Validation of a serum ELISA test for cyathostomin infection in equines.

Cyathostomins are ubiquitous equine nematodes. Infection can result in larval cyathostominosis due to mass larval emergence. Although faecal egg count (FEC) tests provide estimates of egg shedding, these correlate poorly with burden and provide no information on mucosal/luminal larvae. Previous studies describe a serum IgG(T)-based ELISA (CT3) that exhibits utility for detection of mucosal/luminal cyathostomins. Here, this ELISA is optimised/validated for commercial application using sera from horses for which burden data were available. Optimisation included addition of total IgG-based calibrators to provide standard curves for quantification of antigen-specific IgG(T) used to generate a CT3-specific 'serum score' for each horse. Validation dataset results were then used to assess the optimised test's performance and select serum score cut-off values for diagnosis of burdens above 1000, 5000 and 10,000 cyathostomins. The test demonstrated excellent performance (Receiver Operating Characteristic Area Under the Curve values >0.9) in diagnosing infection, with >90% sensitivity and >70% specificity at the selected serum score cut-off values. CT3-specific serum IgG(T) profiles in equines in different settings were assessed to provide information for commercial test use. These studies demonstrated maternal transfer of CT3-specific IgG(T) in colostrum to newborns, levels of which declined before increasing as foals consumed contaminated pasture. Studies in geographically distinct populations demonstrated that the proportion of horses that reported as test positive at a 14.37 CT3 serum score (1000-cyathostomin threshold) was associated with parasite transmission risk. Based on the results, inclusion criteria for commercial use were developed. Logistic regression models were developed to predict probabilities that burdens of individuals are above defined thresholds based on the reported serum score. The models performed at a similar level to the serum score cut-off approach. In conclusion, the CT3 test provides an option for veterinarians to obtain evidence of low cyathostomin burdens that do not require anthelmintic treatment and to support diagnosis of infection.

Equine tapeworm (Anoplocephala spp.) infection: evaluation of saliva- and serum-based antibody detection methods and risk factor analysis in Slovak horse populations.

A lack of accurate information on the prevalence and distribution of Anoplocephala spp. infections on horse farms has led to insufficient attention to tapeworm control and increasing horse anoplocephaloses in Europe. Our study aimed to examine the occurrence of Anoplocephala spp. infection using coprological, serum- and saliva-based antibody detection methods and to analyze the risk factors associated with tapeworm infection in domestic horses in Slovakia. Fecal, serum, and saliva samples were collected from 427 horses from 31 farms in Slovakia. Additionally, a questionnaire study was conducted to collect information on tapeworm distribution on horse farms and analyze risk factors associated with infection. Fecal samples were examined by the mini-FLOTAC and the double centrifugation/combined sedimentation-flotation techniques. Serum and saliva samples were analyzed by ELISA to determine antibody levels against Anoplocephala spp. The effects of variables associated with an individual horse were tested for the positive result of the saliva ELISA test on Anoplocephala spp. Cestode eggs were detected in 1.99% of fecal samples (farm prevalence 12.90%), with no differences between the two coprological methods. Serum-based tapeworm ELISA results revealed that 39.39% of horses tested positive (farm prevalence 83.87%); while saliva-based tapeworm ELISA results revealed 56.95% positive horses (farm prevalence 96.77%). Binary logistic regression analysis revealed four meaningful predictors that significantly impacted the likelihood of detecting tapeworm infection in horses: horse age, pasture size, anthelmintic treatment scheme, and access to pasture. The influences of other variables associated with an individual horse were not significantly associated with detecting tapeworm infection.

The Use of Innovative Diagnostics to Inform Sustainable Control of Equine Helminth Infections.

Helminths are commonly found in grazing equids, with cyathostomin nematodes and the cestode Anoplocephala perfoliata being the most prevalent. Most horses harbour low burdens of these parasites and do not develop signs of infection; however, in a small number of animals, high burdens can accumulate and cause disease. Cyathostomins are associated with a syndrome known as larval cyathostominosis. This occurs when large numbers of larvae emerge from the large intestinal wall. This disease has a case fatality rate of up to 50%. A. perfoliata infection has been associated with various types of colic, with burdens of >20 worms associated with pathogenicity. Anthelmintic resistance is a serious problem in cyathostomins and is emerging in A. perfoliata. Control methods that reduce reliance on anthelmintics now need to be applied, especially as no new dewormer compounds are on the horizon. Sustainable control methods must employ diagnostics to identify horses that require treatment. Coprological tests (faecal egg counts, FECs) have been used for several decades to inform treatment decisions to reduce helminth egg shedding. These tests cannot be used to assess host burdens as FECs do not correlate with cyathostomin or A. perfoliata burdens. In the last decade, new tests have become available that measure parasite-specific antibodies, the levels of which have been shown to correlate with parasite burden. These tests measure antigen-specific IgG(T) and are available in serum (cyathostomin, A. perfoliata) or saliva (A. perfoliata) formats. Tests for other helminths have been developed as research tools and need to be translated to support equine clinicians in practice. A key element of sustainable control strategies is that diagnostics must be used in combination with management approaches to reduce environmental transmission of helminths; this will help limit the proportion of horses harbouring parasite burdens that need to be targeted by treatment. This manuscript provides a review of the development, performance and general utility of various diagnostic methods for informing equine helminth management decisions.

Exploring the transcriptomic changes underlying recombinant vaccine efficacy against Teladorsagia circumcincta in 3-month-old lambs.

Teladorsagia circumcincta is an abomasal parasitic nematode that can cause serious issues in small ruminant production, which are aggravated by drug resistance. Vaccines have been suggested as a feasible, long-lasting alternative for control since adaptation to the host's immune mechanisms by helminths develops at a much slower pace than anthelmintic resistance. Recently, a T. circumcincta recombinant subunit vaccine yielded over a 60% reduction in egg excretion and worm burden and induced strong humoral and cellular anti-helminth responses in vaccinated 3-month-old Canaria Hair Breed (CHB) lambs, but Canaria Sheep (CS) of a similar age were not protected by the vaccine. Here, we compared the transcriptomic profiles in the abomasal lymph nodes of such 3-month-old CHB and CS vaccinates 40 days after infection with T. circumcincta to understand differences in responsiveness at the molecular level. In the CS, differentially expressed genes (DEG) identified were related to general immunity processes such as antigen presentation or antimicrobial proteins and down-regulation of inflammation and immune response through regulatory T cell-associated genes. However, upregulated genes in CHB vaccinates were associated with type-2 oriented immune responses, i.e., immunoglobulin production, activation of eosinophils, as well as tissue structure and wound repair-related genes and protein metabolism pathways such as DNA and RNA processing. These results highlight potentially more optimal timing and orientation of immune responses in CHB sheep compared to CS associated with vaccine-induced protection. The data obtained in this study thus deepens our understanding of variations in responsiveness to vaccination in young lamb and provides insights for vaccine refinement strategies.

Vaccine-induced time- and age-dependent mucosal immunity to gastrointestinal parasite infection.

Individuals vary broadly in their response to vaccination and subsequent challenge infection, with poor vaccine responders causing persistence of both infection and transmission in populations. Yet despite having substantial economic and societal impact, the immune mechanisms that underlie such variability, especially in infected tissues, remain poorly understood. Here, to characterise how antihelminthic immunity at the mucosal site of infection developed in vaccinated lambs, we inserted gastric cannulae into the abomasa of three-month- and six-month-old lambs and longitudinally analysed their local immune response during subsequent challenge infection. The vaccine induced broad changes in pre-challenge abomasal immune profiles and reduced parasite burden and egg output post-challenge, regardless of age. However, age affected how vaccinated lambs responded to infection across multiple immune pathways: adaptive immune pathways were typically age-dependent. Identification of age-dependent and age-independent protective immune pathways may help refine the formulation of vaccines, and indicate specificities of pathogen-specific immunity more generally.

Differences in the protection elicited by a recombinant Teladorsagia circumcincta vaccine in weaned lambs of two Canarian sheep breeds.

Gastrointestinal nematode (GIN) infections are a serious drawback on small ruminant production. Since anthelmintic resistance has extended, optimisation of alternative non-chemical control strategies has attracted interest. Recently, a prototype recombinant vaccine protected immunologically mature sheep from Texel-cross and Canaria Sheep breeds against Teladorsagia circumcincta. The level of protective immunity stimulated by the vaccine varied between individuals and with age. Previous studies suggest that Canaria Hair Breed (CHB) sheep is naturally resistant to GIN infection, with some evidence suggesting that this protection is present in young lambs. Here, we sought to enhance this resistance by immunising three-month-old CHB lambs with a T. circumcincta prototype recombinant vaccine. Following vaccination and a larval challenge period, levels of protection against T. circumcincta infection were compared in CHB lambs with Canaria Sheep (CS) lambs (a breed considered less resistant to GIN). Lambs from the resistant CHB breed appeared to respond more favourably to vaccination, shedding 63% fewer eggs over the sampling period than unvaccinated CHB lambs. No protection was evident in CS vaccinated lambs. At post-mortem, CHB vaccine recipients had a 68% reduction in mean total worm burden, and female worms were significantly shorter and contained fewer eggs in utero compared to unvaccinated CHB lambs. A higher anti-parasite IgG2 level was detected in immunised CHB lambs compared to unvaccinated control CHB animals, with data suggesting that IgA, globular leucocytes, CD45RA+, CD4+ and CD8+ T cells are implicated in this protective response. The development of effective immunity in vaccinated CHB lambs did not reduce lamb growth rate as immunised CHB lambs had a significantly higher average daily weight gain after challenge than their unvaccinated counterparts. Therefore, the protection of CHB lambs was enhanced by immunisation at weaning, suggesting a synergistic effect when combining vaccination with presumed genetic resistance.

Comparative Sensitivity and Specificity of the 7SL sRNA Diagnostic Test for Animal Trypanosomiasis.

Animal trypanosomiasis (AT) is a significant livestock disease, affecting millions of animals across Sub-Saharan Africa, Central and South America, and Asia, and is caused by the protozoan parasites Trypanosoma brucei, Trypanosoma vivax, and Trypanosoma congolense, with the largest economic impact in cattle. There is over-reliance on presumptive chemotherapy due to inadequate existing diagnostic tests, highlighting the need for improved AT diagnostics. A small RNA species, the 7SL sRNA, is excreted/secreted by trypanosomes in infected animals, and has been previously shown to reliably diagnose active infection. We sought to explore key properties of 7SL sRNA RT-qPCR assays; namely, assessing the potential for cross-reaction with the widespread and benign Trypanosoma theileri, directly comparing assay performance against currently available diagnostic methods, quantitatively assessing specificity and sensitivity, and assessing the rate of decay of 7SL sRNA post-treatment. Results showed that the 7SL sRNA RT-qPCR assays specific for T. brucei, T. vivax, and T. congolense performed better than microscopy and DNA PCR in detecting infection. The 7SL sRNA signal was undetectable or significantly reduced by 96-h post treatment; at 1 × curative dose there was no detectable signal in 5/5 cattle infected with T. congolense, and in 3/5 cattle infected with T. vivax, with the signal being reduced 14,630-fold in the remaining two T. vivax cattle. Additionally, the assays did not cross-react with T. theileri. Finally, by using a large panel of validated infected and uninfected samples, the species-specific assays are shown to be highly sensitive and specific by receiver operating characteristic (ROC) analysis, with 100% sensitivity (95% CI, 96.44-100%) and 100% specificity (95% CI, 96.53-100%), 96.73% (95% CI, 95.54-99.96%) and 99.19% specificity (95% CI, 92.58-99.60%), and 93.42% (95% CI, 85.51-97.16% %) and 82.43% specificity (95% CI, 72.23-89.44% %) for the T brucei, T. congolense and T. vivax assays, respectively, under the conditions used. These findings indicate that the 7SL sRNA has many attributes that would be required for a potential diagnostic marker of AT: no cross-reaction with T. theileri, high specificity and sensitivity, early infection detection, continued signal even in the absence of detectable parasitaemia in blood, and clear discrimination between infected and treated animals.

Variability in the Response against Teladorsagia circumcincta in Lambs of Two Canarian Sheep Breeds.

The increasing resistance to anthelmintics has necessitated the exploration of alternative control strategies of gastrointestinal nematode (GIN) infections. A sustainable option is genetic selection based on differences in susceptibility to GIN infection between and within breeds of sheep. Here, three-month-old Canaria Hair breed (GIN-resistant) and Canaria Sheep breed (GIN-susceptible) showed no significant between-breed differences after trickle infection with Teladorsagia circumcincta, whereas considerable individual variability was found in both breeds. Next, data from lambs of both breeds were used to explore the relationships between parasitological variables and T. circumcincta-specific IgA levels, local immune cell populations, and abomasal lymph node gene expression to understand the possible mechanisms underlying resistance. Mucosal IgA levels as well as numbers of globular leukocytes and MHC-II+ cells were associated with protection. Analysis of lymph node gene expression revealed the associations between lower parasite numbers and cumulative fecal egg counts and several immune pathways, such as leukocyte cell adhesion, activation and differentiation of T cells, in particular CD4+ and IL-4 production. The data obtained here may inform on the relationship between phenotypic resistance variability and protective responses at the humoral, cellular, and transcriptomic levels, thus contributing to identifying immune responses in young lambs that could be used as markers for selection.

Cellular and humoral immune responses associated with protection in sheep vaccinated against Teladorsagia circumcincta.

Due to increased anthelmintic resistance, complementary methods to drugs are necessary to control gastrointestinal nematodes (GIN). Vaccines are an environmentally-friendly and promising option. In a previous study, a Teladorsagia circumcincta recombinant sub-unit vaccine was administered to two sheep breeds with different levels of resistance against GIN. In the susceptible Canaria Sheep (CS) breed, vaccinates harboured smaller worms with fewer eggs in utero than the control group. Here, we extend this work, by investigating the cellular and humoral immune responses of these two sheep breeds following vaccination and experimental infection with T. circumcincta. In the vaccinated CS group, negative associations between antigen-specific IgA, IgG2 and Globule Leukocytes (GLs) with several parasitological parameters were established as well as a higher CD4+/CD8+ ratio than in control CS animals, suggesting a key role in the protection induced by the vaccine. In the more resistant Canaria Hair Breed (CHB) sheep the vaccine did not significantly impact on the parasitological parameters studied and none of these humoral associations were observed in vaccinated CHB lambs, although CHB had higher proportions of CD4+ and CD8+ T cells within the abomasal lymph nodes, suggesting higher mucosal T cell activation. Each of the component proteins in the vaccine induced an increase in immunoglobulin levels in vaccinated groups of each breed. However, levels of immunoglobulins to only three of the antigens (Tci-MEP-1, Tci-SAA-1, Tci-ASP-1) were negatively correlated with parasitological parameters in the CS breed and they may be, at least partially, responsible for the protective effect of the vaccine in this breed. These data could be useful for improving the current vaccine prototype.

Characterisation of serum IgG(T) responses to potential diagnostic antigens for equine cyathostominosis.

Cyathostomins are ubiquitous parasitic nematodes of horses. These worms spend substantial periods as intestinal wall stage encysted larvae, which can comprise up to 90% of the total burden. Several million larvae have been reported in individuals. Emergence of these larvae from the gut wall can lead to life-threatening colitis. Faecal egg count tests, increasingly used by horse owners to inform anthelmintic treatments, do not correlate with the intra-host burden of cyathostomins; this represents a key gap in the diagnostic toolbox. Previously, a cyathostomin Gut Associated Larval Antigen was identified as a promising marker for the intra-host stages of infection. Here, cyathostomin Gut Associated Larval Antigen and an additional protein, Cyathostomin Immuno-diagnostic antigen, were investigated to examine their value in providing information on cyathostomin burden. ELISA analyses examined serum IgG(T) responses to recombinant proteins derived from individual cyathostomin species. Receiver Operator Characteristic curve analysis was performed on the ELISA data; proteins with the highest Area Under the Curve values were selected to test protein combinations to investigate which were the most informative in identifying the infection status of individuals. Three cocktail combinations were tested, comprising: (a) Cy-GALA proteins from two species and a Cy-CID protein from a third species (CT3), (b) Cy-GALA proteins from five species (CT5), and (c) all CT5 components, plus a Cy-CID protein from an additional species (CT6). The best predictive values for infection were obtained using CT3 and CT6, with similar values achieved for both. Proteins in CT3 are derived from the most commonly reported species, Cyathostomum catinatum, Cylicocyclus nassatus and Cylicostephanus longibursatus. This combination was selected for future development since it represents a more commercially viable format for a diagnostic test.

A survey of the level of horse owner uptake of evidence-based anthelmintic treatment protocols for equine helminth control in the UK.

Interval treatment control programmes used widely in equine helminth control have favoured the development of anthelmintic resistance worldwide. Best practice guidelines have been designed to address resistance and include the requirement for improved pasture hygiene to break helminth transmission cycles, along with anthelmintic application informed by the results of diagnostic tests to reduce selection pressure for resistance. Using an online questionnaire, this study examined uptake of measures recommended in these guidelines by UK horse owners. The survey comprised 58 questions spanning grazing management, anthelmintic use and use of faecal egg count (FEC) testing to inform treatment decisions. Analysis was carried out using a combination of Chi-square and Mann-Whitney tests. In total, 705 owners responded and, following specific exclusion criteria, the responses of 652 individuals were analysed. The majority of the respondents owned <20 horses on private premises or livery yards in England. The main outputs of the survey were as follows. Overall, 60.9% of respondents used FEC tests to inform the requirement to administer anthelmintics, with macrocyclic lactones the most frequently-used anthelmintics. Of the respondents, 38% obtained advice on anthelmintic choice from their veterinarians; however, many respondents (43.8%) purchased anthelmintics via the internet. Encouragingly, 74.4% of respondents stated that they practiced good pasture hygiene by removing dung from pasture. Generally, there were differences between the responses of participants who based anthelmintic treatments on FEC testing (targeted treatments; TT) and those who practiced calendar-based anthelmintic treatments (interval treatments; IT). Briefly, the "key" findings from the Chi-square analysis included higher levels of satisfaction with the level of knowledge about equine parasites/parasitic diseases and higher levels of concern about anthelmintic resistance from TT-respondents compared to IT-participants. Confusion on the interpretation of quarantine recommendations was identified in this study group and there was poor uptake of testing for anthelmintic effectiveness. Overall, compared to previous reports, this study indicated improved engagement of UK horse owners with some helminth control practices recommended to reduce the spread of anthelmintic resistance. However, a proportion of respondents did not utilise these practices and there were still important gaps in the use of appropriate quarantine and efficacy testing. These identified gaps must be taken into consideration in knowledge dissemination activities in the future.

Impacts of breed type and vaccination on Teladorsagia circumcincta infection in native sheep in Gran Canaria.

Vaccines and genetic resistance offer potential future alternatives to the exclusive use of anthelmintics to control gastrointestinal nematodes (GIN). Here, a Teladorsagia circumcincta prototype vaccine was administered to two sheep breeds which differ in their relative levels of resistance to infection with GIN. Vaccination of the more susceptible Canaria Sheep (CS) breed induced significant reductions in worm length and numbers of worm eggs in utero (EIU) when compared to control CS sheep. In the more resistant Canaria Hair Breed (CHB), although vaccination induced a reduction in all parasitological parameters analysed, differences between vaccinated and control sheep were not statistically significant. Such interactions between sheep breed and vaccination may allow better integrated control of GIN in future.

The rational simplification of a recombinant cocktail vaccine to control the parasitic nematode Teladorsagia circumcincta.

Using data from five independent vaccine trials, which employed a subunit cocktail vaccine containing eight recombinant proteins to protect sheep against Teladorsagia circumcincta, a strategy was developed to simplify antigen complexity of the vaccine. A meta-analysis of data from these five trials demonstrated statistically significant reductions in cumulative faecal egg count and worm burden in vaccinated sheep when compared with those which had received adjuvant only (P = 0.009 and P < 0.0001, respectively). Relationships between antigen-specific antibody levels, antibody avidity and parasitological parameters of efficacy were analysed for each of the eight proteins in these trials. Of these, the strongest correlations between percentage reduction in cumulative faecal egg count and avidity were obtained for the vaccine antigen T. circumcincta apyrase-1 (Tci-APY-1) in relation to either total antigen-specific IgG or IgG1 in sera (P = 0.019 and P = 0.030, respectively). In addition, IgG and IgA within the serum and abomasal mucus of control (parasite challenged) lambs strongly recognised Tci-APY-1 and T. circumcincta metalloproteinase-1 (Tci-MEP-1) but only weakly bound the other six antigens, indicating Tci-APY-1 and Tci-MEP-1 are most effectively recognised by the parasite-induced antibody response. On the basis of these findings, a two-protein vaccine comprising Tci-APY-1 and Tci-MEP-1 was tested in a direct comparison with the original eight-component vaccine. A further group was immunised with Tci-MEP-1 in combination with a mutated form of Tci-APY-1 (mTci-APY-1), which had no enzymatic activity. Across the trial, the mean faecal egg count levels of the eight-antigen recipients were lower than those of the adjuvant only control group (P = 0.013) and the mean FEC of the mTci-APY-1 and Tci-MEP-1 recipients was lower, although not statistically significantly, than that of the adjuvant-only control group (P = 0.093). Mean cumulative faecal egg count levels were reduced by 43% in lambs immunised with mTci-APY-1 plus Tci-MEP-1 compared with the controls (P = 0.079).

A survey of experiences of UK cattle and sheep farmers with anthelmintic prescribers; Are best practice principles being deployed at farm level?

Helminths are common pathogens of ruminants. These are controlled by anthelmintics; however, resistance is a major issue. More sustainable practices need to be deployed to protect anthelmintic efficacy. There is no published information on how farmer/prescriber interactions at anthelmintic purchase shape application of best practice on-farm. To determine experiences of UK ruminant farmers as they relate to anthelmintic purchasing/provision of best practice advice at the point-of-sale, an online survey was distributed. This explored farmer experiences in purchasing anthelmintics from the three UK animal medicines' prescribers (veterinarians, Suitably Qualified Persons [SQPs], veterinary pharmacists) and investigated farmer attitudes to anthelmintic use and resistance. First, farmers were grouped according to whom they purchased anthelmintics from, including those that purchased from > 1 prescriber type. The results indicated there was no significant difference between the type of prescriber from whom farmers bought anthelmintics and route (face-to-face, telephone, online) through which they purchased them. Farmers who bought from veterinarians were more likely to view prescriber knowledge as more important. When asked who influenced purchase, there was a significant difference; farmers tended to be most influenced by the prescriber they purchased from. Those who bought from various types of prescriber were most influenced by veterinarians. Farmers who purchased from veterinarians were significantly more likely to state their seller discussed/recommended faecal egg count (FEC) testing and management strategies to reduce anthelmintic use than those who purchased from SQPs or from > 1 prescribing channel. When grouped according to purchase route, farmers that bought through face-to-face interactions were significantly more likely to state they valued prescriber knowledge of parasites/anthelmintics than farmers that purchased via the telephone or internet. Farmers that purchased online were significantly less likely to consider prescriber advice. Analysis was then applied based on the type of livestock farmed. Generally, sheep farmers undertook FEC testing more than cattle farmers. Relatively few farmers stated they conducted anthelmintic sensitivity testing, with the majority of cattle farmers having not tested at all. This was despite a high level of concern for anthelmintic resistance in all types of farmers. 'Cattle only' farmers viewed convenience factors as more important than those that managed sheep or sheep/cattle. The results suggest that UK farmers that bought anthelmintics from veterinarians were more likely to be exposed to diagnostic-led control advice. Gaps in advice provision, particularly in relation to efficacy testing, were identified in all farmer groups.

An automated high-throughput system for phenotypic screening of chemical libraries on C. elegans and parasitic nematodes.

Parasitic nematodes infect hundreds of millions of people and farmed livestock. Further, plant parasitic nematodes result in major crop damage. The pipeline of therapeutic compounds is limited and parasite resistance to the existing anthelmintic compounds is a global threat. We have developed an INVertebrate Automated Phenotyping Platform (INVAPP) for high-throughput, plate-based chemical screening, and an algorithm (Paragon) which allows screening for compounds that have an effect on motility and development of parasitic worms. We have validated its utility by determining the efficacy of a panel of known anthelmintics against model and parasitic nematodes: Caenorhabditis elegans, Haemonchus contortus, Teladorsagia circumcincta, and Trichuris muris. We then applied the system to screen the Pathogen Box chemical library in a blinded fashion and identified compounds already known to have anthelmintic or anti-parasitic activity, including tolfenpyrad, auranofin, and mebendazole; and 14 compounds previously undescribed as anthelmintics, including benzoxaborole and isoxazole chemotypes. This system offers an effective, high-throughput system for the discovery of novel anthelmintics.

Strongyle egg reappearance period after moxidectin treatment and its relationship with management factors in UK equine populations.

Parasitic nematodes, particularly cyathostomins, are ubiquitous in grazing horses world-wide. Considerable burdens of cyathostomin larvae can encyst in the large intestinal wall. The most recommended treatment against these pathogenic stages is moxidectin. Information is required on how effective moxidectin is against cyathostomin populations in different regions. The objectives here were to determine the efficacy of moxidectin treatment and estimate the strongyle egg reappearance period (ERP) after treatment in several equine populations, to confirm the type of strongyle nematodes present and to identify other (i.e. management) factors associated with shortened ERP. Eight yards were recruited and moxidectin in combination with praziquantel administered to all horses (n=261). Faecal egg count (FEC) analysis was performed at weeks 0, 2, 6, 10 and 12 after treatment to determine efficacy and ERP. The ERP was estimated using two previously published methods. Morphological identification of cultured third stage larvae from the sample population was compared to a Strongylus vulgaris-specific end-point PCR to examine the presence of S. vulgaris in samples before and after treatment. Strongyle egg shedding patterns were also compared to worm management practices at each site. At 2 weeks post-treatment, moxidectin was highly effective (faecal egg count reduction range, 99.9-100%). The strongyle ERP ranged from 6 weeks to >12 weeks depending on the calculation method applied. Only cyathostomin larvae were detected by morphological identification. The results from the coprocultures and PCR showed that S. vulgaris was absent before and after treatment. Analysis revealed that regular faecal removal from pasture was associated with lower average FEC and lower prevalence of egg shedding.

Strongyloides spp. infections of veterinary importance.

This paper reviews the occurrence and impact of threadworms, Strongyloides spp., in companion animals and large livestock, the potential zoonotic implications and future research. Strongyloides spp. infect a range of domestic animal species worldwide and clinical disease is most often encountered in young animals. Dogs are infected with Strongyloides stercoralis while cats are infected with different species according to geographical location (Strongyloides felis, Strongyloides tumefaciens, Strongyloides planiceps and perhaps S. stercoralis). In contrast to the other species, lactogenic transmission is not a primary means of infection in dogs, and S. stercoralis is the only species considered zoonotic. Strongyloides papillosus in calves has been linked to heavy fatalities under conditions of high stocking density. Strongyloides westeri and Strongyloides ransomi of horses and pigs, respectively, cause only sporadic clinical disease. In conclusion, these infections are generally of low relative importance in livestock and equines, most likely due to extensive use of macrocyclic lactone anthelmintics and/or improved hygiene. Future prevalence studies need to include molecular typing of Strongyloides species in relation to different hosts. More research is urgently needed on the potential zoonotic capacity of Strongyloides from dogs and cats based on molecular typing, information on risk factors and mapping of transmission routes.

Investigating interactions between UK horse owners and prescribers of anthelmintics.

Helminths are common pathogens of equids and anthelmintic resistance is a major issue in cyathostomin species and Parascaris equorum. At the heart of mitigating the impact of increasing anthelmintic resistance levels, is the responsible dissemination and use of these medicines following best practice principles. There is a paucity of information on interactions between horse owners and anthelmintic prescribers and how this shapes control. Here, a study was undertaken to determine opinions and experiences of horse owners as they relate to anthelmintics purchase and implementation of best practice control. An online survey was distributed via email and social media to explore owners' experiences of purchasing anthelmintics from United Kingdom prescribers, these being veterinarians, suitably qualified persons (SQPs) and pharmacists. Owner responses (n=494) were analysed statistically to compare answers of respondents grouped according to: (i) from whom they bought anthelmintics (Veterinarians n=60; SQPs n=256; Pharmacists n=42; More than one channel n=136), and (ii) by which route (Face-to-face n=234; Telephone n=31; Online n=226) they purchased. Owners who purchased from veterinarians predominantly did so face-to-face (81.3%), whilst those that bought from SQPs purchased via face-to-face (48.8%) and online (46.0%) interactions. Those who purchased from pharmacists predominantly bought anthelmintics online (76.2%). Participants who bought from veterinarians were more likely to view certain factors (i.e. time to talk to the supplier, supplier knowledge) as more important than those who purchased from other prescribers. Those who purchased from veterinarians were more likely to be recommended faecal egg count (FEC) test analysis; however, there was high uptake of FEC testing across all groups. There was a low uptake of anthelmintic efficacy testing; regardless of the prescriber type from whom anthelmintics were purchased. Those who purchased from veterinarians were more likely to agree that anthelmintics should be sold as veterinary prescription-only medicines. Those who purchased online (regardless of which type of prescriber they bought from) were less likely to consider prescriber advice or knowledge when deciding which product to buy and indicated that sellers were less likely to raise use of anthelmintics for targeting parasites. Across all groups, many owners stated that they were aware of or used non-chemical control measures such as dung removal and diagnostic FEC testing to target treatments. In summary, there were some differences in the type of advice provided at the point of purchase and this was dependent upon whom anthelmintics were purchased from and by which route they were bought.

A survey of UK prescribers' experience of, and opinions on, anthelmintic prescribing practices for livestock and equines.

The aim of this study was to determine practices, attitudes and experiences of UK prescribers of anthelmintics for horses and livestock. A questionnaire was sent by direct email to groups licenced to prescribe these medicines. These were veterinarians, Suitably Qualified Persons (SQPs, registered with the Animal Medicines Training Regulatory Authority) and veterinary pharmacists. The survey was also advertised through social media. It comprised questions relating to demographics, training experiences, current prescribing practices, as well as personal opinions on anthelmintic selection, diagnostics and anthelmintic resistance. A total of 193 veterinarians and 326 SQPs were included in final analysis. Pharmacists were excluded from detailed analysis due to the low numbers that responded (n=3). The results indicated that SQP participants were more likely to receive post-certification parasitology training than the veterinarians, and that both channels consulted similar sources for information about helminths and their control (paper articles in journals, online sources). The SQP participants stated a higher frequency of face-to-face interactions with clients/customers (96.1%) than the veterinarians (76.4%), who stated a higher frequency of telephone interactions (55.1% and 73.5%, respectively). Veterinarians were more likely to state that there were specific factors that limited interactions with their clients (54.1%) than SQPs (19.6%), such as competition from other suppliers. SQP participants considered a wider range of factors as important when deciding on which anthelmintic to recommend (i.e. knowledge of specific parasites, knowledge of specific anthelmintics, discussion of measures to avoid anthelmintic resistance and time to talk with clients/customers); however, the veterinarian participants were more likely to consider the results of diagnostic tests. While discussions about anthelmintic resistance were stated with similar frequency in both groups, less frequent were specific discussions about anthelmintic sensitivity testing. In-house faecal egg count analysis was more likely to be available from those that prescribed anthelmintics for equines alone, compared to prescribers who dispensed anthelmintics for livestock alone or livestock and equines. The SQP participants indicated that they felt a large number of organisations were responsible for ensuring that anthelmintics are used responsibly, whilst veterinarian participants were more likely to place responsibility on the prescribers alone. Taken together, these findings provide an insight into how prescribers of anthelmintics in the UK interact with their clients/customers before and at the point of sale and act as a unique source of information on how best practice advice pertaining to sustainable helminth control is disseminated by the various prescribing channels.

Protection of ewes against Teladorsagia circumcincta infection in the periparturient period by vaccination with recombinant antigens.

Teladorsagiosis is a major production-limiting disease in ruminants in temperate regions throughout the world and one of the key interventions in the management of the disease is the prevention of pasture contamination with Teladorsagia circumcincta eggs by ewes during the periparturient relaxation in immunity which occurs in the period around lambing. Here, we describe the immunisation of twin-bearing ewes with a T. circumcincta recombinant subunit vaccine and the impact that vaccination has on their immune responses and shedding of parasite eggs during a continuous T. circumcincta challenge period spanning late gestation and lactation. In ewes which displayed a clear periparturient relaxation in immunity, vaccination resulted in a 45% reduction in mean cumulative faecal egg count (cFEC, p=0.027) compared to control (immunised with adjuvant only) ewes. Recombinant antigen-specific IgG and IgA, which bound each of the vaccine antigens, were detected in the serum of vaccinated ewes following each immunisation and in colostrum taken from vaccinated ewes post-partum whereas low levels of antigen-specific IgG were detected in serum and colostrum from control ewes. Antigen-specific IgG and IgA levels in blood collected within 48h of birth from lambs largely reflected those in the colostrum of their ewes.