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This study aimed to assess Australian veterinarians’ knowledge, perceptions and treatment strategies for worm control in horses with an online questionnaire. The questionnaire comprised 64 questions covering various aspects of: (i) veterinary practice; (ii) the veterinarian’s knowledge of gastrointestinal nematodes (GINs) and the importance of parasites in different age groups of horses; (iii) the diagnosis and control of worms; (iv) anthelmintics and anthelmintic resistance (AR); (v) grazing management; and (vi) the means of communication and the discussion between veterinarians and their clients regarding worm control. Following a pilot survey, a link for the questionnaire survey was sent to all (n = 1,148) registered members of Equine Veterinarians Australia in April 2020. The response rate for the questionnaire was 10% (118 of 1,148). The findings of this study illustrate veterinarians’ good understanding of aspects of equine parasites, including control. However, respondents mainly recommended frequent, interval-based prophylactic deworming in young horses, and only 40% (96 of 239) diagnosed GIN infections based on faecal egg count (FEC) results in all age groups of horses. Furthermore, only 27% (88 of 330) of the respondents made deworming decisions based on FECs. Most of the respondents recommended macrocyclic lactones (MLs) for all age groups of horses (71%, 481 of 677), and the most frequently used method to calculate the dose of anthelmintics was by estimating the weight of animals visually (53%, 63 of 118). Although the majority of respondents (97%, 115 of 118) perceived AR to be a critical issue in managing worms in horses, 58% (67 of 118) of them were unaware of the status of AR on their clients’ properties. Forty-two percent (50 of 118) of the respondents perceived the presence of AR in worms, including pinworms (16%), strongylins (15%), species of Draschia and Habronema (6%), Strongyloides westeri (2%) and tapeworms (1%). Twenty-seven percent (32 of 118) of the respondents rarely discussed equine worm control practices with their clients. This study provides insights into the perception and worm control practices recommended by Australian veterinarians to manage equine parasites. The findings highlight the importance of continued education and awareness of AR, and the use of non-chemical methods as well as consideration of the legislation of prescription-only use of anthelmintics based on FECs to achieve sustainable control of GINs in Australian horses.

Abstract Background Cyathostomins are the most important and common parasitic nematodes of horses, with > 50 species known to occur worldwide. The frequent and indiscriminate use of anthelmintics has resulted in the development of anthelmintic resistance (AR) in horse nematodes. In this study we assessed the efficacy of commonly used anthelmintics against cyathostomins in Australian thoroughbred horses. Methods Two drug efficacy trials per farm were conducted on two thoroughbred horse farms in the state of Victoria, Australia. In the first trial, the horses on Farm A were treated with single and combinations of anthelmintics, including oxfendazole (OFZ), abamectin (ABM), abamectin and morantel (ABM + MOR), moxidectin (MOX) and oxfendazole and pyrantel (OFZ + PYR), at the recommended doses, whereas the horses on Farm B only received MOX, at the recommended dose. The faecal egg count reduction test (FECRT) was used to determine the efficacy and egg reappearance period (ERP) of anthelmintics. Based on the results of the first trial, the efficacies of MOX and a combination of ABM + MOR were reassessed to confirm their activities against cyathostomins. Results Of the five anthelmintic products tested on Farm A, resistance against OFZ, ABM and OFZ + PYR was found, with efficacies of − 41% (− 195% lower confidence limit [LCL]), 73% (60% LCL) and 82% (66% LCL) at 2 weeks post-treatment, respectively. The FECRT showed high efficacies of MOX and ABM + MOR (100%) at 2 week post-treatment and shortened ERPs for these anthelmintics (ABM + MOR: 4 weeks; MOX: 5 weeks). Resistance to MOX was found on Farm B, with a reduced efficacy of 90% (70% LCL) and 89% (82% LCL) at 2 weeks post-treatment in trials one and two, respectively. Conclusions This study provides the first evidence of MOX- and multidrug-resistant (ABM and combinations of anthelmintics) cyathostomins in Australia and indicates the need for continuous surveillance of the efficacy of currently effective anthelmintics and large-scale investigations to assess the ERP for various anthelmintics. Graphical Abstract

Abstract Background Strongyloides westeri is found in the small intestine of young horses, mainly in foals up to about 16 weeks of age. The main source of infection for foals is through transmammary transmission, and foals can develop acute diarrhoea, weakness, dermatitis and respiratory signs. The epidemiology of S. westeri in Australia is largely unknown. Further, molecular techniques have never been employed for detection of S. westeri in horses. This pilot study aimed to assess the utility of a molecular phylogenetic method for the detection of S. westeri in the faeces of foals. Methods Faecal samples were collected from a foal of less than 2 months of age, and eggs of Strongyloides sp. were detected using the modified McMaster technique. DNA was extracted from purified eggs, and a partial fragment of the small subunit of the nuclear ribosomal DNA (18S) was characterised using polymerase chain reaction, DNA sequencing and phylogenetic methods. Results Microscopic examination of faeces revealed small ellipsoidal eggs typical of Strongyloides sp. The 18S sequence generated by PCR in this study revealed 98.4% identity with that of a reference sequence of S. westeri available from GenBank. Phylogenetic analyses revealed a polyphyletic clustering of S. westeri sequences. Conclusion This is the first study reporting the detection of DNA of Strongyloides sp. in faeces of a foal using a molecular phylogenetic approach targeting the variable region of 18S rDNA. It is anticipated that this study will allow future molecular epidemiological studies on S. westeri in horses. Graphical abstract

Benzimidazoles resistance is widespread in strongyle parasitic nematodes and associated with polym orphisms in the codons 167, 198 and 200 of isotype 1 β-tubulin (tbb-1). In ascarids, benzimidazole (BZ) resistance has rarely been reported and in none of these cases were any of these polymorphisms detected. Here, available genome and transcriptome data from WormBase ParaSite were used to compare the complete β-tubulin reservoirs of Parascaris univalens, Ascaris suum and Ascaris lumbricoides. Adult Parascaris spp. specimens collected in Australia from horses after BZ treatment (susceptible, n = 13) or surviving BZ treatment and collected after ivermectin treatment (resistant, n = 10) were genotyped regarding codons 167, 198 and 200 using Sanger sequencing. Phylogenetic analyses clearly showed that there are no one-to-one ascarid orthologs of strongyle tbb-1 genes. In the reference genomes, as well as phenotypically susceptible and resistant Parascaris spp. from Australia, six out of seven β-tubulin genes showed a BZ-susceptible genotype (F167, E198, F200). The only exception were the testis-specific β-tubulin D genes from all three ascarid species that encode tyrosine at codon 200. This was observed independently of the BZ-susceptibility phenotype of Parascaris spp. These data suggest that different mechanisms lead to BZ resistance in ascarid and strongyle nematodes.