Benzimidazole and Aminoalcohol Derivatives Show in Vitro Anthelmintic Activity Against Trichuris Muris and Heligmosomoides Polygyrus

Background: Infections by gastrointestinal nematodes cause significant economic losses and disease in both human and animals worldwide. The discovery of novel anthelmintic drugs is a crucial point in maintaining control of these parasitic infections.Methods: For this purpose, the aim of the present study was to evaluate the potential anthelmintic activity of three series of compounds against the gastrointestinal nematodes Trichuris muris and Heligmosomoides polygyrusin vitro. The compounds tested are derivatives of benzimidazole, lipidic aminoalcohols and diamines. A primary screening was performed to select those compounds with an ability to inhibit T. muris L1 motility by more than 90% at a single concentration of 100 µM, and then, their respective IC50s were calculated. Those compounds with IC50 lower than 10 µM were also tested against the adult stage of T. muris and H. polygyrus at a single concentration of 10µM.Results: Of the 41 initial compounds screened, only compounds AO14, BZ6 and BZ12 had IC50s lower than 10 µM on T. muris L1 assay, showing IC50 values of 3.30, 8.89 and 4.17 µM, respectively. However, only two of them displayed activity against the adult stage of the parasites: BZ12 killed 81% of adults of T. muris (IC50 of 8.1 µM) and 53% of H. polygyrus while BZ6 killed 100% of H. polygyrus adults (IC50 of 5.3 µM) but only 17% of T. muris. Conclusions: BZ6 and BZ12 could be considered as potential candidates for further in vivo efficacy testing.

Sertraline as a new potential anthelmintic against Haemonchus contortus: toxicity, efficacy, and biotransformation

Haemonchus contortus is a parasitic nematode of ruminants which causes significant losses to many farmers worldwide. Since the drugs currently in use for the treatment of haemonchosis are losing their effectiveness due to the drug-resistance of this nematode, a new or repurposed drug is highly needed. As the antipsychotic drug sertraline (SRT) has been shown to be effective against the parasitic nematodes Trichuris muris, Ancylostoma caninum and Schistosoma mansoni, the aim of the present study was to evaluate the possible effect of SRT on H. contortus. The potential hepatotoxicity of SRT was tested in sheep, a common H. contortus host. In addition, the main metabolic pathways of SRT in H. contortus and the ovine liver were identified. While no effect of SRT on H. contortus egg hatching was observed, SRT was found to significantly decrease the viability of H. contortus adults in drug-sensitive and resistant strains, with its effect comparable to the commonly used anthelmintics levamisole and monepantel. Moreover, SRT in anthelmintically active concentrations showed no toxicity to the ovine liver. Biotransformation of SRT in H. contortus was weak, with most of the drug remaining unmetabolized. Production of the main metabolite hydroxy-SRT did not differ significantly between strains. Other minor metabolites such as SRT-O-glucoside, dihydroxy-SRT, and SRT-ketone were also identified in H. contorts adults. Compared to H. contortus, the ovine liver metabolized SRT more extensively, mainly via desmethylation and glucuronidation. In conclusion, the potency of SRT against H. contortus was proven, and it should be tested further toward possible repurposing.

Un-‘Egg’-Plored: Characterisation of Embryonation in the Whipworm Model Organism Trichuris muris

Trichuris muris, is the murine parasite and widely deployed model for the human whipworm Trichuris trichiura, a parasite that infects around 500 million people globally. Trichuriasis is a classical disease of poverty with a cycle of re-infection due to the continual exposure of humans, particularly children, to infective eggs, which contaminate the soil in endemic areas. Indeed, modelling studies of trichuriasis have demonstrated that the low efficacy rate of current anthelmintics combined with the high possibility of re-infection from the reservoir of infective eggs within the environment, mean that the elimination of morbidity due to trichuriasis is unlikely to occur. Despite the importance of the infective egg stage in the perpetuation of infections, understanding the biology of the Trichuris ova has been neglected for decades. Here we perform experiments to assess the impact of temperature on the embryonation process of T. muris eggs and describe in detail the stages of larval development within these eggs. In keeping with the early works performed in the early 1900s, we show that the embryonation of T. muris is accelerated by an elevation in temperature, up to 37°C above which eggs do not fully develop and become degenerate. We extend these data to provide a detailed description of T. muris egg development with clear images depicting the various stages of development. To the best of our knowledge we have, for the first time, described the presence of birefringent granules within egg-stage larvae, as well as providing a qualitative and quantitative description of a motile larval stage prior to quiescence within the egg. These experiments are the first step towards a better understanding of the basic biology which underlies the process of egg embryonation. With the threat of elevation in global temperatures, the accelerated embryonation rate we observe at higher temperatures may have important consequences for parasite transmission rates and prospective modelling studies. In addition, a deeper understanding of the Trichuris ova may allow the development of novel control strategies targeting the egg stage of Trichuris in the environment as an adjunct to MDA.

Heterogeneous Tfh cell populations that develop during enteric helminth infection predict the quality of type 2 protective response

T follicular helper (Tfh) cells are an important component of the germinal centre (GC)-mediated humoral immunity. Yet, how regulation of Tfh-GC responses impacts on effective responses to helminth infection are poorly understood. Here we show that chronic helminth Trichuris muris infection fails to induce Tfh-GC B cell responses, with Tfh cells expressing T-bet and IFN-γ. In contrast, Tfh cells that express GATA-3 and IL-4 dominate responses to an acute, resolving infection. Accordingly, heightened expression and increased chromatin accessibility of Th1- and Th2 cell-associated genes is observed in chronic and acute induced Tfh cells, respectively. However, both acute and chronic Tfh cell populations retained the capacity to produce IL-21 in spite of the Th-biased response. Blockade of Tfh-GC interactions impaired type 2 immunity, highlighting the protective role of GC-dependent Th2-like Tfh cell responses against helminths. Collectively, these results provide new insights into the protective roles of Tfh-GC responses and identify distinct transcriptional and epigenetic features of Tfh cells that emerge during resolving or chronic helminth infections.

Parasite fauna diversity in Red Fox (Vulpes Vulpes) from natural and anthropized ecosystems of the Republic of Moldova

The study of the diversity of the parasite fauna in the investigated foxes showed a high level of infestation (100%). The taxonomy of parasitofauna in foxes includes 12 parasitic invasions (Isospora canis – 14,3%, Alaria alata – 51,0%, Mesocestoides lineatus – 21,7%, Taeniidae spp – 27,0%, Syphacia obvelata – 17,0%, Strongyloides stercoralis – 13,3%, Toxocara canis – 59,0%, Toxascaris leonina – 65,5%, Ancylostoma caninum – 8,7%, Trichuris vulpis – 26,1%, Trichuris muris – 4,4%, Capilaria hepatica – 35,0%), which belong to 5 classes, 10 families, 11 genera and about 12 species. The share of species from the Sporozoa class is 8.3%, from the Trematoda class - 8.3%, from the Cestoda class - 16.7%, from the Secernentea class - 41.7% and from the Adenophorea class - 25.0%. Analyzing the parasitic species on epidemiological criterion, it was found that 10 species (83.3%) with large spread have zoonotic impact (A. alata, M. lineatus, Taenia spp., S. obvelata, S. ratti, T. canis, T. leonina, A. caninum, C. hepatica, T. vulpis) with a major risk to public health, and the identified invasions (100%) can parasitize domestic animals, as well as game fauna.

Microbial byproducts determine reproductive fitness of free-living and parasitic nematodes

Trichuris nematodes reproduce within the microbiota-rich mammalian intestine, yet microbial byproducts that facilitate the parasite lifecycle are unknown. Here, we report a novel pipeline to identify microbial factors with conserved roles in the reproduction of nematodes. A screen for E. coli mutants that impair C. elegans fertility identified genes in fatty acid biosynthesis and ethanolamine utilization pathways, including fabH and eutN. Trichuris muris eggs displayed defective hatching in the presence of E. coli deficient in fabH or eutN due to reduction in arginine or elevated levels of aldehydes, respectively. Remarkably, T. muris reared in gnotobiotic mice colonized with these E. coli mutants failed to lay viable eggs. These findings indicate that microbial byproducts mediate evolutionarily conserved transkingdom interactions that impact reproductive fitness of distantly-related nematodes.

Trichuris muris infection drives cell-intrinsic IL4R alpha independent colonic RELMα+ macrophages

The intestinal nematode parasite Trichuris muris dwells in the caecum and proximal colon driving an acute resolving intestinal inflammation dominated by the presence of macrophages. Notably, these macrophages are characterised by their expression of RELMα during the resolution phase of the infection. The RELMα+ macrophage phenotype associates with the presence of alternatively activated macrophages and work in other model systems has demonstrated that the balance of classically and alternatively activated macrophages is critically important in enabling the resolution of inflammation. Moreover, in the context of type 2 immunity, RELMα+ alternatively activated macrophages are associated with the activation of macrophages via the IL4Rα. Despite a breadth of inflammatory pathologies associated with the large intestine, including those that accompany parasitic infection, it is not known how colonic macrophages are activated towards an alternatively activated phenotype. Here, we address this important knowledge gap by using Trichuris muris infection, in combination with transgenic mice (IL4Rαfl/fl.CX3CR1Cre) and IL4Rα-deficient/wild-type mixed bone marrow chimaeras. We make the unexpected finding that education of colonic macrophages towards a RELMα+, alternatively activated macrophage phenotype during T. muris infection does not require IL4Rα expression on macrophages. Further, this independence is maintained even when the mice are treated with an anti-IFNγ antibody during infection to create a strongly polarised Th2 environment. In contrast to RELMα, PD-L2 expression on macrophages post infection was dependent on IL4Rα signalling in the macrophages. These novel data sets are important, revealing a surprising cell-intrinsic IL4R alpha independence of the colonic RELMα+ alternatively activated macrophage during Trichuris muris infection.

Trichuris muris Model: Role in Understanding Intestinal Immune Response, Inflammation and Host Defense

Several parasites have evolved to survive in the human intestinal tract and over 1 billion people around the world, specifically in developing countries, are infected with enteric helminths. Trichuris trichiura is one of the world’s most common intestinal parasites that causes human parasitic infections. Trichuris muris, as an immunologically well-defined mouse model of T. trichiura, is extensively used to study different aspects of the innate and adaptive components of the immune system. Studies on T. muris model offer insights into understanding host immunity, since this parasite generates two distinct immune responses in resistant and susceptible strains of mouse. Apart from the immune cells, T. muris infection also influences various components of the intestinal tract, especially the gut microbiota, mucus layer, epithelial cells and smooth muscle cells. Here, we reviewed the different immune responses generated by innate and adaptive immune components during acute and chronic T. muris infections. Furthermore, we discussed the importance of studying T. muris model in understanding host–parasite interaction in the context of alteration in the host’s microbiota, intestinal barrier, inflammation, and host defense, and in parasite infection-mediated modulation of other immune and inflammatory diseases.

Functional Characterization of the Oxantel-Sensitive Acetylcholine Receptor from Trichuris muris

The human whipworm, Trichuris trichiura, is estimated to infect 289.6 million people globally. Control of human trichuriasis is a particular challenge, as most anthelmintics have a limited single-dose efficacy, with the striking exception of the narrow-spectrum anthelmintic, oxantel. We recently identified a novel ACR-16-like subunit from the pig whipworm, T. suis which gave rise to a functional acetylcholine receptor (nAChR) preferentially activated by oxantel. However, there is no ion channel described in the mouse model parasite T. muris so far. Here, we have identified the ACR-16-like and ACR-19 subunits from T. muris, and performed the functional characterization of the receptors in Xenopus laevis oocytes using two-electrode voltage-clamp electrophysiology. We found that the ACR-16-like subunit from T. muris formed a homomeric receptor gated by acetylcholine whereas the ACR-19 failed to create a functional channel. The subsequent pharmacological analysis of the Tmu-ACR-16-like receptor revealed that acetylcholine and oxantel were equally potent. The Tmu-ACR-16-like was more responsive to the toxic agonist epibatidine, but insensitive to pyrantel, in contrast to the Tsu-ACR-16-like receptor. These findings confirm that the ACR-16-like nAChR from Trichuris spp. is a preferential drug target for oxantel, and highlights the pharmacological difference between Trichuris species.