**Effects of fermented strawberry pomace on the cecal microbiota of laying hens

Sequencing and analysis of the cecal microbiota were carried out by Shanghai Personal Biotechnology Co., Ltd. (Shanghai, China). Total microbial DNA was isolated from cecal digesta, and PCR amplification targeted the bacterial 16S rRNA gene V3–V4 region, employing universal primer pairs (forward: 5ʹ-ACTCCTACGGGAGGCA-3ʹ; reverse: 5ʹ-GGACTACHVGGGTWTCTAAT-3ʹ). The resulting PCR amplicons were purified before sequencing and subjected to paired-end sequencing on an Illumina NovaSeq system (Illumina, San Diego, CA, USA). The raw sequences were processed by filtering for quality, removing chimera, and clustering into amplicon sequence variants (ASVs), which using Greengenes as the taxonomic reference database. Indices of α and β diversity were calculated to characterize microbial richness, overall diversity, and between-sample dissimilarity. Differences in genus-level abundance among groups were tested with the Kruskal–Wallis nonparametric test. Discriminant taxa among treatments were screened using LEfSe (LDA score ≥ 2, P < 0.05) in combination with random forest modeling. Finally, KEGG pathway prediction was carried out with PICRUSt2, which reconstructs community functions based on phylogenetic information.