Abstract
The composition of the equine gut microbiome is associated with many aspects of gastrointestinal, respiratory and musculoskeletal health that have been reported in the horse. Scientific studies exploring the microbiome non-intestinal ecological niches in or on horses are lacking. The clinical use of bacterial community profiling in horses is currently limited by cost and by slow analytical workflows.
Most equine microbiome studies have relied on 16S rRNA amplicon sequencing of bacterial DNA, using high-throughput short-read sequencing technologies. This is often provided by an external service due to the cost of Illumina and other sequencers. Analysis of such sequencing files relies upon the researcher to have prior experience of coding-based programs.
To explore the utility of Oxford Nanopore long-read sequencing in the analysis of microbiomes from several anatomical sites of the horse as a quicker and cheaper alternative to short-read sequencing.
Bacterial DNA was extracted from horse (udder) skin swabs, saliva swabs, faecal samples and milk samples. Samples were prepared for Oxford Nanopore long-read sequencing and sequenced using a flow cell on the MinION Mk1D. Sequencing data were analysed using EPI2ME, along with extra analyses on exported taxa abundance data in R.
Diversity measures and taxonomic relative abundance from phylum to family level were comparable to previously published equine studies that used Illumina sequencing. Sequencing data were acquired within 3 days costing around £30 per sample. Long-read sequencing gave accurate taxa assignment for two positive controls included at phylum, class, order and family levels of taxonomic classification.
This work demonstrates that long-read technologies such as Oxford Nanopore MinION sequencing can provide a reliable, quick and cost-effective alternative to short-read Illumina sequencing when characterizing microbial communities from a range of anatomical locations on/in the horse.