Ecological research that unravels the ecological processes of microorganisms is a key topic but is poorly understood (Dini-Andreote et al., 2015; Zhou and Ning, 2017). Based on the niche-based theory and neutral theory, the deterministic and stochastic processes are generally used to describe the ecological processes of the community assembly (Dini-Andreote et al., 2015; Zhou and Ning, 2017). Accumulating evidence also suggests that the two theories are not independent. More specifically, the deterministic and stochastic processes act concurrently to regulate the community assembly (Sloan et al., 2006; Bahram et al., 2016). As a common approach based on Stegen et al. (2012), five ecological processes were included in the deterministic and stochastic processes in driving community assembly, that is, the deterministic process includes the heterogeneous selection and homogeneous selection, while the stochastic process harbors dispersal limitation, homogenizing dispersal, and drift. In detail, heterogeneous selection is defined as the selection derived from different abiotic and biotic variables in the community and results in more dissimilarity among communities, by contrast, homogeneous selection implies the selection under similar abiotic and biotic conditions leading to more similar communities. Dispersal limitation claims that the movement and colonization of individuals in a new habitat are restricted; homogenizing dispersal indicates a high rate of dispersal and effective colonization of individuals among communities, which leads to more similar communities; and drift means that the relative abundance of species randomly change in the community over the time due to stochastic process of inherence, such as speciation, random birth, and death. Recently, the relative roles of deterministic and stochastic processes on the microbial community, as well as the contributions of the five ecological processes in controlling the community assembly of microbes, have been extensively investigated. Using a neutral community model (NCM) and a normalized ratio index (NST), for example, Chen et al. (2021) found that the assembly of the microbial community in termite mounds was strongly driven by the deterministic process. Chen et al. (2019) demonstrated that stochastic processes were sufficient in driving river microeukaryotic communities. In some studies, based on the null model analysis proposed by Stegen et al. (2012), detailed ecological processes driving microbial community assembly have been investigated. For example, Zhang et al. (2021) indicated a determinant role of dispersal limitation in the stochastic process on fungal community assembly in mangrove sediments in southern China. Jiao et al. (2021) found that in the agricultural fields, the fungal community of abundant groups was mainly governed by dispersal limitation in the stochastic process, while the homogeneous selection in the deterministic process was the dominant ecological process in controlling community of the rare fungi. Additionally, for the marine benthic archaeal communities, dispersal limitation in the stochastic process was the main community ecological process (Liu et al., 2020). In the study of the assembly mechanism of bacterial communities on plastisphere in water, it was found that the bacterial communities of plastic balls were mainly driven by the stochastic process of drift and dispersal limitation (Sun et al., 2021). These studies have further clarified the different ecological processes and their relative importance in controlling some microbial communities. However, it is obvious that the microbes in the studies mentioned above focused on either bacteria, archaea, and/or fungi in the water and soil environments, that is, “Free-living” microbes. Thus, an important question remains about ecological processes that underlie the assembly of symbiotic microbes such as mycorrhizal fungi, and it remains uncertain and deserves further exploration since they play a vital role in ecosystems. Mycorrhizae is a typical symbiosis formed between soil fungi of certain groups and host plants. There is increasing evidence that mycorrhizal fungi play important roles in host growth, development, and resistance to stress and can provide host plants with the required nitrogen and phosphorus to increase nutrient absorption (Smith and Read, 2012; van der Heijden et al., 2015). Ectomycorrhizal (EM) fungi are certain groups in mycorrhizal fungi, which could form symbiosis (i.e., ectomycorrhizae) with 30 lineages of angiosperms and gymnosperms, accounting for 2% of terrestrial plants and mostly trees. The ratio of EM host plants is low, but the plants (e.g., Pinaceae, Fagaceae, Betulaceae, and Salicaceae) played important roles in the ecosystems, particularly in the forests (Tedersoo et al., 2020).

A growing number of studies have focused on the relative effects of environmental filtering (e.g., host plant phylogeny, soil factors, and climate factors) and dispersal limitations (e.g., spatial distance) on EM fungal community (Gao et al., 2013, 2015; Tedersoo and Smith, 2013; Wu et al., 2018; Wang et al., 2019, 2021a,b). For example, several studies indicated that EM fungal community was mainly affected by host phylogeny and by spatial distance, soil factors, and climate combination factors (Ishida et al., 2007; Tedersoo et al., 2010; Wang et al., 2019, 2021a,b). These studies mainly reveal the effect of abiotic and abiotic variables on the EM fungal community, but the community assembly from the aspect of ecological processes and their relative importance remains unclear and needs further study.

The Helan Mountains are located in the transitional zone between grassland and desert types of vegetation (Jiang et al., 2007). The research site in the Helan Mountains Range is a unique geographic location that connects the flora and climate of the Inner Mongolian Plateau in Northern China. Picea crassifolia Kom. forests account for 90% of the total forest cover of the Helen Mountains Range, but the EM fungal diversity and ecological processes underlying the community assembly of EM fungi on this plant species have received limited attention. With this in mind, we attempted to provide the first insight into P. crassifolia EM fungal diversity in this nutrient-and water-limited boreal area. We addressed the following questions: (1) How about EM fungal diversity and community composition of P. crassifolia in the Helan Mountains? and (2) What ecological processes determine the community assembly of EM fungi associated with P. crassifolia in the Helan Mountains?

Our finding suggested that a high diversity of EM fungi is associated with P. crassifolia in this study. Similar but relatively lower fungal diversity of EM fungi in the study that also involved in single EM host plant has been reported by Wang et al. (2021a), in which 122 EM fungal OTUs with richness 13.1 ± 2.2 (mean ± SE) were found in 21 root samples of Larix gemelinii. In contrast, Matsuoka et al. (2019) investigated 240 blocks of Castanopsis sieboldii and 365 EM fungal OTUs were found. It has been widely accepted that sample size could affect EM fungal diversity in the studies; indeed, our rarefication curves did not reach a plateau, which implied more field work, and samples could bring more undiscovered fungal OTUs. Additionally, the fungal diversity of EM fungi was also affected by the host plant, soil, and climatic variables, which have been reported in many studies conducted at various habitats (Tedersoo et al., 2013; Miyamoto et al., 2018; Wang et al., 2021a). In our study, EM fungal OTUs richness was significantly different across the three sites, and this may be due to the variations in soil and climatic conditions. Previous studies indicated that warming could decrease EM fungal diversity, and accordingly, the MAT in HLSNS and HLSBS was lower than that in HLWG, which may contribute to a relatively higher fungal diversity in the two sites than in HLWG. Additionally, we suspect that proper precipitation could also support the growth of fungi in the semi-arid forests as HLSNS and HLSBS harbored more MAP than HLWG. The soil variables were not recorded in our study, but their influence on EM fungal diversity has been intensively investigated in many previous studies (Lilleskov et al., 2004; Glassman et al., 2015; Miyamoto et al., 2015; Wu et al., 2018; Otsing et al., 2021).

Sebacina, Tomentella-Thelephora, Wilcoxina, and Amphinema-Tylospora were the most dominant EM fungal lineages of P.crassifolia in the Helan Mountains (accounting for 73.786% of total reads), with Sebacina being the most dominant (accounting for 40.828% of total reads). Recent studies have reported that EM fungal community associated with Larix cajanderi in eastern Siberia was characterized by a high proportion of Suillus-Rhizopogon (Lilleskov et al., 2004; Glassman et al., 2015; Miyamoto et al., 2015; Wu et al., 2018; Otsing et al., 2021), and Tricholoma was the most dominant lineage on Larix gemelinii in the Great Khingan Mountains, Inner Mongolia, China (Wang et al., 2021b). Obviously, these studies and our finding showed inconsistency with previous studies performed in various forests and host plants to a certain extent, in which Tomentella-Thelephora, Sebacina, Inocybe, and Russula-Lactarius were commonly the dominant EM fungal lineages (e.g., Tedersoo et al., 2010; Wu et al., 2018; Wang et al., 2019). Based on the data of abundant EM fungal lineages mentioned above, we found that Tomentella-Thelephora was commonly the abundant lineage in many studies, which showed consistency in EM fungal lineages across studies. However, Tomentella-Thelephora was not always ranked at the number one in abundance, and other lineages, such as Sebacina, Tricholoma, and Suillus-Rhizopogon could be the most abundant ones, which indicated inconsistency or unique characteristic of each study. This further indicated that some unstudied EM plants and habitats may host distinct EM fungal groups and communities, which may be due to the host effect or local fungal species pool, and this speculation needs to be confirmed in further study.

Thus, we can speculate that the variation in the distribution and/or preference for the site of EM fungal OTUs contributes to the difference in EM fungal communities among the three sites in present study. Additionally, the variations in EM fungal communities and distributions/preference of EM fungal OTUs could be fundamentally attributed to the niche difference among fungal OTUs, because different OTUs preferred various habitats in terms of temperature, soil water, and nutrients (e.g., Glassman et al., 2017; Miyamoto et al., 2018); for example, some EM fungal OTUs could adapt well to relative cold environments, while some preferred warm habitats (Miyamoto et al., 2018). Indeed, in our study, the climatic conditions were different across the three sites, and the climatic condition of HLWG is relatively warmer and moist than HLSNS and HLSBS. Additionally, the difference in climatic conditions could result in distinct soil nutrients, which further drive EM fungal community variations. Although soil data were not available in our study, the significant influence of soil parameters on EM fungal communities has been intensively explored in previous studies as different niche requirements for various fungal taxa (Glassman et al., 2017; Wang et al., 2021a,b). Thus, we suspect that the difference in environmental conditions among the three sampling sites could bring various living niches for EM fungi, and different EM fungi lived in proper habitats for their living, hence resulting in significant variations in EM fungal communities across the three sites. Site/fungus preference indicated that the site harbored site-preference OTUs, and some OTUs showed site preference and preferred certain sites. On the one hand, we suspect that the spatial distance strongly limited the dispersal of these EM fungi between sites (species pool) and, thus, was limited in certain sites (Peay and Bruns, 2014). On the other hand, these fungal OTUs may disperse successfully but with narrow niches and, thus, cannot be successfully colonized in the new habitats (Miyamoto et al., 2018). In summary, the results of preference analysis reflected the significant co-occurrence of EM fungi with the site and, thus, contributed to the variations in EM fungal communities among the three sites.

Overall, stochastic processes (74.606–93.891%) played a more important role than deterministic process (6.109–25.394%) in controlling community assembly of EM fungi associated with P. crassifolia in Helan Mountains, in which dispersal limitations (38.945–60.864%) and drift (13.057–50.449%) were most important ecological processes (Figure 6B). The scale of investigations is known to potentially influence the importance of stochastic and deterministic processes in shaping microbial communities (Sun et al., 2021). The lack of significant deterministic processes including homogeneous selection and heterogeneous selection may be due to the fact that this study was conducted at a narrow scale (several km, local scale), and the heterogeneities of environmental conditions may be at low levels (Ning et al., 2020; Sun et al., 2021). A growing body of evidence suggests that the dispersal limitation in stochastic processes has been widely accepted in driving community assembly of soil and root microbes (Beck et al., 2015; Glassman et al., 2015; Wang et al., 2019). The dispersal limitation is derived from the limits of spatial distance on the movement of microbes, thus resulting in a low migration rate of microbes and heterogeneous distribution of microbial community along a spatial distance (Chase, 2003; Segre et al., 2014; Catano et al., 2017; Evans et al., 2017; Ning et al., 2020). Correspondingly, the m-value is 0.007 in our study, which suggested a very low dispersal of EM fungi in present study and supported a strong effect of dispersal limitation on community assembly of EM fungi. Drift mirrors the fluctuation of relative abundances of different fungal taxa within a community over time due to the inherently random process of birth, death, and reproduction (Zhou and Ning, 2017). Interestingly, we found a relatively important role of drift than the other ecological processes at the HLWG site, which may be due to the fact that HLWG harbored a relatively lower size of fungal community when compared with those in the other two study sites, as Gao et al. (2020) suggested that stochastic processes were negatively correlated with fungal community size. That is, the smaller the community, the greater effect of the drift. Deterministic processes including heterogeneous and homogeneous selections are defined as ecological forces that drive community composition to become more dissimilar or similar due to fitness differences among different fungal taxa (Zhou and Ning, 2017). In our study, homogeneous selection played a more important role than heterogeneous selection, and this may be caused by homogeneous environments at small spatial scales in our study in comparison with broad spatial scales. Homogeneous selection played a more role in HLSNS than that in HLSBS and HLWG, and we suspected that this may be caused by more even environmental conditions in site HLSBS, although soil data were unavailable in our study. After removing the site-preference OTUs, the ecological processes underlying community assembly changed to some extent, indicating these OTUs with significant preference influenced community assembly, but the stochastic process always dominantly determined community assembly of EM fungi in the present study, and the underlying mechanisms need further investigation in the future study.

Our study revealed that fungal diversity and ecological processes underlying the community assembly of EM fungi are associated with P. crassifolia for the first time. We found a high diversity of EM fungi on P. crassifolia, that is, 166 OTUs were identified as EM fungi, belonging to 24 lineages. There were obvious differences in EM fungal communities among the three sampling sites, and some abundant EM fungal OTUs preferred specific sites. The EM fungal community assembly was determined by a combination of deterministic and stochastic processes, but dispersal limitation and drift in the stochastic processes were the predominant processes. These findings improved our understanding of EM fungal diversity and ecological processes of EM fungal communities associated with a single host plant in a semi-arid forest.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/Supplementary material.

YF and YW conceived and designed this experiment. JM, SX, YS, and YX collected samples and conducted experiments. XZ participated in the acquisition and analysis of the data. XZ and YW wrote the manuscript. JM and YX participated in the discussion draft of the manuscript. YW, BB, and YF revised the final manuscript. All authors contributed to the article and approved the submitted version.

This study was funded by the National Natural Science Foundation of China (nos. 32260006 and 32260027), Inner Mongolia Natural Science Foundation (nos. 2020MS03001 and 2021BS03027), Science and Technology Project of Inner Mongolia Autonomous Region (no. 2019GG002), and the Youth Science and Technology Talent Support Scheme of Colleges and Universities in Inner Mongolia Autonomous Region (no. NJYT-18-A21).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.