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The eye is host to myriad bacterial, fungal, and viral organisms that likely influence ocular surface physiology in normal and diseased states. The ocular surface mycobiota of horses has not yet been described using NGS techniques. This study aimed to characterize the ocular surface fungal microbiota (mycobiota) in healthy horses in 2 environmental conditions (stalled versus pasture). Conjunctival swabs of both eyes were obtained from 7 adult stallions stabled in an open-air pavilion and 5 adult mares living on pasture. Genomic DNA was extracted from ocular surface swabs and sequenced using primers that target the Internal Transcribed Spacer 1 (ITS1) region of the fungal genome on an Illumina platform. Sequences were processed using Quantitative Insights Into Molecular Ecology (QIIME 2.0) and taxonomy assigned with the Findley et al. 2013 ITS1 database. The most abundant genera identified were Leptosphaerulina (22.7%), unclassified Pleosporaceae (17.3%), Cladosporium (16.2%), Alternaria (9.8%), unclassified Pleosporales (4.4%), unclassified Montagnulaceae (2.9%), Fusarium (2.5%), and Pestalotiopsis (1.4%). Fungal community composition (Jaccard, R = 0.460, p = 0.001) and structure (Bray-Curtis, R = 0.811, p = 0.001) were significantly different between pastured mares and stabled stallions. The ocular surface of pastured mares had significantly increased fungal species richness and diversity compared to stabled stallions (Shannon p = 0.0224, Chao1 p = 0.0118, Observed OTUs p = 0.0241). Relative abundances of Aspergillus (p = 0.005) and Alternaria spp. (p = 0.002) were significantly increased in the mycobiota of pastured mares. This is the first report to describe the mycobiota of the equine ocular surface. Environmental factors such as housing influence the composition, structure, and richness of the equine ocular surface mycobiota., Competing Interests: The authors have declared that no competing interests exist.

Next generation sequencing (NGS) studies have demonstrated a rich and diverse ocular surface-associated microbiota in people that was previously undetected by traditional culture-based methods. The ocular surface microbiome of horses has yet to be investigated using NGS techniques. This study aimed to determine the bacterial composition of the ocular surface microbiome in healthy horses, and to identify whether there are microbial community changes over time and following topical antibiotic use. One eye of 12 horses was treated 3 times daily for 1 week with neomycin-polymyxin-bacitracin ophthalmic ointment. Contralateral eyes served as untreated controls. The inferior conjunctival fornix of both eyes was sampled at baseline prior to initiating treatment (day 0), after 1 week of treatment (day 7), and 4 weeks after concluding treatment (day 35). Genomic DNA was extracted from ocular surface swabs and sequenced using primers that target the V4 region of bacterial 16S rRNA. At baseline, the most abundant phyla identified were Proteobacteria (46.1%), Firmicutes (24.6%), Actinobacteria (12.6%), and Bacteroidetes (11.2%). The most abundant families included Pasteurellaceae (13.7%), Sphingomonadaceae (7.9%), an unclassified Order of Cardiobacteriales (7.7%), and Moraxellaceae (4.8%). Alpha and beta diversity measurements were unchanged in both treatment and control eyes over time. Overall, the major bacterial taxa on the equine ocular surface remained stable over time and following topical antibiotic therapy., Competing Interests: The authors have declared that no competing interests exist.

Next generation sequencing (NGS) studies in healthy equine eyes have shown a more diverse ocular surface microbiota compared to culture-based techniques. This study aimed to compare the bacterial ocular surface microbiota in both eyes of horses with unilateral ulcerative keratitis (UK) with controls free of ocular disease. Conjunctival swabs were obtained from both ulcerated eyes and unaffected eyes of 15 client-owned horses with unilateral UK following informed consent, as well as from one eye of 15 healthy horses. Genomic DNA was extracted from the swabs and sequenced on an Illumina platform using primers that target the V4 region of bacterial 16S rRNA. Data were analyzed using Quantitative Insights Into Molecular Ecology (QIIME2). The ocular surface of ulcerated eyes had significantly decreased species richness compared with unaffected fellow eyes (Chao1 q = 0.045, Observed ASVs p = 0.045) with no differences in evenness of species (Shannon q = 0.135). Bacterial community structure was significantly different between either eye of horses with UK and controls (unweighted UniFrac: control vs. unaffected, p = 0.03; control vs. ulcerated, p = 0.003; unaffected vs. ulcerated, p = 0.016). Relative abundance of the gram-positive taxonomic class, Bacilli, was significantly increased in ulcerated eyes compared with controls (q = 0.004). Relative abundance of the taxonomic family Staphylococcaceae was significantly increased in ulcerated and unaffected eyes compared with controls (q = 0.030). The results suggest the occurrence of dysbiosis in infected eyes and reveal alterations in beta diversity and taxa of unaffected fellow eyes. Further investigations are necessary to better understand the role of the microbiome in the pathophysiology of ocular surface disease. [ABSTRACT FROM AUTHOR]