Update on the prevalence of anthelmintic resistance in gastrointestinal nematodes of sheep in New Zealand

Abstract

Extract

Recently, a comparison was made between the prevalence of anthelmintic resistance, determined using faecal egg count reduction tests (FECRT), on randomly selected sheep farms in a systematic national survey (Waghorn et al. 2006 Waghorn, TS, Leathwick, DM, Rhodes, AP, Lawrence, KE, Jackson, R, Pomroy, WE, West, DM and Moffat, JR. 2006. Prevalence of anthelmintic resistance on sheep farms in New Zealand. New Zealand Veterinary Journal, 54: 271–277. [Taylor & Francis Online], [Web of Science ®] , [Google Scholar]) and that derived from similar case material submitted to a veterinary pathology laboratory on a more ad-hoc basis (McKenna 2008 McKenna, PB. 2008. An examination of the relative reliability of laboratory case submissions in determining the prevalence of anthelmintic resistance in sheep nematodes in New Zealand and the possible influence of test analysis methodology on such data. New Zealand Veterinary Journal, 56: 55–59. [Taylor & Francis Online], [Web of Science ®] , [Google Scholar]). The results of that comparison showed that while there were some differences between them, there were considerable similarities in the prevalence figures obtained from either source. Those similarities, which were particularly evident in terms of the overall pattern of involvement of the various worm genera and the types of anthelmintic concerned, led to the conclusion that FECRT case submissions to veterinary laboratories may offer a useful source of information regarding changes in the prevalence of anthelminticresistant sheep nematodes in New Zealand (McKenna 2008 McKenna, PB. 2008. An examination of the relative reliability of laboratory case submissions in determining the prevalence of anthelmintic resistance in sheep nematodes in New Zealand and the possible influence of test analysis methodology on such data. New Zealand Veterinary Journal, 56: 55–59. [Taylor & Francis Online], [Web of Science ®] , [Google Scholar]). Accordingly, the present study was undertaken to make further use of this material, to try to ascertain what, if any, such changes may have taken place over the last few years.     

A larval development test for the detection of anthelmintic resistance in nematodes of sheep

First stage larvae of a number of species of parasitic nematodes of sheep have been shown to develop to third stage larvae in the presence of a defined medium consisting of Earle's balanced salt solution and yeast extract. A larval development test, based on this culture technique, was used as a screen for detecting the presence of anthelmintic resistance in nematodes of sheep. It was found to be sensitive and simple to use and also appeared capable of detecting resistance to any of the main anthelmintic groups. Available anthelmintic sensitive and resistant strains of Haemonchus contortus and Ostertagia circumcincta showed differences in development when incubated in the presence of either thiabendazole, levamisole and ivermectin. These differences were expressed as the minimum inhibitory concentration required to prevent larval development over the incubation period.     

A microlarval development assay for the detection of anthelmintic resistance in sheep nematodes.

A microlarval development test for the detection of anthelmintic resistance in nematodes is described. Haemonchus contortus, Teladorsagia circumcincta and Trichostrongylus colubriformis eggs were cultured to third stage larvae in the presence of Earle's balanced salt solution, yeast extract and bacteria in a total volume of 150 microliters. Good dose-response data were obtained with thiabendazole, levamisole, pyrantel tartrate and ivermectin allowing the determination of the 50 per cent lethal concentration and of resistance factors when resistant strains were available. The test was found to be accurate, sensitive, easy to carry out and applicable to the routine detection of resistance.     

The prevalence of anthelmintic resistance in sheep nematodes on the central tablelands of New South Wales

A survey was undertaken to assess the prevalence of anthelmintic resistance in sheep nematode populations on 40 commercial farms distributed throughout the central tablelands of New South Wales. Representatives of the 2 major groups of broad spectrum anthelmintics with different modes of action (thiabendazole and levamisole) were used at the manufacturer's recommended dose rates. Efficacy was assessed on the basis of the reduction in faecal strongyle egg counts 7 days after treatment. An efficacy of less than 90% using both anthelmintics was obtained on 4 farms. Thiabendazole had an efficacy of less than 90% on a further 21 farms and levamisole had an efficacy of less than 90% on an additional 4 farms. There was no evidence of anthelmintic resistance on 8 farms, while the remaining 3 had insignificant parasite burdens. Based on larval cultures from faeces, Ostertagia and Trichostrongylus were the most significant species in resistant populations. Haemonchus burdens were sporadic and levels of resistance relatively low. Nematodirus burdens were widespread but no evidence of resistance was detected.     

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.     

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.     

The anthelmintic efficacy of netobimin against naturally acquired gastrointestinal nematodes in sheep

The broad-spectrum anthelmintic efficacy of netobimin (SCH 32481, Schering Corporation) was evaluated using 30 cross-bred spring lambs with naturally acquired infections of gastrointestinal nematodes. Three groups of 10 animals each were allotted into either control (given a tap water drench as a placebo) or 7.5 and 20 mg kg-1 dosage groups (given the netobimin as an oral drench). Seven to fourteen days post-treatment, animals were necropsied and nematodes recovered by standard techniques. Examination of fecal samples taken on dates of necropsy showed median egg production was reduced in treated animals (61.98% with 7.5 mg kg-1 and 100% with 20 mg kg-1). The compound was highly effective in removal of adult nematodes representing a number of genera and species of trichostrongyloids at the 7.5 and 20 mg kg-1 dose levels (shown, respectively, below). These included Ostertagia spp., with O. circumcincta, O. trifurcata, O. ostertagi and Teladorsagia davtiani (96.20%; 100%), Trichostrongylus spp., with T. axei, T. vitrinus and T. colubriformis (100%; 98.72%), Nematodirus spp., with N. spathiger, N. filicollis and N. battus (100% both levels) and Haemonchus contortus (100% both levels). High efficacies against other species of nematodes (at both dose levels) were not statistically significant (Cooperia spp., Chabertia ovina and Oesophagostomum venulosum). At 20 mg kg-1, netobimin significantly reduced populations of early and late fourth stage larvae of Ostertagia spp. by 100%. The overall efficacy (all life stages included) was 90.16% at 7.5 mg kg-1 and 98.77% at 20 mg kg-1 dose levels. No adverse reactions or signs of toxicosis were observed.     

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.     

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.     

The rapid appearance of anthelmintic resistance on a sheep farm

A sudden outbreak of perinatal mortality in piglets occurred between May and August 1991 in a 60-sow piggery near Palmerston North. None of the piglets born to the 30 sows which farrowed during this period of time survived more than a few days and all live-born piglets appeared weak. Three newborn live piglets were humanely killed and at necropsy had marked diffuse subcutaneous oedema, the only gross and histological anomalies detected in the extensive range of tissues examined. Serological tests of sows were negative for leptospiral, porcine herpes viral (Aujeszky's), porcine parvoviral, encephalomyocarditis viral and Brucelh suis infections. No viral agents were recovered on cell culture from pooled piglet tissues. Liver selenium levels were normal, but Vitamin E values (0.8, 0.8 and 0.6 μmol/kg respectively) seemed low according to the recently established range of 0.9-1.4 μmol/kg for normal unsuckled piglets in New Zealand⁽¹⁾. No Vitamin E/selenium deficiency syndromes had been recognised previously in this herd.     

The role of combination anthelmintic formulations in the sustainable control of sheep nematodes

Combinations of anthelmintics with a similar spectrum of activity and different mechanisms of action and resistance are widely available in several regions of the world for the control of sheep nematodes. There are two main justifications for the use of such combinations: (1) to enable the effective control of nematodes in the presence of single or multiple drug resistance, and (2) to slow the development of resistance to the component anthelmintic classes. Computer model simulations of sheep nematode populations indicate that the ability of combinations to slow the development of resistance is maximised if certain prerequisite criteria are met, the most important of which appear to concern the opportunity for survival of susceptible nematodes in refugia and the pre-existing levels of resistance to each of the anthelmintics in the combination. Combinations slow the development of a resistant parasite population by reducing the number of resistant genotypes which survive treatment, because multiple alleles conferring resistance to all the component anthelmintic classes must be present in the same parasite for survival. Individuals carrying multiple resistance alleles are rarer than those carrying single resistance alleles. This enhanced efficacy leads to greater dilution of resistant genotypes by the unselected parasites in refugia, thus reducing the proportion of resistant parasites available to reproduce with other resistant adults that have survived treatment. Concerns over the use of anthelmintic combinations include the potential to select for resistance to multiple anthelmintic classes concurrently if there are insufficient parasites in refugia; the potential for shared mechanisms of resistance between chemical classes; and the pre-existing frequency of resistance alleles may be too high on some farms to warrant the introduction of certain combinations. In conclusion, anthelmintic combinations can play an important role in resistance management. However, they are not a panacea and should always be used in accordance with contemporary principles for sustainable anthelmintic use.