Plant NB-LRR proteins confer robust protection against microbes and metazoan parasites by recognizing pathogen-derived avirulence (Avr) proteins that are delivered to the host cytoplasm. Microbial Avr proteins usually function as virulence factors in compatible interactions; however, little is known about the types of metazoan proteins recognized by NB-LRR proteins and their relationship with virulence. In this report, we demonstrate that the secreted protein RBP-1 from the potato cyst nematode Globodera pallida elicits defense responses, including cell death typical of a hypersensitive response (HR), through the NB-LRR protein Gpa2. Gp-Rbp-1 variants from G. pallida populations both virulent and avirulent to Gpa2 demonstrated a high degree of polymorphism, with positive selection detected at numerous sites. All Gp-RBP-1 protein variants from an avirulent population were recognized by Gpa2, whereas virulent populations possessed Gp-RBP-1 protein variants both recognized and non-recognized by Gpa2. Recognition of Gp-RBP-1 by Gpa2 correlated to a single amino acid polymorphism at position 187 in the Gp-RBP-1 SPRY domain. Gp-RBP-1 expressed from Potato virus X elicited Gpa2-mediated defenses that required Ran GTPase-activating protein 2 (RanGAP2), a protein known to interact with the Gpa2 N terminus. Tethering RanGAP2 and Gp-RBP-1 variants via fusion proteins resulted in an enhancement of Gpa2-mediated responses. However, activation of Gpa2 was still dependent on the recognition specificity conferred by amino acid 187 and the Gpa2 LRR domain. These results suggest a two-tiered process wherein RanGAP2 mediates an initial interaction with pathogen-delivered Gp-RBP-1 proteins but where the Gpa2 LRR determines which of these interactions will be productive.

Gastrointestinal strongyles are a major threat to horses' health and welfare. Given that strongyles inhabit the same niche as the gut microbiota, they may interact with each other. These beneficial or detrimental interactions are unknown in horses and could partly explain contrasted susceptibility to infection between individuals. To address these questions, an experimental pasture trial with 20 worm-free female Welsh ponies (10 susceptible (S) and 10 resistant (R) to parasite infection) was implemented for 5 months. Fecal egg counts (FEC), hematological and biochemical data, body weight and gut microbiological composition were studied in each individual after 0, 24, 43, 92 and 132 grazing days. R and S ponies displayed divergent immunological profiles and slight differences in microbiological composition under worm-free conditions. After exposure to natural infection, the predicted R ponies exhibited lower FEC after 92 and 132 grazing days, and maintained higher levels of circulating monocytes and eosinophils, while lymphocytosis persisted in S ponies. Although the overall gut microbiota diversity and structure remained similar during the parasite infection between the two groups, S ponies exhibited a reduction of bacteria such as Ruminococcus, Clostridium XIVa and members of the Lachnospiraceae family, which may have promoted a disruption of mucosal homeostasis at day 92. In line with this hypothesis, an increase in pathobionts such as Pseudomonas and Campylobacter together with changes in several predicted immunological pathways, including pathogen sensing, lipid metabolism, and activation of signal transduction that are critical for the regulation of immune system and energy homeostasis were observed in S relative to R ponies. Moreover, S ponies displayed an increase in protozoan concentrations at day 92, suggesting that strongyles and protozoa may contribute to each other's success in the equine intestines. It could also be that S individuals favor the increase of these carbohydrate-degrading microorganisms to enhance the supply of nutrients needed to fight strongyle infection. Overall, this study provides a foundation to better understand the mechanisms that underpin the relationship between equines and natural strongyle infection. The profiling of horse immune response and gut microbiota should contribute to the development of novel biomarkers for strongyle infection.

Cholinergic agonists such as levamisole and pyrantel are widely used as anthelmintics to treat parasitic nematode infestations. These drugs elicit spastic paralysis by activating acetylcholine receptors (AChRs) expressed in nematode body wall muscles. In the model nematode Caenorhabditis elegans, genetic screens led to the identification of five genes encoding levamisole-sensitive-AChR (L-AChR) subunits: unc-38, unc-63, unc-29, lev-1 and lev-8. These subunits form a functional L-AChR when heterologously expressed in Xenopus laevis oocytes. Here we show that the majority of parasitic species that are sensitive to levamisole lack a gene orthologous to C. elegans lev-8. This raises important questions concerning the properties of the native receptor that constitutes the target for cholinergic anthelmintics. We demonstrate that the closely related ACR-8 subunit from phylogenetically distant animal and plant parasitic nematode species functionally substitutes for LEV-8 in the C. elegans L-AChR when expressed in Xenopus oocytes. The importance of ACR-8 in parasitic nematode sensitivity to cholinergic anthelmintics is reinforced by a ‘model hopping’ approach in which we demonstrate the ability of ACR-8 from the hematophagous parasitic nematode Haemonchus contortus to fully restore levamisole sensitivity, and to confer high sensitivity to pyrantel, when expressed in the body wall muscle of C. elegans lev-8 null mutants. The critical role of acr-8 to in vivo drug sensitivity is substantiated by the successful demonstration of RNAi gene silencing for Hco-acr-8 which reduced the sensitivity of H. contortus larvae to levamisole. Intriguingly, the pyrantel sensitivity remained unchanged thus providing new evidence for distinct modes of action of these important anthelmintics in parasitic species versus C. elegans. More broadly, this highlights the limits of C. elegans as a predictive model to decipher cholinergic agonist targets from parasitic nematode species and provides key molecular insight to inform the discovery of next generation anthelmintic compounds.

cIvermectin and moxidectin are the most widely administered anthelmintic macrocyclic lactones (MLs) to treat human and animal nematode infections. Their widespread and frequent use has led to a high level of resistance to these drugs. Although they have the same mode of action, differences in terms of selection for drug resistance have been reported. Our objective was to study and compare changes occurring upon ivermectin or moxidectin selection in the model nematode Caenorhabditis elegans. C. elegans worms were submitted to stepwise exposure to increasing doses of moxidectin. The sensitivity of moxidectin-selected worms to MLs was determined in a larval development assay and compared with those of wild-type and ivermectin-selected strains. Selection with either ivermectin or moxidectin led to acquired tolerance to ivermectin, moxidectin, and eprinomectin. Importantly, moxidectin was the most potent ML in both ivermectin-and moxidectin-selected strains. Interestingly, this order of potency was also observed in a resistant Haemonchus contortus isolate. In addition, ivermectin-and moxidectin-selected strains displayed constitutive overexpression of several genes involved in xenobiotic metabolism and transport. Moreover, verapamil potentiated sensitivity to ivermectin and moxidectin, demonstrating that ABC transporters play a role in ML sensitivity in ML-selected C. elegans strains. Finally, both ivermectin-and moxidectin-selected strains displayed a dye-filling-defective phenotype. Overall, this work demonstrated that selection with ivermectin or moxidectin led to cross-resistance to several MLs in nematodes and that the induction of detoxification systems and defects in the integrity of amphidial neurons are two mechanisms that appear to affect the responsiveness of worms to both ivermectin and moxidectin.T he broad-spectrum anthelmintic macrocyclic lactones (MLs) are most commonly used in veterinary medicine to treat diseases caused by gastrointestinal nematodes and external parasites in livestock (1, 2). Ivermectin (IVM) was the first ML approved for use in animals and remains today the sole ML registered for use in humans, mainly to treat onchocerciasis through mass chemotherapy. Another ML, moxidectin (MOX), was subsequently commercialized for the veterinary market and is currently being evaluated for possible use against human onchocerciasis (3). Inevitably, the intensive use of these compounds has led to the emergence of resistance in small ruminant, cattle, and some human nematode parasites (4-7). Discovering the mechanisms by which resistance to MLs occurs remains an important challenge today.There is consistent evidence that ATP-binding-cassette (ABC) transporters such as P-glycoproteins (Pgps) play an important role in multidrug resistance (MDR) in many organisms, including several nematode species. Gene expression levels of ABC transporters or allele frequencies were modified after ML selection (8-13), and they are involved in the tolerance of Caenorhabditis elegans (9, 14-16) and parasitic nematodes s...

These data implicate a role for polyamines in ethanol withdrawal-induced neurotoxicity and suggest that inhibiting the actions of polyamines on NMDARs may be neuroprotective under these conditions.

Resistance to the anthelmintic macrocyclic lactone ivermectin (IVM) has a great impact on the control of parasitic nematodes. The mechanisms by which nematodes adapt to IVM remain to be deciphered. We have identified NHR-8, a nuclear hormone receptor involved in the xenobiotic response in Caenorhabditis elegans , as a new regulator of tolerance to IVM. Loss-of-function nhr-8(ok186) C . elegans mutants subjected to larval development assays and electropharyngeogram measurements, displayed hypersensitivity to IVM, and silencing of nhr-8 in IVM-resistant worms increased IVM efficacy. In addition, compared to wild-type worms, nhr-8 mutants under IVM selection pressure failed to acquire tolerance to the drug. In addition, IVM-hypersensitive nhr-8(ok186) worms displayed low transcript levels of several genes from the xenobiotic detoxification network and a concomitant low Pgp-mediated drug efflux activity. Interestingly, some pgp and cyp genes known to impact IVM tolerance in many nematode species, were down regulated in nhr-8 mutants and inversely upregulated in IVM-resistant worms. Moreover, pgp-6 overexpression in nhr-8(ok186) C . elegans increased tolerance to IVM. Importantly, NHR-8 function was rescued in nhr-8(ok186) C . elegans with the homolog of the parasitic nematode Haemonchus contortus , and silencing of Hco-nhr-8 by RNAi on L2 H . contortus larvae increased IVM susceptibility in both susceptible and resistant H . contortus isolates. Thus, our data show that NHR-8 controls the tolerance and development of resistance to IVM in C . elegans and the molecular basis for this relates to the NHR-8-mediated upregulation of IVM detoxification genes. Since our results show that Hco - nhr-8 functions similarly to Cel - nhr-8 , this study helps to better understand mechanisms underlying failure in drug efficacy and open perspectives in finding new compounds with NHR-8 antagonist activity to potentiate IVM efficacy.

An elicitation exercise was conducted to collect and identify pressing questions concerning the study of helminths in livestock, to help guide research priorities. Questions were invited from the research community in an inclusive way. Of 385 questions submitted, 100 were chosen by online vote, with priority given to open questions in important areas that are specific enough to permit investigation within a focused project or programme of research. The final list of questions was divided into ten themes. We present the questions and set them briefly in the context of the current state of knowledge. Although subjective, results provide a snapshot of current concerns and perceived priorities in the field of livestock helminthology, and we hope will stimulate ongoing or new research efforts.

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