2024-2025, Goat intestinal fecal microbiota sequencing

The collected fecal samples were sent to Novogene for amplicon sequencing analysis of the V3-V4 region of the bacterial 16S rRNA gene. The sequencing results were used to construct libraries and perform quantitative analysis. Based on the required effective library concentration and data volume, quantified libraries were pooled and sequenced on the Illumina MiSeq platform. Raw data were processed using Fastp software to filter and remove chimeric sequences, yielding final high-quality valid data. Additionally, denoising was performed with DADA2 in QIIME2 software (version QIIME2-20202) to obtain Amplicon Sequence Variants (ASVs). Taxonomic annotation was conducted using the QIIME2 software. The relative abundance of microbial taxa was calculated as follows: at a given taxonomic level (phylum, class, order, family, or genus), the relative abundance of a specific species was defined as the number of tags corresponding to an ASV divided by the total number of tags clustered from all ASVs in the sample. The H group represents fecal samples from healthy goats, while the C.p group represents fecal samples from clinically abscess-affected goats.

本研究采集的粪便样本被送至诺禾致源（Novogene），用于开展细菌16S核糖体RNA基因（16S rRNA gene）V3-V4可变区的扩增子测序（amplicon sequencing）分析。测序结果被用于构建文库并开展定量分析。根据所需的有效文库浓度与数据量，将定量后的文库进行混合，并在Illumina MiSeq测序平台（Illumina MiSeq）上完成测序。原始数据经Fastp软件（Fastp）处理，通过过滤及嵌合序列移除操作，最终得到高质量的有效数据。此外，借助QIIME2软件（版本号QIIME2-20202）中的DADA2工具进行降噪处理，以获取扩增子序列变异体（Amplicon Sequence Variants，ASVs）。分类学注释采用QIIME2软件完成。微生物类群的相对丰度计算方式如下：在指定的分类学水平（门、纲、目、科或属）下，某一特定类群的相对丰度定义为该扩增子序列变异体（ASV）对应的标签数，除以样本中所有ASVs聚类得到的总标签数。H组代表健康山羊的粪便样本，C.p组则代表临床脓肿感染山羊的粪便样本。