title: sludge
design: PCR amplification of the bacterial 16S rRNA gene was carried out using primers targeting the V3-V4 variable region, 338F (-ACTCCTACGGGAGGCAGCA-) / 806R (-GGACTACHVGGGTWTCTAAT-) (Dennis et al., 2013). A 2% agarose gel electrophoresis was used to select mixed PCR products. The AxyPrep gel extraction kit (AXYGEN, USA) was used to remove non-specific amplification products. The purified samples were subjected to amplicon sequencing of all samples using the Illumina MiSeq platform by Majorbio Biopharm Biotechnology Co., Ltd (Shanghai, China). The raw sequences were assembled, and low-quality sequences and chimeras were removed. Sequence numbers of each sample were clustered, and the assembled sequences were classified into operational taxonomic units (OTUs) using Usearch software with a 97% similarity threshold. The microbial communities at various taxonomic levels were annotated using the Ribosomal Database Project classifier based on the Silva database.