Vaginal and Uterine Microbiota of Healthy Maiden Mares during Estrus

Abstract

This descriptive cross-sectional study compared the microbiota of the uterus, vagina, clitoral fossa (CF), and perineal skin in healthy maiden mares during estrus. Twelve synchronized, healthy maiden mares (3-4 years old) from one single recipient mare herd were included. Microbial communities were characterized by amplifying the V3-V4 region of the 16S rRNA gene using the Illumina MiSeq platform. The uterine and vaginal microbiota had significantly lower richness (Chao-1) than the skin (p < 0.05). The uterine and vagina bacterial composition was similar in presence and abundance and could be differentiated from that of the CF and perineal skin. The microbial composition (Jaccard and Bray-Curtis distances) significantly differed across body-site locations (p < 0.05), which explained approximately 14% and 19% of the variation in microbial composition for Jaccard and Bray-Curtis distances, respectively. Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes were the dominant taxa in the uterus and vagina, with higher proportions of Proteobacteria in the vaginal samples compared to the uterine samples. Streptococcaceae and Staphylococcaceae were present in high abundance in the uterine and vaginal samples, while Lactobacillaceae were not (<10%). We demonstrate that the uterine and vaginal microbiota of healthy maiden mares during estrus is similar but both distinct from that of the CF and perineal skin.

Keywords: cervix; clitoral fossa; fertility; metritis.

Figure 1

Alpha diversity metrics of uterine, vaginal, clitoral fossa (CF), and perineal skin samples from healthy mares during estrus. Indices measured were (A) observed ASV count, (B) Chao1 (richness), (C) Shannon’s index, and (D) Fisher’s alpha. Different letters for the same alpha diversity measure represent a significant difference (p < 0.05).

Figure 2

Principal coordinates analysis distances ((A) Jaccard and (B) Bray–Curtis dissimilarities) of the uterine, vaginal, clitoral fossa (CF), and perineal skin samples from healthy mares during estrus. Samples are colored according to site location.

Figure 3

(A) Bacterial relative abundance for each mare and swab location. Relative abundance was calculated based on taxonomically classifiable bacteria at each taxonomic rank analyzed. Bacteria were grouped at the phylum level, and bacteria under 5% relative abundance for each swab location were grouped for clearer visualization. (B) Bacterial relative abundance for each mare and swab location. Relative abundance was calculated based on taxonomically classifiable bacteria at each taxonomic rank analyzed. Bacteria were grouped at the family level, and bacteria under 10% relative abundance for each swab location were grouped for clearer visualization.

Figure 3

(A) Bacterial relative abundance for each mare and swab location. Relative abundance was calculated based on taxonomically classifiable bacteria at each taxonomic rank analyzed. Bacteria were grouped at the phylum level, and bacteria under 5% relative abundance for each swab location were grouped for clearer visualization. (B) Bacterial relative abundance for each mare and swab location. Relative abundance was calculated based on taxonomically classifiable bacteria at each taxonomic rank analyzed. Bacteria were grouped at the family level, and bacteria under 10% relative abundance for each swab location were grouped for clearer visualization.

Figure 4

Differential abundance analysis of bacterial families for each mare swab location against air swab abundance. Differential abundances were calculated using DESeq2, and average log2 fold change is compared to air swab abundance. Only bacteria that were significantly more/less abundant in mare swab locations (p < 0.01) compared to air swabs are shown. Bacterial families were colored based on phyla taxonomy, and dot shape represents the mare swab location.