Nematode Haemonchus Contortus Research Articles

Genome-wide RNA interference of the nhr gene family in barber's pole worm identified members crucial for larval viability in vitro

Nuclear hormone receptors (NHRs) are emerging target candidates against nematode infection and resistance. However, there is a lack of comprehensive information on NHR-coding genes in parasitic nematodes. In this study, we curated the nhr gene family for 60 major parasitic nematodes from humans and animals. Compared with the free-living model organism Caenorhabditis elegans, a remarkable contraction of the nhr family was revealed in parasitic species, with genetic diversification and conservation unveiled among nematode Clades I (10–13), III (16–42), IV (33–35) and V (25–64). Using an in vitro biosystem, we demonstrated that 40 nhr genes in a blood-feeding nematode Haemonchus contortus (clade V; barber's pole worm) were responsive to host serum and one nhr gene (i.e., nhr-64) was consistently stimulated by anthelmintics (i.e., ivermectin, thiabendazole and levamisole); Using a high-throughput RNA interference platform, we knocked down 43 nhr genes of H. contortus and identified at least two genes that are required for the viability (i.e., nhr-105) and development (i.e., nhr-17) of the infective larvae of this parasitic nematode in vitro. Harnessing this preliminary functional atlas of nhr genes for H. contortus will prime the biological studies of this gene family in nematode genetics, infection, and anthelmintic metabolism within host animals, as well as the promising discovery of novel intervention targets.

An alkaloidal fraction of Annona tomentosa possesses anthelmintic activity and induces surface modifications in Haemonchus contortus

The nematode Haemonchus contortus is, as a parasite, responsible for most mortality of small ruminants, causing significant economic losses. Numerous plant-derived compounds have exhibited promising anthelmintic activities against this nematode. Notably, the Annona genus stands out for demonstrated anthelmintic effects by extracts from several of its species against different nematodes. This study aimed to assess the effect of an Annona tomentosa fraction, rich in alkaloids, on H. contortus. This fraction, named Alk.F, is derived from the methanolic extract of the plant’s stem bark. Chemical characterization of Alk.F was performed by liquid chromatography coupled with mass spectrometry. Among the nine predominant peaks obtained, seven alkaloids were identified: reticuline, reticuline N-oxide, reticuline N-oxide isomer, cyclanoline, asimilobine, tetrahydropalmatine and anonaine. Alk.F inhibited the larval development of H. contortus with an IC50 of 0.026 mg/mL, inhibited larval exsheathment with an IC50 of 0.38 mg/mL, and displayed low hemolytic activity towards sheep erythrocytes. Furthermore, atomic force microscopy revealed that Alk.F altered adhesive forces and the height profile on the surface of H. contortus larvae. In conclusion, A. tomentosa alkaloids alter the cuticle structure of H. contortus, inhibiting larval development and exsheathment, thus offering possibilities for contributing to the development of new anthelmintic drugs.

Ethnopharmacology and anthelmintic screening of some plants used in traditional medicine in the Far-North of Cameroon

Background: The use of traditional medicinal plants in Cameroon for the treatment of intestinal helminthiasis is a common practice among traditional healers and livestock breeders. However, the use of ethnopharmaceutical products for their anthelmintic activities against Haemonchus contortus has been poorly addressed in the Far-North region of Cameroon, therefore justifying the need to bridge this gap. Methods: An ethno-pharmacological survey was conducted among livestock breeders and traditional healers in the Far-North region of Cameroon using a questionnaire to determine different plants used for the treatment of gastrointestinal helminths. Thereafter, aqueous and hydroethanolic extracts of the most used plant (frequency >2%) were evaluated for their anthelmintic activity against the gastrointestinal nematode Haemonchus contortus in vitro. Results: Seventy traditional healers and 36 livestock breeders were surveyed with 22 medicinal plants found to be involved in the treatment of intestinal helminthiasis in the Far-North Region of Cameroon. These plant species are distributed in 15 families with the Fabaceae being the most used (22.6%). The roots (39.6%) and leaves (36.8%) of the plants were the most frequently used parts mainly in the form of decoction (45.3%) and maceration (29.3%). Aristolochia baetica (20.8%) was the most used plant. Eleven plants were selected and all exhibited significant (p ˂ 0.001) anthelmintic activity on adult Haemonchus contortus. The highest anthelmintic activity was observed with the hydroethanolic extract irrespective of the plant species. The hydroalcoholic and aqueous extracts of Tephrosia pedicellata, Aristolochia baetica, and Abelmochus esculentus showed the highest anthelmintic activity with 100% mortality at 1 mg/mL after 24 hours of incubation. Conclusion: This study allowed us to list the anthelmintic plants used in traditional medicine in the Far North of Cameroon, with the anthelmintic screening sustaining the traditional use of these active plants for the control of helminthiasis in humans and animals.

Phytochemical Profiling Studies of Alkaloids and Coumarins from the Australian Plant Geijera parviflora Lindl. (Rutaceae) and Their Anthelmintic and Antimicrobial Assessment.

Phytochemical profiling followed by antimicrobial and anthelmintic activity evaluation of the Australian plant Geijera parviflora, known for its customary use in Indigenous Australian ceremonies and bush medicine, was performed. In the present study, seven previously reported compounds were isolated including auraptene, 6'-dehydromarmin, geiparvarin, marmin acetonide, flindersine, and two flindersine derivatives from the bark and leaves, together with a new compound, chlorogeiparvarin, formed as an artefact during the isolation procedure and isolated as a mixture with geiparvarin. Chemical profiling allowed for a qualitative and quantitative comparison of the compounds in the leaves, bark, flowers, and fruit of this plant. Subsequently, a subset of these compounds as well as crude extracts from the plant were evaluated for their antimicrobial and anthelmintic activities. Anthelmintic activity assays showed that two of the isolated compounds, auraptene and flindersine, as well as the dichloromethane and methanol crude extracts of G. parviflora, displayed significant activity against a parasitic nematode (Haemonchus contortus). This is the first report of the anthelmintic activity associated with these compounds and indicates the importance of such fundamental explorations for the discovery of bioactive phytochemicals for therapeutic application(s).

In vitro evaluation of the potential of mites of the family Macrochelidae (Acari: Mesostigmata) as macrobiological agents against the nematode Haemonchus contortus (Strongylida: Trichostrongylidae)

Small ruminants (sheep and goats) constantly suffer from endoparasitoses caused by gastrointestinal nematodes. Among these, the species Haemonchus contortus (Rudolphi, 1803) is considered to be the one of greatest importance within sheep farming. This nematode is difficult to control due to its resistance to most commercial anthelmintics. The aim of the present study was to assess the potential of macrochelid mites as macrobiological agents for controlling endoparasitoses of sheep caused by the nematode, H. contortus. For this, novel in vitro methodology was used, in which assessments were made not only of the predatory ability but also the population growth of mite species (Macrocheles merdarius, Macrocheles robustulus and Holostaspella bifoliata) when offered larvae of the nematode, H. contortus. The predatory ability of the mites, M. merdarius and H. bifoliata were efficient regarding their predatory ability against H. contortus nematode larvae. The mite, M. merdarius exhibited the highest predation rate with mean distribution values for the treated group of 18656 ± 10091 and for the control group of 1178 ± 712 (P < 0.0001). The species, H. bifoliata presented the highest population growth rate, with a percentage acarid recovery rate of 263% in relation to the number added initially. The data from this in vitro predation experiment suggest that, M. merdarius and H. bifoliata showed promise as macrobiological agents for controlling gastrointestinal endoparasitoses of sheep caused by the nematode, H. contortus given that both species reduced the population of this helminth by more 70% and the number of mites recovered was three times greater than the number added.

Application of the SeDeM system for the preparation of antiparasitic tablets from mesquite flour for use in sheep.

The use of herbal medicine to treat various diseases is becoming increasingly important as an alternative therapy. Numerous plants have been traditionally used for different purposes, including antiparasitic in humans and animals. Diseases caused by gastrointestinal parasites in ruminants, especially by the nematode Haemonchus contortus, cause large economic losses to the producers, whether by complications of the diseases or the cost of treatment. The main way of handling nematodiasis is by administering anthelmintic drugs, but their excessive use has the disadvantage of causing drug resistance; therefore, an alternative is the use of herbal medicine for this purpose. Mesquite (Prosopis spp.) has been used in Mexico to treat gastrointestinal diseases attributed to helminths. The present study aimed to characterize the rheological properties of mesquite flour using the SeDeM Expert System to determine its suitability for tablet production by direct compression. Direct compression technology facilitates the tableting process by reducing manufacturing costs. The results of the present study indicate that mesquite flour can be processed by direct compression. The latter could allow the manufacturing of economic tablets to treat infections by H. contortus in ruminants.

Flubendazole carbonyl reduction in drug-susceptible and drug-resistant strains of the parasitic nematode Haemonchus contortus: changes during the life cycle and possible inhibition

Carbonyl-reducing enzymes (CREs) catalyse the reduction of carbonyl groups in many eobiotic and xenobiotic compounds in all organisms, including helminths. Previous studies have shown the important roles of CREs in the deactivation of several anthelmintic drugs (e.g., flubendazole and mebendazole) in adults infected with the parasitic nematode Haemonchus contortus, in which the activity of a CRE is increased in drug-resistant strains. The aim of the present study was to compare the abilities of nematodes of both a drug-susceptible strain (ISE) and a drug-resistant strain (IRE) to reduce the carbonyl group of flubendazole (FLU) in different developmental stages (eggs, L1/2 larvae, L3 larvae, and adults). In addition, the effects of selected CRE inhibitors (e.g., glycyrrhetinic acid, naringenin, silybin, luteolin, glyceraldehyde, and menadione) on the reduction of FLU were evaluated in vitro and ex vivo in H. contortus adults. The results showed that FLU was reduced by H. contortus in all developmental stages, with adult IRE females being the most metabolically active. Larvae (L1/2 and L3) and adult females of the IRE strain reduced FLU more effectively than those of the ISE strain. Data from the in vitro inhibition study (performed with cytosolic-like fractions of H. contortus adult homogenate) revealed that glycyrrhetinic acid, naringenin, mebendazole and menadione are effective inhibitors of FLU reduction. Ex vivo study data showed that menadione inhibited FLU reduction and also decreased the viability of H. contortus adults to a similar extent. Naringenin and mebendazole were not toxic at the concentrations tested, but they did not inhibit the reduction of FLU in adult worms ex vivo.

PCR-RFLP study on drug resistance of Haemonchus contortus to benzimidazole in sheep, North region of Iran

Haemonchosis is one of the most important parasitic diseases of the gastrointestinal tract of small ruminants in different parts of Iran and worldwide. While the disease poses major economic problems in the livestock industry, there are many reports on parasites’ resistance to benzimidazoles necessita­ting to study of the level of this drug resistance in pathogens. The aim of this study was to evaluate the drug resistance of Haemonchus contortus to benzimidazole using molecular method in sheep from the North region of Iran. In this study, the resistance to benzimidazole was investigated using the PCR-RFLP method in the nematode Haemonchus contortus from sheep slaughtered in the northern region of Iran (Gilan and Mazandaran provinces). The samples examined in this study were evaluated between April 2020 and September 2021 from a total of 2400 sheep by determining the age groups of &lt;2 years, 2-4 years, and &gt;4 years. Three hundred Haemonchus contortus nematodes were randomly selected and investigated by means of PCR-RFLP and using TaaI endonuclease enzyme (SNP) in the beta-tubulin gene, responsible for drug resistance of Haemonchus contortus to benzimidazole. Haemonchus contortus was found in 66.8% and 60.5% of sheep in Gilan and Mazandaran. The results of the study showed that the drug sensitivity rate was 24% (72/300), the drug resistance rate was 54% (162/300) and 22% (66/300) of the samples had both resistant and sensitive alleles. Presented data showed a high level of prevalence of drug resistance in the nematode Haemonchus contortus in sheep in the northern region of Iran, which requires special attention to control the development of this resistance and the epidemic of haemonchosis in ruminants.

Anthelmintic efficacity of balanites Aegyptiaca (l.) Delile (Zygophyllaceae) almonds on three developmental stages of haemonchus contortus

Gastrointestinal nematodes are a major constraint on small ruminant farming in Burkina Faso. To overcome this, rural farmers use the Balanites aegyptiaca plant to combat gastrointestinal parasites on their family farms. The aim of this study was therefore to investigate the in vitro anthelmintic efficacy of the aqueous extract of B. aegyptiaca almonds powder against the parasitic nematode Haemonchus contortus. To this end, in vitro tests were carried out to evaluate the biological activity of the aqueous extract using five increasing concentrations over the three life stages of H. contortus by performing the tests (i) inhibition of egg hatch, (ii) inhibition of larva development and (iii) mortality of adult female worms. The results showed 21.22-80.4 %, 47.80-88.57 % and 22.20-100 % of ovicide, larvicide and vermicide rates, respectively. The effects were dose and exposition time dependent. The median lethal inhibition concentrations (IC50) were estimated to be 2.20 and 0.84 mg/ mL for the inhibitionof egg and the inhibition of larva, respectively. It can be concluded that B. aegyptiaca almonds powder has anthelmintic activity against H. contortus. However, this activity needs to be verified in vivo to reassure farmers about its use in small ruminant raising.

PSII-5 Evaluating Haemonchus Contortus Resistance in Hair Sheep Lambs Under Divergent Selection for Reduced Fecal Egg Count

Abstract The gastrointestinal nematode (GIN) Haemonchus contortus is a major threat to global sheep production, a problem exacerbated by increasing levels of anthelmintic resistance. Genetic selection of sheep for reduced fecal egg count (FEC) is hypothesized to improve parasite resistance. This study measured the biological response and worm burden of Dorper x White Dorper (DWD) with high (average = +37.8) or low (average = -12.0) post-weaning FEC estimated breeding values (EBV) and Barbados Blackbelly x Mouflon (BBM) following oral inoculation with 10,000 H. contortus larvae. Lambs (aged 8 months) were grouped by breed and EBV (DWD+, DWD-, BBM) and treatment (inoculated or non-inoculated control) into 6 groups (n = 4/grp). At d 0 and 21, blood samples were collected to measure packed-cell volume (PCV) and serum IgG via enzyme linked immunosorbent assay and at d 21 FEC were additionally recorded. Lambs were subsequently harvested and adult H. contortus counts for each lamb were captured from the abomasum. Data analysis was performed in SAS v 9.4 and significance was set at P &amp;lt; 0.05. In the inoculated lambs, no difference was observed in PCV at d 21, but DWD+ had a greater mean FEC (1,400 + 297 epg) than both DWD- (263 + 167; P = 0.0157) and BBM (50 + 21; P = 0.004). Total serum IgG (ug/mL) was greater in DWD- than DWD+ lambs at both d 0 (686.37 + 50.7 v. 486.15 + 51.25; P = 0.0321) and d 21 (287.16 + 14.33 v. 197.24 + 12.82; P = 0.0034). In BBM, there was no difference in IgG from d 0 (172.80 + 22.44) to d 21 (185.08 ± 4.25). While BBM had reduced IgG at d 0 than both DWD groups, there was no difference between BBM and DWD+ at d 21 (P = 0.4026). Following harvest, inoculated DWD+ had greater worm burden than DWD- (1,330 + 76 v. 818 + 165; P = 0.0308) and both DWD groups were greater than BBM (263 + 120; P = 0.0003). These data confirm previous findings that BBM are more resistant to H. contortus than DWD, and selection for decreased FEC EBV decreased fecal egg output and worm burden within the DWD breed. Future studies will include RNA-sequencing of abomasal epithelial tissue from inoculated and control lambs in an effort to elucidate gene expression differences between divergently selected DWD and BBM sheep infected with H. contortus.

Study of a Mexican isolate of Arthrobotrys musiformis (Orbiliales): Predatory behavior and nematocidal activity of liquid culture filtrates against Haemonchus contortus (Trichostrongylidae), protein profile and myco-constituent groups

A Mexican isolate of the nematophagous fungus Arthrobotrys musiformis was obtained from a soil sample from the Chapultepec ecological reserve zone, in Cuernavaca, Morelos, Mexico. This isolate demonstrated an important predatory activity (74.9%) against the parasitic nematode Haemonchus contortus (L3) and its fungal liquid culture filtrates (LCF) grown in two media showed the following highest nematocidal activities (NA): In Czapek-DoxBroth (CzDoxB) 80.66% and potato-dextrose broth (PDB) 49.84%. Additionally, two major compounds derived from carboxylic acids and two derivates from alkane group were identified by GC–MS. These compounds have been associated to many biological activities. On the other hand, the protein profile analysis by SDS-electrophoresis followed by a zymogram revealed a 10 kDa protein with protease activity. This study provides important information for future experiments focused to explore the potential use of this protein as well as the identified bioactive compounds presents in the LCF as potential candidates against sheep haemonchosis.

Growth Performance and Ruminal Fermentation in Lambs with Endoparasites and In Vitro Effect of Medicinal Plants

We investigated growth performance and ruminal fermentation associated with gastrointestinal nematode (GIN) Haemonchus contortus in lambs and in vitro ruminal fermentation of mallow, chamomile, fumitory, wormwood (Herbmix), and chicory using inoculum from GIN-infected lambs. Twelve lambs were equally divided into two groups: uninfected animals (CON) and animals infected (INF) with approximately 5000 third-stage larvae derived from the MHCo1 strain of GIN H. contortus. Two lambs per group were killed on days 48, 49, and 50 after infection and ruminal content was collected separately from each lamb. Batch cultures of ruminal fluid from CON and INF were incubated for 24 h in vitro with 0.25 g meadow hay, Herbmix, and chicory using an in vitro gas production technique. Daily weight gain was relatively lower in the INF than the CON group, but not significantly (72.6 vs. 130.1 g/day). The ruminal populations of protozoa, bacteria, total Archaea, Methanobacteriales, and Methanomicrobiales were significantly higher in the INF than in the CON group. The substrates affected the concentrations of n-butyrate, iso-butyrate, n-valerate, iso-valerate, ammonia-N, total gas, and methane (p &lt; 0.001) in vitro. GIN infection affected fermentation and microbial population in the rumens of the lambs, and chicory was a promising substrate to modulate ruminal fermentation in vitro.

Impact of Haemonchus contortus infection on feed intake, digestion, liveweight gain, and enteric methane emission from Red Maasai and Dorper sheep

A study was conducted with Red Maasai and Dorper lambs to evaluate the effects of infection with the gastrointestinal nematode (GIN) Haemonchus contortus on feed intake, liveweight gain (LWG), feed energy and nitrogen partitioning, and enteric methane (CH4) emissions. Six- to seven-month-old Red Maasai (n=12) and Dorper (n=12) lambs were randomly allocated to three treatments (n=8, four lambs per breed) in a 2×3 factorial cross-over study over two periods (P1 and P2) of 36 days each. The treatments consisted of three combinations of GIN infection and feeding level: Infected + ad libitum feeding (I-adlib), uninfected + ad libitum feeding (Un-adlib), and uninfected + restricted feeding (Un-restd), across the two breeds. Lambs in the I-adlib group were trickle-infected daily with 1,000 L3 stage larvae of H. contortus for four consecutive days (Days 1–4), whereas lambs in the other experimental treatments were kept GIN free. The feed intake was measured daily. Liveweight (LW), faecal egg counts (FEC), and packed cell volume (PCV) were measured on Day 1 and weekly thereafter. On Days 29–33 total faecal and urine outputs were determined in metabolic crates. The lambs were then housed in respiration chambers for three consecutive days (Days 34–36). There was a washout period of 21 days before P2 started. Uninfected lambs (Un-adlib and Un-restd) had undetectable FEC throughout the study. On Day 36, FEC did not differ between the breeds (P&amp;gt;0.05). Infected lambs (I-adlib) had lower PCV than uninfected (Un-adlib and Un-restd) lambs on day 36. Neither breed nor infection influenced feed and nutrient intake, but as expected, restricted-fed lambs had a lower intake (P&amp;lt;0.05). The LWG of Un-adlib lambs was significantly higher than that of I-adlib and Un-restd lambs (P&amp;lt;0.05), whereas there was no breed difference (P&amp;gt;0.05). Neither breed nor infection affected feed digestibility, nitrogen retention or energy metabolisability (P&amp;gt;0.05). However, feed restriction decreased feed intake, LWG and N retention, whereas feed digestibility and energy metabolisability were unaffected. Neither daily CH4 emissions nor yield (per unit of feed intake) were affected by experimental infection, but Un-restd lambs had lower CH4 emissions per day. Red Maasai lambs had consistently lower daily CH4 emissions and yields than Dorper (P&amp;lt;0.01). This study confirmed the relative resistance of indigenous sheep (Red Maasai) to H. contortus infection, but the increased CH4 emission and yield due to GIN observed in other studies was not confirmed. Further investigations are needed to test whether in environments with multiple stress factors, local or indigenous breeds or their crossbreeds with exotic breeds may be better equipped to sustain production and simultaneously have a reduced carbon footprint than purebred exotic breeds.

Anthelmintic activity of European fern extracts against Haemonchus contortus

Most drugs used in the treatment of helminthiasis in humans and animals have lost their efficacy due to the development of drug-resistance in helminths. Moreover, since anthelmintics, like many pharmaceuticals, are now recognized as hazardous contaminants of the environment, returning to medicinal plants and their products represents an environmentally friendly way to treat helminthiasis. The goal of the present study was to test the anthelminthic activity of methanol extracts of eight selected European ferns from the genera Dryopteris, Athyrium and Blechnum against the nematode Haemonchus contortus, a widespread parasite of small ruminants. Eggs and adults of H. contortus drug-susceptible strain ISE and drug-resistant strain WR were isolated from experimentally infected sheep. The efficacy of fern extracts was assayed using egg hatch test and adults viability test based on ATP-level measurement. Among the ferns tested, only Dryopteris aemula extract (0.2 mg/mL) inhibited eggs hatching by 25% in comparison to control. Athyrium distentifolium, Dryopteris aemula and Dryopteris cambrensis were effective against H. contortus adults. In concentration 0.1 mg/mL, A. distentifolium, D. aemula, D. cambrensis significantly decreased the viability of females from ISE and WR strains to 36.2%, 51.9%, 32.9% and to 35.3%, 27.0%, 23.3%, respectively in comparison to untreated controls. None of the extracts exhibited toxicity in precise cut slices from ovine liver. Polyphenol’s analysis identified quercetin, kaempferol, luteolin, 3-hydroxybenzoic acid, caffeic acid, coumaric acid and protocatechuic acid as the major components of these anthelmintically active ferns.

An initial investigation of the predatory activity of the phoretic mites of dung beetles, Macrocheles sp. (Mesostigmata: Macrochelidae), on the gastrointestinal nematode of sheep Haemonchus contortus (Strongylida: Trichostrongylidae)

Worldwide increasing levels of anthelmintic resistance are pushing farmers and health advisors towards a more sustainable management of gastrointestinal nematodes (GIN), one of the greatest threats to the viability of grazing small ruminant production systems, and to these animals’ welfare. These Strongyloidea develop freely in livestock dung, where they are exposed to predatory organisms that can eventually break their cycle. The aim of this study was to investigate the effect of phoretic mites of dung beetles Macrocheles sp. (Macrochelidae) on the gastrointestinal nematode Haemonchus contortus (Rudolphi, 1803) that has a free-living larval stage that evolves in sheep feces. Direct observations under a stereomicroscope confirmed that the studied Macrocheles mites may have an active predatory behavior on the third-stage (L3) larvae of H. contortus. In order to quantify the effects of the mites’ predation on the survival of L3 nematode larvae, we designed experimental units, replicated across five trials, and containing free-living stages of the nematode either in the presence or absence of mites. The number of H. contortus larvae that reached the third-stage was significantly reduced in the presence of mites, despite a variable intensity of the predatory effects between trials. This is the first report of a predatory activity of phoretic mites of dung beetles’ on a livestock gastrointestinal nematode. This first step encourages further studies in order to have a better quantification of this interaction, and to investigate to which extent the phoretic mites of dung beetles can impact H. contortus populations in natural conditions.

Venom composition and bioactive RF-amide peptide toxins of the saddleback caterpillar, Acharia stimulea (Lepidoptera: Limacodidae)

Limacodidae is a family of lepidopteran insects comprising >1500 species. More than half of these species produce pain-inducing defensive venoms in the larval stage, but little is known about their venom toxins. Recently, we characterised proteinaceous toxins from the Australian limacodid caterpillar Doratifera vulnerans, but it is unknown if the venom of this species is typical of other Limacodidae. Here, we use single animal transcriptomics and venom proteomics to investigate the venom of an iconic limacodid, the North American saddleback caterpillar Acharia stimulea. We identified 65 venom polypeptides, grouped into 31 different families. Neurohormones, knottins, and homologues of the immune signaller Diedel make up the majority of A.stimulea venom, indicating strong similarities to D. vulnerans venom, despite the large geographic separation of these caterpillars. One notable difference is the presence of RF-amide peptide toxins in A. stimulea venom. Synthetic versions of one of these RF-amide toxins potently activated the human neuropeptide FF1 receptor, displayed insecticidal activity when injected into Drosophila melanogaster, and moderately inhibited larval development of the parasitic nematode Haemonchus contortus. This study provides insights into the evolution and activity of venom toxins in Limacodidae, and provides a platform for future structure-function characterisation of A.stimulea peptide toxins.

Refugia or reservoir? Feral goats and their role in the maintenance and circulation of benzimidazole-resistant gastrointestinal nematodes on shared pastures.

Gastrointestinal nematodes threaten the productivity of grazing livestock and anthelmintic resistance has emerged globally. It is broadly understood that wild ruminants living in sympatry with livestock act as a positive source of refugia for anthelmintic-susceptible nematodes. However, they might also act as reservoirs of anthelmintic-resistant nematodes, contributing to the spread of anthelmintic resistance at a regional scale. Here, we sampled managed sheep and cattle together with feral goats within the same property in New South Wales, Australia. Internal transcribed spacer 2 (ITS-2) nemabiome metabarcoding identified 12 gastrointestinal nematodes (Cooperia oncophora, Cooperia punctata, Haemonchus contortus, Haemonchus placei, Nematodirus spathiger, Ostertagia ostertagi, Teladorsagia circumcincta, Oesophagostomum radiatum, Oesophagostomum venulosum, Trichostrongylus axei, Trichostrongylus colubriformis and Trichostrongylus rugatus). Isotype-1 β-tubulin metabarcoding targeting benzimidazole resistance polymorphisms identified 6 of these nematode species (C. oncophora, C. punctata, H. contortus, H. placei, O. ostertagi and T. circumcincta), with the remaining 3 genera unable to be identified to the species level (Nematodirus, Oesophagostomum, Trichostrongylus). Both ITS-2 and β-tubulin metabarcoding showed the presence of a cryptic species of T. circumcincta, known from domestic goats in France. Of the gastrointestinal nematodes detected via β-tubulin metabarcoding, H. contortus, T. circumcincta, Nematodirus and Trichostrongylus exhibited the presence of at least one resistance genotype. We found that generalist gastrointestinal nematodes in untreated feral goats had a similarly high frequency of the benzimidazole-resistant F200Y polymorphism as those nematodes in sheep and cattle. This suggests cross-transmission and maintenance of the resistant genotype within the wild ruminant population, affirming that wild ruminants should be considered potential reservoirs of anthelmintic resistance.

The Haemonchus contortus LGC-39 subunit is a novel subtype of an acetylcholine-gated chloride channel

The nematode genome exhibits a vast array of Cys-loop receptors that are activated by a diverse set of neurotransmitters and anthelmintic drugs such as ivermectin and levamisole. While many Cys-loop receptors have been functionally and pharmacologically characterized, there remains a large subset of orphan receptors where the agonist remains unknown. We have identified an orphan Cys-loop receptor, LGC-39, from the parasitic nematode Haemonchus contortus that is a novel type of cholinergic-sensitive ligand-gated chloride channel. This receptor groups outside of the acetylcholine-gated chloride channel family, in the previously named GGR-1 (GABA/Glycine Receptor-1) group of Cys-loop receptors. We found that LGC-39 forms a functional homomeric receptor when expressed in Xenopus laevis oocytes and is activated by several cholinergic ligands including acetylcholine, methacholine and surprisingly, atropine with an EC50 for atropine on the low μM range. A homology model was generated which revealed some key features of the LGC-39 ligand-binding pocket that may explain some of the elements important for atropine recognition of the LGC-39 receptor. Overall these results suggest that the GGR-1 family (now called LGC-57) of Cys-loop receptors includes novel acetylcholine-gated chloride channel subtypes and may represent important future drug targets.

Short-chain dehydrogenases in Haemonchus contortus: changes during life cycle and in relation to drug-resistance

Short-chain dehydrogenases/reductases (SDRs) regulate the activities of many hormones and other signaling molecules and participate in the deactivation of various carbonyl-bearing xenobiotics. Nevertheless, knowledge about these important enzymes in helminths remains limited. The aim of our study was to characterize the SDR superfamily in the parasitic nematode Haemonchus contortus. Genome localization of SDRs was explored, and phylogenetic analysis in comparison with SDRs from free-living nematode Caenorhabditis elegans and the domestic sheep (Ovis aries, a typical host of H. contortus) was constructed. The expression profile of selected SDRs during the life cycle along with differences between the drug-susceptible and drug-resistant strains, were also studied. Genome sequencing enabled the identification of 46 members of the SDR family in H. contortus. A number of genes have no orthologue in the sheep genome. In all developmental stages of H. contortus, SDR1, SDR3, SDR5, SDR6, SDR14, and SDR18 genes were the most expressed, although in individual stages, huge differences in expression levels were observed. A comparison of SDRs expression between the drug-susceptible and drug-resistant strains of H. contortus revealed several SDRs with changed expression in the resistant strain. Specifically, SDR1, SDR12, SDR13, SDR16 are SDR candidates related to drug-resistance, as the expression of these SDRs is consistently increased in most stages of the drug-resistant H. contortus. These findings revealing several SDR enzymes of H. contortus warrant further investigation.

Survival of Nematode Larvae Strongyloides papillosus and Haemonchus contortus under the Influence of Various Groups of Organic Compounds

Many chemically synthesized xenobiotics can significantly inhibit the vitality of parasitic nematodes. However, there is yet too little research on the toxicity of such contaminating compounds toward nematodes. Compounds that are present in plants are able to inhibit the vitality of parasitic organisms as well. According to the results of our laboratory studies of toxicity, the following xenobiotics caused no decrease in the vitality of the larvae of Strongyloides papillosus and Haemonchus contortus: methanol, propan-2-ol, propylene glycol-1,2, octadecanol-1, 4-methyl-2-pen-tanol, 2-ethoxyethanol, butyl glycol, 2-pentanone, cyclopentanol, ortho-dimethylbenzene, dibutyl phthalate, succinic anhydride, 2-methylfuran, 2-methyl-5-nitroimidazole. Strong toxicity towards the nematode larvae was exerted by glutaraldehyde, 1,4-diethyl 2-methyl-3-oxobutanedioate, hexylamine, diethyl malonate, allyl acetoacetate, tert butyl carboxylic acid, butyl acrylate, 3-methyl-2-butanone, isobutyraldehyde, methyl acetoacetate, ethyl acetoacetate, ethyl pyruvate, 3-methylbutanal, cyclohexanol, cyclooctanone, phenol, pyrocatechin, resorcinol, naphthol-2, phenyl ether, piperonyl alcohol, 3-furoic acid, maleic anhydrid, 5-methylfurfural, thioacetic acid, butan-1-amine, dimethylformamide, 1-phenylethan-1-amine, 3-aminobenzoic acid. Widespread natural compounds (phytol, 3-hydroxy-2-butanone, maleic acid, oleic acid, hydroquinone, gallic acid-1-hydrate, taurine, 6-aminocaproic acid, glutamic acid, carnitine, ornithine monohydrochloride) had no negative effect on the larvae of S. papillosus and H. contortus. A powerful decrease in the vitality of nematode larvae was produced by 3,7-dimethyl-6-octenoic acid, isovaleric acid, glycolic acid, 2-oxopentanedioic acid, 2-methylbutanoic acid, anisole, 4-hydroxy-3-methoxybenzyl alcohol, furfuryl alcohol. The results of our studies allow us to consider 28 of the 62 compounds we studied as promising for further research on anti-nematode activity in manufacturing conditions.