The pharmacology of anthelmintics in livestock

The recent development of ivermectin, netobimin, closantel, triclabendazole and clorsulon as well as new ways of using established anthelmintics has significantly improved the potential for parasite control in livestock. It is of interest to note that potential control of F. hepatica has been particularly improved with four of these new anthelmintic drugs being fasciolicidal. In addition to an appreciation of the spectrum of action of the new anthelmintics, a considerable amount of information on the pharmaco-kinetic behaviour, metabolism and mode of action of these drugs has been published. New information on pharmacology of established anthelmintics has provided new insights into the biology of parasites, drug resistance and the options for improved chemotherapy by manipulation of pharmacokinetics and metabolism. Much recent pharmacological research has been concentrated on the BZs and this has been considered here in an attempt to provide a better understanding of this significant group of anthelmintics. The development of new delivery systems, such as intraruminal slow release devices, is potentially useful with most anthelmintics. So far, this new approach to parasite control has been applied most successfully with morantel tartrate.     

The biochemical basis of anthelmintic action and resistance

The most commonly used modern anthelmintics include the benzimidazoles, the nicotinic agonists. praziquantel, triclabendazole and the macrocyclic lactones. These drugs interfere with target sites that are either unique to the parasite or differ in their structural features from those of the homologous counterpart present in the vertebrate host. The benzimidazoles exert their effect by binding selectively and with high affinity to the beta-subunit of helminth microtubule protein. The target site of the nicotinic agonists (e.g. levamisole, tetrahydropyrimidines) is a pharmacologically distinct nicotinic acetylcholine receptor channel in nematodes. The macrocyclic lactones (e.g. ivermectin, moxidectin) act as agonists of a family of invertebrate-specific inhibitory chloride channels that are activated by glutamic acid. The primary mode of action of other important anthelmintics (e.g. praziquantel, triclabendazole) is unknown. Anthelmintic resistance is wide-spread and a serious threat to effective control of helminth infections, especially in the veterinary area. The biochemical and genetic mechanisms underlying anthelmintic resistance are not well understood, but appear to be complex and vary among different helminth species and even isolates. The major mechanisms helminths use to acquire drug resistance appear to be through receptor loss or decrease of the target site affinity for the drug. Knowledge on the mechanisms of drug action and resistance may be exploitable for the development of new drugs and may provide information on ways to overcome parasite resistance, respectively.     

Unresolved issues in anthelmintic pharmacology for helminthiases of humans

Helminth infections are an important constraint on the health and development of poor children and adults. Anthelmintic treatment programmes provide a safe and effective response, and increasing numbers of people are benefitting from these public health initiatives. Despite decades of clinical experience with anthelmintics for the treatment of human infections, relatively little is known about their clinical pharmacology. All of the drugs were developed initially in response to the considerable market for veterinary anthelmintics in high- and middle-income countries. In contrast, the greatest burden caused by these infections in humans is in resource-poor settings and as a result there has been insufficient commercial incentive to support studies on how these drugs work in humans, and how they should best be used in control programmes. The advent of mass drug administration programmes for the control of schistosomiasis, lymphatic filariasis, onchocerciasis and soil-transmitted helminthiases in humans increases the urgency to better understand and better monitor drug resistance, and to broaden the currently very narrow range of available anthelmintics. This provides fresh impetus for developing a comprehensive research platform designed to improve our understanding of these important drugs, in order to bring the scientific knowledge base supporting their use to a standard equivalent to that of drugs commonly used in developed countries. Furthermore, a better understanding of their clinical pharmacology will enable improved therapy and could contribute to the discovery of new products.     

Importance of microbial natural products and the need to revitalize their discovery

Microbes are the leading producers of useful natural products. Natural products from microbes and plants make excellent drugs. Significant portions of the microbial genomes are devoted to production of these useful secondary metabolites. A single microbe can make a number of secondary metabolites, as high as 50 compounds. The most useful products include antibiotics, anticancer agents, immunosuppressants, but products for many other applications, e.g., antivirals, anthelmintics, enzyme inhibitors, nutraceuticals, polymers, surfactants, bioherbicides, and vaccines have been commercialized. Unfortunately, due to the decrease in natural product discovery efforts, drug discovery has decreased in the past 20 years. The reasons include excessive costs for clinical trials, too short a window before the products become generics, difficulty in discovery of antibiotics against resistant organisms, and short treatment times by patients for products such as antibiotics. Despite these difficulties, technology to discover new drugs has advanced, e.g., combinatorial chemistry of natural product scaffolds, discoveries in biodiversity, genome mining, and systems biology. Of great help would be government extension of the time before products become generic.     

Anthelmintic resistance: the state of play revisited

Helminthosis is one of the major constraints in the successful wool and mutton industry throughout the world. Anthelmintic Resistance (AR) is said to have been established when previously effective drug ceases to kill exposed parasitic population at the therapeutically recommended dosages. Anthelmintic resistance is almost cosmopolitan in distribution and it has been reported in almost all species of domestic animals and even in some parasites of human beings. Some of the most important species of parasites of small ruminants in which AR has been reported include: Haemonchus spp., Trichostrongylus spp. Teladorsagia spp., Cooperia spp. Nematodirus spp., and Oesophagostomum spp. All the major groups of anthelmintics have been reported for development of variable degrees of resistance in different species of gastrointestinal nematodes. This paper describes the global scenario of prevalence and methods used for detection of AR in small ruminants. Different mechanisms and contributory factors for the development of AR are discussed. Various options and alternate strategies for the control and/or delay in the onset of AR are suggested in the light of available information.     

The sensory amphidial structures of Caenorhabditis elegans are involved in macrocyclic lactone uptake and anthelmintic resistance

Parasitic nematodes represent formidable pathogens of humans, livestock and crop plants. Control of these parasites is almost exclusively dependent on a small group of anthelmintic drugs, the most important of which belong to the macrocyclic lactone class. The extensive use of these drugs to control the ubiquitous trichostrongylid parasites of grazing livestock has resulted in the emergence of both single and multi-drug resistance. The expectation is that this resistance will eventually occur in the human parasites such as the common and debilitating soil transmitted nematodes and vector-borne filarial nematodes. While the modes of action of anthelmintics such as ivermectin, have been elucidated, notably in the model nematode Caenorhabditis elegans, the molecular nature of this resistance remains to be fully determined. Here we show that the anterior amphids play a key role in ivermectin uptake and mutations in these sensory structures result in ivermectin resistance in C. elegans. Random genetic mutant screens, detailed analysis of existing amphid mutants and lipophilic dye uptake indicate that the non-motile ciliated amphid neurons are a major route of ivermectin ingress; the majority of the mutants characterised in this study are predicted to be involved in intraflagellar transport. In addition to a role in ivermectin resistance, a subset of the amphid mutants are resistant to the non-related benzimidazole class of anthelmintics, raising the potential link to a multi-drug resistance mechanism. The amphid structures are present in all nematodes and are clearly defined in a drug-sensitive strain of Haemonchus contortus. It is predicted that amphidial drug uptake and intraflagellar transport may prove to be significant in the development of single and multi-drug resistance in the nematode pathogens of veterinary and human importance.     

A research agenda for helminth diseases of humans: intervention for control and elimination

Recognising the burden helminth infections impose on human populations, and particularly the poor, major intervention programmes have been launched to control onchocerciasis, lymphatic filariasis, soil-transmitted helminthiases, schistosomiasis, and cysticercosis. The Disease Reference Group on Helminth Infections (DRG4), established in 2009 by the Special Programme for Research and Training in Tropical Diseases (TDR), was given the mandate to review helminthiases research and identify research priorities and gaps. A summary of current helminth control initiatives is presented and available tools are described. Most of these programmes are highly dependent on mass drug administration (MDA) of anthelmintic drugs (donated or available at low cost) and require annual or biannual treatment of large numbers of at-risk populations, over prolonged periods of time. The continuation of prolonged MDA with a limited number of anthelmintics greatly increases the probability that drug resistance will develop, which would raise serious problems for continuation of control and the achievement of elimination. Most initiatives have focussed on a single type of helminth infection, but recognition of co-endemicity and polyparasitism is leading to more integration of control. An understanding of the implications of control integration for implementation, treatment coverage, combination of pharmaceuticals, and monitoring is needed. To achieve the goals of morbidity reduction or elimination of infection, novel tools need to be developed, including more efficacious drugs, vaccines, and/or antivectorial agents, new diagnostics for infection and assessment of drug efficacy, and markers for possible anthelmintic resistance. In addition, there is a need for the development of new formulations of some existing anthelmintics (e.g., paediatric formulations). To achieve ultimate elimination of helminth parasites, treatments for the above mentioned helminthiases, and for taeniasis and food-borne trematodiases, will need to be integrated with monitoring, education, sanitation, access to health services, and where appropriate, vector control or reduction of the parasite reservoir in alternative hosts. Based on an analysis of current knowledge gaps and identification of priorities, a research and development agenda for intervention tools considered necessary for control and elimination of human helminthiases is presented, and the challenges to be confronted are discussed.     

Efficacy evaluation of anthelmintics: which methods to use in the field?

Efficacy evaluation of anthelmintics is very important even in veterinary practice due to the existence of acquired resistance, poor quality of anthelmintics in several region of the world, and nedds for quarantine strategy prior introduction of new animals. Clearly, flock evaluation can be opposed to individually based evaluations. I propose that individually based evaluations should be preferred in field condition controls: although they are more sophisticated on a statistical basis, the availability of a software for calculations renders the proposal realistic.     

The New Anthelmintic Tribendimidine is an L-type (Levamisole and Pyrantel) Nicotinic Acetylcholine Receptor Agonist

Background

Intestinal parasitic nematodes such as hookworms, Ascaris lumbricoides, and Trichuris trichiura are amongst most prevalent tropical parasites in the world today. Although these parasites cause a tremendous disease burden, we have very few anthelmintic drugs with which to treat them. In the past three decades only one new anthelmintic, tribendimidine, has been developed and taken into human clinical trials. Studies show that tribendimidine is safe and has good clinical activity against Ascaris and hookworms. However, little is known about its mechanism of action and potential resistance pathway(s). Such information is important for preventing, detecting, and managing resistance, for safety considerations, and for knowing how to combine tribendimidine with other anthelmintics.

Methodology/Principal Findings

To investigate how tribendimidine works and how resistance to it might develop, we turned to the genetically tractable nematode, Caenorhabditis elegans. When exposed to tribendimidine, C. elegans hermaphrodites undergo a near immediate loss of motility; longer exposure results in extensive body damage, developmental arrest, reductions in fecundity, and/or death. We performed a forward genetic screen for tribendimidine-resistant mutants and obtained ten resistant alleles that fall into four complementation groups. Intoxication assays, complementation tests, genetic mapping experiments, and sequencing of nucleic acids indicate tribendimidine-resistant mutants are resistant also to levamisole and pyrantel and alter the same genes that mutate to levamisole resistance. Furthermore, we demonstrate that eleven C. elegans mutants isolated based on their ability to resist levamisole are also resistant to tribendimidine.

Conclusions/Significance

Our results demonstrate that the mechanism of action of tribendimidine against nematodes is the same as levamisole and pyrantel, namely, tribendimidine is an L-subtype nAChR agonist. Thus, tribendimidine may not be a viable anthelmintic where resistance to levamisole or pyrantel already exists but could productively be used where resistance to benzimidazoles exists or could be combined with this class of anthelmintics.     

The cholinomimetic morantel as an open channel blocker of the <Emphasis Type="Italic">Ascaris suum</Emphasis> ACR-16 nAChR

Nematode parasite infections pose a significant threat in human and veterinary medicine. At least a third of the world’s population is at risk from nematode parasite infections. These infections not only cause health problems, but also cause loss of livestock production and hence, economic losses. Anthelmintic drugs are the mainstay by which control of nematode parasite infections is achieved. Many of the currently available anthelmintics act on nicotinic acetylcholine receptors (nAChRs). However, the detailed mode of action (MOA) of these anthelmintics is not clearly understood. Elucidation of the MOA of anthelmintics is highly desirable; an in-depth knowledge of the MOA will better inform on mechanisms of resistance development and on ways to slow down or overcome resistance. The cholinomimetic anthelmintic, morantel, has a complex MOA involving the activation and block of levamisole-sensitive single nAChR channels (L-type nAChR or L-nAChR). More recently, morantel has been demonstrated to activate Haemonchus contortus and Parascaris equorum ACR-26/ACR-27 nAChRs expressed in Xenopus laevis oocytes. Previous studies in our laboratory, however, have shown morantel does not activate the nicotine-sensitive nAChR (N-type nAChR or N-nAChR), Ascaris suum ACR-16 (Asu-ACR-16). In this study, we used two-electrode voltage-clamp (TEVC) electrophysiology to investigate the inhibitory effects of morantel, on expressed Asu-ACR-16 nAChRs in X. laevis oocytes. Our results show that morantel acts as a non-competitive antagonist on Asu-ACR-16. This non-competitive antagonism by morantel was further demonstrated to be voltage-sensitive. We conclude based on our findings that morantel is a non-competitive voltage-sensitive open channel blocker of Asu-ACR-16.     

Integrated control of nematode infections in cattle: a reality? A need? A future?

Helminth infections are a major cause of production loss in cattle. Great progress has been achieved in the design of control strategies for these infections. Control is based mainly on the use of anthelmintics, and these have become more potent and easier to administer. However, the most effective control is possible only through the integration of different approaches. Moreover, an increasing number of disadvantages of chemotherapy/prophylaxis--biological, economical and environmental--have been suggested. In sheep, the high incidence of anthelmintic resistance has simply forced veterinarians/producers to adopt alternative control strategies; in cattle, no real need for deviation from the actual control programmes seems to exist. Therefore, the following questions are discussed: (1) Based on the distribution of cattle worldwide, what are the target parasites? (2) Can we continue to rely on control based mainly on the use of (highly effective) anthelmintics? (3) What are the prospects for non-chemical control? (4) Who will develop and implement integrated control systems? (5) In the case of parasite control in Western Europe, has it been efficient and can/need it be changed? (6) How can we integrate helminth control in the general design of herd disease control?     

Anthelmintic resistance in nematodes: extent, recent understanding and future directions for control and research

Resistance has now been reported to all of the broad spectrum anthelmintic types currently available, namely to the benzimidazoles, levamisole/morantel and to ivermectin. The problem causes most concern for parasite control in sheep, but anthelmintic resistance has also been reported in nematodes of horses, goats, pigs and more recently cattle. Our understanding of the factors which select rapidly for resistance has increased and programmes of worm control which minimize selection for anthelmintic resistance are being developed and tested. One of the greatest problems encountered in attempting to reduce the selection for overt drug resistance is the need for more sensitive tests for developing resistance. In the long term, new approaches to chemotherapy and to overcoming anthelmintic resistance problems will arise from improving our understanding of the modes of action of, and mechanisms of resistance to, anthelmintics at the level of the receptor proteins and their genes.     

Anthelmintic resistance in human helminths: Learning from the problems with worm control in livestock

During the past decade, the prevalence of anthelmintic resistance in some economically important helminths of sheep, goats and horses has increased dramatically. In some regions of Australia, South America and South Africa, anthelmintic resistance has become a serious threat to the survival of the sheep industry. Mass treatment programmes and exclusive reliance on anthelmintics for worm control in livestock are amongst the most important reasons for the development of anthelmintic resistance. In this article, Stanny Geerts, Gerald Coles and Bruno Gryseels draw the attention to a number of errors that have occurred in the control of helminths in livestock and that should be avoided in the control of worms in humans.     

The beta-tubulin genes of two Strongyloides species

The World Health Organization is sponsoring major treatment programs with the aim of controlling helminth infection throughout the tropical world. Prominent among the anthelmintics recommended for use in these programs are drugs in the benzimidazole (BZ) class. Resistance to these drugs has been associated with polymorphisms in the beta-tubulin gene. We have cloned and sequenced the beta-tubulin genes of Strongyloides stercoralis and Strongyloides ratti and have proceeded to develop a protocol for genotyping single worms for polymorphisms in beta-tubulin. Our findings indicate that S. ratti has a single beta-tubulin gene, making DNA sequence analysis of a single larva PCR product a feasible means of studying BZ resistance in these species. Our genotyping test allows the identification of polymorphisms at codons 167, 198, and 200 in the Strongyloides beta-tubulin gene, thus enabling survey for BZ resistant genotypes.     

Recent findings on the genetics of gastro-intestinal nematode resistance in ruminants

The control of helminthiases in ruminants raised in open pasture has been mainly undertaken by using prophylactic measures in the environment, but these are often inadequate due to incorrect application. With the appearance of anthelmintics, the strategy for controlling these parasitoses, passed to pharmacological treatments which became effective in reducing their impact. However, the frequent and incorrect utilisation of these molecules resulted in resistance to anthelmintics and the presence of chemical residues in animal products for human consumption. Anthelmintic resistance is widespread throughout the world, heterogeneous and probably underestimated. This has encouraged the introduction of homeopathic agents and products derived from plants whose effectiveness has not been scientifically assessed. It is well known that it is possible to detect differences in resistance to the most important parasites between breeds. In Europe, it has been reported that some ovine autochthonous breeds, Scottish Blackface and Lacaune, showed higher resistance. The implementation of breeding strategies aimed at obtaining animals with naturally low susceptibility to nematode infestations could therefore play an increasingly important role. Standard animal breeding techniques have been largely successful in improving the performance of domestic animals in the last century. Standard quantitative selection requires field data on: i) individual phenotype performance; ii) expected covariance among animals due to blood relationship between them. The whole process of predicting the breeding value of animals in order to select subsequently the genetically superior parents of the next generation is entirely based on sophisticated computations (BLUP-animal model). In sheep, the main objective is always selecting for milk yield and sometimes, in addition, milk composition. However, due to the evolution of the EU agricultural policy and consumer demand in terms of healthy and organic food, more attention is now being given to traits related to health (resistance to EST, mastitis or parasitic diseases). Some studies conducted in New Zealand and Australia showed that nematode resistance is genetically controlled with high heritabilities and quite low genetic correlations with production traits. In this sense, some studies showed that it is possible to decrease the number of parasites in the framework of a traditional breeding programme. However, in most situations, this trait is not extensively recorded due to the high cost of individual recording. Therefore, it would be useful to implement breeding strategies based on the knowledge of the genes involved in this trait expression. Traditionally, two approaches are available to locate a gene: i) genome scan; ii) candidate gene approach. The candidate gene approach attempts to link general resistance to some particular genes. To date, genetic resistance against parasites is considered to be linked with the MHC and IgE genes. Furthermore, several gene detection studies based on the genome scan approach for this trait are currently being carried out on both crossed experimental populations (fat x lean Blackface lines and Sarda x Lacaune) and pure breeds (Churra). The preliminary results seem promising as to the use of marker assisted or genotype assisted selection for this trait, which is difficult and expensive to measure on a population scale.     

Influence of host nutrition on the development and consequences of nematode parasitism in ruminants.

Control of gastrointestinal nematodes of ruminants is based largely on use of anthelmintics combined, where practical, with pasture management. The increasing prevalence of resistance to anthelmintics has led to the search for alternative sustainable control strategies. Here, we consider how nutrition, as a short-term alternative, can influence the host--parasite relationship in ruminants, using gastrointestinal nematode infections of sheep as the model system. Nutrition can affect the ability of the host to cope with the consequences of parasitism and to contain and eventually to overcome parasitism. It can also affect the parasite population through the intake of antiparasitic compounds.     

The nervous systems of helminths as targets for drugs.

Processes that critically differentiate parasitic helminths and their hosts are obvious candidates for chemotherapeutic intervention. The recognition that neurobiology distinguishes helminths from their vertebrate hosts is due in part to the fact that several efficacious anthelmintics, derived generally from empirical screening, have been found to act selectively on the neuromuscular system of these parasites. In addition, basic physiological and pharmacological research has revealed considerable differences in the ways in which helminths and their hosts transmit information in the nervous system and respond to it in innervated tissues. Unfortunately, most of these differences have yet to be exploited in chemotherapy. The topics for this review include an analysis of mechanistic aspects of the pharmacology of anthelmintics that act on neuromuscular systems and a consideration of the prospects for discovery of novel drugs that act on this system.     

Selection studies on anthelmintic resistant and susceptible populations of Trichostrongylus colubriformis of sheep

Waller P. J., Dobson R. J., Donald A. D., Griffiths D. A. and Smith E.F. 1985. Selection studies on anthelmintic resistant and susceptible populations of Trichostrongylus colubriformis of sheep. International Journal for Parasitology15: 669–676. A T. colubriformis population (BCK), formerly resistant to benzimidazole anthelmintics, but now highly resistant to levamisole after 6 years exposure to this drug alone in the field, was passed through 12 generations in the laboratory in three separate lines exposed either to selection with thiabendazole or levamisole, or to no selection. Another population (McM) not previously exposed to these anthelmintics was treated similarly in two lines, selected with thiabendazole or not selected.Selection with thiabendazole resulted in a return of benzimidazole resistance in the BCK line which occurred faster than in the McM line, but a similar level of resistance was reached in each by the twelfth generation. Resistance ratios in both selected lines compared with the unselected McM line were less than 20: 1, and only 1.5 times the recommended dose rate of thiabendazole was required to remove more than half of the resistant population. This suggests that a polygenic vigour tolerance rather than a specific resistance had been selected.In the case of levamisole resistance, the BCK population was found to contain two distinct subpopulations, one susceptible and the other highly resistant. Resistance ratios for the highly resistant subpopulation were greater than 4000: 1, implying a specific resistance controlled by a major gene. During the 12 generations of levamisole selection, the proportion of resistant phenotypes fluctuated about an average level of 70%, suggesting that susceptibility alleles were being maintained in the population through superior heterozygote fitness. This conclusion is supported by a significant decline in levamisole resistance in the absence of levamisole selection. Moreover, thiabendazole selection hastened the reversion to levamisole suceptibility.The results provide support for the reintroduction of a benzimidazole anthelmintic to control this helminth population, and for a slow rotation in the use of drugs with different modes of action.     

Emodepside and SL0-1 potassium channels: a review

Nematode parasites infect humans and domestic animals; treatment and prophylaxis require anthelmintic drugs because vaccination and sanitation is limited. Emodepside is a more recently introduced cyclooctadepsipeptide drug that has actions against GI nematodes, lungworm, and microfilaria. It has a novel mode of action which breaks resistance to the classical anthelmintics (benzimidazoles, macrocyclic lactones and cholinergic agonists). Here we review studies on its mode of action which suggest that it acts to inhibit neuronal and muscle activity of nematodes by increasing the opening of calcium-activated potassium (SLO-1) channels.