Comparing microbial communities between gastric compartments in Hanwoo steers using 16S rRNA amplicon sequencing data

초록

Introduction: In ruminants, the rumen and abomasum differ in physicochemical conditions and harbor distinct microbial communities that affect digestion and host metabolism. 16S rRNA amplicon sequencing is a standard culture-independent method for profiling these communities, but the raw reads must undergo several bioinformatic steps before taxonomic composition and diversity can be compared between samples. This technical protocol describes a reproducible QIIME 2 and R workflow for comparing the rumen (RUM) and abomasum (ABO) microbial communities of Hanwoo steers using 16S rRNA amplicon sequencing data while accounting for the pairing between samples from the same animal. Practice: The workflow is demonstrated on paired RUM and ABO samples from 62 Hanwoo steers. It comprises primer and adapter trimming, DADA2 denoising and ASV inference, SILVA taxonomic classification, and rarefaction-based diversity estimation in QIIME 2. This is followed by paired alpha and beta diversity testing, relative abundance profiling, and differential abundance analysis in R. Summary: The protocol offers a reproducible framework for comparing microbial communities between paired sample sites. The same steps can be adapted to other paired or compartment-based microbiome comparisons and can be extended to other animal species. This is relevant to animal breeding and genomics studies in which microbiome measures are analyzed alongside traits such as feed efficiency and carcass traits.

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