Abstract: Background – Taxonomic classification of the equine fecal microbiome using untargeted approaches results in differences in taxonomic resolution and characterization. The development of targeted assays may provide validated reference intervals to complement untargeted approaches allowing comparisons across methods.Hypothesis/Objectives – Compare taxonomic data derived from 16S rRNA gene (16S) and shallow shotgun sequencing (SSS), and targeted qPCR assays performed on one sample set.Animals – Feces from 28 horses (n=17 non-hospitalized, healthy horses; n=11 hospitalized horses with colitis).Methods – DNA was extracted from feces and submitted for 16S and SSS. qPCR for taxa previously found to be discriminatory between states of gastrointestinal health or disease was performed. Taxonomic abundance from 16S and SSS was compared using a Mann-Whitney test and correlations between sequencing and qPCR were made with a Spearman’s test.Results – SSS identified a greater number of taxa at all taxonomic levels. There were significant differences in the abundance of 9 out of 10 phyla, including those important in gastrointestinal health or disease; Bacteroidetes was significantly less abundant by SSS compared to 16S and Verrucomicrobia was not detected by SSS. Spearman’s correlation was significant (p< 0.05) between qPCR and SSS (Blautia, Escherichia coli, Firmicutes, Lactobacillus, Ruminococcus, C. scindens and Streptococcus) and qPCR and 16S (Akkermansia, Lactobacillus, Ruminococcus, and Streptococcus).Conclusions and clinical importance – While SSS has higher resolution at the genus and species level, 16S identified taxa critical to gastrointestinal health/disease. Targeted qPCR assays can be used to validate taxa identified with either untargeted method.
