Meta-analysis of cyathostomin species-specific prevalence and relative abundance in domestic horses from 1975-2020: emphasis on geographical region and specimen collection method.

BACKGROUND: Cyathostomins infect virtually all horses, and concomitant infections with 10 or more species per horse is standard. Species-specific knowledge is limited, despite potential species bias in development of disease and anthelmintic resistance. This is the first meta-analysis to examine effects of geographical region and cyathostomin collection method on reported composition of cyathostomin communities. METHODS: Thirty-seven articles published in English in 1975 or later, in which adults of individual species were systematically enumerated, were included. Seven regions; North America, South America, eastern Europe, western Europe, northern Europe, southern Africa, and Oceania, and three cyathostomin collection methods; (i) standard necropsy recovery from the large intestine, (ii) critical test collection from post-treatment feces and necropsy, and (iii) diagnostic deworming recovery solely from post-treatment feces, were considered. Generalized mixed linear models analyzed the effects of region and collection method on species-specific prevalence and relative abundance. Species richness was analyzed by mixed linear models. RESULTS: Definitively, the most prevalent and relatively abundant species were Cylicocyclus nassatus (prevalence = 93%, relative abundance = 20%), Cylicostephanus (Cys.) longibursatus (93%, 20%), and Cyathostomum catinatum (90%, 16%). A bias toward horses with high infection intensities and cyathostomin collection from feces resulted in North American critical tests and eastern European diagnostic deworming overestimating the species-specific prevalence and underestimating the relative abundance of rare/uncommon species compared to respective intra-regional standard necropsies. North American critical tests underestimated species richness due partially to identification key errors. Inter-regional standard necropsy comparisons yielded some species-specific regional differences, including a significantly higher Cys. longibursatus prevalence and relative abundance in North America (92%, 33%) than in eastern Europe (51%, 7%) (P > 0.0001). Localization of critical tests to North America and diagnostic deworming to Eastern Europe precluded expansive 'region by collection method' interaction analyses. CONCLUSION: We provide substantial data to inform study design, e.g. effect and study size, for cyathostomin research and highlight necessity for method standardization and raw data accessibility for optimal post-factum comparisons.

Determination of the specific gravity of eggs of equine strongylids, Parascaris spp., and Anoplocephala perfoliata.

Given the ever-increasing levels of anthelmintic resistance in livestock parasites globally, it is recommended to use parasite fecal egg counts to make treatment decisions and to evaluate treatment efficacy. The consensus in equine parasitology is to use a flotation medium with a specific gravity (SG) of ≥ 1.20 to float the main parasite egg types of interest in egg counting techniques. However, the density of common equine endoparasite eggs has been sparsely investigated. Equine tapeworm eggs are known to be particularly difficult to determine and count in fecal samples. It is unknown whether this could be because of differences in egg density. The aim of this study was to provide estimates of relative densities for equine ascarid, strongyle, and tapeworm eggs. Six aqueous glucose-salt solutions with specific gravities ranging from 1.06 to 1.16 were made and placed from most to least dense into thirteen 15 mL centrifuge tubes. Concentrated aqueous suspensions of the three types of endoparasite eggs were placed on top of each tube. These tubes were then centrifuged at 800 g for 20 min and each layer of flotation solution was carefully pipetted and transferred to a McMaster egg counting slide. Egg type and count were recorded for each specific gravity layer. Each egg was assigned a specific gravity based on the specific gravity layer it was observed in. In a second trial of this study, five similar flotation media were made ranging from 1.02 to 1.10 and were used in four subsequent replicates. In total between the two trials, the mean egg SGs of Anoplocephala perfoliata (n = 3811), Parascaris spp. (n = 3478), and strongylid type eggs (n = 9291) were 1.0636 (95% confidence interval (CI): 1.0629-1.0642), 1.0903 (95% CI: 1.0897-1.0909), and 1.0453 (95% CI: 1.0448-1.0458), respectively. The three egg types were statistically different from each other (p < 0.0001). This is the first time that the specific gravity of equine strongylid and Anoplocephala perfoliata eggs has been determined. With a tapeworm egg density demonstrated to be between that of strongylids and Parascaris spp., the poor recovery of tapeworm eggs in equine fecal samples must have other explanations.

Anthelmintic therapy of equine cyathostomin nematodes - larvicidal efficacy, egg reappearance period, and drug resistance.

Cyathostomins are ubiquitous in grazing horses across the world, and anthelmintic resistance has been reported with increasing levels over past decades. The aims of the present study were (i) to investigate the efficacy against encysted larval stages of moxidectin (0.4 mg/kg) and fenbendazole (10 mg/kg daily for five consecutive days) and compare these regimens at 2 and 5 weeks post-treatment, (ii) to investigate individual cyathostomin species associated with shortened egg reappearance periods, and (iii) to document species exhibiting decreased susceptibility to the evaluated compounds. Thirty-six ponies were allocated to treatment groups with half euthanatized 2 weeks post-treatment, and the remainder necropsied after 5 weeks. Luminal and mucosal worm counts were conducted and strongyle egg counts were determined at weekly intervals. At 2 weeks, mean reductions of early L3s were 50.4% and 73.8% for fenbendazole and moxidectin, respectively. At 5 weeks, the respective efficacies were 51.3% and 71.8%. Two week efficacies against late L3s and L4s (LL3s/L4s) were 70.8% and 74.6% for fenbendazole and moxidectin, respectively, whereas very low numbers were found in all three groups at 5 weeks. None of the mucosal counts were significantly different between treatment groups. Fenbendazole and moxidectin reduced luminal worm counts by 93.2% and 98.3% at 2 weeks following administration, with moxidectin group adult counts being significantly lower than the other two groups (P < 0.0001). Both treatment groups had increased counts 3 weeks later (P = 0.0415). A moxidectin ERP of 4 weeks was associated with surviving luminal L4s, and adult species contributing to this were Cyathostomum catinatum, Cylicostephanus longibursatus, Cylicocyclus ashworthi and Cylicocyclus nassatus. This study documented (i) larvicidal efficacy of fenbendazole much lower than historical standards, (ii) survival of luminal immatures (L4) following moxidectin administration, and (iii) new information about cyathostomin species associated with these phenomena.

Objective evaluation of two deworming regimens in young Thoroughbreds using parasitological and performance parameters.

Parasitic helminths of equids are capable of causing ill-thrift, clinical disease, and death. Although young horses are the most susceptible to parasitic disease and are the most intensively treated cohort, deworming regimens are rarely evaluated within this age group. This study objectively evaluated the impact of deworming regimen on fecal egg counts (FECs), growth rates, and body-condition scores in young Thoroughbreds. Forty-eight Thoroughbred foals from three central Kentucky farms were randomly allocated to two treatment groups: an interval dose program receiving bi-monthly rotations of pyrantel pamoate and ivermectin and a daily deworming group receiving daily rations of pyrantel tartrate feed additive throughout the study, oxibendazole at two months of age, and moxidectin treatments at 9.5 and 16.5 months of age. Pre- and post-treatment eggs per gram of feces (EPGs) of Parascaris spp. and strongyle family parasites, gel/paste dewormer efficacies, and monthly weights and body condition scores were collected. Ascarid and strongyle FECs were not significantly different between groups but were significantly influenced by horse age with strongyle counts continually increasing and ascarid counts peaking at 4.5 months of age. Reduced strongyle efficacies of ivermectin and moxidectin were observed on two farms with consistently low pyrantel pamoate efficacies on all three farms. Ivermectin also exhibited reduced ascarid efficacy. Average daily gain did not differ significantly between groups and was only significantly influenced by age, mirroring average daily gain reference data for Kentucky Thoroughbreds born in 2013. Body condition scores also did not differ between groups, remaining in the optimal range (5-6) for the duration of the study. Management practices resulting in growth rates matching the reference data and in optimal body condition scores compensate for the negative impacts of parasitism even in cases of reduced drug efficacy. Performance parameters can provide useful information in cases of suboptimal parasite control.

Evaluation of Baermann apparatus sedimentation time on recovery of Strongylus vulgaris and S. edentatus third stage larvae from equine coprocultures.

Traditional methods of diagnosing equine Strongylinae infections require culturing feces, sedimenting the culture media in Baermann apparatuses, collecting the sediment, and morphologically identifying recovered third stage larvae. However, this method is plagued by low negative predictive values. This study evaluated sedimentation time within the Baermann apparatus by comparing larval recovery from the traditionally collected sediment, "sediment 1", and from the usually discarded remaining fluid contents, "sediment 2", of the Baermann apparatus after 12, 24, and 48 h. A grand total of 147,482 larvae were recovered and examined. Sedimentation time did not significantly influence total larval recovery. At all three durations, significantly more Cyathostominae and Strongylus vulgaris larvae were covered from sediment 1 than from sediment 2. However, less than 60% of all recovered Strongylus edentatus were recovered from sediment 1. As 95% of S. vulgaris larvae were always recovered from sediment 1, the need for collection and examination of the remaining fluid contents of the Baermann apparatus is obviated when performing coprocultures for diagnosis of S. vulgaris infections, and sedimentation for 12h is adequate. Approximately 70% of Cyathostominae were recovered in sediment 1 at all durations, suggesting that 12h of sedimentation is adequate, although there is a need for future research to evaluate the risk of selection bias at differing sedimentation times among individual cyathostomin species. In contrast to S. vulgaris, collecting and examining the entire contents of the Baermann apparatus may be necessary when an increased diagnostic sensitivity and negative predictive value is desired in diagnosing S. edentatus infections as only 38-61% of larvae were recovered from sediment 1 portion of the Baermann apparatus. This information will allow researchers and practitioners to make more informed decisions in choosing appropriate larval recovery techniques, balancing recovery, time, and effort.

Parascaris univalens--a victim of large-scale misidentification?

The equine ascarid parasite Parascaris equorum is well known as a ubiquitous parasite infecting foals. A sibling species, Parascaris univalens, was first described over 130 years ago, but very little attention has been given to its existence and possible implications for anthelmintic resistance, clinical disease, or host age spectrum. P. univalens only possesses one germ line chromosome pair as opposed to two for P. equorum, but the two species are otherwise considered morphologically identical. For the present study, live worms obtained from the University of Kentucky parasitology horse herd were dissected and identified using karyotyping techniques. With no exception, all specimens (n = 30) were identified to be P. univalens. Further, the karyotyping technique was adapted to ascarid eggs derived from fecal samples and carried out on samples collected from 25 Thoroughbred foals from three farms in Central Kentucky. P. equorum was not identified among these, whereas P. univalens was found in 17 samples, with the remaining being inconclusive. The mitochondrial genome was sequenced, assembled, and annotated from one male worm identified as P. univalens, and comparison with available sequence reads labeled as P. equorum revealed only 0.16% nucleotide differences. However, it is unlikely that the sequences available in public databases have been unequivocally identified to species level by karyotyping. Taken together, these data suggest that P. univalens is likely the main species now observed in equines and that perhaps the designation Parascaris spp. should be used unless cytological characterization has confirmed the species.