Simultaneous resistance to two or more broad-spectrum anthelmintics by gastrointestinal nematode parasites of sheep and goats

The detection, by means of faecal egg count reduction tests and larval cultures, of two cases of multiple anthelmintic resistance in goats and two in sheep, is reported. The former cases appeared to involve resistance to all three broad-spectrum drench groups (benzimidazole, levamisole, ivermectin) by mainly Ostertugia spp., the latter resistance to two of them (benzimidazole, levamisole) bv Trichostrondus spp. (New Zealand Veterinary Journal 38, 114–117, 1990)     

The effectiveness of benzimidazole-levamisole combination drenches in the presence of resistance to both benzimidazole and levamisole anthelmintics in New Zealand sheep

Examination of 25 cases of multiple benzimidazole and levamisole resistance, identified in sheep by faecal egg count reduction testing at the Batchelar, Lincoln and Invermay Animal Health Laboratories, showed that benzimidazole-levamisole combinations provided effective control in eight (47%) of 17 cases in which they were tested. Overall, the use of combination drenches resulted in average improvements in faecal egg count reductions of 25.6% (p<0.01) and 23.2% (p<0.05), respectively, over those achieved by the use of benzimidazole or levamisole drenches alone. The results suggest that instances of multiple resistance in which combination drenches might be effective are unlikely to be predictable either by identification of the parasites involved, or from the levels of benzimidazole or levamisole resistance present.     

Anthelmintic resistance in Australian sheep nematode populations

SUMMARY The resistance status of gastro–intestinal nematodes to anthelmintics was evaluated on 881 sheep farms throughout Australia during 1991–92. Resistance was shown to be widespread. Overall, 85% of farms had sheep infected with nematodes resistant to benzimidazole, 65% to levamisole and 34% to combination (benzimidazole + levamisole) products. Resistance to ivermectin was not detcted. On only 9% of farms did all anthelmintic groups reduce egg counts by greater than or equal to 95%. The culture of faeces from untreated sheep showed Telodorsagia circumcincta, Trichostrongylus spp, Chabertia ovina and Haemonchus contortus to be the principal species. The nature and prevalence of resistance was not significantly correlated with stocking rate. However, resistance to combination products was almost twice as prevalent on farms in areas with an average annual rainfall of greater than 500 mm.     

Anthelmintic resistance in the field: Changes in resistance status of parasitic populations in response to anthelmintic treatment

Changes in anthelmintic resistance in nematode parasites were monitored in sheep grazing on 2 separate farms, but with the same anthelmintic treatment program, over 16 years. High levels of benzimidazole resistance emerged in Ostertagia and Trichostrongylus spp populations on both farms following 9 years of continuous use of this class of drug. Subsequently, variations in the levels of resistance occurred for the same species between farms and between species on the same farm. A change to levamisole for 2 years resulted in a significant reversion towards benzimidazole susceptibility, but a concomitant rise in levamisole resistance, in Ostertagia on one farm. However, benzimidazole resistance increased rapidly following the re-introduction of oxfendazole into the anthelmintic treatment program. Results from both farms illustrate the pitfalls of using one anthelmintic class for an extended period and provide indirect support for the alternation of anthelmintic classes at approximately yearly intervals.     

Anthelmintic resistance in nematode parasites of goats

Cases of anthelmintic resistance on 3 goat farms in Gippsland were investigated. On the first farm Haemonchus contortus, Trichostrongylus colubriformis and Ostertagia (Teladorsagia) circumcincta were found to be resistant to fenbendazole, but the first 2 species were fully susceptible to levamisole. On the second farm a population of T. colubriformis, resistant to concurrent full doses of levamisole and a benzimidazole, was found to retain this resistance when transferred to sheep. On the third farm, heavy mortality due to Ostertagia and Trichostrongylus spp burdens was arrested only by the use of concurrent full doses of levamisole and oxfendazole; no single anthelmintic was found to be effective. A disturbing rise in the incidence of clinical helminthiosis, often accompanied by anthelmintic resistance, occurred on goat farms in Gippsland early in 1984.     

Efficacy of monepantel and anthelmintic combinations against multiple-resistant Haemonchus contortus in sheep, including characterisation of the nematode isolate

Three experiments defined the resistance profile of a population of Haemonchus contortus, which was shown to express multiple resistances to the benzimidazole, levamisole, macrocyclic lactone and salicylanilide anthelmintic classes when given as a registered combination. Study 1 was a faecal egg count reduction (FECR) test and the efficacies for the anthelmintics were monepantel, 100%; abamectin+levamisole+oxfendazole, 40.0%; and abamectin+levamisole+oxfendazole+naphthalophos, 100%. No larvae were recovered from the post-treatment cultures for monepantel or the 4-way treatment, and for the 3-way treatment the culture was 100% Haemonchus spp. Efficacies in Study 2 were calculated from mean post-mortem nematode burdens of H. contortus and were levamisole+oxfendazole, 3.1%; abamectin+levamisole+oxfendazole, 5.0%; ivermectin, 0.4%; moxidectin, 28.4% and closantel, 70.2%. Study 3 was also a FECR test that resulted in efficacies of 100% for monepantel and 83.0% for a formulated 4-way combination of abamectin+levamisole+albendazole+closantel. Larvae recovered from the post-treatment culture for the combination-treated sheep were all Haemonchus spp. Multi-resistant parasites such as examined in these studies are a continuing challenge to be managed by farmers and their advisors. Control programs must be planned and well-managed, and should include on-farm testing for anthelmintic resistance, monitoring of nematode burdens (by FEC and larval culture) to determine appropriate treatment times and the management of pastures to reduce the overall parasite challenge. This should be in balance with the generation, use and maintenance of drug-susceptible nematode populations in refugia.     

Multiple species of nematodes resistant to ivermectin and a benzimidazole-levamisole combination on a sheep farm in New Zealand

AIM: To confirm the presence of multiple anthelmintic resistance on a sheep farm in New Zealand. METHODS: Three groups of 10 weaned Romney-cross lambs were treated either with an oral dose of ivermectin (0.2 mg/kg), or a benzimidazole/levamisole (BZ/LEV) combination (4.75 albendazole and 7.5 mg/kg levamisole), or were left untreated. Ten days later, animals were necropsied, and adult worms recovered and identified from the abomasa and small intestines. Pre- and post-treatment faecal nematode egg counts (FEC) were recorded, and larval cultures were performed. RESULTS: In a faecal egg count reduction test (FECRT), adjusted to reflect pre- and post-treatment larval culture results, ivermectin resistance was detected in Teladorsagia (Ostertagia), Trichostrongylus and Haemonchus spp, while BZ/LEV combination- resistant Teladorsagia and Trichostrongylus spp were also present. Adult worm counts confirmed these results, and identified the species involved as Teladorsagia circumcincta, Trichostrongylus colubriformis and H. contortus. CONCLUSION: Multiple, multi-generic anthelmintic resistance was confirmed on a sheep property in New Zealand. This included the first confirmed case of ivermectin resistance in T. colubriformis from sheep in New Zealand.     

Anthelmintic resistance in sheep and goat farms on Peninsular Malaysia.

The faecal egg count reduction test (FECRT) was conducted on 39 sheep farms and 9 goat farms located in Peninsular Malaysia. The anthelmintic groups used in these tests were the benzimidazoles, levamisole, the benzimidazole/levamisole combination, macrocyclic lactones and closantel. Results indicated that the prevalence of resistance to the benzimidazole group was high, with approximately 50% of the sheep farms and 75% of the goat farms having resistant nematode parasite populations present. Resistance to levamisole, closantel and ivermectin was also detected. Differentiation of the infective larvae derived from faecal cultures indicated that by far the most predominant parasite species was Haemonchus contortus.     

Lack of reversion of a benzimidazole resistant strain of Haemonchus contortus after six years of levamisole usage

The resistance level of Haemonchus contortus was studied on a farm where benzimidazole resistance had been detected in 1980 and where, since 1982, only levamisole had been used for worm control. Egg development tests gave LD50 values of 0.18, 0.19 and 0.21 microgram ml-1 thiabendazole for samples taken from ewes in 1988 compared with 0.21 microgram ml-1 in 1981. Studies on sheep experimentally infected with larvae cultured from the faeces of these ewes confirmed the continued resistance of H contortus to benzimidazoles, and demonstrated the absence of benzimidazole resistance in other nematode species together with the susceptibility of all species to levamisole.     

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.     

Multiple anthelmintic resistance in a goat herd

Anthelmintic resistance was monitored over a 30 month period within a goat herd in eastern Virginia, USA. Resistance to ivermectin, levamisole and benzimidazole drugs was detected in Haemonchus contortus using the fecal egg count reduction test (FECRT). When levamisole use was discontinued for 1 year, susceptibility to levamisole appeared to return. Although a single treatment with fenbendazole was able to reduce fecal egg counts by only 50%, two doses administered in a 12 h interval increased efficacy to 92%, however, confidence intervals indicated that resistance was still present. When fecal egg counts were determined the following year after several treatment using this protocol, the efficacy of fenbendazole had fallen again to 57% reduction in fecal egg counts. The predominant genus present in cultured composite fecal samples was Haemonchus. Trichostrongylus, Cooperia and Teladorsagia were also present in smaller numbers.     

Field studies investigating anthelmintic resistance in young cattle on five farms in New Zealand

AIM: To investigate the efficacy of pour-on anthelmintics against field strains of parasitic nematodes in young cattle on five farms in New Zealand. METHODS: Faecal nematode egg count (FEC) reduction (FECR) tests were carried out on five calf-rearing farms using pour-on formulations of levamisole, ivermectin, eprinomectin, and the simultaneous administration of levamisole and ivermectin. Faecal samples were collected per rectum before treatment and about 7, 14, 21 and 28 days after treatment, for FEC and faecal nematode larval culture. RESULTS: Resistance (i.e. <95% reduction in FEC) of Cooperia oncophora to ivermectin and eprinomectin was identified on all five farms. There was limited evidence of possible emerging resistance in Ostertagia spp to ivermectin but not eprinomectin, in short-tailed larvae of Cooperia spp to ivermectin and eprinomectin, and in Trichostrongylus spp to ivermectin, eprinomectin and levamisole used separately. Levamisole was effective against C. oncophora, but had variable efficacy against Ostertagia spp in the calves in this study. Simultaneous treatment with levamisole and ivermectin pour-on formulations were effective against all genera on all farms. CONCLUSIONS: To effectively manage roundworm parasites in their calves farmers need to be aware of the resistance status of the parasites on their farms. Levamisole is likely to be an effective anthelmintic on most farms at times of the year when the impact of Ostertagia spp is not high. Simultaneous administration of levamisole and ivermectin pour-on anthelmintics to cattle is likely to control both ML-resistant C. oncophora and stages of Ostertagia spp that are not controlled by levamisole alone.     

THE ISOLATION OF A FIELD STRAIN OF HAEMONCHUS CONTORTUS IN QUEENSLAND SHOWING MULTIPLE ANTHELMINTIC RESISTANCE

SUMMARY Following the apparent failure of levamisole to control infections of Haemonchus contortus in sheep at Lawes in south eastern Queensland, a strain of this parasite was isolated at the Animal Research Institute, Yeerongpilly. This strain was used to infect sheep at Yeerongpilly and the Merrindale Research Station, Victoria where four experiments to classify the resistance pattern of the parasite were carried out. Resistance to thiabendazole was first suspected in 1969, and these experiments confirmed that resistance to this drug was still present. They also showed that a strong degree of resistance had been developed to both levamisole and morantel tartrate. Other benzimidazole anthelmintics and also the organophosphorus compound naphthalophos were only moderately effective against the original isolate but rafoxanide, nitroxynil and phenothiazine were almost 100% effective. Other highly effective chemicals were disophenol and closantel. After passaging the strain for four generations with both levamisole and albendazole, resistance to both naphthalophos and the newer benzimidazole anthelmintics increased dramatically. This is the first report of a field strain of H. contortus exhibiting resistance to benzimidazole, non-benzimidazole and organophosphorus anthelmintics.     

Prevalence of anthelmintic resistance on 62 beef cattle farms in the North Island of New Zealand

AIM: To establish the prevalence of anthelmintic resistance in parasitic nematodes on a random sample of beef cattle herds in the North Island of New Zealand. METHODS: A cross-sectional prevalence study was conducted using a standardised faecal nematode egg count (FEC) reduction (FECR) test (FECRT) for ivermectin, levamisole and albendazole on 60 calves on each of 62 farms in the North Island chosen at random from farms that conformed with the selection criteria. 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 that completed the FECRT, 4/61 (7%) showed > or =95% reduction in FEC for all anthelmintics tested. Resistance to ivermectin was evident on 56/61 (92%) farms, to albendazole on 47/62 (76%) farms, and to both ivermectin and albendazole on 45/61 (74%) farms. Resistance to levamisole was evident on only 4/62 (6%) farms. The parasites most prevalent in resistant populations cultured were Cooperia spp. On 45/61 (74%) farms where Cooperia spp were present in sufficient numbers, resistance to both ivermectin and albendazole was evident. No cases of levamisole-resistant Cooperia spp were detected. Resistance of Ostertagia spp to ivermectin was evident on 4/45 (9%) farms, to albendazole on 15/46 (35%) farms, and to levamisole on 4/46 (9%) farms. CONCLUSION: Anthelmintic resistance in parasitic nematodes of cattle is common in the North Island of New Zealand. Beef farmers need to be aware of the risks posed by anthelmintic resistance, and routine FECR testing is recommended to ensure optimal productivity and to guide decision-making when purchasing anthelmintics to be used on-farm.     

Field studies investigating anthelmintic resistance in young cattle on five farms in New Zealand

AIM: To investigate the efficacy of pour-on anthelmintics against field strains of parasitic nematodes in young cattle on five farms in New Zealand.

METHODS: Faecal nematode egg count (FEC) reduction (FECR) tests were carried out on five calf-rearing farms using pour-on formulations of levamisole, ivermectin, eprinomectin, and the simultaneous administration of levamisole and ivermec- tin. Faecal samples were collected per rectum before treatment and about 7, 14, 21 and 28 days after treatment, for FEC and faecal nematode larval culture.

RESULTS: Resistance (i.e. <95% reduction in FEC) of Cooperia oncophora to ivermectin and eprinomectin was identified on all five farms. There was limited evidence of possible emerging resistance in Ostertagia spp to ivermectin but not eprinomectin, in short-tailed larvae of Cooperia spp to ivermectin and eprinomec- tin, and in Trichostrongylus spp to ivermectin, eprinomectin and levamisole used separately. Levamisole was effective against C. oncophora, but had variable efficacy against Ostertagia spp in the calves in this study. Simultaneous treatment with levamisole and ivermectin pour-on formulations were effective against all genera on all farms.

CONCLUSIONS: To effectively manage roundworm parasites in their calves farmers need to be aware of the resistance status of the parasites on their farms. Levamisole is likely to be an effective anthelmintic on most farms at times of the year when the impact of Ostertagia spp is not high. Simultaneous administration of levamisole and ivermectin pour-on anthelmintics to cattle is likely to control both ML-resistant C. oncophora and stages of Ostertagia spp that are not controlled by levamisole alone.     

Anthelmintic resistance in cattle nematodes on 13 south-west Victorian properties

Objective To report the level of anthelmintic resistance on 13 commercial cattle properties in south-west Victoria, Australia. Procedure Between 2006 and 2009 worm egg count reduction tests were conducted on calves on the 13 properties. Samples were collected 10–14 days post anthelmintic treatment and worm egg counts and larval differentiation tests were conducted. Resistance was defined if there was less than 95% reduction (lower confidence limit <90%) in the faecal worm egg count for the particular genus. Results The percentage of properties with anthelmintic resistance in at least one species was 54% for benzimidazole (BZ), 100% for levamisole (LEV) and for ivermectin (IVM) it was 100% for the half-dose (0.1 mg/kg) and 62% for the full dose (0.2 mg/kg). A substantial frequency of resistance was detected in Ostertagia ostertagi to BZ (5/11), LEV (3/3) and IVM (5/11), in Trichostrongylus spp. to BZ (4/7) and in Cooperia spp. to IVM (6/11). No resistance to LEV was detected in Trichostrongylus or Cooperia spp. Suspected IVM-resistant Trichostrongylus spp. and BZ-resistant Cooperia spp. were only detected on one property each. Conclusion This is the first Australian report of macrocyclic lactone-resistant O. ostertagi in the refereed literature. The frequency of resistance in O. ostertagi to BZ, LEV and IVM and in Trichostrongylus spp. to BZ in the present study appears higher than levels detected in the 2004–05 New Zealand survey, whereas the resistance frequency in Cooperia spp. to IVM and BZ was less.     

MULTIPLE AND MULTIGENERIC ANTHELMINTIC RESISTANCE ON A SHEEP FARM IN KENYA

The anthelmintic efficacy of benzimidazoles (albendazole, fenbendazole and oxfendazole), levamisole, oral ivermectin and closantel was evaluated on a farm in Kenya using faecal egg count reduction test, larval cultures and a controlled slaughter trial. The results of this study indicated simultaneous resistance of Haemonchus contortus against benzimidazoles, levamisole and ivermectin, and of Trichostrongylus colubriformis and Oesophagostomum spp. against levamisole on the same farm. Ivermectin resistance developed to 47% within 15 months of first use. Closantel was effective against the benzimidazoles, levamisole and ivermectin resistant H. contortus.     

Prevalence of anthelmintic resistance on 62 beef cattle farms in the North Island of New Zealand

AIM: To establish the prevalence of anthelmintic resistance in parasitic nematodes on a random sample of beef cattle herds in the North Island of New Zealand.

METHODS: A cross-sectional prevalence study was conducted using a standardised faecal nematode egg count (FEC) reduction (FECR) test (FECRT) for ivermectin, levamisole and albendazole on 60 calves on each of 62 farms in the North Island chosen at random from farms that conformed with the selection criteria. 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 that completed the FECRT, 4/61 (7%) showed ≥95% reduction in FEC for all anthelmintics tested. Resistance to ivermectin was evident on 56/61 (92%) farms, to albendazole on 47/62 (76%) farms, and to both ivermectin and albendazole on 45/61 (74%) farms. Resistance to levamisole was evident on only 4/62 (6%) farms. The parasites most prevalent in resistant populations cultured were Cooperia spp. On 45/61 (74%) farms where Cooperia spp were present in suffi cient numbers, resistance to both ivermectin and albendazole was evident. No cases of levamisole-resistant Cooperia spp were detected. Resistance of Ostertagia spp to ivermectin was evident on 4/45 (9%) farms, to albendazole on 15/46 (35%) farms, and to levamisole on 4/46 (9%) farms.

CONCLUSION: Anthelmintic resistance in parasitic nematodes of cattle is common in the North Island of New Zealand. Beef farmers need to be aware of the risks posed by anthelmintic resistance, and routine FECR testing is recommended to ensure optimal productivity and to guide decision-making when purchasing anthelmintics to be used on-farm.     

Further studies on anthelmintic resistance in sheep at an organised farm in arid region of Rajasthan

Faecal egg count reduction test (FECRT) and in vitro egg hatch assay (EHA) was performed on Indian Karakul (3/4 crosses of Karakul with Malpura, Marwari and Sonadi) transported from arid region campus, Bikaner, where resistance was reported in 1996. FECRT revealed that fenbendazole and levamisole were 100% effective in reducing the egg counts. LC50 value on EHA was 0.074 +/- 0.015 microg thiabendazole ml(-1). The faecal culture examination revealed the presence of Haemonchus contortus only. It was concluded that H. contortus was fully susceptible to both benzimidazole and levamisole.     

The detection of anthelmintic resistance in nematodes of veterinary importance

Before revised World Association for the Advancement of Veterinary Parasitology (WAAVP) guidelines on the detection of anthelmintic resistance can be produced, validation of modified and new methods is required in laboratories in different parts of the world. There is a great need for improved methods of detection of anthelmintic resistance particularly for the detection of macrocyclic lactone resistance and for the detection of resistant nematodes in cattle. Therefore, revised and new methods are provided here for the detection of anthelmintic resistance in nematodes of ruminants, horses and pigs as a basis for discussion and with the purpose that they are evaluated internationally to establish whether they could in the future be recommended by the WAAVP. The interpretation of the faecal egg count reduction test has been modified and suggestions given on its use with persistent anthelmintics and continuous release devices. An egg hatch test for benzimidazole (BZ) resistance is described. A microagar larval development test for the detection of benzimidazole and levamisole resistance provides third stage larvae for the identification of resistant worms. The sensitivity of these two tests can be increased by using discriminating doses rather than LD(50) values. Details are given of a PCR based test for the analysis of benzimidazole resistance in strongyles of sheep and goats, horses and cattle. Although promising for ruminant trichostrongyles, quantitative determination of gene frequency using real time PCR requires further development before PCR tests will be used in the field. Apart from faecal egg count reduction tests there are currently no satisfactory tests for macrocylic lactone resistance despite the great importance of this subject. Except for treatment and slaughter trials there are no validated tests for fasciolicide resistance or for the detection of resistance in cestodes.