Species diversity and within-host tropism for mixed equine strongyle infections using a cytochrome c oxidase subunit I metabarcoding approach.

Strongyles, especially non-migratory Strongylidae, are the most common equine gut parasites, and typically occur in mixed infections with 10 - 20 species per host. Current knowledge on strongyle species prevalence and within-host tropism is sparse. Herein species composition of mixed strongyle infections of 12 naturally infected untreated young horses, based on strongyle eggs extracted from faeces, cultured third stage larva and luminal worms collected from ventral and dorsal colon was examined. Species were identified using a cytochrome c oxidase I (COI) metabarcoding approach allowing differentiation of operational taxonomic units (OTU) in small strongyle species complexes. In total, 16 small strongyle and one large strongyle species were identified. Eleven small strongyles were detected in all equines, including three cryptic species: Cylicostephanus calicatus OTU II, Cylicostephanus minutus OTU II and III. Coherence of detection rate, alpha and beta diversity showed high agreeance between data obtained using DNA from faecal eggs and cultured larvae. Diversity, but not observed richness, was higher in eggs and larvae compared to luminal worms (Inverse Simpson index, Shannon index, all P < 0.05). Bray-Curtis and Jaccard dissimilarity showed overlapping beta diversity clusters for eggs and larva, while ventral and dorsal colon samples clustered separately and differed significantly according to PERMANOVA (P < 0.001). Five small strongyle species showed significantly higher occurrence in the ventral than dorsal colon, including cryptic species Cylicostephanus calicatus OTU II. This study provides novel prevalence data for five cryptic species, documents species tropism within intestinal compartments and demonstrates high strongyle species composition agreement between faecal eggs and cultured larvae.