*2.1. Field Description*

The study region is located in Wanglang National Nature Reserve (32◦49–33◦02 N, 103◦55–104◦10 E; 2300–4983 m a.s.l.), Pingwu County, Sichuan, southwestern China, which is distributed on the eastern Qinghai-Tibet Plateau and in the upper reaches of Yangtze River. The area has a semihumid climate typical of the Danba-Songpan region. The annual precipitation is 859.9 mm, and the annual mean temperature is 2.9 ◦C, with maximum and minimum mean temperatures of 12.7 ◦C (in July) and −6.1 ◦C (in January), respectively [35]. The subalpine forest consists of abundant plant species, including mosses, herbs, shrubs, deciduous broadleaved trees and coniferous trees.

#### *2.2. The Method of Selection of Study Plots and Their Characteristics*

Wanglang subalpine coniferous region seriously damaged by long-term the commercial logging of natural forests below 2700 m since the 1950s and natural disturbance such as the 1976 Songpan-Pingwu earthquake [38,39]. Forest logging in the area ceased in 1962. Therefore, the Wanglang Nature Reserve has formed many secondary forests in different stages of succession and primary forests that have never been destroyed [35,38–40]. We considered their distribution based on forest disturbance history and natural recovery process and interviews with local supervisors, We defined the forests of S1 to S5 succession stages by referring to the succession division of Zhang et al. [35], and defined the primary coniferous forests which have never been destroyed to S6 succession stage. According to Zhang et al. [35], the S1 successional stage was dominated by shrubs (e.g., *Salix* spp., *Ulmus pumila* L. and *Hippophae rhamnoides* L.); the S2 successional stage was dominated by the deciduous broadleaved species *Betula* spp. and *Populus* spp.; the S3 successional stage was characterized by mixed forests dominated by deciduous broadleaved species (e.g., *Betula* spp.) and coniferous species (e.g., *Abies faxoniana* Rehd.); the S4 successional stage was dominated by middle-age coniferous species with a diameter at breast height (DBH) less than 20 cm (e.g., *Larix gmelinii* (Rupr.) Kuzen. and *Sabina saltuaria* Rehd.); the S5 successional stage was dominated by mature coniferous plants with a DBH less than 40 cm and more than 20 cm (e.g., *A. faxoniana* Rehd.); and the S6 succession stage was characterized as primary coniferous forest dominated by *Picea purpurea* Mast (Table S1). The traits of the six successional stages, including the coordinates, altitudes, directions, canopy covers, soil names and dominant species are shown in Table S1.

#### *2.3. Soil Sampling Method*

Soils across the subalpine forest successional series were sampled in August 2019. Replicate plots (10 m × 20 m) were established in each successional stage as follows: 12 plots in S1, 9 plots in S2, 9 plots in S3, 6 plots in S4, 6 plots in S5 and 6 plots in S6. After removing the litter layers, nine soil samples in each plot were randomly collected from the top 20 cm of the soil profiles by a stainless steel corer (5-cm diameter) and mixed. Moreover, the topsoil temperature (T) of each plot was measured by a portable soil thermometer (CEM,

DT-131, Shenzhen, China). The roots and stones were removed from the soil samples, which were then divided into two subsamples. One subsample was stored in a bag on ice, immediately transported to a laboratory, and stored at −80 ◦C for high-throughput sequencing for microbial community diversity analysis [31]. The other subsample was transported to the laboratory and air-dried for soil property analysis.

#### *2.4. Soil Agrochemical and Chemical Properties Analyses*

After being passed through a 1-mm sieve, the air-dried soil samples were used for chemical analyses. Soil pH was measured at a soil: water ratio of 1:2.5 ( *m*:*v*) with a digital pH meter (FE20K, Mettler-Toledo, Greifensee, Switzerland). The concentrations of total nitrogen (TN) in the soils were determined from a milled sample by combustion at 950 ◦C using an elemental analyzer (Vario EL Ш, Elementar, Langenselbold, Germany). Soil organic carbon (SOC) was measured by the potassium dichromate oxidation method [41]. The C:N ratio (C:N) was calculated as the mass ratio of SOC and TN [42].

#### *2.5. Soil DNA Extraction, PCR Amplification and High-Throughput Sequencing*

Total soil DNA was extracted from 0.5 g soil samples using the FastDNA Spin Kit for Soil (MP Biomedicals, Santa Ana, CA, USA) according to the instructions of the manufacturer. The V4 hypervariable region of the 16S rDNA [43] and the internal transcribed spacer (ITS1 and ITS2) region of rDNA [44] were selected as sequencing targets for bacteria and fungi, respectively. The library was built using the NEBNext® Ultra ™ DNA Library Prep Kit for Illumina (NEB, Ipswich, MA, USA) at Novogene Company (Beijing, China). Sequencing was performed using a paired\_end sequencing strategy with barcode connectors based on the Illumina MiSeq PE250 platform (Illumina Inc., San Diego, CA, USA). The barcode sequences were spliced with FLASH (v 1.2.7) software to obtain the raw tags [45]. The clean tags were obtained by quality filtering the raw tags using QIIME (v1.9.0) software [46,47]. Chimeric sequences were removed using the UCHIME algorithm [48]. Operational taxonomic unit (OTU) clustering for all the effective tags with 97% identity was performed by UPARSE (v 7.0.1001) software after discarding singletons [49]. The SSUrRNA database of SILVA132 [50] and Mothur software, and the UNIT (v 7.2) database [51] and QIIME (v 1.9.0) [52] software were selected as species annotation analysis for 16S rDNA and ITS, respectively.
