*2.1. Study Area*

This study area was in Lushan Nature Reserve in Jiangxi province, encompassing a total area of 30.2 km2. The region has a subtropical monsoon climate, an annual average rainfall of 2070 mm, an annual average temperature of 11.6 ◦C, a maximum temperature of 31.1 ◦C, and a minimum temperature of −16.7 ◦C. The average annual fog days are 191 days, and the frost periods are 150 days. In recent years, many moso bamboo forests have been expanding into the surrounding Japanese cedar forests, in areas over 800 m above sea level. Moreover, the number of moso bamboo plants has increased sharply, which greatly affects the forest landscape pattern and the stability of the forest ecosystem in the protected area.

#### *2.2. Experimental Design and Sample Collection*

Complete randomized experimental design was used in this study. Japanese cedar forests with different mixing rate of moso bamboo were selected to represent the expanding stage of moso bamboo into Japanese cedar plantations. Specifically, the research area was divided into four treatments, including Japanese cedar forests (115◦5717" E, 29◦3242" N), altitude 980 m; mixed 1 (30% moso bamboo and 70% Japanese cedar) (115◦5721" E, 29◦3246" N), altitude 980 m; mixed 2 (60% moso bamboo and 40% Japanese cedar) (115◦5712" E, 29◦3244" N), altitude 960 m; and moso bamboo forests (115◦5712" E, 29◦3244" N), altitude 950 m. Each treatment including four replications, and each of them were separated by at least 500 m in the studied area. In July 2020, soils were collected from four 20 × 20 m sample plots. A five-point sampling method was adopted, and a soil drill (diameter of 5 cm) was used to take soil samples from the 0–20-cm soil layer, which were transported back to the laboratory in an icebox. The soil samples from the five points were mixed evenly by plots, the visible stones and plant residues (such as roots, stems and leaves) were carefully removed from the fresh samples, and the samples were then passed through a 2-mm sieve for the experiment. At the same time, the litter of Japanese cedar and moso bamboo were collected and brought back to the laboratory.

#### *2.3. Soil and Litter Chemical Characteristic Analyses*

We removed part of the soil sample for air drying, one part for the determination of soil pH, and another part for the determination of soil total organic carbon (TOC) and total nitrogen (TN), after screening with a pore sieve of 0.149 mm. Some of the remaining fresh samples were placed at 4 ◦C and measured for available nitrogen (AN = NH4+-N + NO3−-N) soon after, while the other samples were placed in a −80 ◦C refrigerator for microbiological measurements [30]. Soil pH was calculated using the electric electrode method (water: soil = 2.5:1) (Metter Toledo, Shanghai, China). Moso bamboo and Japanese cedar litter was dried at 60 ◦C and weighed, and the litter carbon (LC) and litter total nitrogen (LN) contents were determined after passing through a 0.149-mm aperture sieve. TOC and LC were determined by the potassium dichromic (H2SO4–K2Cr2O7) method; and soil TN and LN, NO3−-N, and NH4+-N were determined by automatic analyzer (Smart Chem 200, Westco, Rome, Italy) after being extracted by 2 mol L−<sup>1</sup> KCl solution [30].

#### *2.4. DNA Extraction and Gene Sequencing*

DNA (Deoxyribonucleic acid) was extracted from 0.5 g of fresh soil samples, and the DNA concentration and purity (0.8% agarose gel) were monitored. The primers used for bacterial 16S rRNA were 338F: 5-ACTCCTACGGGAGGCAGCA-3, 806R: 5-GGACTACHVGGGTWTCTAAT-3 for the V4 region. The soil fungal ITS1 region was detected by 18S rRNA, and PCR amplification was performed. The primer sequences were ITS5F: 5-GGAAGTAAAAGTCGTAACAAGG-3 and ITS2R: 5-GCTGCGTTCTTCATCGATGC-3. Approximately 2 μL of template DNA, 1 μL of each forward and reverse primer, and 15 μL of Phusion High-Fidelity PCR Master Mix (New England Biolabs, Ipswich, MA USA) were used for the PCR reactions. The thermal-cycling program was as follows: after the

components required for the PCR (Polymerase Chain Reaction) reaction were configured, the template DNA was fully denatured at 98 ◦C for 30 s on the PCR instrument, following which the amplification cycle was entered. In each cycle, the template was denatured by maintaining it at 98 ◦C for 15 s, following which the temperature was reduced to 50 ◦C for 30 s, allowing for the primer and template to be fully annealed. At 72 ◦C for 30 s, the primer was extended on the template to synthesize DNA, completing a cycle. This cycle was repeated 25 to 27 times, resulting in a large accumulation of amplified DNA fragments. Finally, the product was kept at 72 ◦C for 5 min to complete the extension and was stored at 4 ◦C. The amplification results were electrophoresed by 2% agarose gel. The target fragment was cut and then recovered with an Axygen gel recovery kit. The samples were sequenced using the Illumina NovaSeq platform for high-throughput sequencing by Shanghai Personalbio Technology (Shanghai, China).
