Cyathostomin species richness and alpha diversity at egg reappearance following fecal egg count-based selective ivermectin treatment

Cyathostomin nematodes are ubiquitous in grazing horses. The overuse of anthelmintics has led to increasing problems with drug resistance to benzimidazoles and pyrantel. Although macrocyclic lactones (ML), such as ivermectin, generally retain the highest efficacy, reduced ML efficacy has increasingly been reported. A shortened egg reappearance period (ERP) is frequently used as an indicator of reduced susceptibility. However, the species-specific response dynamics after ivermectin treatment are inadequately characterized in cyathostomins. The objective of this study was to compare the cyathostomin community composition before ivermectin treatment and at egg reappearance (ER). We used an ITS2 rDNA metabarcoding protocol followed by next-generation sequencing. Samples were collected from 35 horses before ivermectin treatment and 11 at ER on Swedish farms. DNA was extracted from L3 larval cultures, and the nematode community was sequenced using the PacBio platform. Species identification was based on amplicon sequence variants using a curated ITS2 reference database and BLAST verification. Statistical analyses were conducted to evaluate species richness, alpha diversity indices and their correlation with fecal egg count (FEC) and horse age. Ivermectin treatment was 100% effective, and treatment did not alter the species richness (median four species at week 0 and three at ER, p = 0.71) or two alpha diversity indices at ER (Inverse Simpson, p = 0.41; Shannon-Weiner, p = 0.29). Cylicocyclus nassatus and Cyathostomum catinatum were the two dominant species both before treatment and at ER (weeks 6-8). Minor shifts in the relative abundance of less prevalent species were observed at ER. In pre-treatment samples, no statistically significant associations were detected between species richness and either FEC or species richness and age. The species Cylicocyclus insigne was not detected at ER after ivermectin treatment. At egg reappearance following FEC-based ivermectin treatment, cyathostomin species richness and alpha diversity did not differ detectably from pretreatment.