This randomised study aimed to assess and compare the efficacy of treatment protocols containing levamisole, ivermectin, or moxidectin against Capillaria spp. in naturally infected European hedgehogs (Erinaceus europaeus) presented to a British wildlife rehabilitation centre. Faecal analysis, consisting of wet mount and flotation, was performed for 229 hedgehogs weighing ≥200g. Animals testing positive for Capillaria spp. (81%), excluding pregnant females, were randomly allocated a treatment protocol. Initially, hedgehogs (n = 50) received one of six ‘pilot’ protocols, whereas the remaining animals (n = 97) received one of three ‘main’ protocols. Faecal analysis was repeated on day 8 and day 12 after treatment initiation. Efficacy of each treatment was assessed based on Capillaria reduction rate (CRR), weight gain, presence of respiratory clinical signs, and outcome. Pilot protocols containing only moxidectin had a significantly lower CRR (≥28.1%) compared to those with levamisole or ivermectin (≥86.6%), whereas the main protocols containing levamisole had a significantly higher CRR (≥93.0%) compared to those containing only ivermectin (≥69.3%). Clinical parameters did not differ significantly between treatments, but animals with respiratory clinical signs at the end of the trial were significantly more likely to have lower CRR and test positive for Crenosoma striatum. C. striatum often appeared refractory to treatment, and managing these infections requires additional anthelmintic therapy. Based on the formulations and dosages trialled, moxidectin is not recommended for treating capillariosis in European hedgehogs, whereas levamisole given orally for two consecutive days at 25–35 mg/kg is suggested as the treatment of choice.

Levamisole (Lms) is an anthelminthic drug with immunomodulatory activity. Chagas disease (CD) is caused by Trypanosoma cruzi and there is very low access to the drugs available, benznidazole (Bz) and nifurtimox, both far from ideal. In a drug-repurposing strategy to test potential activity as antiparasitic and immunomodulatory agent for CD, Lms was assayed on acute T. cruzi murine infection, alone and in co-administration with Bz. During protocol standardization, 100 and 10 mpk of Bz given for five consecutive days resulted in parasitaemia suppression and 100% animal survival only with the highest dose. Flow cytometry showed that both optimal (100 mpk) and suboptimal (10 mpk) doses of Bz equally decreased the plasma levels of cytokines commonly elevated in this acute infection model. Lms alone (10–0.5 mpk) did not decrease parasitaemia nor mortality rates. Co-administration was investigated using the suboptimal dose of Bz and different doses of Lms. While Bz 10 mpk did not alter parasitaemia, the combo partially reduced it but only slightly promoted animal survival. This effect could be related to Th1-response modulation since interleukin-6 and interferon-γ were higher after treatment with the combo.

Parasites reduce host fitness and consequently impose strong selection pressures on their hosts. It has been hypothesized that parasites are scarcer and their overall effect on hosts is weaker at higher latitudes. Although Antarctic birds have relatively low numbers of parasites, their effect on host fitness has rarely been investigated. The effect of helminth parasitism on growth rate was experimentally studied in chinstrap penguin (Pygoscelis antarctica) nestlings. In a total of 22 two-nestling broods, 1 nestling was treated with anthelminthics (for cestodes and nematodes) while its sibling was left as a control. Increased growth rate was predicted in de-wormed nestlings compared to their siblings. As expected, 15 days after treatment, the experimental nestlings had increased body mass more than their siblings. These results show a non-negligible negative effect of helminth parasites on nestling body condition that would presumably affect future survival and thus fitness, and it has been suggested there is a strong relationship between body mass and mortality in chinstrap penguins.

The cyclooctadepsipeptide PF1022A derived from the fungus, Mycelia sterilia, is characterized by a broad spectrum of activity against different parasitic gastrointestinal nematodes of livestock. In the present work the anthelmintic activity of PF1022A against Heligmosomoides bakeri, a widely used laboratory model was studied. Albendazole, ivermectin and levamisole served as reference. In vitro, PF1022A showed low activity on embryonation but significantly inhibited egg hatch (10 and 100 μg/ml), whereas albendazole (10 and 100 μg/ml) revealed statistically significant inhibitions of both embryonation and egg hatch. PF1022A (1–100 μg/ml) completely inhibited larval movement at most examination points. Comparable significant anthelmintic activity on the larval stages of H. bakeri was observed with levamisole (48–100%), while slightly lower activities were observed with ivermectin (20–92%) and albendazole (0–87%) at 1–100 μg/ml. PF1022A and levamisole significantly inhibited motility and egg release of adult worms, while albendazole and ivermectin failed to demonstrate activity. Significant worm burden reductions were achieved with PF1022A, levamisole and ivermectin in vivo. For example, at 0·125 mg/kg PF1022A a worm burden reduction of 91·8% was observed. The use of drug combinations did not further enhance the in vitro and in vivo activity of PF1022A. In conclusion, further investigations are warranted with PF1022A, as the drug is characterized by significant larvicidal and nematocidal activity in vitro and in vivo.

The Consortium for Anthelmintic Resistance and Susceptibility (CARS) brings together researchers worldwide, with a focus of advancing knowledge of resistance and providing information on detection methods and treatment strategies. Advances in this field suggest mechanisms and features of resistance that are shared among different classes of anthelmintic. Benzimidazole resistance is characterized by specific amino acid substitutions in beta-tubulin. If present, these substitutions increase in frequency upon drug treatment and lead to treatment failure. In the laboratory, sequence substitutions in ion-channels can contribute to macrocyclic lactone resistance, but there is little evidence that they are significant in the field. Changes in gene expression are associated with resistance to several different classes of anthelmintic. Increased P-glycoprotein expression may prevent drug access to its site of action. Decreased expression of ion-channel subunits and the loss of specific receptors may remove the drug target. Tools for the identification and genetic analysis of parasitic nematodes and a new online database will help to coordinate research efforts in this area. Resistance may result from a loss of sensitivity as well as the appearance of resistance. A focus on the presence of anthelmintic susceptibility may be as important as the detection of resistance.

Albion mice (Balb c/nut) experimentally infected with Heligmosomoides polygyrus were treated with levamisole hydrochloride (‘Nemicide’—ICI Pharmaceutical PLC) by oral drenching or subcutaneous injection at 5, 10 and 20 mg kg −1. Faecal egg counts monitored for two three days after dosing and post-mortem worm counts were used to assess the efficacy of these treatments. The lowest dose rate gave poor clearance of adult worms but at 10 and 20 mg kg−1, 91 and 96% reduction in worm burden were achieved. Reduction in post-dosing faecal egg counts were variable. Neither method of administration offered particular advantage in terms of efficacy. Faecal egg count data gave inconsistent differences and at necropsy, worm counts were lower in mice dosed orally but not significantly so.

The inhibition of acetylcholinesterase secretion by male and female Heligmosomoides polygyrus was tested on worms taken from experimentally infected mice and maintained for 3 days in vitro in levamisole. The dose inhibiting 50% of enzyme secretion (ID 50) of male worms was twice the ID 50 for female worms. A similar difference was observed in vivo between the dose of levamisole removing 50% (LD 50) of male and female worms from the mouse. Acetylcholinesterase secretion by worms and ID 50 were tested in vitro at 3-weekly intervals from 3 to 21 weeks post infection (WPI). Acetylcholinesterase secretion was always significantly higher for male than for female worms. A decrease of ID 50, correlated with the age of the worms was observed: from 1·5 to 0·5 μg/ml for males and from 0·7 to 0·1 μg/ml for females. These results were confirmed in vivo by a higher efficacy of the anthelminthic at 21 than at 4 WPI.

Three groups of anthelmintic drugs act directly and selectively on muscle membrane receptors of parasitic nematodes. These groups of anthelmintics are: (1) The Nicotinic Agonists(levamisole, pyrantel, morantel and oxantel) that act on acetylcholine receptors of nematode somatic muscle; (2) The GAB A Agonist, piperazine, that acts on nematode muscle GABA receptors; and (3) The Avermectins that open glutamate gated Cl- channels on nematode pharyngeal muscle. The electrophysiology and pharmacology of muscle and neuromuscular transmission the nematode parasite, Ascaris suum, is outlined and effects of anthelmintics that interfere with transmission described. Resistance to anthelmintics has appeared in some parasitic nematodes but the mechanisms of this resistance remain to be determined

A study to compare effects of mass, targeted and selective chemotherapy with levamisole (Ketrax) as an action for the control of Ascaris lumbricoides was carried out in three communities in rural Oyo State, Nigeria. Selective treatment was applied in one village by treating the most heavily infected 20% of the inhabitants, targeted treatment in the second village involved children aged 2–15 years, while mass treatment was offered to all inhabitants excluding infants under 1 year and pregnant women in the third village. Recommended doses of levamisole were given in the villages, as described above, at 3-monthly intervals during a period of 1 year. Prevalence and intensity (epg) of A. lumbricoides infection were determined immediately before and 3 months after the period of intervention using a modified Kato-Katz technique. In the selective treatment village, no significant differences were found between the pre- and post-treatment egg counts (mean(±S.D.) epg 6776±10791 versus 4259±10909 respectively) of A. lumbricoides in the total population. In the targeted treatment village, significant differences were recorded in pre- and post-treatment egg counts for the total population (9057 ± 15 797 versus 2579±6529) among the children alone (10935±20094 versus 992±3175) and among the untreated adults (7742±9782 versus 4561±8798). In the mass treatment village, significant differences in pre- and post-treatment egg count values were also recorded (11 907±17220 versus 1489±5165). The intensity of Trichuris trichiura and hookworm infections among the villagers before and after intervention were not observed to have changed significantly regardless of selective, targeted or mass treatment.

A study to compare the effects of different frequencies of targeted chemotherapy with levamisole (Ketrax: JAGAL Pharma, Lagos, Zeneca Pharmaceuticals UK) as an action for the control of Ascaris lumbricoides was carried out in 3 communities in rural Oyo State, Nigeria. The targeted group comprised children, aged 5 to 15 years, attending primary school within their village. Treatment was provided within the school with the assistance of the school teachers. Three frequencies of targeted treatment were offered. In one village targeted treatment was provided on 1 occasion in 1 year, in another village at two 6-monthly intervals and in the third village every 4 months. Prevalence and intensity (e.p.g.) of A. lumbricoides infection were determined immediately before and after the period of intervention using a modified Kato–Katz technique. In the villages which received treatment once and at 6-monthly intervals, a reduction in post-treatment intensity of A. lumbricoides was observed in the total population but this failed to attain statistical significance. In contrast, within the village which received 4-monthly targeted chemotherapy, a significant reduction in post-treatment intensity of A. lumbricoides was observed in the total population and in the targeted children. In general, reductions in the intensity of A. lumbricoides after intervention were not particularly pronounced in untreated children (aged 0–4 years) even in the 4-monthly targeted village, whereas in untreated adults, reductions approached statistical significance in villages which received targeted treatment once and at 4-monthly intervals.

Anthelmintic resistance has compromised the control of nematode parasites in several animal-based industries. Studies of resistance have not only improved our understanding of this phenomenon but also shed light on physiological systems of parasitic helminths. In addition, research on molecular aspects of anthelmintic resistance may provide selectable markers for use in future transfection studies with helminths. Several anthelmintics act on helminth neuromuscular systems. Drugs such as levamisole are cholinergic agonists and, based on pharmacological studies, levamisole-resistant nematodes appear to have altered acetylcholine receptors. It is likely that anticholinesterase anthelmintics share cross resistance with levamisole. Ivermectin appears to be a glutamate agonist. In vitro studies of ivermectin-resistant nematodes suggest that IVM receptors are located on pharyngeal and somatic muscle. The free-living nematode Caenorhabditis elegans may provide a model for anthelmintic resistance. It has been useful in cloning drug receptors from parasites but differences between its life history and habitat compared with parasitic nematodes may limit its usefulness for studying resistance in these parasites.

Anthelmintic resistance is a serious problem in veterinary medicine and appears to be developing in some helminths of importance to human health. Anthelmintic drugs remain the principal means of control of helminth infections in animals and humans and the continued dependence on these pharmaceuticals will continue to impose selection pressure for resistance development. Our ability to detect anthelmintic resistance before control breaks down and to monitor the spread of anthelmintic resistance is quite limited. We are currently dependent on biological methods which are not sufficiently sensitive to detect low levels of drug resistance and are particularly difficult to perform on helminth parasites of humans. There is a serious need for new molecular markers for detecting and monitoring for anthelmintic resistance. The problem of anthelmintic resistance is already very serious in nematode parasites of livestock. In addition, there should be great concern about possible anthelmintic resistance development and the lack of tools and efforts for monitoring it as part of the major worldwide programmes to control helminth parasites in people. An international Consortium has been formed to develop Anthelmintic Resistance Single nucleotide polymorphism markers (CARS). Discussions within the Consortium have addressed the need for such markers, the current state of knowledge about them, possible mechanisms of anthelmintic resistance and approaches and constraints to the development of markers. Summaries of the state of the art in these areas are presented in a series of papers in this Special Issue of Parasitology.

Here we review molecular information related to resistance to the cholinergic anthelmintics in nematodes. The amount of molecular information available varies between the nematode species, with the best understood so far being C. elegans. More information is becoming available for some other parasitic species. The cholinergic anthelmintics act on nematode nicotinic acetylcholine receptors located on somatic muscle cells. Recent findings demonstrate the presence of multiple types of the nicotinic receptors in several nematodes and the numerous genes required to form these multimeric proteins. Not only are the receptors the product of several genes but they are subject to modulation by several other proteins. Mutations altering these modulatory proteins could alter sensitivity to the cholinergic anthelmitics and thus lead to resistance. We also discuss the possibility that resistance to the cholinergic anthelmintics is not necessarily the result of a single mutation but may well be polygenic in nature. Additionally, the mutations resulting in resistance may vary between different species or between resistant isolates of the same species. A list of candidate genes to examine for SNPs is presented.

A cDNA-AFLP (cDNA-Amplified Fragment Length Polymorphism)-based strategy has been used to identify levamisole (LEV) resistance markers in the nematode Haemonchus contortus. Transcript profiles of adult nematodes from two LEV-resistant and two susceptible isolates were compared. Among the 17 280 transcript-derived fragments (TDFs) amplified, 26 presented a polymorphic pattern between resistant and susceptible nematodes: 11 TDFs were present in both resistant isolates and absent from both susceptible isolates whereas 15 TDFs were present in both susceptible isolates and absent from both resistant isolates. 8 TDFs specifically present in resistant isolates were cloned and sequenced. Some of these TDFs could represent novel genes, as their sequences presented no homologies in databases. Interestingly, specific expression of one candidate (HA17) in resistant nematodes from different isolates was confirmed by RT-PCR experiments. The finding that HA17 expression correlates with LEV resistance in three H. contortus isolates vs five susceptible isolates strongly suggest that we identified a new potential marker of LEV resistance. This differential approach at the transcriptome level could be of great interest for the identification of the molecular mechanism involved in this phenotype.

The problem of anthelmintic resistance prevents efficient control of parasites of livestock and may soon compromise human parasite control. Research into the mechanisms of resistance and the quest for diagnostic tools to aid control has required research that focuses on field resistance. On the other hand, resistant worms, including those kept in the laboratory, provide useful tools for studying drug action, especially at neuromuscular targets in worms. While the needs and directions of these research aims overlap, this review concentrates on research on drug targets. In this context, resistance is a useful tool for site of action confirmation. For example, correlations between molecular expression studies and resistance assays conducted on whole worms can strengthen claims for sites of anthelmintic action. Model systems such as Caenorhabditis elegans have been very useful in understanding targets but give a limited picture as it is now clear that resistance mechanisms in this worm are different from those in parasites. Accordingly, research on parasites themselves must also be performed. Resistant isolates of the sheep nematode parasite Haemonchus contortus are the most widely used for this purpose as in vivo, in vitro, physiological and molecular studies can be performed with this species. Neuromuscular target sites for the anthelmintics levamisole and ivermectin are the best studied and have benefited most from the use of resistant worm isolates. Resistance to praziquantel and the newer chemical groups should provide new tools to explore targets in the future.

Here we review recent studies on the mode of action of the cholinergic anthelmintics (levamisole, pyrantel etc.). We also include material from studies on the free living nematode Caenorhabditis elegans. The initial notion that these drugs act on a single receptor population, while attractive, has proven to be an oversimplification. In both free living and parasitic nematodes there are multiple types of nicotinic acetylcholine receptor (nAChR) on the somatic musculature. Each type has different (sometimes subtly so) pharmacological properties. The implications of these findings are: (1) combinations of anthelmintic that preferentially activate a broad range of nAChR types would be predicted to be more effective; (2) in resistant isolates of parasite where a subtype has been lost, other cholinergic anthelmintics may remain effective. Not only are there multiple types of nAChR, but relatively recent research has shown these receptors can be modulated; it is possible to increase the response of a parasite to a fixed concentration of drug by altering the receptor properties (e.g. phosphorylation state). These findings offer a potential means of increasing efficacy of existing compounds as an alternative to the costly and time consuming development of new anthelmintic agents.

This paper reviews the immunomodulatory effects (immunosuppression or immunoactivation) of various anthelmintics including levamisole, fenvalerate, dieldrin, carbofuran, aminocarb, thiabendazole, fenbendazole, oxfendazole and ivermectin. The induced modulation of immune function may occur via direct and/or indirect mechanisms. The immunomodulatory effects of these anti-parasitics have been studied in a variety of bacterial (e.g. brucellosis, salmonellosis, paratuberculosis, mastitis), viral (e.g. infectious bovine rhinotracheitis, Herpes, foot and mouth disease), parasitic (e.g. onchocerciasis, coccidiosis, ascariasis, schistosomiasis) and neoplastic diseases. Some antiparasitics have also been used to boost immunity in a number of human diseases including leprosy, Hodgkin's disease, rheumatoid arthritis, and in adjuvanted therapy of colorectal cancer. The ability to stimulate the immune response of animals offers a new means of disease intervention. Future research on immunomodulatory effects of anti-parasitics, for humans and domestic farm animals, will provide additional methods of treating immunosuppressed subjects. The immunopotentiating or immunosuppressing activity of anti-parasitics will dictate whether co-administration of vaccines and anthelmintics or administration of vaccines during the window of immunoactivation is justified or not.

It is pointed out that two of the three major groups of anthelmintic act by opening membrane ion-channels. It is appropriate, therefore, to use electrophysiological methods to study the properties of the sites of action of these drugs and the changes in the properties of these receptor sites associated with resistance. This paper describes the use of the patch- clamp technique to observe the currents that flow through the levamisole-activated channels as they open and close in levamisole-sensitive and levamisole-resistant isolates. It was found that, on average, the proportion of time the channels are open, is less in the resistant isolate. The patch-clamp technique also showed that the ion-channels are heterogeneous and that one of the subtypes is lost with the appearance of resistance. The use of the current clamp technique is illustrated to record a site of action of ivermectin in the pharyngeal muscle of Ascaris.

This paper reviews sites of action of anthelmintic drugs including: (1) levamisole and pyrantel, which act as agonists at nicotinic acetylcholine receptors of nematodes; (2) the avermectins, which potentiate or gate the opening of glutamate-gated chloride channels found only in invertebrates; (3) piperazine, which acts as an agonist at GABA gated chloride channels on nematode muscle; (4) praziquantel, which increases the permeability of trematode tegument to calcium and results in contraction of the parasite muscle; (5) the benzimidazoles, like thiabendazole, which bind selectively to parasite β-tubulin and prevents microtubule formation; (6) the proton ionophores, like closantel, which uncouple oxidative phosphorylation; (7) diamphenethide and clorsulon, which selectively inhibit glucose metabolism of Fasciola and; (8) diethylcarbamazine, which appears to interfere with arachidonic acid metabolism of filarial parasites and host. The review concludes with brief comments on the development of anthelmintics in the future.