design: The V4 region of the 16S rRNA gene was amplified using the primers 515f and 806r (forward: 5-GTGCCAGCMGCCGCGGTAA-3, reverse: 5-GACTACHVGGGTWTCTAATCC-3), and indexes were added using the Nextera XT DNA Library Preparation Kit (Illumina, Inc., USA). Sequencing was performed on a MiSeq platform (Illumina, Inc., USA).