High-level pyrantel resistance in the hookworm Ancylostoma caninum

While anthelmintic resistance is now a widely recognized issue in the livestock industries, its existence within companion animal medicine has been rarely established conclusively. We undertook a placebo-controlled in vivo trial to measure the efficacy of pyrantel embonate against pooled isolates of the hookworm Ancylostoma caninum from Brisbane, Australia. A statistically significant fall in adult worm burden was observed among dogs in the pyrantel treatment group compared to the control dogs (178.0+/-24.5 versus 239.7+/-14.0; p=0.02), equating to an efficacy of just 25.7% (95% CI, 15.0-35.1%), as based upon reduction in mean worm burden. Analysis of faecal egg count trends through the course of the study revealed that egg counts rose in both control and pyrantel-treated dogs, with a greater rise observed in the latter group (11.6+/-8.3% versus 17.3+/-7.6%; p=0.04), despite the decrease in adult worm numbers in this group. Our results indicate that high-level anthelmintic resistance does occur in companion animal medicine, and highlight the need for greater vigilance and more judicious use of anthelmintics in small animal practice. They further indicate that the faecal egg count reduction test needs to be used with caution with this parasite.     

Pyrantel in small animal medicine: 30 years on

Pyrantel, a tetrahydropyrimidine nicotinic agonist anthelmintic, has been used in companion animal medicine since the 1970s to control two important nematode groups, the hookworms and the roundworms. Given the zoonotic potential of these parasites, pyrantel has served a dual role in helping to protect the health of both companion animals and the public for more than 30 years. This review describes the history and mechanism of action of this drug, and collates evidence that resistance to pyrantel has developed in at least one canine nematode, the hookworm Ancylostoma caninum. The role of in vitro diagnosis tests in managing anthelmintic resistance in companion animal parasites is discussed, as are management practices that may reduce the rate at which resistance develops.     

Anthelmintic treatment in horses: the extra-label use of products and the danger of under-dosing

Anthelmintic products form the basis of helminth control practices on horse stud farms at present. Regular evaluation of the efficacy of these products is advisable, as it will provide information on the worm egg reappearance period and the resistance status in the worm population. The aim of this study was to evaluate the efficacy of doramectin, pyrantel pamoate, ivermectin and moxidectin on a Thoroughbred stud farm in the Western Cape Province, South Africa. The study also compared the anthelmintic efficacy of two moxidectin formulations administered at their recommended dosages (an injectable, at 0.2 mg/kg, not registered for horses, and an oral gel at 0.4 mg/kg, registered for horses). Two mixed-sex groups of 30 yearlings and 40 weaners were tested in 2001 and 2002, respectively, divided into 3 and 4 groups of equal size. In 2001, moxidectin was one of 3 drugs administered orally and at a dose rate of 0.4 mg/kg. In 2002, pyrantel pamoate and ivermectin were orally administered at 19 and 0.2 mg/kg. Moxidectin and doramectin (the latter not registered for horses) were administered by intramuscular injection at a dose of 0.2 mg/kg, the dosage registered for other host species. The faecal egg count reduction test was used to determine the anthelmintic efficacies in both years. Each animal acted as its own control and the arithmetic mean faecal egg count and lower 95% confidence limit was calculated for each of the groups. A 100% reduction in the faecal egg counts and a 100% lower 95% confidence limit was recorded for moxidectin (0.4 mg/kg) in 2001. In 2002, a 99% and 96% reduction was recorded for pyrantel pamoate and ivermectin, respectively. In the same year doramectin and moxidectin (both injectable and given at 0.2 mg/kg) did not have any effect on worm egg counts. Of the 4 drugs tested in 2002, only pyrantel pamoate recorded lower 95% confidence limits above 90%.     

Effects of artemisinin and Artemisia extracts on Haemonchus contortus in gerbils (Meriones unguiculatus)

Anthelmintic resistance is a worldwide concern in small ruminant industry and new plant-derived compounds are being studied for their potential use against gastrointestinal nematodes. Mentha piperita, Cymbopogon martinii and Cymbopogon schoenanthus essential oils were evaluated against developmental stages of trichostrongylids from sheep naturally infected (95% Haemonchus contortus and 5% Trichostrogylus spp.) through the egg hatch assay (EHA), larval development assay (LDA), larval feeding inhibition assay (LFIA), and the larval exsheathment assay (LEA). The major constituent of the essential oils, quantified by gas chromatography for M. piperita oil was menthol (42.5%), while for C. martinii and C. schoenanthus the main component was geraniol (81.4% and 62.5%, respectively). In all in vitro tests C. schoenanthus essential oil had the best activity against ovine trichostrongylids followed by C. martini, while M. piperita presented the least activity. Cymbopogon schoenanthus essential oil had LC(50) value of 0.045 mg/ml in EHA, 0.063 mg/ml in LDA, 0.009 mg/ml in LFIA, and 24.66 mg/ml in LEA. The anthelmintic activity of essential oils followed the same pattern in all in vitro tests, suggesting C. schoenanthus essential oil could be an interesting candidate for nematode control, although in vivo studies are necessary to validate the anthelmintic properties of this oil.     

Efficacy of 3 anthelmintics in communally grazed sheep as reflected by faecal egg count reduction tests in a semi-arid area of South Africa

A survey was conducted on the occurrence of anthelmintic resistance of nematodes in communally grazed sheep in a semi-arid area near Mafikeng, South Africa, from January to March 2006. In the herds belonging to twelve smallholder sheep farmers, the efficacies of albendazole, levamisole and closantel were tested by faecal egg count reduction tests where 80 % efficacy was considered the cut off for anthelmintic resistance. The results of the faecal egg count reduction tests showed more than 80% efficacy with all the drugs used in most cases, but there were notable exceptions. In 1 case, closantel had an efficacy of 72%, albendazole had an efficacy of 68% and levamisole showed efficacies of 58%, 60% and 75% respectively on 4 farms. The occurrence of anthelmintic resistance in this farming sector is of concern and steps should be taken to prevent its further spread and development to avoid a situation developing as on numerous commercial sheep farms in South Africa where resistance is common.     

The in vitro diagnosis of anthelmintic resistance in cyathostomins

Cyathostomins are the primary parasitic pathogens of equids. For over 40 years, these nematodes have been controlled using broad spectrum anthelmintics. Three classes of anthelmintic are currently available for this use but, unfortunately, resistance to each of these has now been recorded in cyathostomin populations. As part of an optimal strategy to control cyathostomin infections in the field, it will be important to identify drug-resistant worms at as early a stage as possible. This objective needs to be supported by methodologies that will allow the accurate comparison of anthelmintic resistance in different nematode populations. At present, the faecal egg count reduction test is considered the most suitable method for initial screening for anthelmintic resistance in equine nematode populations. However, in its current state, this test lacks sensitivity. It is also costly and time-consuming to perform. Laboratory-based techniques, such as the egg hatch assay, larval development assay, larval migration inhibition assay and the larval feeding inhibition assay offer alternative options for assessing anthelmintic resistance in nematode populations. All of these tests have been investigated for their utility in measuring drug resistance in sheep nematode populations and some have proven useful. The egg hatch assay, larval development assay and larval migration inhibition assay have been investigated for use in measuring levels of drug resistance in equine nematode populations. However, at best, the results obtained thus far indicate that these tests require further refinement.     

An evaluation of techniques used for the detection of anthelmintic resistance in nematode parasites of domestic livestock

In vitro and in vivo techniques used for the detection of anthelmintic resistance are reviewed in terms of their versatility, reliability, accuracy, cost and simplicity. The faecal egg count reduction test is considered the most suitable for field screening of resistance but the larval development assay is likely to prove a valuable adjunct in such investigations. Other in vitro techniques, such as egg hatch tests, larval motility and tubulin binding assays are recommended as suitable only for research investigations. Similarly, in vivo procedures such as the critical and controlled anthelmintic efficacy tests are considered to be appropriate only for specific research purposes because of the costs involved in such studies.     

Reduced efficacy of anthelmintics in young compared with adult horses

Studies on a Thoroughbred breeding farm in Ohio from 1982 to 1988 demonstrated the value of three anthelmintic pastes (ivermectin, oxibendazole, pyrantel pamoate) in controlling benzimidazole resistant cyathostomes (small strongyles) in adult horses. However, a comparison of drug efficacy in suppressing faecal egg counts for the full period between treatments showed a significant reduction in efficacy of all drugs in yearling horses compared with adults. Mean faecal egg counts of adult horses were generally kept below 100 eggs per gram (epg) of faeces when using oxibendazole or pyrantel pamoate at four to five week intervals and ivermectin at eight week intervals. By contrast, mean counts of young horses rose as high as 655 epg (oxibendazole), 729 epg (pyrantel pamoate) and 852 epg (ivermectin) within the same time period after treatment. Individual counts of treated yearlings sometimes exceeded 3,000 epg. Three distinct mechanisms appeared to be involved in the poor results in young horses. These were 1) anthelmintic refuge, 2) anthelmintic resistance, and 3) anthelmintic avoidance.     

Evidence for multiple anthelmintic resistance in two nematode parasite genera on a Saanen goat dairy

Multiple resistance was demonstrated within the experimental milking goat herd at the Ruakura Agricultural Centre. The development of this resistance was probably exacerbated by the atypical animal management and parasite control programmes under which the dairy was managed. Resistance by Teladorsagia (Ostertagia) spp. to members of the three broad-spectrum families of anthelmintic is reported for the first time. Worm counts in a controlled slaughter trial demonstrated multiple anthelmintic resistance in a mixed population of Teladorsagia (T. circumcincta and T. trifurcata) and Trichostrongylus spp. The efficacies of oxfendazole, morantel citrate and ivermectin in reducing the Teladorsagia population were 43.8%, 75.6% and 93.1%, respectively. Ivermectin removed more than 99% of the small intestinal Trichostrongylus spp. but oxfendazole and morantel were less effective, removing 37.9% and 87.7%, respectively. Only treatment with ivermectin totally eliminated faecal nematode egg counts 8 days after treatment. The lack of agreement between faecal nematode egg depressions and worm counts in lambs given ivermectin suggests that the drug may have affected the fecundity of the surviving nematode population or the assay lacked the sensitivity required. These data suggest that the sole use of faecal egg depression would be inappropriate for accurate assessment of anthelmintic effectiveness.     

Efficacy of ivermectin and pyrantel pamoate combined in a chewable formulation against heartworm, hookworm, and ascarid infections in dogs.

Eight trials were conducted in dogs to document the efficacy of ivermectin (6 micrograms/kg of body weight) and pyrantel pamoate (5 mg of active pyrantel/kg) in a beef-based chewable formulation against Dirofilaria immitis, Ancylostoma caninum, Uncinaria stenocephala, Toxocara canis, and Toxascaris leonina. Three studies involved induced infection with D immitis, and 5 studies involved induced or natural infection with hookworms and ascarids. In 3 intestinal parasite trials, the efficacy of the combination chewable tablet was compared with each of its components. Results indicated that 1 component did not interfere with the activity of the other. In 1 heartworm and 2 intestinal parasite trials, the efficacy of pyrantel, ivermectin/pyrantel combination, or ivermectin with pyrantel dosage of 10 mg/kg was evaluated. The ivermectin/pyrantel combination was 100% effective in preventing development of D immitis larvae. Efficacy of the combined product against T canis, Toxascaris leonina, A caninum, and U stenocephala was 90.1, 99.2, 98.5, and 98.7%, respectively. In the intestinal parasite trials, each individual component was found not to interfere with the anthelmintic action of the other. Increasing the dosage of pyrantel to 10 mg/kg (2 x that in the combination) did not interfere with the efficacy of ivermectin against heartworm or increase the activity of pyrantel against intestinal parasites.     

Anthelmintic Resistance on an Organized Sheep Farm in India

Monitoring anthelmintic resistance in strongyle nematodes by the faecal egg count reduction test and a commercial larval development assay on an organized sheep farm in the semi-arid area of Rajasthan revealed the emergence of resistance to benzimidazoles and rafoxanide and a potential risk of the development of levamisole/tetramisole resistance. A benzimidazole/levamisole combination, avermectins and closantel were each found to be efficacious.     

Prevalence of anthelmintic resistant nematodes in sheep in south-east England

The prevalence of anthelmintic resistant nematodes among 52 commercial flocks in south-east England was investigated by comparing the faecal egg counts of groups of lambs before, and seven days after, treatment with thiabendazole and levamisole. Evidence of thiabendazole resistance was found on seven farms. in each case Haemonchus contortus was the only species of nematode involved. In vitro egg hatch assays carried out for isolates from these farms gave ED50 estimates of 0.065 to 0.332 micrograms thiabendazole/ml compared with estimates of 0.027 to 0.031 micrograms thiabendazole/ml for a known susceptible strain of H contortus assayed at the same time. In a series of slaughter trials, there was a 17 to 85 per cent reduction, as compared with controls, in the mean worm burdens of groups of lambs infected with these isolates and killed seven days after treatment with thiabendazole, confirming their resistance to this anthelmintic.     

A Field Survey on Anthelmintic Resistance in Equine Small Strongyles in Norway

A field survey at 17 stables involving 221 horses was performed to evaluate the presence of anthelmintic resistance in the equine small strongyles (cyathostomes). The horses were allocated into treatment groups, and resistance to fenbendazole (FBZ), pyrantel pamoate (PYR) and ivermectin (IVM) was tested by the faecal egg count reduction test (FECR-test). Faecal samples were collected at the time of treatment, 14 days post treatment and 90 days post treatment.Resistance to FBZ, which was defined as a faecal egg count reduction <95%, was found in 14 out of 17 stables. In 2 of the 14 stables the egg reductions were close to the limit of 95%, 91 and 93%, respectively. In 1 stable the egg reductions indicated resistance to PYR as well as detection of resistance to FBZ, 94% reduction for PYR and 85% for FBZ. No signs of resistance were detected to IVM.The investigation was performed in late autumn and winter, and due to the climatic conditions and cleaning procedures in the stables no reinfection took place during this period. The faecal egg count reduction from treatment till day 90 post treatment was used as an expression of the effect of PYR and IVM on the early stage (hypobiotic), late third stage and fourth stage larvae in the gut wall. This was justified because there was no reinfection and because the 14 day post treatment egg counts were zero or close to zero for the PYR and IVM treatment groups. The effects of PYR and IVM on the larval stages were compared and no statistically significant differences were found.     

Use of two in vitro methods for the detection of anthelmintic resistant nematode parasites on Slovak sheep farms

A survey of the prevalence of anthelmintic resistant nematode populations was conducted on 32 sheep farms in the Slovak Republic. In vitro egg hatch test and larval development tests were used for the detection of resistance to benzimidazole anthelmintics and they were compared with in vivo faecal egg count reduction tests. There was agreement in the declaration of resistance between the faecal egg count reduction test and both in vitro tests. The presence of resistant populations was determined on two farms using egg hatch test. In both farms, the LD(50) values were higher than 0.1 microg TBZ/ml, indicating resistance. By using LD(99) values it might be possible to reveal relatively small proportion of resistant larvae in the population. The prevalence of benzimidazole resistance has not change on Slovak sheep farms during last decade.     

Condensed tannins act against cattle nematodes

The use of natural plant anthelmintics was suggested as a possible alternative control of gastrointestinal nematodes (GIN) in ruminants. Direct anthelmintic effects of tannin-containing plants have already been shown in sheep and goat GIN. These anthelmintic properties are mainly associated with condensed tannins. In the present study, we evaluated possible in vitro effects of three tannin-containing plants against bovine GIN. Effects of Onobrychis viciifolia, Lotus pedunculatus and Lotus corniculatus condensed tannin (CT) extracts on Cooperia oncophora and Ostertagia ostertagi were determined by a larval feeding inhibition assay (LFIA) and a larval exsheathment assay (LEA). In the LFIA, all three plant extracts significantly inhibited larval feeding behaviour of both C. oncophora and O. ostertagi first stage larvae in a dose-dependent manner. The L. pedunculatus extract, based on EC(50) (effective concentration for 50% inhibition), was the most effective against both nematodes, followed by O. viciifolia and L. corniculatus. The effect of CT extracts upon larval feeding behaviour correlates with CT content and procyanidin/prodelphidin ratio. Larval exsheathment of C. oncophora and O. ostertagi L3 larvae (third stage larvae) was also affected by CT extracts from all three plants. In both in vitro assays, extracts with added polyvinylpolypyrrolidone, an inhibitor of tannins, generated almost the same values as the negative control; this confirms the role of CT in the anthelmintic effect of these plant extracts. Our results, therefore, indicated that tannin-containing plants could act against cattle nematodes.     

Resistance to levamisole and cross-resistance between pyrantel and levamisole in Oesophagostomum quadrispinulatum and Oesophagostomum dentatum of pigs

Two strains of Oesophagostomum spp., consisting of both O. quadrispinulatum and O. dentatum, were subjected to a controlled in vivo assay for resistance to levamisole and pyrantel by comparison with susceptible isolates. One strain (LEM) was recently isolated from a commercial herd, where sows showed high numbers of strongyle eggs in faeces within 2 weeks of farrowing and following treatment with levamisole at the manufacturer's recommended dose rate 1 week before farrowing. Levamisole had been used as the sole anthelmintic for treatment for at least 7 years on this farm. Treatment with pyrantel in this herd also indicated cross-resistance to this drug. A mixed population of O. quadrispinulatum and O. dentatum of this strain was subjected to controlled in vivo assays. Faecal egg count reduction (FECR) was found to be -573.3% (P greater than 0.05) and worm count reductions (WCR) of O. quadrispinulatum and O. dentatum were estimated as 44.5% (P greater than 0.05) and 96.4% (P less than 0.001), respectively. Treatment with pyrantel showed that cross-resistance existed to this drug, with FECR of 10.4% (P greater than 0.05) and WCR of 64.5% (P greater than 0.05) and 90.7% (P less than 0.05) for O. quadrispinulatum and O. dentatum, respectively. Another strain (VJ) was isolated from another commercial pig herd, which was dosed with pyrantel citrate four times a year for at least 8 years. This strain showed resistance to pyrantel, with FECR of 43.8% (P greater than 0.05) and WCR of 65.9% (P greater than 0.05) and 49.4% (P greater than 0.05) for O. quadrispinulatum and O. dentatum, respectively. However, both species were susceptible to levamisole. Our results suggested that selection with levamisole gave rise to levamisole resistance and automatically conferred resistance to pyrantel, whereas selection with pyrantel only resulted in resistance to this drug alone. These findings are discussed in relation to the location of the two species of Oesophagostomum in the large intestine of pigs and the mode of action of this class of anthelmintics.     

Characterisation of two triple resistant field isolates of Teladorsagia from Scottish lowland sheep farms

The anthelmintic resistance status of two field isolates derived from farms (farm A and B) located near Edinburgh were examined using both controlled efficacy tests (CET) and faecal egg count reduction tests (FECRT). Efficacies against fenbendazole (FBZ), levamisole (LEV) and ivermectin (IVM) and, for one isolate, against combinations of these anthelmintics and moxidectin were determined in na?ve lambs, artificially infected with the isolates and treated with the compounds at the manufacturers recommended dose rates. (FBZ, 5mg/kg bodyweight (BW); LEV, 7.5mg/kg BW; IVM, 0.2mg/kg BW; Moxidectin (MOX) 0.2mg/kg BW). In both field isolates, the predominant species found pre-treatment and the only species found post-treatment was Teladorsagia circumcincta. Resistance to FBZ, LEV and IVM was confirmed in CET and FECRT on farm A and to the latter two compounds on farm B, which had a history of benzimidazole resistance and where TBZ resistance was also demonstrated using an egg hatch assay (EHA). For the farm A isolate CET efficacies against FBZ; IVM; LEV; FBZ + IVM; FBZ + LEV; FBZ, LEV + IVM and MOX were 59, 60, 88, 94,93, 92 and 98%, respectively. The CET efficacies for the farm B isolate were 51% and 72% for LEV and IVM, respectively.     

A comparison of in vitro tests and a faecal egg count reduction test in detecting anthelmintic resistance in horse strongyles.

This study reports a comparison between faecal egg count reduction test (FECRT), egg hatch assay (EHA) and larval development assay (LDA) for detecting anthelmintic resistance in equine strongyles. Resistance to benzimidazoles was demonstrated in 33 of 42 (79%) farms tested by FECRT and in 32 (62%) of the 52 farms tested by EHA. As the reference strain used was not fully susceptible to benzimidazoles it was not possible to determine the level of resistance by LDA. Pyrantel resistance was indicated on three of 15 farms by faecal egg count reduction. Resistance was also indicated by LDA for one of these farms. In addition resistance was indicated by LDA on two more farms that were not tested by FECRT. Further testing is needed to confirm if these findings are truly indicative of resistance. Generally, correlations between the tests were poor and it was not possible to use the outcome of one test to predict the outcome of another.     

Management of drug-resistant cyathostominosis on a breeding farm in central North Carolina

REASONS FOR PERFORMING STUDY: Possible anthelmintic resistance on a breeding farm where a rapid rotation anthelmintic programme had been implemented for 9 years was investigated. Cyathostomins resistant to fenbendazole and pyrantel were documented by faecal worm egg count reduction test (FWECRT). OBJECTIVES: To 1) manage small strongyle transmission in a herd of horses in which resistance to both pyrantel pamoate and fenbendazole was identified and thereby reduce the risk of clinical disease in the individual animal, 2) monitor the change in resistance patterns over time and 3) monitor the efficacy of ivermectin over the study period. METHODS: Targeted ivermectin treatment of horses on the farm was instituted for mature horses with faecal worm egg counts (FWEC) > 200 eggs/g (epg) and for horses < age 2 years with FWEC > 100 epg. RESULTS: Over a 30 month period, targeted ivermectin treatment achieved acceptable control in mares, as judged by FWEC, and improved control of patent cyathostome infection in consecutive foal crops. Egg reappearance time (ERT) after treatment with ivermectin was < 8 weeks in mares and foals more frequently in the second year of the study than in the first year. Numbers of anthelmintic treatments were reduced by 77.6 and 533% in the mare and foal group, respectively. CONCLUSIONS: Targeted ivermectin treatment may be an economically viable method of managing multiple drug resistant cyathostominosis. POTENTIAL RELEVANCE: Use of ivermectin should be monitored closely for development of resistance.     

Impaired efficacy of ivermectin against Parascaris equorum, and both ivermectin and pyrantel against strongyle infections in trotter foals in Finland

In order to assess the resistance situation against macrocyclic lactones in Parascaris equorum and against tetrahydropyrimidine derivatives in strongyles in Finnish trotter horses, 112 foals on 18 farms, mostly 1 year old, were examined for these parasites with a modified McMaster faecal flotation method. P. equorum positive foals (n=24) were given ivermectin orally at a dose of 200 μg/kg b.w., while strongyle positive but P. equorum negative foals (n=38) received pyrantel embonate orally at a dose of 19 mg/kg. Sixteen P. equorum infected foals, treated with ivermectin, also harboured strongyles. During the anthelmintic treatment visit to the farm, Faecal Egg Count Reduction Test (FECRT) reference (first) samples were collected. Fourteen days later, the second sampling (reduction samples) was done. The FECR was calculated for each foal/parasite combination. The reduction efficacies of ivermectin against P. equorum (mean 52%, calculated from the individual egg count reductions) and pyrantel against strongyles (43%) were strongly indicative of widespread resistance. Also indication of ivermectin resistance among strongyles was seen. The widespread use of anthelmintics for Finnish horses obviously has resulted in resistance, as has happened elsewhere, too.