This study quantified the extent of anthelmintic resistance (AR) in ascarid and strongylid nematodes against commonly used anthelmintics in Australian Thoroughbred horses. Faecal egg count reduction tests (FECRTs, n = 86) and egg reappearance period (ERP) tests were conducted on 22 farms across Australia. Faecal egg counts (FECs) were determined using the modified McMaster technique, and percent faecal egg count reduction (% FECR) was calculated using the Bayesian hierarchical model and hybrid Frequentist/Bayesian analysis method. The results were interpreted using old (published in 1992) and new (2023) research guidelines of the World Association for the Advancement of Veterinary Parasitology (WAAVP). The species composition of strongylid nematodes was detected utilising a DNA-metabarcoding method using pre- and post-treatment samples. Resistance was observed in strongylid nematodes to commonly used single-active and combination anthelmintics, including ivermectin (IVM %FECR range: 82%-92%; 95% lower credible interval (LCI) range: 80%-90%), abamectin (ABM: 73%-92%; 65%-88%), moxidectin (MOX: 89%-91%; 84%-89%), oxfendazole (OFZ: 0%-56%; 0%-31%) and its combination with pyrantel (OFZ + PYR: 0%-82%; 0%-78%). Resistance in Parascaris spp. was observed to IVM (10%-43%; 0%-36%), ABM (0%; 0%) and MOX (0%; 0%). When the new thresholds recommended by the WAAVP were used, AR was detected in six additional FECRTs for strongylids and three more tests for Parascaris spp., introducing resistance to OFZ and OFZ + PYR in the latter. Shortened ERPs (4-6 weeks) of strongylids were observed in 31 FECRTs in which AR was not detected at 2 weeks post-treatment for all the anthelmintics tested. Among cyathostomins, Cylicocyclus nassatus, Cylicostephanus longibursatus and Coronocyclus coronatus were the most prevalent species at 2 weeks post-treatment, whereas the main species appearing at five weeks following treatments with macrocyclic lactones were Cylicocyclus nassatus, Cylicostephanus longibursatus and Cylicocyclus ashworthi. After treatment with OFZ + PYR, the latter three, plus Coronocyclus coronatus and Cyathostomum catinatum, were detected at 5 weeks post-treatment. Overall, the study highlights the prevalence of AR in both ascarids and strongylid nematodes against commonly used anthelmintic products to control worms in Australian horses. The results indicate that ML combination products provided acceptable efficacy at 2 weeks. However, ERP calculations suggest that products work less effectively than previously measured. It is suggested to regularly monitor the efficacy of the anthelmintics and consider changing the worm control practices to better manage worms and AR in Australian horses.

This study reports the spatial and temporal distribution of ascarid and strongylid nematodes in Thoroughbred horses by age category across different climatic zones in Australia over an 18-month period. Faecal samples (n = 2046) from individual horses were analysed using the modified McMaster technique for faecal egg counts (FECs). Strongylids were identified using PCR-directed next-generation sequencing of the second internal transcribed spacer (ITS-2) of the nuclear ribosomal DNA. Yearlings had the highest prevalence (82%) of strongyle eggs followed by weanlings (79%), foals (58%), wet mares (49%) and dry mares (46%). For Parascaris spp., foals had the highest prevalence (35%) followed by weanlings (21%) and yearlings (10%). The highest mean FECs for Parascaris spp. were observed in foals (525 eggs per gram [EPG] of faeces) while those for strongyles were in yearlings (962 EPG). Among horses that were classified as adults at the time of sampling, 77% (860 of 1119) of mares were low (i.e., < 250 EPG) strongyle egg-shedders. Mean strongyle FEC counts were highest in the Mediterranean (818 EPG) followed by summer (599 EPG), winter (442 EPG), and non-seasonal (413 EPG) rainfall zones. Twenty-six nematode species were detected, with Cylicostephanus longibursatus (26.5%), Cylicocyclus nassatus (23.7%) and Coronocyclus coronatus (20.5%) being the most frequently detected species. Their richness and relative abundance varied with horse age, season and climatic zone. In addition, Strongylus equinus and Triodontophorus spp. (T. brevicauda and T. serratus) were also detected. This comprehensive study elucidates spatial (climatic zone) and temporal (i.e., seasonal) trends in prevalence and burdens of intestinal nematodes in Australian horses using non-invasive conventional and molecular methods. The information presented in this study is crucial for developing integrated management strategies to control horse parasites in farmed horses.

This review is aimed to (i) appraise the literature on the use of molecular techniques for the detection, quantification and differentiation of gastrointestinal helminths (GIH) of equids, (ii) identify the knowledge gaps and, (iii) discuss diagnostic prospects in equine parasitology. Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines for systematic reviews, we retrieved 54 studies (horses: 50/54; donkeys and zebras: 4/54) from four databases. Polymerase chain reaction (PCR) was employed in all of the studies whereas PCR amplicons were sequenced in only 18 of them. Other techniques used (including modifications of PCR) were reverse line blot, quantitative (q)PCR, restriction fragment length polymorphism, nestedPCR, PCR-directed next-generation sequencing, Southern blotting, single strand conformation polymorphism, PCR-enzyme linked immunosorbent assay, matrix-assisted laser desorption/ionisation-time of flight and random amplification of polymorphic DNA. Most of the studies (53/54) used nuclear ribosomal RNA (including the internal transcribed spacers, intergenic spacer, 5.8 S, 18 S, 28 S and 12 S) as target loci while cytochrome c oxidase subunit 1 and random genomic regions were targeted in only three and one studies, respectively. Overall, to date, the majority of molecular studies have focused on the diagnosis and identification of GIHs of equids (i.e. species of Anoplocephala, Craterostomum, cyathostomins, Oesophagodontus, Parascaris, Strongylus, Strongyloides and Triodontophorus), with a recent shift towards investigations on anthelmintic resistance and the use of highthroughput nemabiome metabarcoding. With the increasing reports of anthelmintic resistance in equid GIHs, it is crucial to develop and apply techniques such as advanced metabarcoding for surveillance of parasite populations in order to gain detailed insights into their diversity and sustainable control. To the best of our knowledge, this is the first systematic review that evaluates molecular investigations published on the diagnosis and quantification of equid GIHs and provides useful insights into important knowledge gaps and future research directions in equid molecular parasitology.

Summary Background The natural progression of equine glandular gastric disease (EGGD) in the absence of treatment has not yet been described in the literature, nor has the prevalence in a teaching herd population been reported. Objectives The aims of this study were to determine the prevalence of disease in a teaching population over the study period (2019–2021) and to observe the changes over time in disease severity of naturally occurring diseases (not experimentally induced) without medical intervention. Methods Twenty‐one horses underwent an initial gastroscopy and a repeat gastroscopy between 14 and 731 days later. Gastroscopy data were graded quantitatively and described qualitatively. Prevalence and 95% confidence intervals (CI) were calculated. The changes over time were determined by comparing initial and repeat gastroscopies. Results The prevalence from initial, repeat and total number of gastroscopies was 62% (95% CI: 40.8–79.3), 71% (95% CI: 50.0–86.2) and 67% (95% CI: 51.6–79), respectively. The changes over time included worsening of disease in 29% of horses (95% CI: 13.8–50.0), improvement of disease to a lower grade in 24% (95% CI: 10.6–45.1), no change in grade in 38% (95% CI: 20.8–59.1), and complete resolution of disease to grade 0 in 10% (95% CI: 2.7–28.9). Main limitations Limitations included a maximum of two gastroscopies per horse given COVID‐19 restrictions on data collection, and highly varied interval times between initial and repeat gastroscopies. Conclusion In conclusion, there is a high prevalence of disease in this teaching herd. The changes over time in naturally occurring diseases without medical intervention might include worsening, improvement, no change or resolution of disease.

The study presents the results of a cross-sectional survey to describe the epidemiology of ascarid and strongylid nematodes in horses, the impact of diverse climatic conditions on parasite diversity and the levels of faecal egg shedding in different age groups of managed Thoroughbred horses. Individual faecal samples (n = 1377) collected from 62 Thoroughbred farms across four climatic zones in Australia were analysed using the modified McMaster technique for faecal egg counts (FECs) and strongylid nematodes were identified utilising PCR-directed next-generation sequencing (NGS) of the second internal transcribed spacer of the nuclear ribosomal DNA (ITS -2). Across all age groups, the prevalence of ascarid and strongylid nematodes was 12% (95% confidence interval 10-14%) and 72% (70-74%), respectively. Based on strongylid FECs, yearlings had the highest prevalence (89%) followed by weanlings (83%), foals (79%), wet mares (61%), dry mares (59%) and stallions (54%). However, for Parascaris spp., foals had the highest prevalence (46%) followed by weanlings (32%) and yearlings (13%). The highest mean FECs for Parascaris spp. were observed in foals (418 eggs per gram [EPG] of faeces) while those for strongylids were in yearlings (1002 EPG). Of the adult horses (mares and stallions), 67% (489 of 729) and 11% (77 of 729) were low (i.e., <= 250 EPG) and moderate (i.e., 251-500 EPG) strongylid egg-shedders, respectively. Strongylid egg shedding varied across climatic zones, with the highest mean FECs in the summer rainfall (723 EPG) followed by non-seasonal rainfall (629 EPG), winter rainfall (613 EPG), and Mediterranean (606 EPG) rainfall zones. Twenty-three nematode species were detected using NGS, with Cylicostephanus longibursatus (28%), Cylicocyclus nassatus (23%) and Coronocyclus coronatus (23%), being the most abundant species. Three species of Strongylus (i.e., S. vulgaris, S. equinus and S. edentatus) were also detected. The nemabiome composition, species richness and relative abundance varied within horse age and between climatic zones. These empirical findings provide a comprehensive understanding of the prevalence of parasites within horse populations and the multifaceted factors that influence their occurrence, thereby allowing for the formulation of tailored strategies aimed at parasite control in domestic horses.