Parasitic Nematode Larvae Research Articles

Direct and indirect effects of fire on parasites in an African savanna

Abstract Fires in grassy ecosystems consume vegetation and initiate high‐quality regrowth, which results in pyric herbivory when mammalian grazers concentrate feeding in recent burns. For environmentally transmitted parasites with transmission mechanisms linked to vegetation structure, fire should exert direct effects on parasites, as well as indirect effects resulting from subsequent enhanced herbivory, which can affect parasite input and exposure to environmental conditions. We combined an experimental manipulation with observational data in the Serengeti National Park to investigate the direct and indirect effects of fire on parasites. We assessed the direct effects of fire by measuring changes in parasitic nematode larvae in the grass layer before and after fire on paired experimental burned and control plots. To investigate indirect effects linked to pyric herbivory, we sampled herbivore dung, grass biomass, ground temperature and larval densities every month for 5 months following fire in seven pairs of burned and unburned monitoring plots. Finally, to assess if fire‐driven changes to larval densities affected host infection burdens, we collected faecal samples from a key host, Grant's gazelle (Nanger granti), each month for 5 months to estimate within‐host parasite burdens. Fire killed all larvae and increased grazer dung inputs by 40% for 2 months following fire. Dung inputs after fire led to larval parasite recolonization of burned patches, but intense herbivory kept grass short and larval densities were associated with changes in ground temperature linked to grass biomass and ambient temperature. Grant's gazelles had lower parasite burdens when sampled in areas with higher compared to lower burned area fraction. Fire and pyric herbivory change the densities of larval parasites in the environment and divide the landscape into burned and unburned regions with distinct infection risks for local herbivores. The indirect effects quantified here represent a novel finding with major implications for all grazing systems impacted by fire.

Compatibility study of Duddingtonia flagrans conidia and its crude proteolytic extract

This study aimed to evaluate the concomitant use of the nematophagous fungus Duddingtonia flagrans (AC001) and its protease-rich crude extract for the in vitro control of Panagrellus sp., Haemonchus spp., and Trichostrongylus spp. The nematicidal tests were carried out on larvae of the free-living nematode Panagrellus sp. and infective larvae of the gastrointestinal parasitic nematodes of domestic ruminants (Haemonchus spp. and Trichostrongylus spp). Five experimental groups were set: (1) one control group (G1) and (4) four treated groups –G2 – active crude extract; G3 – denatured crude extract; G4 – fungus, and G5 – fungus + active extract. Plates were incubated at 28 ºC for 24 h followed by the recovery of the larvae using the Baermann technique. The results showed a lower recovery of Panagrellus sp. larvae in the experimental groups compared to the control group, as follows: 52 % (G2), 16 % (G3), 46 % (G4), and 77 % (G5). An even greater reduction (77 ± 5 %) occurred in the group (G5). In addition, the authors observed lower averages of L3 of Haemonchus spp. and Trichostrongylus spp. in the experimental groups compared to the control group, as follows: 59 % (G2), 0 % (G3), 86 % (G4), and 76 % (G5). In turn, there was a difference (p < 0.01) between (G5) and (G2). The results this study indicate a positive effect from the compatible use of the D. flagrans fungus and its enzymatic crude extract (protease), which has been demonstrated here for the first time and with potential field applications for further designs.

Identifikasi Nematoda Usus Pada Sayuran Lalapan Yang Dijual Di Pasar Tradisional dan Pasar Modern Di Kota Medan

This study aims to determine whether there is contamination of intestinal Nematode parasites found in fresh vegetables sold in Traditional Markets and Modern Markets in Medan City. This research method uses a descriptive method with a total sampling technique to be studied. Where the data that has been obtained and obtained is described or described as it is without intending to make conclusions that apply to the public or generalizations. Then the data can be made in tabular form in accordance with the Laboratory Standards. The results showed that from examining samples from the Brayan market, Sambu market, MMTC Pancing market, Sukaramai Denai market and Simpang Limun market, it was found that 5 samples of cabbage vegetables were contaminated with eggs and larvae of intestinal Nematode parasites and 5 samples of lettuce were contaminated with eggs and larvae of Nematode parasites. intestines. The types of parasites found were Ascaris lumbricoides eggs and Ancylostoma duodenale and Necator americanus larvae. Meanwhile, the samples from Brastagi Plaza, Carefour, Irian Supermarket, Maju Bersama Supermarket, and Cemara Fruit and Vegetable Market showed that 2 samples of lettuce were contaminated with Ascaris lumbricoides worm eggs, and the samples of cabbage and basil vegetables did not find larvae or basil.

Anisakid parasites (Nematoda: Anisakidae) in 3 commercially important gadid fish species from the southern Barents Sea, with emphasis on key infection drivers and spatial distribution within the hosts.

Northeast Arctic cod, saithe and haddock are among the most important fisheries resources in Europe, largely shipped to various continental markets. The present study aimed to map the presence and distribution of larvae of parasitic nematodes in the Anisakidae family which are of socioeconomic and public health concern. Fishes were sourced from commercial catches during winter or spring in the southern Barents Sea. Samples of fish were inspected for nematodes using the UV-press method while anisakid species identification relied on sequencing of the mtDNA cox2 gene. Anisakis simplex (s.s.) was the most prevalent and abundant anisakid recorded, occurring at high infection levels in the viscera and flesh of cod and saithe, while being less abundant in haddock. Contracaecum osculatum (s.l.) larvae, not found in the fish flesh, showed moderate-to-high prevalence in saithe, haddock and cod, respectively. Most Pseudoterranova spp. larvae occurred at low-to-moderate prevalence, and low abundance, in the viscera (Pseudoterranova bulbosa) and flesh (Pseudoterranova decipiens (s.s.) and Pseudoterranova krabbei) of cod, only 2 P. decipiens (s.s.) appeared in the flesh of saithe. Body length was the single most important host-related factor to predict overall abundance of anisakid larvae in the fish species. The spatial distribution of Anisakis larvae in the fish flesh showed much higher abundances in the belly flaps than in the dorsal fillet parts. Trimming of the flesh by removing the belly flaps would reduce larval presence in the fillets of these gadid fish species by 86–91%.

Efficient and Scalable Process to Produce Novel and Highly Bioactive Purified Cytosolic Crystals from Bacillus thuringiensis.

ABSTRACTBacillus thuringiensis (Bt) is a Gram-positive soil bacterium that is widely and safely applied in the environment as an insecticide for combatting insect pests that damage crops or are disease vectors. Dominant active ingredients made by Bt are insect-killing crystal (Cry) proteins released as crystalline inclusions upon bacterial sporulation. Some Bt Cry proteins, e.g., Cry5B (formally Cry5Ba1), target nematodes (roundworms) and show exceptional promise as anthelmintics (cures for parasitic nematode diseases). We have recently described inactivated bacteria with cytosolic crystal(s) (IBaCC) in which bioactive Bt Cry crystals (containing Cry5B) are fully contained within the cytosol of dead bacterial ghosts. Here, we demonstrate that these IBaCC-trapped Cry5B crystals can be liberated and purified away from cellular constituents, yielding purified cytosolic crystals (PCC). Cry5B PCC contains ~95% Cry5B protein out of the total protein content. Cry5B PCC is highly bioactive against parasitic nematode larvae and adults in vitro. Cry5B PCC is also highly active in vivo against experimental human hookworm and Ascaris infections in rodents. The process was scaled up to the 100-liter scale to produce PCC for a pilot study to treat two foals infected with the ascarid Parascaris spp. Single-dose Cry5B PCC brought the fecal egg counts of both foals to zero. These studies describe the process for the scalable production of purified Bt crystals and define a new and attractive pharmaceutical ingredient form of Bt Cry proteins.IMPORTANCE Bacillus thuringiensis crystal proteins are widely and safely used as insecticides. Recent studies have shown they also can cure gastrointestinal parasitic worm (nematode) infections when ingested. However, reproducible, scalable, and practical techniques for purifying these proteins have been lacking. Here, we address this severe limitation and present scalable and practical methods for large-scale purification of potently bioactive B. thuringiensis crystals and crystal proteins. The resultant product, called purified cytosolic crystals (PCC), is highly compatible with ingestible drug delivery and formulation. Furthermore, there are growing applications in agriculture and insect control where access to large quantities of purified crystal proteins is desirable and where these methods will find great utility.

Epidemiology of Trichinella infection in wild boar from Spain and its impact on human health during the period 2006-2019.

Trichinellosis is a notifiable zoonotic disease caused by parasitic nematode larvae belonging to the genus Trichinella. Domestic pig and wild boar are important hosts within the natural cycle of T. spiralis, the last one being an animal whose populations have experienced an important growth. Therefore, this paper studies the prevalence of Trichinella infection in wild boar in Spain, as well as its relation with hunting and its impact on public health during the period 2006–2019. For this purpose, different sources of information were consulted and analyzed depending on the autonomous communities of Spain and years. During the fourteen years of study, the number of wild boars hunted and the number of cases of Trichinella infection in them increased (from 172 cases in 2006 to 421 in 2019), although prevalence values remained low as the number of animals analyzed also increased. On the other hand, trichinellosis in humans tended to decrease (from a peak of 107 cases in 2007 to 11 cases in 2019). Nevertheless, the numbers of both wild boars and humans infected with Trichinella in Spain are among the highest in Europe, and this emphasizes the importance of food safety, sanitary controls of game meat and citizen awareness campaigns, which prevent the spread of Trichinella through the human population.

The ‘nuclear option’ revisited: Confirmation of Ss-daf-12 function and therapeutic potential in Strongyloides stercoralis and other parasitic nematode infections

Mechanisms governing morphogenesis and development of infectious third-stage larvae (L3i) of parasitic nematodes have been likened to those regulating dauer development in Caenorhabditis elegans. Dauer regulatory signal transduction comprises initial G protein-coupled receptor (GPCR) signaling in chemosensory neurons of the amphidial complex that regulates parallel insulin- and TGFβ-like signaling in the tissues. Insulin- and TGFβ-like signals converge to co-regulate steroid signaling through the nuclear receptor (NR) DAF-12. Discovery of the steroid ligands of DAF-12 opened a new avenue of small molecule physiology in C. elegans. These signaling pathways are conserved in parasitic nematodes and an increasing body of evidence supports their function in formation and developmental regulation of L3i during the infectious process in soil transmitted species. This review presents these lines of evidence for G protein-coupled receptor (GPCR), insulin- and TGFβ-like signaling in brief and focuses primarily on signaling through parasite orthologs of DAF-12. We discuss in some depth the deployment of sensitive analytical techniques to identify Δ7-dafachronic acid as the natural ligand of DAF-12 homologs in Strongyloides stercoralis and Haemonchus contortus and of targeted mutagenesis by CRISPR/Cas9 to assign dauer-like regulatory function to the NR Ss-DAF-12, its coactivator Ss-DIP-1 and the key ligand biosynthetic enzyme Ss-CYP-22a9. Finally, we present published evidence of the potential of Ss-DAF-12 signaling as a chemotherapeutic target in human strongyloidiasis.

Monacrosporium sinense and Pochonia chlamydosporia for the biological control of bovine infective larvae in Brachiaria brizantha pasture

Ecologically correct solutions are necessary to minimize the use of anthelmintics in the control of cattle parasitic diseases. This study evaluated the effects of oral administration pellets containing the fungal combination Monacrosporium sinense and Pochonia chlamydosporia in the biological control of infective larvae (L3) of parasitic gastrointestinal nematodes of cattle grazed in pasture. Twelve crossbred bovines (Holstein × Zebu), seven to nine years old, after dewormed, were randomly divided into two groups (group treated with the combination of fungi and control group) and placed in separate paddocks of Brachiaria brizantha that were naturally infested with L3. Feces samples were collected from the animals to determine the parasitic load and pasture samples were collected from the soil to determine the L3 infestation. The recovery of L3 from the pasture of the treated group was 91.7% and 86.4% less than in the pasture of the control group, respectively for samples collected at 20 cm and 40 cm from the faecal pats. The values of eggs per gram of feces in the months of May 2019 and January 2020 were lower (p ≤ 0.05) in the treated group with the fungal combination. The combined administration of M. sinense and P. chlamydosporia reduced the L3 infestation on pasture and in animal parasitic load, making it a tool for controlling bovine gastrointestinal parasitic nematodes.

Immune serum-activated human macrophages coordinate with eosinophils to immobilize Ascaris suum larvae.

Helminth infection represents a major health problem causing approximately 5 million disability-adjusted life years worldwide. Concerns that repeated anti-helminthic treatment may lead to drug resistance render it important that vaccines are developed but will require increased understanding of the immune-mediated cellular and antibody responses to helminth infection. IL-4 or antibody-activated murine macrophages are known to immobilize parasitic nematode larvae, but few studies have addressed whether this is translatable to human macrophages. In the current study, we investigated the capacity of human macrophages to recognize and attack larval stages of Ascaris suum, a natural porcine parasite that is genetically similar to the human helminth Ascaris lumbricoides. Human macrophages were able to adhere to and trap Asuum larvae in the presence of either human or pig serum containing Ascaris-specific antibodies and other factors. Gene expression analysis of serum-activated macrophages revealed that CCL24, a potent eosinophil attractant, was the most upregulated gene following culture with Asuum larvae in vitro, and human eosinophils displayed even greater ability to adhere to, and trap, Asuum larvae. These data suggest that immune serum-activated macrophages can recruit eosinophils to the site of infection, where they act in concert to immobilize tissue-migrating Ascaris larvae.

The Viability of Haemonchus Contortus (Nematoda, Strongylida) and Strongyloides Papillosus (Nematoda, Rhabditida) Larvae Exposed to Various Flavourings and Source Materials Used in Food Production

Abstract The objective of this study is to evaluate the viability of larval nematodes of ungulates under the infl uence of 14 fl avourings and source materials approved for use in and on foods. Minimum LD50 for L3 Strongyloides papillosus (Wedl, 1856) were observed when using Cinnamaldehyde, α-Terpineol and Benzyl alcohol, for L1–2 S. papillosus under the infl uence of Benzyl alcohol, Cinnamaldehyde, L-Linalool and Benzyl acetate, for L3 Haemonchus contortus (Rudolphi, 1803) under the influence γ-Undecalactone and Cinnamaldehyde. Ethyl acetate, β-Ionone и D-Limonene had the lowest effect on all the studied stages of development of the nematodes: larvae did not die during 24 hours even at 1 % concentration of these substances. Further experiments with usage of mixtures of the substances studied in this article will contribute to the development of preparations with a stronger effect on larvae of nematode parasites of the digestive system of vertebrate animals.

Nematocidial activity of aqueous solutions of plants of the families Cupressaceae, Rosaceae, Asteraceae, Fabaceae, Cannabaceae and Apiaceae

In natural ecosystems of animals, introduction of larvae of parasitic nematodes into the litter layer from excrement facilitates their migration and search of new host vertebrate animals. In such conditions they are constantly affected by biologically active substances of the abundant species of plants which grow in pastures. Currently, the influence of substances present in the above-ground part of plants on the vitality of larvae of helminths in the environment remains unstudied. In this article, we present the results of our research on the nematocidial activity in vitro in the aqueous solutions of 21 species of plants distributed in the territory of Steppe Ukraine: Sanguisorba officinalis L., Rosa canina L., Crataegus sanguinea Pall., Crataegus pentagyna Waldst. &amp; Kit. ex Willd., Armeniaca vulgaris Lam., Taraxacum officinale F. H. Wigg., Iva xanthiifolia Nutt., Artemisia campestris L., Arctium minus (Hill) Bernh., Ambrosia artemisiifolia L., Cannabis sativa L., Humulus lupulus L., Melilotus officinalis (L.) Pall., Vicia cracca L., Lotus ucrainicus Klok., Onobrychis arenaria (Kit.) DC., Foeniculum vulgare Mill., Eryngium planum L., Conium maculatum L., Juniperus communis L., Thuja occidentalis L. The level of vitality of nematode larvae of the Strongylida (Haemonchus contortus (Rudolphi, 1803)) and Rhabditida (Strongyloides papillosus (Wedl, 1856)) orders varied depending on the species of plant, and also five experimental concentrations in aqueous solutions tested in seven replications. The most notable nematocidial effect was exerted by Taraxacum officinale F. H. Wigg. – we observed death of larvae of third stage development L3 H. contortus and larvae of first-third stages L1–3 S. papillosus at 24 h exposure to 3% aqueous solution. Three percent aqueous solutions of S. officinalis and A. artemisiifolia displayed nematocidial properties only against S. papillosus: death of L1–3 S. papillosus was observed. Aqueous solutions of R. canina, A. vulgaris, A. minus, H. lupulus, V. cracca, L. ucrainicus, O. arenaria, E. planum, C. maculatum, J. communis, Th. occidentalis had lethal effect only on non-invasive larvae (larvae of the first and second stage L1–2) of S. papillosus. They displayed no nematocidial properties towards invasive larvae of H. contortus and S. papillosus. At exposure to aqueous solutions of the rest of the studied species of plants, over 50% of L3 H. contortus and L1–3 S. papillosus larvae remained alive. The determined patterns allow us to state that while living in the litter and soil in the root zone of plants nematode larvae undergoa negative influence caused by some plant species.

Efficacy of Origanum syriacum Essential Oil against the Mosquito Vector Culex quinquefasciatus and the Gastrointestinal Parasite Anisakis simplex, with Insights on Acetylcholinesterase Inhibition.

Developing effective and eco-friendly antiparasitic drugs and insecticides is an issue of high importance nowadays. In this study, we evaluated the anthelminthic and insecticidal potential of the leaf essential oil obtained from Origanum syriacum against the L3 larvae of the parasitic nematode Anisakis simplex and larvae and adults of the mosquito Culex quinquefasciatus. Tests on A. simplex were performed by standard larvicidal and penetration assays, while mosquito toxicity was assessed relying on larvicidal, tarsal contact, and fumigation tests. To shed light on the possible mode of action, we analyzed the oil impact as acetylcholinesterase (AChE) inhibitor. This oil was particularly active on L3 larvae of A. simplex, showing a LC50 of 0.087 and 0.067 mg mL−1 after 24 and 48 h treatment, respectively. O. syriacum essential oil was highly effective on both larvae and adults of C. quinquefasciatus, showing LC50 values of 32.4 mg L−1 and 28.1 µg cm−2, respectively. Its main constituent, carvacrol, achieved larvicidal LC50(90) of 29.5 and 39.2 mg L−1, while contact toxicity assays on adults had an LC50(90) of 25.5 and 35.8 µg cm−2, respectively. In fumigation assays, the LC50 was 12.1 µL L−1 after 1 h and decreased to 1.3 µL L−1 in 24 h of exposure. Similarly, the fumigation LC50 of carvacrol was 8.2 µL L−1 after 1 h of exposure, strongly decreasing to 0.8 µL L−1 after 24 h of exposure. These results support the folk usage of Lebanese oregano as an antiparasitic agent, providing new insights about its utilization for developing new effective and eco-friendly nematocidal and insecticidal products.

Association and predatory capacity of fungi Pochonia chlamydosporia and Arthrobotrys cladodes in the biological control of parasitic helminths of bovines.

Nematophagous fungi are used in the biological control of the parasitic helminths of animals and plants. The association of ovicidal and predator nematophagous fungi may present a complementary and increased action on the biological control of helminths. Joint growth compatibility and predation tests were carried out on infective larvae of nematode parasites of bovines with the nematophagous fungus ovicide Pochonia chlamydosporia and the nematophagous fungus predator Arthrobotrys cladodes. The tests of antagonism in direct confrontation, antibiosis and the effect of volatile metabolites between the isolates of P. chlamydosporia and A. cladodes indicated the viability of joint growth of these fungi. The association of the fungi P. chlamydosporia and A. cladodes presented a higher predatory capacity of infective larvae of the parasitic nematodes of bovines when compared to the predation of the fungi used alone. Therefore, under laboratory conditions, the fungi studied presented growth compatibility and the association of these increased the nematicidal activity against parasitic helminths of cattle.

The nematophagous fungus Duddingtonia flagrans reduces the gastrointestinal parasitic nematode larvae population in faeces of orally treated calves maintained under tropical conditions—Dose/response assessment

This research assessed the dose/response to Duddingtonia flagrans chlamydospores (Df-C) administered to calves naturally infected with gastrointestinal nematodes (GINs), and its effect in reducing the infective larvae (L3) population in faeces in a farm in the Mexican tropics. Forty zebu calves, between 6 and 12 months of age, were randomly distributed into four groups of 10 calves. One control and three groups treated with different oral doses of Df-C based on their body weight (BW) were established as follows: group 1 (control); group 2, 0.250 × 106Df-C per kg/BW; group 3, 0.5 × 106Df-C per kg/BW and group 4, 1 × 106Df-C per kg/BW. The fungal doses were administered daily for 10 days. Every group was confined to individual pens, and they received a nutritional regime based on Buffel grass, concentrated supplement and water ad libitum. Every third day, starting one week before treatments, faeces were taken from the rectum of each animal to determine the number of eggs per g of faeces (epg) through the McMaster technique. Four coprocultures of 20 g each from each individual faecal sample were prepared and incubated for 14 days. The efficacy of the treatments was based on the mean of the GIN L3 recovered from coprocultures of the different groups. Data were analysed using a completely randomized design through an ANOVA analysis, followed by a Duncan multiple range test. The efficacy of treatments was expressed as the larval reduction rate. High variation in the epg in the different groups along the experiment was recorded. The reduction in the GIN L3 population was observed from the 4 to 11 day post-treatment in the three assessed doses. Results in group 2 (lowest fungal dose), showed 88.5, 57.6, 55.9 and 30% (58% overall reductions) in the GIN L3 in the faeces of animals 4, 7, 9 and 11 days post-treatment, respectively. In group 3 (medium fungal dose), 95.8, 80.4, 63.4 and 52.7% GIN L3 reductions (73% overall reduction) were recorded, respectively. At the highest Df-C dose used (1 × 106 per kg/BW), the results were 88.9, 78.0, 59.3 and 67.3% (73.5% overall reduction), respectively (p < 0.05). The species of identified nematodes through L3 morphometric and molecular taxonomy were Cooperia spp. and H. contortus. From the three Df-C assessed doses, the medium dose (0.5 × 106Df-C per kg/BW) was sufficient to substantially reduce the GIN L3 in zebu calves maintained under conditions in the Mexican tropics.

Using the fungus Arthrobotrys cladodes var. macroides as a sustainable strategy to reduce numbers of infective larvae of bovine gastrointestinal parasitic nematodes.

Research in the area of sanitation in ruminant production has focused on discovery of potential agents for biological control of helminths with nematophagous fungi and has provided evidence of success. The antagonistic potential of the fungus Arthrobotrys cladodes var. macroides on infective larvae of bovine gastrointestinal parasitic nematodes was evaluated by scanning electron microscopy. Additionally, an in vivo test of the resistance to digestive processes and viability of the fungus was carried out using a formulation based on sodium alginate administered orally in cattle. Production of conidia and chlamydospores was high. In in vitro tests, the number of infective nematode larvae was reduced 68.7% by the fungus in the treated group compared to the control group. The interaction between the fungus and the nematodes was confirmed by scanning electron microscopy. Plates containing fecal samples collected after oral administration of 100 g of pellets containing the A. cladodes fungus showed that the fungus survived passage through the gastrointestinal tract of ruminants, grew on agar, formed traps and preyed on L3 larvae of gastrointestinal parasites. The results of the present study provide a new opportunity for alternative, environmentally safe control of ruminant nematodes.

Field evaluation of Duddingtonia flagrans IAH 1297 for the reduction of worm burden in grazing animals: Tracer studies in sheep

The aim of these studies was to determine the reduction in pasture infectivity likely to be achieved by the supplementation of grazing sheep with BioWorma®, a product containing the chlamydospores of the nematophagous fungus Duddingtonia flagrans strain IAH 1297. Four placebo-controlled trials were conducted between 2009 and 2013 in sheep in different climatic regions of New South Wales and Queensland, Australia and across several seasons. The effectiveness of BioWorma was assessed by total worm counts in tracer sheep placed in paddocks grazed by parasitised sheep which were fed a daily supplement with and without BioWorma under group-feeding conditions. Further proof of concept was obtained by assessing the worm burdens and weight gains of the parasitised sheep, as well as the number of anthelmintic (“salvage”) treatments required when faecal egg counts exceeded a threshold level.Significant reductions ranging from 57 to 84% (P < 0.05) in worm burdens of the tracer sheep placed in the paddock grazed by BioWorma treated sheep were obtained in all four trials, compared to the Control group. In two of the studies the treatment effect was greater at the end of the trial, indicating that pasture infectivity in the Control paddocks had risen considerably. The main nematodes encountered were Haemonchus spp., Trichostrongylus spp., and Teladorsagia spp. (including multi-resistant strains) and significant reductions were demonstrated for each of these species.Given the results of the four trials it can be concluded that supplementation of pastured sheep with BioWorma was effective in reducing the numbers of parasitic nematode larvae ingested by tracer sheep. It is considered that these levels of reduced pasture larvae would result in productivity increases in grazing sheep and reduce the requirement for intervention with anthelmintic chemicals. Therefore, use of BioWorma will provide an alternative means for control of gastrointestinal nematode (GIN) parasites on pasture.

WITHDRAWN: The nematophagous fungus Duddingtonia flagrans reduces the gastrointestinal parasitic nematode larvae population in faeces of orally treated calves maintained under tropical conditions. Dose/Response assessment

Abstract This research assessed the dose/response to Duddingtonia flagrans chlamydospores (Df-C) administered to calves naturally infected with gastrointestinal nematodes (GINs), and its effect in reducing the infective larvae (L3) population in faeces in a farm in the Mexican tropics. Forty zebu calves, between 6 and 12 months of age, were randomly distributed into four groups of 10 calves. One control and three groups treated with different oral doses of Df-C based on their body weight (BW) were established as follows: group 1 (control); group 2, 0.250 × 106 Df-C per kg/BW; group 3, 0.5 × 106 Df-C per kg/BW and group 4, 1 × 106 Df-C per kg/BW. The fungal doses were administered daily for 10 days. Every group was confined to individual pens, and they received a nutritional regime based on Buffel grass, concentrated supplement and water ad libitum. Every third day, starting one week before treatments, faeces were taken from the rectum of each animal to determine the number of eggs per g of faeces (epg) through the McMaster technique. Four coprocultures of 20 g each from each individual faecal sample were prepared and incubated for 14 days. The efficacy of the treatments was based on the mean of the GIN L3 recovered from coprocultures of the different groups. Data were analysed using a completely randomized design through an ANOVA analysis, followed by a Duncan multiple range test. The efficacy of treatments was expressed as the larval reduction rate. High variation in the epg in the different groups along the experiment was recorded. The reduction in the GIN L3 population was observed from the 4 to 11 day post-treatment in the three assessed doses. Results in group 2 (lowest fungal dose), showed 88.5, 57.6, 55.9 and 30% (58% overall reductions) in the GIN L3 in the faeces of animals 4, 7, 9 and 11 days post-treatment, respectively. In group 3 (medium fungal dose), 95.8, 80.4, 63.4 and 52.7% GIN L3 reductions (73% overall reduction) were recorded, respectively. At the highest Df-C dose used (1 × 106 per kg/BW), the results were 88.9, 78.0, 59.3 and 67.3% (73.5% overall reduction), respectively (p

Motility in the L3 stage is a poor phenotype for detecting and measuring resistance to avermectin/milbemycin drugs in gastrointestinal nematodes of livestock

Motility is a commonly used in vitro phenotype for assessing anthelmintic activity of candidate compounds, and for detecting anthelmintic resistance in nematodes. Third-stage larvae (L3) of parasitic nematodes are commonly used in motility-based assays because L3 are simple to obtain and can remain viable in storage for extended periods. To improve the measurement of motility of microscopic stages of nematodes, our laboratory developed the Worminator, which quantitatively measures motility of parasites. Using the Worminator, we compared the dose-response characteristics of several avermectin/milbemycin (AM) compounds using L3 from both AM-susceptible and AM-resistant Cooperia spp. (abamectin, doramectin, eprinomectin, ivermectin, moxidectin) and Haemonchus contortus (eprinomectin, ivermectin, moxidectin). Concentrations tested with the Worminator ranged from 0.156 to 40 μM. Differences in EC50 between AM-susceptible and AM-resistant isolates of Cooperia spp. and Haemonchus contortus were small, with resistance ratios ranging from 1.00 to 1.34 for Cooperia spp., 0.99 to 1.65 for Haemonchus contortus. Larval migration inhibition assays were conducted using the same isolates and were equally ineffective for detection of resistance with resistance ratios less than 2.0. These results contrast with those of the Larval Development Assay where we obtained a resistance ratio of 16.48 using the same isolates of Haemonchus contortus. Moreover, even at the highest concentration tested (40 μM), 100% inhibition of motility was never achieved and EC50 for Worminator assays were more than 100× higher than peak plasma levels achieved in vivo following treatment. These data demonstrate that dose-response characteristics for inhibition of motility in L3 of gastrointestinal nematodes of livestock do not significantly differ for AM-susceptible and AM-resistant isolates. These data challenge the suitability of motility as a phenotype for detecting and measuring resistance to AM drugs in gastrointestinal nematodes of livestock.

Signaling in Parasitic Nematodes: Physicochemical Communication Between Host and Parasite and Endogenous Molecular Transduction Pathways Governing Worm Development and Survival.

Signaling or communication between host and parasite may occur over relatively long ranges to enable host finding and acquisition by infective parasitic nematode larvae. Innate behaviors in infective larvae transmitted from the soil that enhance the likelihood of host contact, such as negative geotaxis and hypermotility, are likely mediated by mechanoreception and neuromuscular signaling. Host cues such as vibration of the substratum, elevated temperature, exhaled CO2, and other volatile odorants are perceived by mechanosensory and chemosensory neurons of the amphidial complex. Beyond this, the molecular systems that transduce these external cues within the worm are unknown at this time. Overall, the signal transduction mechanisms that regulate switching between dauer and continuous reproductive development in Caenorhabditis elegans, and doubtless other free-living nematodes, have provided a useful framework for testing hypotheses about how the morphogenesis and development of infective parasitic nematode larvae and the lifespan of adult parasites are regulated. In C. elegans, four major signal transduction pathways, G protein-coupled receptor signaling, insulin/insulin-like growth factor signaling, TGFβ-like signaling and steroid-nuclear hormone receptor signaling govern the switch between dauer and continuous development and regulate adult lifespan. Parasitic nematodes appear to have conserved the functions of G-protein-coupled signaling, insulin-like signaling and steroid-nuclear hormone receptor signaling to regulate larval development before and during the infective process. By contrast, TGFβ-like signaling appears to have been adapted for some other function, perhaps modulation of the host immune response. Of the three signal transduction pathways that appear to regulate development in parasitic nematodes, steroid-nuclear hormone signaling is the most straightforward to manipulate with administered small molecules and may form the basis of new chemotherapeutic strategies. Signaling between parasites and their hosts' immune systems also occurs and serves to modulate these responses to allow chronic infection and down regulate acute inflammatory responses. Knowledge of the precise nature of this signaling may form the basis of immunological interventions to protect against parasitism or related lesions and to alleviate inflammatory diseases of various etiologies.

Identification of Peptide Mimics of a Glycan Epitope on the Surface of Parasitic Nematode Larvae.

Phage display was used to identify peptide mimics of an immunologically protective nematode glycan (CarLA) by screening a constrained C7C peptide library for ligands that bound to an anti-CarLA mAb (PAB1). Characterisation of these peptide mimotopes revealed functional similarities with an epitope that is defined by PAB1. Mimotope vaccinations of mice with three selected individual phage clones facilitated the induction of antibody responses that recognised the purified, native CarLA molecule which was obtained from Trichostrongylus colubriformis. Furthermore, these mimotopes are specifically recognised by antibodies in the saliva of animals that were immune to natural polygeneric nematode challenge. This shows that antibodies to the PAB1 epitope form part of the mucosal polyclonal anti-CarLA antibody response of nematode immune host animals. This demonstrates that the selected peptide mimotopes are of biological relevance. These peptides are the first to mimic the PAB1 epitope of CarLA, a defined larval glycan epitope which is conserved between many nematode species.