Evaluation of tests for anthelmintic resistance in cyathostomes

Resistance, especially to the anthelmintic benzimidazoles (BZ), has been reported in horse cyathostomes world-wide. Diagnosis of resistance has traditionally been made by faecal egg count reduction (FECR) trials, however, this technique has limitations. Some of the shortcomings may be resolved by refining the test or by using an in vitro test. FECR tests and the larval development assay (LDA) were performed on adult horses held on 15 different horse properties across a wide geographical area of NSW, Australia. FECR were measured before and 10-14 after days treatment with oxibendazole (OBZ), morantel (MOR) or ivermectin (IVM) at recommended dose rates. Eight properties were rejected following low pre-treatment egg counts, leaving seven in the study. On these, the majority of larvae recovered from faecal cultures were cyathostomes. Using a definition of resistance as a FECR of <90%, resistance to OBZ was present on six properties and to MOR on two properties. Resistance to IVM was not detected. An alternative method of calculating FECR based on individual horse egg counts pre- and post-treatment was developed and results from the same properties compared with the results of the LDA. For example, for the BZ, correlation coefficients of values of lethal concentration to kill 50% of population (LC50) on LDA and FECR percentages were -0.536 before and -0.704 after OBZ treatment. We conclude that the LDA has the potential to be a single visit test for detection of anthelmintic resistance in horse cyathostomes but requires further investigation and standardisation.     

Prevalence of anthelmintic resistant cyathostomes on horse farms

OBJECTIVE: To determine prevalence of anthelmintic resistance in cyathostome nematodes of horses in the southern United States. DESIGN: Cross-sectional study. ANIMALS: 786 horses on 44 farms and stables in Georgia, South Carolina, Florida, Kentucky, and Louisiana. PROCEDURE: Fecal egg count (FEC) reduction tests were performed on 44 large farms and stables. Horses on each farm were treated with an oral paste formulation of fenbendazole, oxibendazole, pyrantel pamoate, or ivermectin at recommended label dosages. A mixed linear model was fitted to the percentage reduction in FEC, accounting for differences among farms, states, ages, treatments, and treatment by state interactions. RESULTS: By use of a conservative measure of resistance (< 80% reduction), the percentage of farms with anthelmintic-resistant cyathostomes was 97.7%, 0%, 53.5%, and 40.5% for fenbendazole, ivermectin, oxibendazole, and pyrantel pamoate, respectively. Mean percentage reductions in FEC for all farms were 24.8%, 99.9%, 73.8%, and 78.6% for fenbendazole, ivermectin, oxibendazole, and pyrantel pamoate, respectively. Pairwise contrasts between states for each treatment revealed that in almost all instances, there were no significant differences in results between states. CONCLUSIONS AND CLINICAL RELEVANCE: The prevalence of resistance found in this study was higher than that reported previously, suggesting that anthelmintic resistance in equine cyathostomes is becoming a major problem. Furthermore, data from these 5 southern states, which are geographically and physiographically distinct, were remarkably similar. This suggests that drug resistance in cyathostomes is highly prevalent throughout the entire southern United States and probably nationwide.     

Evaluation of prevalence and clinical implications of anthelmintic resistance in gastrointestinal nematodes in goats

OBJECTIVE: To determine prevalence of resistance to all anthelmintics that are commonly used to treat gastrointestinal nematodes (GINs) in goats. DESIGN: Prospective study. ANIMALS: 777 goats. PROCEDURE: On each farm, goats were assigned to 1 of 5 treatment groups: untreated controls, albendazole (20 mg/kg [9.0 mg/lb], p.o., once), ivermectin (0.4 mg/kg [0.18 mg/lb], p.o., once), levamisole (12 mg/kg [5.4 mg/lb], p.o., once), or moxidectin (0.4 mg/kg, p.o., once), except on 3 farms where albendazole was omitted. Fecal samples were collected 2 weeks after treatment for determination of fecal egg counts (FECs), and percentage reductions were calculated by comparing data from anthelmintic-treated and control groups. Nematode populations were categorized as susceptible, suspected resistant, or resistant by use of guidelines published by the World Association for the Advancement of Veterinary Parasitology. RESULTS: Resistance to albendazole was found on 14 of 15 farms, and resistance to ivermectin, levamisole, and moxidectin was found on 17, 6, and 1 of 18 farms, respectively. Suspected resistance to levamisole and moxidectin was found on 4 and 3 farms, respectively. Resistance to multiple anthelmintics (albendazole and ivermectin) was found on 14 of 15 farms and to albendazole, ivermectin, and levamisole on 5 of 15 farms. Mean overall FEC reduction percentages for albendazole, ivermectin, levamisole, and moxidectin were 67, 54, 94, and 99%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Anthelmintic resistance in GINs of goats is highly prevalent in the southern United States. The high prevalence of resistance to multiple anthelmintics emphasizes the need for reexamination of nematode control practices.     

Survey of anthelmintic resistance in Western Australian sheep flocks 1. Prevalence

Between July 1981 and December 1983, 116 randomly selected sheep farms in the south west of Western Australian were surveyed for resistance to anthelmintics. A faecal worm egg count reduction test was carried out on each farm. Anthelmintics tested were thiabendazole (44 mg/kg) and levamisole (7.5 mg/kg) given by intra-ruminal injection and comparisons were made with an untreated group on each farm. Successful tests were carried out on 84 farms and 68% of these had resistant worms present. The prevalence of thiabendazole resistant populations was for H. contortus 18%; Teladorsagia, 41% and Trichostrongylus, 48% and for levamisole resistant populations H. contortus, 10%; Teladorsagia, 41%; Trichostrongylus, 24%, and Nematodirus, 10%. Multiple resistant populations were found on 17% of farms. Although the distribution of nematode genera varied between the 400 to 750 mm and the greater than 750 mm rainfall zones there was no significant difference in the prevalence of resistance between zones. About one third of resistant populations were severely resistant (less than 60% reduction). It is likely that resistant worms were present on many farms without causing clinical disease and continued anthelmintic selection pressure will result in further development of resistance.     

Prevalence of anthelmintic resistance on sheep farms in New Zealand

AIM: To establish the prevalence of anthelmintic resistance in parasitic nematodes on sheep farms in New Zealand.

METHODS: A cross-sectional prevalence study was conducted, using a standardised faecal nematode egg count (FEC) reduction (FECR) test (FECRT) for ivermectin, at a full (0.2 mg/kg) and half (0.1 mg/kg) dose rate, and albendazole, levamisole and albendazole-levamisole in combination, on 60 lambs (n=10 per group) on farms selected from throughout New Zealand. Farms that conformed with selection criteria were chosen at random (n=80) or with a history of suspected resistance to macrocy- clic lactone (ML) anthelmintics (n=32). Resistance to an an- thelmintic was inferred when there was <95% reduction in FEC 7-10 days after treatment. Larval cultures were performed for all control groups and for treated groups for which resistance was evident.

RESULTS: Of the farms randomly selected, 36% showed ≥95% FECR for all anthelmintics tested; resistance to ivermectin at 0.1 and 0.2 mg/kg liveweight was evident on 36% and 25% of these farms, respectively. Resistance to both ivermectin (0.2 mg/kg) and levamisole was evident on 8/80 (10%) farms, to ivermectin and albendazole on 10/80 (13%) farms, and to iver- mectin, levamisole and albendazole on 6/80 (8%) farms. The prevalence of resistance to a half dose of ivermectin tended to be more prevalent on farms with a history of suspected ML resistance (p=0.06). Resistance to albendazole was seen across all the main parasite genera, and to levamisole in Nematodirus, Ostertagia (= Teladorsagia) and Trichostrongylus species. Resistance to ivermectin was dominated by Ostertagia spp, although Cooperia, Nematodirus and Trichostrongylus species were also implicated.

CONCLUSION: Anthelmintic resistance in parasitic nema-todes of sheep is common in New Zealand. Not only was resistance to albendazole and levamisole common, but resistance to the ML, ivermectin, was at a higher prevalence than expected. Sheep farmers and advisors in New Zealand need to re-evaluate the way they manage parasites, and more research is urgently needed if the steady decline in anthelmintic susceptibility is to be halted.     

Prevalence of anthelmintic resistance on sheep farms in New Zealand

AIM: To establish the prevalence of anthelmintic resistance in parasitic nematodes on sheep farms in New Zealand. METHODS: A cross-sectional prevalence study was conducted, using a standardised faecal nematode egg count (FEC) reduction (FECR) test (FECRT) for ivermectin, at a full (0.2 mg/kg) and half (0.1 mg/kg) dose rate, and albendazole, levamisole and albendazole-levamisole in combination, on 60 lambs (n=10 per group) on farms selected from throughout New Zealand. Farms that conformed with selection criteria were chosen at random (n=80) or with a history of suspected resistance to macrocyclic lactone (ML) anthelmintics (n=32). Resistance to an anthelmintic was inferred when there was <95% reduction in FEC 7-10 days after treatment. Larval cultures were performed for all control groups and for treated groups for which resistance was evident. RESULTS: Of the farms randomly selected, 36% showed > or =95% FECR for all anthelmintics tested; resistance to ivermectin at 0.1 and 0.2 mg/kg liveweight was evident on 36% and 25% of these farms, respectively. Resistance to both ivermectin (0.2 mg/kg) and levamisole was evident on 8/80 (10%) farms, to ivermectin and albendazole on 10/80 (13%) farms, and to ivermectin, levamisole and albendazole on 6/80 (8%) farms. The prevalence of resistance to a half dose of ivermectin tended to be more prevalent on farms with a history of suspected ML resistance (p=0.06). Resistance to albendazole was seen across all the main parasite genera, and to levamisole in Nematodirus, Ostertagia (= Teladorsagia) and Trichostrongylus species. Resistance to ivermectin was dominated by Ostertagia spp, although Cooperia, Nematodirus and Trichostrongylus species were also implicated. CONCLUSION: Anthelmintic resistance in parasitic nematodes of sheep is common in New Zealand. Not only was resistance to albendazole and levamisole common, but resistance to the ML, ivermectin, was at a higher prevalence than expected. Sheep farmers and advisors in New Zealand need to re-evaluate the way they manage parasites, and more research is urgently needed if the steady decline in anthelmintic susceptibility is to be halted.     

In vitro field screening for anthelmintic resistance in strongyles of sheep and horses

Simplified in vitro field methods are described for the detection and assay of benzimidazole-resistance in sheep trichostrongylids and horse strongyles. Worm eggs are recovered from fresh faeces, within 1 hour of collection, by flotation in sugar solution. The separated eggs are then incubated for 20–24 hours at 27–30° C in solutions of thiabendazole in distilled water ranging from 0.1 to 1.1 p.p.m. for sheep trichostrongylids and from 0.05 to 0.5 p.p.m. for horse strongyles. Under controlled conditions, eggs from thiabendazole-susceptible individuals of both sheep and horse nematodes rarely hatch at thiabendazole concentrations of 0.1 p.p.m. Eggs from resistant individuals will hatch at 0.1 p.p.m. and above. Semi-quantitative estimates of the level of resistance can be determined by measuring the % egg-hatch at varying concentrations of thiabendazole. A field method for selecting test animals with low egg-counts, and an in vitro method for the culture of eggs or first-stage larvae to third stage for identification are described.     

Antimicrobic and anthelmintic resistance.

Antimicrobial and anthelmintic resistance are growing issues for the equine practitioner. The development of antimicrobial or anthelmintic resistance is a source of significant concern because of increased frequency of treatment failures and increased treatment costs. In addition, antimicrobial resistance may have important consequences for public health. Only through judicious use can the efficacy of antimicrobials and anthelmintics be prolonged. This article discusses the development of resistance and suggestions for control.     

Benzimidazole resistance in equine cyathostomes in Slovakia

The present study included 19 stud farms, including 243 horses, that were investigated for the occurrence of anthelmintic resistant cyathostomes. The number of horses on the farms varied from nine to more than 100, and horses of all ages were included. A minimum of seven horses were used for faecal egg count reduction (FECR) tests. The anthelmintics included were: fenbendazole (paste formulation), ivermectin (paste formulation) and pyrantel (powder). Resistance to benzimidazoles was detected on 14 farms, with FECR values ranging from 65.1 to 86.3%. Larval cultures after fenbendazole treatment revealed exclusively cyathostome larvae. Ivermectin was tested on eight farms and proved to be effective on all. Pyrantel was tested on two farms and FECR test indicated high efficacy (92-97%). Egg hatch assay (EHA) results showed that mean concentrations of thiabendazole that inhibited hatching in 50% of the eggs (ED(50)) in resistant populations were over 0.1 microg ml(-1). The results of our study suggest widespread resistance to fenbendazole in equine cyathostomes in Slovakia, and possible strategies to delay anthelmintic resistance are discussed briefly.     

Larval development assay for detection of anthelmintic resistance in cyathostomins of Swedish horses

The aim of this study was to investigate the suitability of a larval development assay (LDA) for the determination of anthelmintic resistance in cyathostomin nematode populations of the horse. In addition, comparison of results between geographic regions, types of horse establishment, and the use of anthelmintics in Sweden, was established. Seventy horse herds from different parts of Sweden were sampled, and strongyle eggs from the faeces of 54 of those were investigated by an LDA (DrenchRite). The following anthelmintics were tested: thiabendazole (TBZ), levamisole (LEV), ivermectin monosaccharide (IVM-MS), ivermectin aglycone (IVM-AG) and pyrantel (PYR). The LC50 values for TBZ and LEV were generally lower than those previously reported in other LDA studies on horse nematodes. This could be related to the infrequent use of these compounds for the past 20 years in Sweden. In this study, there was a great variation within and between assay plates that could not be explained. Still the LC50 values differed significantly between the regions for all anthelmintics, except for pyrantel. The highest LC50s were observed in parasite populations from the south of Sweden. There were no significant differences between riding schools and studs. Limitations of this technique exist, namely the lack of established cut-off values for susceptible and resistant populations and interpretation problems related to multi-species infections. Although there are advantages with LDA such as the possibility of testing several compounds simultaneously without interference with the deworming programmes on the farms, we conclude that LDA currently is not a reliable alternative to the faecal egg count reduction test (FECRT).     

The anthelmintic efficacy of cambendazole in horses

The anthelmintic activity of cambendazole 31% paste was evaluated in a controlled trial against naturally acquired gastro-intestinal parasites of the horse. Four groups of five adult horses were treated at dose rates of 0, 15, 20 and 25 mg/kg. The three dose levels of cambendazole tested were efficacious against the large strongyles Strongylus edentatus and S. vulgaris, the stomach worm Trichostrongylus axei, the pinworms Oxyuris equi and Probstmayria vivipara and all the species of small strongyles present. No efficacy was demonstrated against the tapeworm Anoplocephala perfoliata or the horse bots Gasterophilus intestinalis or G. nasalis     

A field evaluation of anthelmintics for control of cyathostomes of horses in Brazil

The anthelmintic efficacy of ivermectin (IVM), piperazine citrate (PPZ) and fenbendazole (FBZ) alone, and combinations of FBZ with varying dosages of PPZ, was evaluated for the control of strongyles of Mangalarga equids. In all cases of positive strongly egg counts per gram of faeces (EPG) before and after treatment, pure populations of cyathostome larvae with eight gut cells were consistently observed in faecal cultures. The faecal egg count reductions (FECR) were evaluated between Day 7 and Day 56 post-treatment. Significant differences (P less than or equal to 0.05) were found between Day 7 and Day 49 when comparing the mean EPG values of IVM with those of the other anthelmintics. PPZ alone and the combination of FBZ (7.5 mg kg-1) and PPZ (55 mg base kg-1) did not show significant differences during the trial; both groups exhibiting an FECR of greater than 90% between Day 7 and Day 30. Horses treated with the combinations of FBZ (7.5 mg kg-1) and PPZ at either 40 or 25 mg base kg-1, and with FBZ alone at either 7.5 or 15 mg kg-1, showed an FECR of less than 90% 2 weeks after treatment. The emergence of a degree of resistance of strongyles to FBZ and PPZ in a segment of the cyathostomes in this population is suggested.