Chicory (Cichorium intybus) reduces cyathostomin egg excretion and larval development in grazing horses.

Cyathostomins are the most prevalent parasitic nematodes of grazing horses. They are responsible for colic and diarrhea in their hosts. After several decades of exposure to synthetic anthelmintics, they have evolved to become resistant to most compounds. In addition, the drug-associated environmental side-effects question their use in the field. Alternative control strategies, like bioactive forages, are needed to face these challenges. Among these, chicory (Cichorium intybus, Puna II cultivar (cv.)) is known to convey anthelmintic compounds and may control cyathostomins in grazing horses. To challenge this hypothesis, we measured fecal egg counts and the rate of larval development in 20 naturally infected young saddle horses (2-year-old) grazing either (i) a pasture sown with chicory (n = 10) or (ii) a mesophile grassland (n = 10) at the same stocking rate (2.4 livestock unit (LU)/ha). The grazing period lasted 45 days to prevent horse reinfection. Horses in the chicory group mostly grazed chicory (89% of the bites), while those of the control group grazed mainly grasses (73%). Cyathostomins egg excretion decreased in both groups throughout the experiment. Accounting for this trajectory, the fecal egg count reduction (FECR) measured in individuals grazing chicory relative to control individuals increased from 72.9% at day 16 to 85.5% at the end of the study. In addition, larval development in feces from horses grazed on chicory was reduced by more than 60% from d31 compared to control individuals. Using a metabarcoding approach, we also evidenced a significant decrease in cyathostomin species abundance in horses grazing chicory. Chicory extract enriched in sesquiterpenes lactones was tested on two cyathostomins isolates. The estimated IC50 was high (1 and 3.4 mg/ml) and varied according to the pyrantel sensitivity status of the worm isolate. We conclude that the grazing of chicory (cv. Puna II) by horses is a promising strategy for reducing cyathostomin egg excretion and larval development that may contribute to lower the reliance on synthetic anthelmintics. The underpinning modes of action remain to be explored further.

Comparison of two molecular barcodes for the study of equine strongylid communities with amplicon sequencing.

Basic knowledge on the biology and epidemiology of equine strongylid species still needs to be improved to contribute to the design of better parasite control strategies. Nemabiome metabarcoding is a convenient tool to quantify and identify species in bulk samples that could overcome the hurdle that cyathostomin morphological identification represents. To date, this approach has relied on the internal transcribed spacer 2 (ITS-2) of the ribosomal RNA gene, with a limited investigation of its predictive performance for cyathostomin communities. Using DNA pools of single cyathostomin worms, this study aimed to provide the first elements to compare performances of the ITS-2 and a cytochrome c oxidase subunit I (COI) barcode newly developed in this study. Barcode predictive abilities were compared across various mock community compositions of two, five and 11 individuals from distinct species. The amplification bias of each barcode was estimated. Results were also compared between various types of biological samples, i.e., eggs, infective larvae or adults. Bioinformatic parameters were chosen to yield the closest representation of the cyathostomin community for each barcode, underscoring the need for communities of known composition for metabarcoding purposes. Overall, the proposed COI barcode was suboptimal relative to the ITS-2 rDNA region, because of PCR amplification biases, reduced sensitivity and higher divergence from the expected community composition. Metabarcoding yielded consistent community composition across the three sample types. However, imperfect correlations were found between relative abundances from infective larvae and other life-stages for Cylicostephanus species using the ITS-2 barcode. While the results remain limited by the considered biological material, they suggest that additional improvements are needed for both the ITS-2 and COI barcodes.

Species interactions, stability, and resilience of the gut microbiota - Helminth assemblage in horses.

The nature and strength of interactions entertained among helminths and their host gut microbiota remain largely unexplored. Using 40 naturally infected Welsh ponies, we tracked the gut microbiota-cyathostomin temporal dynamics and stability before and following anthelmintic treatment and the associated host blood transcriptomic response. High shedders harbored 14 species of cyathostomins, dominated by Cylicocyclus nassatus. They exhibited a highly diverse and temporal dynamic gut microbiota, with butyrate-producing Clostridia likely driving the ecosystem steadiness and host tolerance toward cyathostomins infection. However, anthelmintic administration sharply bent the microbial community. It disrupted the ecosystem stability and the time-dependent network of interactions, affecting longer term microbial resilience. These observations highlight how anthelmintic treatments alter the triangular relationship of parasite, host, and gut microbiota and open new perspectives for adding nutritional intervention to current parasite management strategies.

Effect of sainfoin (Onobrychis viciifolia) on cyathostomin eggs excretion, larval development, larval community structure and efficacy of ivermectin treatment in horses.

Alternative strategies to chemical anthelmintics are needed for the sustainable control of equine strongylids. Bioactive forages like sainfoin (Onobrychis viciifolia) could contribute to reducing drug use, with the first hints of in vitro activity against cyathostomin free-living stages observed in the past. We analysed the effect of a sainfoin-rich diet on cyathostomin population and the efficacy of oral ivermectin treatment. Two groups of 10 naturally infected horses were enrolled in a 78-day experimental trial. Following a 1-week adaptation period, they were either fed with dehydrated sainfoin pellets (70% of their diet dry matter) or with alfalfa pellets (control group) for 21-days. No difference was found between the average fecal egg counts (FECs) of the two groups, but a significantly lower increase in larval development rate was observed for the sainfoin group, at the end of the trial. Quantification of cyathostomin species abundances with an ITS-2-based metabarcoding approach revealed that the sainfoin diet did not affect the nemabiome structure compared to the control diet. Following oral ivermectin treatment of all horses on day 21, the drug concentration was lower in horses fed with sainfoin, and cyathostomin eggs reappeared earlier in that group. Our results demonstrated that short-term consumption of a sainfoin-rich diet does not decrease cyathostomin FEC but seems to slightly reduce larval development. Consumption of dehydrated sainfoin pellets also negatively affected ivermectin pharmacokinetics, underscoring the need to monitor horse feeding regimes when assessing ivermectin efficacy in the field.

Genetic × environment variation in sheep lines bred for divergent resistance to strongyle infection.

Drug-resistant parasites threaten livestock production. Breeding more resistant hosts could be a sustainable control strategy. Environmental variation linked to animal management practices or to parasite species turnover across farms may however alter the expression of genetic potential. We created sheep lines with high or low resistance to Haemonchus contortus and achieved significant divergence on both phenotypic and genetic scales. We exposed both lines to chronic stress or to the infection by another parasite Trichostrongylus colubriformis, to test for genotype-by-environment and genotype-by-parasite species interactions respectively. Between-line divergence remained significant following chronic stress exposure although between-family variation was found. Significant genotype-by-parasite interaction was found although H. contortus-resistant lambs remained more resistant against T. colubriformis. Growth curves were not altered by the selection process although resistant lambs were lighter after the second round of divergence, before any infection took place. Breeding for resistance is a sustainable strategy but allowance needs to be made for environmental perturbations and worm species.

Ivermectin treatment in lactating mares results in suboptimal ivermectin exposure in their suckling foals.

The management of equine strongyles has become problematic over the last decade because of an increased prevalence of drug-resistant isolates worldwide. Therapeutic options are therefore limited, leaving macrocyclic lactones as the most often effective drug class. However, their lipophilic properties result in a long-lasting elimination that could favour drug resistance selection. As a result, ivermectin treatment in lactating mares could promote suboptimal exposure of their foal parasites to ivermectin, thereby selecting for more resistant worms. To test for this putative transfer, we selected two groups of six foal-mare pairs, one group of mares receiving ivermectin and the other being left untreated. We compared faecal egg count trajectories in foals from the two groups and quantified plasma ivermectin concentrations in ivermectin treated mares and their foals during seven days. Our results showed limited but sustained plasmatic exposure of foals associated with non-significant faecal egg count reduction (P = 0.69). This suggests that ivermectin treatment in lactating mares results in suboptimal exposure to the drug in their foal.

Effects of dietary yeast strains on immunoglobulin in colostrum and milk of sows.

The ban of antibiotic growth promoters in pig diet required the development of alternative strategies and reinforced the importance of maternal immunity to protect neonates from intestinal disorders. Milk from sows fed active dry yeasts during gestation and lactation exhibited higher immunoglobulin (Ig) and protein content in milk at day 21 of lactation. In this study, we investigated whether the administration of Saccharomyces cerevisiae strains of various origins (Sc01, Sc02, Sb03) to sows during late gestation and lactation could induce higher Ig content in colostrum and milk. Results show that yeast supplementation did not increase significantly sow body weight at days 112 of gestation and 18 of lactation as well as piglet body weight gain from birth to weaning. In contrast, the IgG level in colostrum was increased in comparison with the control group when sows were supplemented with Sc01 at both 0.05 and 0.5% (p<0.05) and Sb03 at 0.5% (p<0.01). During the lactation, the level of milk IgG remained significantly higher in comparison with the control group when sows were supplemented with Sc02 at 0.05% and 0.5% and with Sb03 at 0.5%. Furthermore, in comparison with the control sows, the level of milk IgA was significantly maintained in sows supplemented with the 3 yeast strains at 0.05%. The incidence of piglet diarrhoea was decreased in groups Sc01 at both 0.05% and 0.5% and Sc02 at 0.05%. Thus, these results show that the 3 yeast strains display immunostimulatory effects on maternal immunity, but only Sc01 supplementation at 0.05% allowed jointly the increase of IgG level in colostrum, the maintenance of IgA level in milk and the decrease of piglet diarrhoea incidence. This stimulation of maternal immunity could be associated with a better systemic (colostrum IgG) and local (milk IgA) protection of neonates and suggests that dietary yeasts may have stimulated the local gut immune system of sows.