Amplicon Seq (NGS v3-v4) of   Acanthopagrus schlegelii gut microbial community: AH Group Sample5
The PowerFecal DNA Isolation Kit (MoBio Laboratories, Carlsbad, CA, United States) was employed to extract DNA from intestinal samples, in accordance with the manufacturer's instructions, with six samples selected from each group. Extraction negative controls were included to monitor for potential contamination. The quantities and qualities of the extracted DNA were verified through a NanoDrop 2000 (ThermoFisher, Wilmington, DE, United States) and gel electrophoresis. The V3V4 region of bacterial 16S rRNA gene was amplified by PCR using the primers of ~20 ng with the following primer order: forward primers: 5-ACTCCTACGGGAGGCAGCA-3; reverse primers: 5-GGACTACHVGGGTWTCTAAT-3 (Pan et al., 2021). The PCR negative controls (no-template controls) were also included in the amplification step. The high-throughput sequencing was performed by the Shanghai Personal Biotechnology Co., Ltd. (Shanghai, China) on an Illumina platform (Illumina, San Diego, CA, US) with a PE250 strategy in accordance with the previously described method (Wang et al., 2020).