3 A comparison of the equine fecal microbiome within different horse populations

Abstract

The equine fecal microbiome may vary across horse populations due to the diversity of the habitual diet. The purpose of this study was to assess and compare the microbial population of different horse populations, specifically the differences between feral versus domesticated populations. Samples were collected from 3 different populations of horses: horses from the Shackleford Banks (n = 24), a feral horse population living on the Outer Banks of North Carolina who eat native grasses such as Spartina marsh and island grasses; horses from the NCSU Equine Educational Unit (n = 18) that are predominantly kept on cool season mixed pastures and may be supplemented with hay and concentrates from time to time; and finally, privately owned horses (n = 36) that are fed mixed diets consisting of pasture, hay and concentrates. Horses were monitored and samples were collected immediately following a void by swabbing the middle of the void. Swabs were placed in a tube containing 500 uL DNA/RNA shield (Zymo Research, Irvine, CA) and were sent to the Emerging Technology Center (Purina Animal Nutrition, Gray Summit, MO) where they were stored at −80°C and then the V3 and V4 regions of the 16S rRNA gene were sequenced following the Illumina 16S Protocol (San Diego, CA). Samples were processed, filtered and trimmed through DADA2 using the QIIME2 pipeline. Statistical analysis was performed in R(Version 4.1.1) and a P-value of ≤0.05 was considered significant. After processing to eliminate samples with low sampling depth (<20,085), 78 total samples across the 3 populations were analyzed. For the results, when testing α diversity with Shannon's Index, a Kruskal-Wallis rank sum test revealed a significant difference between all populations (P = 0.01). There was a visual distinction between the Shackleford Banks population compared with the others when utilizing Bray-Curtis to assess β diversity. Additionally, an apparent significant difference between all populations using the PERMANOVA UniFrac test (P < 0.001) was observed. The 3 most predominant bacterial phylum seen across all populations were Firmicutes, Bacteroidetes and Spirochaetes. The top 5 phyla observed in the Shackleford Banks population were Firmicutes, Bacteroidetes, Spirochaetes, Kiritimatiellaeota and Fibrobacteres. Based on the results, there is a distinctive separation in microbial diversity between these horse populations, specifically between the Shackleford Banks horses versus the NCSU and privately owned horses. This separation is likely due to the habitual diet of these specific horse populations influencing the composition of their microbiome within the hindgut.