it is still incomplete.
Except the microbial diversity itself, previous studies rev-
ealed that ecosystem functions were also regulated by
microbial community composition, which drive by different
community assembly (Knelman and Nemergut, 2014;
Metcalf et al., 2016; Graham and Stegen, 2017). Commu-
nity assembly patterns, based on a null-modelling frame-
work, can be divided into stochastic and deterministic
processes (Vellend, 2010; Stegen et al., 2013). Stochastic
assembly processes (including ecological drift and dis-
persal limitation) can lead to spatially complex community
structures (Hubbell, 2001). The deterministic assembly
process (homogeneous and variable selection) can typi-
cally cause beta diversity (taxonomic turnover between
two sites or communities) associated with environmental
characteristics (Stegen et al., 2013, 2015; Zhou and
Ning, 2017). Indeed, microbial community assembly mech-
anisms have enriched our knowledge of ecological pro-
cesses in soil ecosystems (Stegen et al., 2013; Zhou and
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17582229, 2022, 1, Downloaded from https://ami-journals.onlinelibrary.wiley.com/doi/10.1111/1758-2229.13033 by Univ of Sao Paulo - Brazil, Wiley Online Library on [21/07/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
Ning, 2017; Tripathi et al., 2018). However, how microbial
community assembly processes regulate and govern soil
multifunctionality performance has been mostly
unexplored. Therefore, the next frontier is to now empiri-
cally test whether changes in the microbial community
assembly process, as indicated by both stochastic and
deterministic changes, are crucial for the way the microbial
community affects soil ecosystem multifunctionality.
Forest ecosystems are richer in nutrients and microbial
diversity than other terrestrial ecosystems (e.g., cultivated
land, grassland and desert). Nevertheless, few studies have
revealed the relationships between microbial diversity and
soil multifunctionality in forest ecosystems (e.g., Li
et al., 2019). To provide more evidence and deepen our
understanding of the linkages among microbial biodiversity,
community assembly processes and soil multifunctionality
in subtropical forest ecosystems, soil samples were
obtained from eight sites along the elevational gradients of
Mount Lu, Jiangxi Province, South China. To assess bacte-
rial and fungal diversity and community assembly pro-
cesses, we performed high-throughput DNA sequencing of
taxonomic marker genes and 16S rRNA V3-V4 and fungal
internal transcribed spacer (ITS) regions. To evaluate soil
multifunctionality, we measured 10 microbial-mediated
carbon-, nitrogen-, phosphorus- and sulfur-related enzyme
activities. In this study, we intend to address the following:
(i) how bacterial and fungal biodiversity (alpha and beta
diversity) affects soil multifunctionality in subtropical forest
ecosystems and (ii) how bacterial and fungal community
assembly processes regulate soil multifunctionality.
Results
Soil enzyme activities
Soil enzyme activities showed an inconsistent pattern
across elevation gradients (Figs 1A and S1). Briefly, the
activities of β-D-xylosidase and α-D-glucopyranosidase
exhibited a hump-shaped elevational pattern. The activi-
ties of cellulose, invertase and urease were greater at
elevations lower than 400 m. The enzyme activities asso-
ciated with C-, N-, S- and P-cycling were mostly positive
correlated. For example, invertase was positively related
to enzymes other than acid phosphatase, while β-D-
glucosidase was positively correlated with β-D-
cellobiosidase. However, acid phosphatase was nega-
tively correlated with most enzyme activities (Fig. S2). As
shown in Fig. S3, acid phosphatase had a positive corre-
lation with soil pH and a negative correlation with SWC
(soil water content), TC (soil total carbon), TN (soil total
nitrogen), SOC (soil organic carbon) and AP (soil avail-
able phosphorus). pH showed a negative association
with most soil enzyme activities. SWC, TC, TN and AP
had a positive influence on most soil enzyme activities,

Fig. 1. Heat map for the changes in multiple enzyme activities (A) and microbial diversity (B) along elevational gradients. The capital letters C, N,
P and S show the respective C, N, P and S-cycle enzyme categories.
except for β-D-glucosidase, sulfatase, β-D-cellobiosidase
and N-acetyl-glucosaminidase.
Soil bacterial and fungal communities
The taxonomic distribution of bacteria at the phylum
level showed higher relative abundances for
Acidobacteria (28.1%–52.8%), Proteobacteria (22.5%–
31.7%), Actinobacteria (13.2%–29.7%) and Chloroflexi
(0.9%–15.9%) across the examined soils (Fig. 2A).
Basidiomycetes (20.3%–92.3%), Ascomycetes (9.7%–
52.9%) and Mortierellomycota (1.2%–30.5%) were the
most abundant fungal phyla, especially in EG951
(87.1%) and EG1000 (92.3%) (Fig. 2B). The analysis of
similarity demonstrated that the elevational gradient
was a significant driver of bacterial (p < 0.001) and fun-
gal (p < 0.001) community composition. Principal com-
ponent analysis (PCA) revealed that both bacterial and
fungal community structures were very divergent with
elevation in the mountain forest soils (Fig. 2C and D). In
addition, the total explained variables of the first two
axes by PCA were 39.38% and 42.65% for the bacterial
and fungal communities, respectively. The Mantel test
showed that all the measured soil variables significantly
impacted both the bacterial (except for AP) and fungal
community structures (Table S1).
© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd., Environmental Microbiology Reports, 14, 85–95
17582229, 2022, 1, Downloaded from https://ami-journals.onlinelibrary.wiley.com/doi/10.1111/1758-2229.13033 by Univ of Sao Paulo - Brazil, Wiley Online Library on [21/07/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
Bacterial drive soil multifunctionality 87
Relationship among microbial diversity and soil variables
and multifunctionality
As shown in Fig. 1B, microbial alpha diversity (based on
the Shannon index) was significantly different across
elevational gradients in mountain forest soils. Bacterial
diversity was positively regulated by soil pH but nega-
tively related to soil TC and SOC (Fig. S4). In contrast,
fungal diversity was only positively related to soil AP but
negatively correlated with soil carbon (TC and SOC)
(Fig. S4). The averaging approach showed that soil multi-
functionality was significantly and negatively (R2 = 0.36,
p = 0.002) related to bacterial diversity (Fig. 3A),
whereas a marginal negative (R2 = 0.15, p = 0.060) cor-
relation was found with fungal diversity (Fig. 3B). The
multiple-threshold approach indicated that bacterial and
fungal diversity was negatively correlated with a larger
number of functions working at a high functional level
(Fig. 3C–F). In addition, bacterial and fungal diversities
were found to be negatively related to each enzyme
activity, except for urease and acid phosphatase
(Fig. S2). Interestingly, all the measured C-related
enzyme activities were negatively associated with bacte-
rial and fungal diversity. Correlation analysis revealed a
negative linkage between soil ecosystem multi-
functionality and soil pH and a positive relationship with
soil water content, carbon and nitrogen, and available
phosphorus (Fig. S5).
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88 S. Han et al.

Fig. 2. Taxonomic compositions of bacterial (A) and fungal (B) communities at the phylum level determined by the proportional frequencies of
16S rRNA and ITS sequences. Principal component analysis (PCA) ordinations based on Euclidean distances show the changes in bacterial
(C) and fungal (D) community composition.

Relationship between microbial ecological processes
and multifunctionality
We further assessed the phylogenetic and taxonomic
turnover between sampling sites by comparing βNTI and
RCbray metrics to examine whether and how microbial
community assembly processes vary across elevational
gradients. We found that the soil bacteria phylogenetic
turnover among most (
60%) elevational gradient pairs
was different from the null expectation (jβNTIj > 2;
Fig. 4A and C). These results suggest that deterministic
processes, specifically variable selection (51.81%), may
have dominated the bacterial community assembly
among most elevational sites, whereas dispersal limita-
tion (24.64%) may play a central role in leading to taxo-
nomic turnover among other sites. In sharp contrast, soil
fungal phylogenetic turnover among most elevational
gradient pairs (
87%) was not significantly different
from the null expectation (Fig. 4B and D). These results
indicated that stochastic processes might have played a
key role in causing fungal phylogenetic turnover among

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Bacterial drive soil multifunctionality 89

Fig. 3. The relationships between bacterial (A) and fungal (B) Shannon diversity and ecosystem multifunctionality, with diversity effects for a
range of ecosystem multifunctionality thresholds. Effect of bacterial (C, D) and fungal (E, F) Shannon diversity on the number of functions above
the thresholds. Lines represent the slope between diversity and the number of functions more significant than or equal to a threshold value rang-
ing from 5% to 99% of maximum for each function. The dotted curves indicate the changes in the number of functions per unit increase in the
diversity of bacteria.

most elevational gradients, with dispersal limitation between βNTI and pairwise differences in all six soil vari-
(60.51%) tending to be the dominant driver of the ables across the entire elevational gradients indicated
observed taxonomic turnover. The correlation analysis that bacterial βNTI was significantly (p < 0.001)

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 17582229, 2022, 1, Downloaded from https://ami-journals.onlinelibrary.wiley.com/doi/10.1111/1758-2229.13033 by Univ of Sao Paulo - Brazil, Wiley Online Library on [21/07/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
90 S. Han et al.

Fig. 4. Patterns of bacterial (A) and fungal (B) βNTI across elevational gradients. Horizontal dashed blue lines indicate upper and lower signifi-
cance thresholds at βNTI = +2 and 2 respectively. Contributions of ecological assembly processes governing bacterial (C) and fungal
(D) community turnover.

correlated with the variation in soil SWC, pH, TC, TN
and SOC; however, fungal βNTI had no significant
(p > 0.05) correlations with those factors (Fig. S6).
These results suggested that soil variables accounted
for relatively larger and smaller variations in bacterial
and fungal βNTI, respectively, between samples across
the elevational gradients. Furthermore, to assess the
relationship between bacterial and fungal community
assembly processes and multifunctionality, correlation
rates were calculated. We found that bacterial βNTI,
rather than fungalβNTI, were profoundly linked to the
variation in multifunctionality (Fig. 5A and B). These
observations imply that soil variables could regulate
multifunctionality by shaping bacterial phylogenetic turn-
over and taxonomic turnover rather than those of fungi.
Reliable predictors of soil multifunctionality
The random forest models explained 85.7% of the vari-
ance in soil multifunctionality, and bacterial βNTI and
biodiversity were the first two key predictors (Fig. 6).
All measured soil variables (except AP) played an
essential role in predicting forest soil ecosystem multi-
functionality, and soil carbon (TC and SOC) was the
most important abiotic factor. These observations were
consistent with the Pearson correlation analysis
(Fig. S5). In addition, fungal diversity was also a signifi-
cant factor in predicting multifunctionality, while fungal
βNTI was not.
Discussion
The aims of this study were to decipher the relationship
among the microbial diversity and community assembly
processes and soil ecosystem multifunctionality that can
better predict the changes in soil multifunctionality, rather
than reveal the microbial community differences along
altitudes in a subtropical mountain forest ecosystem. Fur-
thermore, this study provides substantial evidence that
bacterial rather than fungal diversity and community
assembly determine soil multifunctionality. Therefore, our
study provides a better understanding of the importance
of bacteria and fungi participating in biogeochemical
cycling and ecosystem functioning.

© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd., Environmental Microbiology Reports, 14, 85–95

Fig. 5. Linear regressions between pairwise bacterial (A) and fungal (B) community phylogenetic turnover (βNTI) and pairwise differences in the
soil ecosystem multifunctionality index.
Fig. 6. Main predictors of ecosystem multifunctionality. The figure
shows the random forest mean predictor importance (% increase in
mean square error) of soil variable drivers, and microbial Shannon
diversity and βNTI on ecosystem multifunctionality. The significance
levels of each predictor are as follows: *p < 0.05 and **p < 0.01.
Bacterial and fungal diversity negatively predicts soil
multifunctionality
In this study, it is interesting to note that soil multi-
functionality was negatively associated with both bacte-
rial and fungal diversity, and the soil ecosystem
multifunctionality was lower at higher threshold levels
(Fig. 3). These findings are inconsistent with the findings
from most of the previous studies, which showed a posi-
tive association between microbial biodiversity and eco-
system multifunctionality (Bardgett and Van der
Putten, 2014; Jing et al., 2015; Delgado-Baquerizo
et al., 2016c, 2017; Li et al., 2019; Chen et al., 2020).
Our results suggested that there may be a higher poten-
tial functional redundancy in this forest ecosystem and
that the reduction in soil biodiversity does not yield
decreased ecosystem functioning. This is contrary to the
previous view on the lack of functional redundancy
between bacterial diversity and ecosystem functionality
(Mori et al., 2016; Delgado-Baquerizo et al., 2016a). Our
study also indicated that bacterial rather than fungal
© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd., Environmental Microbiology Reports, 14, 85–95
17582229, 2022, 1, Downloaded from https://ami-journals.onlinelibrary.wiley.com/doi/10.1111/1758-2229.13033 by Univ of Sao Paulo - Brazil, Wiley Online Library on [21/07/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
Bacterial drive soil multifunctionality 91
diversity dominates soil multifunctionality in this subtropi-
cal forest soil (Figs 3 and 6), notably different from
boreal forest soil, which showed that fungal rather than
bacterial richness contributes to multifunctionality
(Li et al., 2019). Therefore, rational interpretations must
understand and disentangle the factors that regulate
microbial diversity and ecological functions in forest
ecosystems.
The majority of biogeochemical processes are driven by a
range of energy-transducing metabolic pathways, each of
which has been detected in diverse microbial clades
(Falkowski et al., 2008; Hawley et al., 2017). Other studies
have shown that higher microbial diversity does not always
guarantee simultaneously high levels of ecosystem functions
(Zavaleta et al., 2010; Gamfeldt et al., 2013; Byrnes
et al., 2014). The plausible reason is that a species that sup-
ports a special function at a high level could reduce its ability
to support other functions. In addition, richer nutrients are
available in this subtropical forest soil. However, nutrient
enrichment could induce microbial dormancy in salt marsh
sediments (Kearns et al., 2016). It is likely that more dormant
or inactive microorganisms are present, which may explain
the higher diversity, but do not display corresponding special
ecological functions in this forest soil. This hypothesis could
be supported by the negative relationship between biodiver-
sity and individual soil enzyme activities. It is accepted that
less than 10%–20% of microbes are metabolically active
and capable of driving soil biogeochemical processes
(Lennon and Jones, 2011). Those currently dormant or inac-
tive have lower metabolic capacities but could serve as a
‘seed bank’ of soil microbial propagules and play critical
roles in the resilience of the soil community in the face of
environmental disturbances (Prosser et al., 2007; Lennon
and Jones, 2011). Hence, the potential functional redun-
dancy in the current study suggests that increasing species
diversity may not always lead to enhanced ecosystem func-
tions, especially in soils with higher biodiversity and resource
availability. Another explanation might be that higher micro-
bial diversity results in metabolic overlap (Hester
et al., 2019); therefore, reducing diversity may lead to higher

92 S. Han et al.
functions, e.g., decomposition (Bell et al., 2013). Our findings
may also reveal that soil ecosystem multifunctionality
depends on microbial diversity to varying degrees.
The bacterial ecological process, rather than that of
fungal, contributes to soil multifunctionality
Various and significant influences have been demon-
strated concerning the roles of bacterial and fungal
alpha diversity in ecosystem multifunctionality due to the
dissimilarities of bacterial and fungal community structure
(Zheng et al., 2019). Our results showed that deterministic
processes (e.g., variable selection) play a crucial role in
shaping the phylogenetic and taxonomic turnover of bac-
teria, while stochastic processes (e.g., dispersal limitation)
play central roles in regulating the phylogenetic and taxo-
nomic turnover of fungi along elevational gradients. Con-
sistently, bacterial βNTI was significantly impacted by soil
variables – e.g., SWC, SOC, TC, TN and pH – whereas
soil variables had a lesser influence on fungal phyloge-
netic turnover (Fig. S6). Accordingly, we detected a signif-
icant contribution of bacterial βNTI and soil factors to
predict multifunctionality (Fig. 6). These results indicated
that soil factors along elevational gradients impose selec-
tion pressures that drive the turnover of bacterial commu-
nity structure and thereby govern soil multifunctionality.
Indeed, Powell et al. (2015) demonstrated that bacterial
rather than fungal community turnover was highly driven
by habitat turnover. The differences in soil variables along
the elevational gradients provide spatially heterogeneous
habitats for microbes, affecting the ecological functions
that microbes mediate. Under deterministic variable selec-
tion, three possible mechanisms may explain the contribu-
tion of the bacterial community to multifunctionality across
the forest. First, bacterial species can support multi-
functionality through their functional traits and environ-
mental conditions, e.g., soil variables. Second, higher
phylogenetic and taxonomic turnover of bacterial species
may result in niche complementarity and thus lead to
greater multifunctionality. Finally, bacterial species can
yield ecological interactions, including nutrient availability
for multiple ecosystem functioning. In contrast, stochastic
processes can account for more diversified functional
traits of microbes, but at the cost of suppressing their
ecosystem function (Gamfeldt et al., 2013; Knelman and
Nemergut, 2014; Bao et al., 2020). This reason may
partly explain why the fungal ecological process did not
sustain soil multifunctionality in this study. Together, the
ecological process appear to be significantly impacted by
elevational gradients and indirectly related to soil multi-
functionality by changing microbial community diversity
and structure.
© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd., Environmental Microbiology Reports, 14, 85–95
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Microbial taxonomic composition and multifunctionality
The ecological process can impact ecosystem functioning
because variations in microbial taxonomic composition
also affect species diversity and abundance distributions
(McGill et al., 2007; Chen et al., 2020). Of particular inter-
est is the fact that the soil multifunctionality was less
affected by the dominant bacteria (e.g., Acidobacteria,
Proteobacteria and Actinobacteria) and fungal phyla
(e.g., Basidiomycota and Ascomycota) because no signifi-
cant relationship was detected between soil multi-
functionality and the dominant phyla of bacteria and fungi
(Table S2). Only WPS2, which has a low relative abun-
dance, has a positive relationship with multifunctionality.
This result agrees with that from Chen et al. (2020), who
indicated that rare microbial taxa, not abundant species,
are crucial drivers of ecosystem multifunctionality in long-
term fertilized soils. However, in our study, the dominant
phyla of bacteria and fungi exhibited apparent relationships
with some individual soil enzyme activities (Fig. S7). For
example, Proteobacteria exhibited a negative association
with sulfatase, N-acetyl-glucosaminidase, β-D-xylosidase
and invertase; Chloroflexi, Gemmatimonadetes and
Ascomycota also exhibited a negative association with cel-
lulase. It is likely that ecological function is not sensitive to
the presence and abundance of special microbial groups
at more significant diversities. Therefore, diverse soil
microorganisms might unite together in response to soil
ecosystem multifunctionality. No single clade is the sole
representative of any of those functions. For example, the
degradation of organic matter from complex to simple
structures requires many distinct C degradation-related
enzymes secreted by different microorganisms (Hooper
et al., 2000; Wardle et al., 2004; Polizeli et al., 2005). In
short, most metabolic functions are not monophyletic
(Aguilar et al., 2004; Martiny et al., 2013), and microbial
decomposers are particularly important drivers of ecosys-
tem functioning (Soliveres et al., 2016).
In summary, to our knowledge, this study is the first to
reveal the linkage between microbial diversity and assem-
bly mechanisms and soil multifunctionality in a subtropical
forest ecosystem. We demonstrate that bacterial and fungal
diversities are negatively related to soil multifunctionality,
although bacterial diversity is significant. The mechanisms
underlying microbial community assembly differ between
bacteria and fungi in this mountain forest soil. For soil bac-
teria, variable selection (deterministic) dominated across
elevational sites, and bacterial community assembly
governed multifunctionality. In contrast, for soil fungi, dis-
persal limitation (stochastic) appeared to dominate across
the forest soils but did not determine the soil multi-
functionality. These findings improve our understanding of
how bacterial and fungal community diversities and assem-
bly processes predict multifunctionality, and this allows us

to execute more efficient ways to maintain soil microorgan-
isms and their functions in subtropical forest ecosystems.
Experimental procedures
Detailed information regarding the experimental proce-
dures can be found in the Appendix S1.
Acknowledgements
This work was supported by National Natural Science Foun-
dation of China (42020104003 and 41830756) and National
Key Research and Development Program of China
(2018YFE0105600).
Author contributions
SH designed the study; SH collected soil samples, per-
formed experiments and statistical analyses with ST and
AW; SH wrote the draft with WC and QH; WC: resources,
supervision, writing-review and editing; QH: resources,
supervision, writing-review and editing. All authors read
and approved the final manuscript.
Data availability statement
The 16S rRNA and ITS sequences were uploaded to
NCBI Sequence Read Archive (SRA) under accession
numbers PRJNA647425 and PRJNA647426
respectively.
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Supporting Information
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Appendix S1: Supplementary Information.