Detecting microbial communities and pathogens in gastrointestinal tracts of commercial broilers in Thailand using high-throughput sequencing technology with different bioinformatic pipelines
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Abstract
High-throughput sequencing is widely applied to explore microbial communities and detect multiple pathogens simultaneously. The 16S rRNA amplicon sequencing is gaining attention in the poultry industry to monitor animal gut health and pathogens, especially Salmonella. A bioinformatics approach for the accurate characterization of the microbial communities obtained via high-throughput genetic sequences is crucial. This study provides a comparison between the commonly used bioinformatics tools QIIME2 and Mothur for the amplicon-based microbiome analysis of commercial broilers in Thailand and a mock community. We conducted QIIME2 and Mothur with the implementation of amplicon sequence variants (ASVs) and operational taxonomic units (OTUs) for grouping amplicon sequences as a unit of an organism, respectively. The two pipelines provided similar microbial profiles at all taxonomic levels. Most bacteria in the ileum samples belonged to the genera Lactobacillus and Romboutsia. However, different results were found in Salmonella detection. Based on the SILVA138 reference database and the same samples, Mothur provided the taxonomic assigned OTUs of the genus Salmonella, whereas QIIME2 could assign the taxonomy for the identified ASVs only at the level of the family Enterobacteriaceae. We recommend further annotating the ASVs of the family Enterobacteriaceae using a phylogenetic tree and basic local alignment search tool (BLAST) to discriminate the genus Salmonella. This study demonstrates that caution is required for Salmonella detection when performing 16S rRNA amplicon sequencing analysis for the best interpretation output of the tools.
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