Major Gastrointestinal Nematodes Research Articles

The identification of small molecule inhibitors with anthelmintic activities that target conserved proteins among ruminant gastrointestinal nematodes.

Gastrointestinal nematode (GIN) infections are a major concern for the ruminant industry worldwide and result in significant production losses. Naturally occurring polyparasitism and increasing drug resistance that potentiate disease outcomes are observed among the most prevalent GINs of veterinary importance. Within the five major taxonomic clades, clade Va represents a group of GINs that predominantly affect the abomasum or small intestine of ruminants. However, the development of effective broad-spectrum anthelmintics against ruminant clade Va GINs has been challenged by a lack of comprehensive druggable genome resources. Here, we first assembled draft genomes for three clade Va species (Cooperia oncophora, Trichostrongylus colubriformis, and Ostertagia ostertagi) and compared them with closely related ruminant GINs. Genome-wide phylogenetic reconstruction showed a relationship among ruminant GINs structured by taxonomic classification. Orthogroup (OG) inference and functional enrichment analyses identified 220 clade Va-specific and Va-conserved OGs, enriched for functions related to cell cycle and cellular senescence. Further transcriptomic analysis identified 61 taxonomically and functionally conserved clade Va OGs that may function as drug targets for new broad-spectrum anthelmintics. Chemogenomic screening identified 11 compounds targeting homologs of these OGs, thus having potential anthelmintic activity. In in vitro phenotypic assays, three kinase inhibitors (digitoxigenin, K-252a, and staurosporine) exhibited broad-spectrum anthelmintic activities against clade Va GINs by obstructing the motility of exsheathed L3 (xL3) or molting of xL3 to L4. These results demonstrate valuable applications of the new ruminant GIN genomes in gaining better insights into their life cycles and offer a contemporary approach to discovering the next generation of anthelmintics.IMPORTANCEGastrointestinal nematode (GIN) infections in ruminants are caused by parasites that inhibit normal function in the digestive tract of cattle, sheep, and goats, thereby causing morbidity and mortality. Coinfection and increasing drug resistance to current therapeutic agents will continue to worsen disease outcomes and impose significant production losses on domestic livestock producers worldwide. In combination with ongoing therapeutic efforts, advancing the discovery of new drugs with novel modes of action is critical for better controlling GIN infections. The significance of this study is in assembling and characterizing new GIN genomes of Cooperia oncophora, Ostertagia ostertagi, and Trichostrongylus colubriformis for facilitating a multi-omics approach to identify novel, biologically conserved drug targets for five major GINs of veterinary importance. With this information, we were then able to demonstrate the potential of commercially available compounds as new anthelmintics.

Mucosal Immunity of Major Gastrointestinal Nematode Infections in Small Ruminants Can Be Harnessed to Develop New Prevention Strategies.

Gastrointestinal parasitic nematode (GIN) infections are the cause of severe losses to farmers in countries where small ruminants such as sheep and goat are the mainstay of livestock holdings. There is a need to develop effective and easy-to-administer anti-parasite vaccines in areas where anthelmintic resistance is rapidly rising due to the inefficient use of drugs currently available. In this review, we describe the most prevalent and economically significant group of GIN infections that infect small ruminants and the immune responses that occur in the host during infection with an emphasis on mucosal immunity. Furthermore, we outline the different prevention strategies that exist with a focus on whole and purified native parasite antigens as vaccine candidates and their possible oral-nasal administration as a part of an integrated parasite control toolbox in areas where drug resistance is on the rise.

A new diagnostic approach to fast-track and increase the accessibility of gastrointestinal nematode identification from faeces: FECPAKG2 egg nemabiome metabarcoding

Effective gastrointestinal nematode management in livestock industries is becoming increasingly difficult due to the rise of anthelmintic resistance and changes in the temporal and geographical distribution of major gastrointestinal nematodes. Underpinning the response to these challenges is the need for a fast-tracked diagnostic identification technique, making it easier for livestock producers to make informed gastrointestinal nematode management decisions. The traditional ‘gold-standard’ approach, larval culture followed by morphological differentiation, is laborious and potentially inaccurate. We developed a new diagnostic approach to identify gastrointestinal nematodes that integrates a remote-location digital faecal egg count platform, FECPAKG2, with internal transcribed spacer 2 (ITS2) nemabiome metabarcoding. The technique involves a quick and simple protocol to harvest concentrated strongyle eggs from the FECPAKG2 cassette utilising a repurposed pipette tip, followed by DNA isolation and Illumina next generation amplicon sequencing. The gastrointestinal nematode compositions and alpha diversity generated by our FECPAKG2 egg nemabiome metabarcoding approach was not significantly different to traditional morphological larval differentiation and nemabiome metabarcoding of larval and faecal samples. We demonstrated that storing FECPAKG2 harvested eggs in either DNA isolation lysis buffer or 80% ethanol (v/v) had no impact on gastrointestinal nematode identification outcomes for at least 60 days; enabling the transport of biological samples from their remote origins to a molecular diagnostic facility for nemabiome metabarcoding, in the absence of a cold chain. We discovered that sustained gastrointestinal nematode egg embryonation in the lysis buffer storage solution lead to higher yields of DNA compared with ethanol-stored gastrointestinal nematode eggs or faeces with gastrointestinal nematode eggs. Taking advantage of an already well-established platform such as FECPAKG2, and providing the livestock producers that use it with the option to identify gastrointestinal nematodesin their samples and contribute to large-scale gastrointestinal nematode distribution and/or anthelmintic resistance surveys, is an important future direction for the FECPAKG2 egg nemabiome metabarcoding approach.

Study on Prevalence of Major Gastrointestinal Nematodes of Sheep in Wayu Tuka and Diga District, Oromia Regional State

Aim This study was devised to be conducted to determine the prevalence and identify the major gastrointestinal nematode parasites based on fecal examination. Methods A cross-sectional study was carried out to determine the prevalence and associated factors with ovine gastrointestinal nematode infestation by fecal examination of 384 sheep from Wayu Tuka and Diga district, Eastern Oromia regional state. Results Out of the total 384 sampled sheep, 169 (44.0%) had been infected with gastrointestinal nematode parasite. Haemonchus species were the most frequently (20.8%) recovered nematode eggs followed by Trichostrongylus (13.0%) and Nematodirus (10.2%). There was no significant difference (p>0.05) in prevalence between age groups and sex. Sheep with poor body condition had a significantly higher prevalence of gastrointestinal nematode parasite (p<0.05) than those sheep in moderate or good body condition. There was no significant association between the gastrointestinal nematode infection in animals of different ages and sex groups. Conclusion The study shows that the gastrointestinal nematode parasite was a major important health problem and impact on the production of sheep in the study area. Therefore, a detailed study should be conducted to identify the parasite at the species level and special consideration should be taken on the management of sheep in poor body condition to reduce the burden of gastrointestinal nematodes.

Nematocidal Effects of a Coriander Essential Oil and Five Pure Principles on the Infective Larvae of Major Ovine Gastrointestinal Nematodes In Vitro.

The anthelmintic effects of extracted coriander oil and five pure essential oil constituents (geraniol, geranyl acetate, eugenol, methyl iso-eugenol, and linalool) were tested, using larval motility assay, on the third-stage larvae (L3s) of Haemonchus contortus, Trichostrongylus axei, Teladorsagia circumcincta, Trichostrongylus colubriformis, Trichostrongylus vitrinus and Cooperia oncophora. Coriander oil and linalool, a major component of tested coriander oil, showed a strong inhibitory efficacy against all species, except C. oncophora with a half maximal inhibitory concentration (IC50) that ranged from 0.56 to 1.41% for the coriander oil and 0.51 to 1.76% for linalool. The coriander oil and linalool combinations conferred a synergistic anthelmintic effect (combination index [CI] <1) on larval motility comparable to positive control (20 mg/mL levamisole) within 24 h (p < 0.05), reduced IC50 values to 0.11–0.49% and induced a considerable structural damage to L3s. Results of the combined treatment were validated by quantitative fluorometric microplate-based assays using Sytox green, propidium iodide and C12-resazurin, which successfully discriminated live/dead larvae. Only Sytox green staining achieved IC50 values comparable to that of the larval motility assay. The cytotoxicity of the combined coriander oil and linalool on Madin–Darby Canine Kidney cells was evaluated using sulforhodamine-B (SRB) assay and showed no significant cytotoxic effect at concentrations < 1%. These results indicate that testing essential oils and their main components may help to find new potential anthelmintic compounds, while at the same time reducing the reliance on synthetic anthelmintics.

Increasing importance of anthelmintic resistance in European livestock: creation and meta-analysis of an open database.

Helminth infections are ubiquitous in grazing ruminant production systems, and are responsible for significant costs and production losses. Anthelmintic Resistance (AR) in parasites is now widespread throughout Europe, although there are still gaps in our knowledge in some regions and countries. AR is a major threat to the sustainability of modern ruminant livestock production, resulting in reduced productivity, compromised animal health and welfare, and increased greenhouse gas emissions through increased parasitism and farm inputs. A better understanding of the extent of AR in Europe is needed to develop and advocate more sustainable parasite control approaches. A database of European published and unpublished AR research on gastrointestinal nematodes (GIN) and liver fluke (Fasciola hepatica) was collated by members of the European COST Action “COMBAR” (Combatting Anthelmintic Resistance in Ruminants), and combined with data from a previous systematic review of AR in GIN. A total of 197 publications on AR in GIN were available for analysis, representing 535 studies in 22 countries and spanning the period 1980–2020. Reports of AR were present throughout the European continent and some reports indicated high within-country prevalence. Heuristic sample size-weighted estimates of European AR prevalence over the whole study period, stratified by anthelmintic class, varied between 0 and 48%. Estimated regional (country) prevalence was highly heterogeneous, ranging between 0% and 100% depending on livestock sector and anthelmintic class, and generally increased with increasing research effort in a country. In the few countries with adequate longitudinal data, there was a tendency towards increasing AR over time for all anthelmintic classes in GIN: aggregated results in sheep and goats since 2010 reveal an average prevalence of resistance to benzimidazoles (BZ) of 86%, macrocyclic lactones except moxidectin (ML) 52%, levamisole (LEV) 48%, and moxidectin (MOX) 21%. All major GIN genera survived treatment in various studies. In cattle, prevalence of AR varied between anthelmintic classes from 0–100% (BZ and ML), 0–17% (LEV) and 0–73% (MOX), and both Cooperia and Ostertagia survived treatment. Suspected AR in F. hepatica was reported in 21 studies spanning 6 countries. For GIN and particularly F. hepatica, there was a bias towards preferential sampling of individual farms with suspected AR, and research effort was biased towards Western Europe and particularly the United Kingdom. Ongoing capture of future results in the live database, efforts to avoid bias in farm recruitment, more accurate tests for AR, and stronger appreciation of the importance of AR among the agricultural industry and policy makers, will support more sophisticated analyses of factors contributing to AR and effective strategies to slow its spread.

Assessment of biophysical properties of Haemonchus contortus from different life cycle stages with atomic force microscopy

Atomic force microscopy (AFM) was used in this work to investigate the ultrastructural and mechanical characteristics of Haemonchus contortus, the major gastrointestinal nematode that infects small ruminants worldwide. The biophysical characterization of this species is extremely important in order to reveal mechanisms of action of drugs and to classify its ultrastructure and biomechanical properties. High-resolution topographic images by AFM as well as data on biomechanical properties of cuticles were obtained at different stages of H. contortus. The results reveal details of the mechanical and structural properties of this nematode never observed before for nematodes parasite with other microscope techniques. Qualitative and quantitative reductions in the elasticity of the larvae stage egg were compared with those of the morulae stage, and the increased adhesion of unsheathed L3 were compared with the same stage of sheathed larvae. The results presented here open possibilities for understanding the mechanisms of drug and biomolecular actions that can be used to control infections caused by H. contortus.

Validation of ITS-2 rDNA nemabiome sequencing for ovine gastrointestinal nematodes and its application to a large scale survey of UK sheep farms

We have validated ITS-2 rDNA nemabiome next-generation amplicon sequencing to determine relative species abundance of gastrointestinal nematode species in ovine fecal samples. In order to determine species representation biases, ITS-2 rDNA amplicon sequencing was applied to mock communities or field populations with known proportions of L3 for eight of the major ovine gastrointestinal nematode species: Teladorsagia circumcincta, Trichostrongylus vitrinus, Haemonchus contortus, Cooperia curticei, Trichostrongylus axei, Trichostrongylus colubriformis, Chabertia ovina and Oesophagostumum venulosum. Correction factors, calculated from this data, were shown to reduce species representation biases when applied to an independent set of field samples of known composition. We compared ITS-2 rDNA amplicon sequencing data that was generated from harvested eggs, freshly hatched L1 or L3 larvae following fecal culture and no statistically significant differences were found for the more abundant parasite species. We then applied the validated ITS-2 rDNA nemabiome amplicon sequencing assay to a set of archived L1 gastrointestinal nematode populations, collected in 2008 from fecal samples from 93 groups of 20 ewes and 61 groups of 20 lambs derived from 99 UK sheep farms. The presence of the major gastrointestinal nematode species had previously been determined on this large sample set by species-specific PCR. We show how the ITS-2rDNA amplicon sequencing data provided much more detailed information on species abundance than the previous species-specific PCR. This new data represents the most comprehensive overview of the relative abundance of the major gastrointestinal nematode species across UK sheep farms to date. Substantial variance in the relative abundance of both T. circumcincta and T. vitrinus between farms was revealed with the former species being of statistically significantly higher abundance in all three regions sampled (England, Scotland and Wales). The data also revealed that the relative abundance of T. circumcinta was statistically significantly higher in ewes than in lambs with the opposite pattern being the case for T. vitrinus. The nemabiome sequencing data also clearly illustrated the sporadic nature and skewed distribution of H. controtus across UK sheep farms as well as a higher relative abundance on farms from England compared to Wales and Scotland. The nemabiome survey also provides the first widescale data on the relative abundance of the two major large intestinal nematodes C. ovina and O. venulosum. This work validates ITS-2 rDNA nemabiome sequencing for use in sheep and illustrates the power of the approach for large scale surveillance of ovine gastrointestinal nematodes.

Resistance of Black Bengal goat to Haemonchus contortus

Artificial challenge trial and field trial were carried out in two populations of Black Bengal goat in Bangladesh to evaluate the resistance to Haemonchus Contortus-a major gastrointestinal nematode in goat. Artificial challenge trial revealed that there was no difference in susceptibility of H. contortus in Black Bengal goat of hilly region (BBH) and Black Bengal goat of western region (BBW). In absence of parasitic infection, growth rate of kids of both populations were almost equal. Field trial was conducted with kids of BBW population to find out the within breed variation. Numerically, there was variation among the individuals for parasitic load however the results were statistically insignificant (p &gt; 0.05). At individual level, the parasite load ranged from 100 to 400 eggs per gram (epg). The location of villages, age and sex of kids did not influence on the parasitic load of kids. There was no interaction for location, age and sex for body weight at day 8. The Packed Cell Volume and Hemoglobin values differed significantly (p &lt; 0.01) due to age of kids at day 8 and 28 after deworming. The results indicate that Black Bengal goat can be considered as resilience to H. contortus.&#x0D; Progressive Agriculture 30 (1): 55-64, 2019

A detection of benzimidazole resistance-associated SNPs in the isotype-1 \u03b2-tubulin gene in Haemonchus contortus from wild blue sheep (Pseudois nayaur) sympatric with sheep in Helan Mountains, China

BackgroundBenzimidazole (BZ) resistance is an increasingly serious problem due to the excessive use of this anthelmintic for controlling Haemonchus contortus, which is one of the major gastrointestinal nematodes infecting small ruminants worldwide. Three known single nucleotide polymorphisms (SNPs), F167Y (TAC), E198A (GCA) and F200Y (TAC), in the isotype-1 β-tubulin gene of H. contortus are associated with BZ resistance. Comprehending the spread and origins of BZ resistance-associated SNPs has important implications for the control of this nematode.ResultsTwenty-seven adult H. contortus were harvested from wild blue sheep (Pseudois nayaur), small wild ruminants sympatric with domestic ruminants, inhabiting the Helan Mountains, China, to monitor the status of BZ resistance. In addition, 20 adult H. contortus from domestic sheep sympatric with this wild ruminant and 36 isotype-1 β-tubulin haplotype sequences of H. contortus (two of these haplotypes, E198A3 and E198A4, possessed resistance-associated SNP E198A (GCA) from domestic ruminants in eight other geographical regions of China were used to further define the origins of BZ resistance-associated SNPs within the worms collected from blue sheep. The BZ resistance-associated SNP E198A was detected, whereas SNPs F167Y (TAC) and F200Y (TAC) were not found within the worms collected from blue sheep, and the frequency of homozygous resistant E198A (GCA) was 7.40%. The evolutionary tree and network showed consistent topologies for which there was no obvious boundary among the worms from the wild and domestic hosts, and two haplotypes (E198A1 and E198A2) possessing E198A from the wild blue sheep had two different independent origins. E198A1 had the same origin with E198A3 but E198A2 had a different origin with them. Population genetic analyses revealed a low level of Fst values (ranging from 0 to 0.19749) between all H. contortus worm groups in China.ConclusionsResults of the current study of the three BZ resistance-associated SNPs of H. contortus from wild blue sheep suggested that only E198A (GCA) was present within the worms collected from the wild ruminants and had multiple independent origins.

Prevalence of Major Gastrointestinal Nematode and Degree of Parasite Infestation in Sheep of Bako Agricultural Research Center Community Based Breeding Program Project Small Holder Farms at Horro District

Prevalence of Major Gastrointestinal Nematode and Degree of Parasite Infestation in Sheep of Bako Agricultural Research Center Community Based Breeding Program Project Small Holder Farms at Horro District

Droplet digital polymerase chain reaction (ddPCR) as a novel method for absolute quantification of major gastrointestinal nematodes in sheep

In this paper, we present for the first time a new tool, based on Droplet Digital™ Polymerase Chain Reaction (ddPCR), for absolute quantification of key genera of gastrointestinal (GI) nematode parasites of grazing livestock. Four combinations of primers/probe sets targeting the internal transcribed spacer region 2 (ITS2) of the ribosomal RNA gene array were designed using the Primer3 software, following in silico analysis of nucleotide sequences from nematodes of interest downloaded from common databases. The amplified regions include both a universal region for detection of any strongylid gastrointestinal parasite and three different genus specific regions, making it possible to differentiate between the most important GI nematodes of sheep in Sweden: Haemonchus, Teladorsagia and Trichostrongylus. Analysis of samples containing serial dilutions and different mixtures of genomic DNA extracted from different species of adult worms proved useful in assessment of different threshold settings with the QuantaSoft software. Analysis of template DNA from these worms indicated that ddPCR is a viable choice for detection and absolute quantification of the different genera and also in samples with multiple species. Interpretation of the ddPCR results was straightforward and choice of analytical approach had little influence on the final results. Thus, the results obtained in the different analytical approaches seemed to be robust and the concentrations determined were uniform. Furthermore, the linear range of the Haemonchus ddPCR assay was similar to that of real-time PCR (qPCR). Taken together, our data confirm the suitability of ddPCR for detection and absolute quantification of three major sheep pathogens when tested on larval cultures from pooled ovine faeces. The results also indicate that ddPCR can be a useful complement to applications based on conventional egg counting methods such as the faecal egg reduction test (FECRT).

English

A study by qualitative fecal examination of 384 fecal samples (201 sheep and 183 goats) was conducted from November 2011 to April 2012 with the objective to determine the major gastrointestinal (GIT) nematodes of small ruminants and their prevalence in sheep and goats in Guto Gida District. The study showed that 186 (92.5%) sheep and 150 (81.97%) goats were found to harbor eggs of GIT nematodes. Both sheep and goats were infected with identical parasites species, but with different level of infection. The six genera of nematodes were identified with prevalence of 21.87, 14.87, 12.5, 10.67, 11.19 and 7.29% for Haemonchus, Trichostrongylus, Trichuris, Oesophagostomum, Bunostumum and Strongloides, respectively. There was a significant difference (p<0.05) in the prevalence of GIT nematodes between sex and species of animals but not for different age group. The study showed that GIT nematodes are major problems of small ruminants in the study area. Therefore, comprehensive study on GIT nematodes, cost effective control strategy and awareness creation to the farmers should be instituted in the area. Key words: Sheep, goats gastrointestinal nematodes, Guto Gida District, Nekemte.

Pharmacokinetics and anthelmintic efficacy of injectable eprinomectin in goats

The objective of this study was to assess the pharmacokinetics and the anthelmintic efficacy of eprinomectin (Eprecis® 20mg/mL) following subcutaneous administration to goats. Forty non-lactating female Alpine goats aged between one and three years and weighing between 32.7 and 59.5kg, were randomly allocated to one of the following groups (8 animals per group): two groups were not infected and were treated at a dose of either 0.2 or 0.4mg/kg BW of eprinomectin, two groups were experimentally infected with nematodes and treated at 0.2 or 0.4mg/kg BW of eprinomectin similarly and one group was infected and left untreated (control). Infection consisted in a single and simultaneous administration of 5000 Haemonchus contortus and 12,500 Trichostrongylus colubriformis infective larvae. No local or general adverse reaction was visually observed whatever the dose rate. The maximal plasma concentrations (Cmax) were 20.68±12.85 vs 39.79±17.25μg/L and the plasma bioavailabilities (AUC) 83.45±34.75 vs 169.37±43.44μg*d/L for 0.2 vs 0.4mg/kg respectively. The efficacy against H. contortus and T. colubriformis was 97.8 and 98.7% at 0.2mg/kg and 98.4% and >99.9% at 0.4mg/kg respectively. The differences in worm burdens between the two dose rates were only significant for T. colubriformis. These results indicate that injectable eprinomectin is a potent anthelmintic against the two major gastrointestinal nematodes in goats. Additional information is needed regarding pharmacokinetics in lactating goats and milk residues.

Variability in fecal egg count of Haemonchus contortus infection to native goat breeds of China and Bangladesh under natural grazing condition

Haemonchus contortus is a major gastrointestinal nematode affecting goat in China and Bangladesh like many countries in the world. The aim of this study was to evaluate the susceptibility of Haemonchus contortus infection in different goat breeds/populations in China and Bangladesh under the natural grazing condition. Fecal Egg of Haemonchus contortus was counted from 430 goats in both countries using McMaster’s technique. Body weight, Pack Cell Volume and Hemoglobin value were measured for Bangladeshi goats. Statistical analysis was done after transforming data into log10 (n+1), where n is the number of egg per gram feces and analysis of variance was done by using Generalized Linear Model procedures of computer package of SAS. Prevalence of parasite and Fecal Egg Count varied significantly (P&lt;0.01) in all six goat populations. The highest prevalence (88.89%) of Haemonchus contortus infection was found in Enshi Black and the highest parasitic load (527.74 ±78.13 epg) was found in Chinese hybrid goat population. Most of the individuals, except Chinese hybrid goats, had Fecal Egg Count less than 300 epg. Fecal Egg Count between sexes within breed of goat did not differ significantly (P&gt;0.05). Black Bengal goat in the hilly region of Bangladesh was the least susceptible to Haemonchus contortus infection in term of Fecal Egg Count. Our study suggests that Chinese goat breeds were more susceptible to Haemonchus contortus infection as compared to Black Bengal goat of Bangladesh under natural grazing condition. Further studies on immune-genetics aspects of goats might be helpful to find out the actual causes of such differences and to develop disease resistant breed of goat.Progressive Agriculture 27 (4): 473-481, 2016

Prevalence of ovine gastrointestinal nematodes in Meskan district, Gurage zone, Southern Ethioipa

A cross sectional study was conducted with the objectives of determining the prevalence and risk factors associated with ovine major gastrointestinal nematodes in Meskan district, Gurage zone from November 2014 to April 2015. A fresh faecal sample of approximately 10 g was collected directly from the rectum of 350 sheep by using gloved fingers. All statistical analysis was performed using statistical software packages for social science (SPSS) version 20. From the total 350 local sheep breed examined, 212 (60.6%) were found infected with different types of gastro-intestinal nematodes. Strongyles were the most frequently 95 (27.1%) recovered nematode eggs followed by Strongyloides 38 (10.9%), Trichuris species 13 (3.7%) and Ascaris spps 2 (0.6%). There was significant association (p 0.05) in prevalence between body condition and different study sites of the subject area. Due to its important health problem and impact on production in the study area, emphasis should be given for the control and prevention of gastrointestinal nematode infection with further studies on genus and species identification by using further laboratory techniques. Keywords : Nematode, Meskan distric, Gastrointestinal nematode, Prevalence, Sheep

Prevalence, intensity and risk factors of infestation with major gastrointestinal nematodes in equines in and around Shashemane, Southern Ethiopia.

Prevalence, intensity and risk factors of major gastrointestinal nematode infestation in equines were studied through a cross-sectional survey in 384 equids from October 2013 to April 2014 in and around Shashemane, southern Ethiopia. Three hundred and fifteen equids (82%) were demonstrated harbouring one or more gastrointestinal (GIT) nematodes using the faecal flotation technique. The prevalence of GIT nematode infestation was 73.4, 85 and 86.5% for horses, mules and donkeys, respectively. The identified nematodes were strongyle type (73.4%), Parascaris equorum (21.4%) and Oxyuris equi (4.4%). Species of equines had a significant (χ (2) = 9.35, P < 0.01) association with the occurrence of GIT nematode infestation. Donkeys were two times (OR = 2.3, 95% CI 1.27-4.28, P < 0.01) more likely getting GIT nematode infestation than horses. Moreover, donkeys had the highest mean faecal egg counts (1831.2 egg per gram (EPG)) followed by mules (915.7 EPG) and horses (772.5 EPG). There was a significant association (P < 0.05) between mean EPG and body condition score in each equine species. In conclusion, this study provides information which might help in designing upcoming control strategies to control nematode infestation in equines. Moreover, suitable tropical climatic conditions, low level of management and owners' awareness, and poor animal health services are expected to contribute for high nematode infestation. Therefore, emphasis should be given to awareness creation about the strategic deworming, animal welfare and management.

In vitro activity of essential oils of free and nanostructured Melaleuca alternifolia and of terpinen-4-ol on eggs and larvae of Haemonchus contortus.

Haemonchus contortus is one of the major gastrointestinal nematodes responsible for significant economic and production losses of sheep. Diseases caused by this species lack effective anthelmintic products, and the search for new compounds to replace synthetic anthelmintics has been extensive. The present investigation assesses the in vitro activity of the essential oil of melaleuca (Melaleuca alternifolia), both free (TTO) and nanostructured (nanoTTO), and terpinen-4-ol (terp-4-ol) on eggs and larvae of H. contortus. Tests of egg hatching (EHT) and inhibition of larval migration (LMIT) were used to assess the in vitro efficacy of TTO, nanoTTO and terp-4-ol. Using EHT, at a concentration of 3.5 mg/ml, 100% inhibition occurred using TTO and terp-4-ol, with LC50 values of 0.43 and 0.63 mg/ml, and LC90 values of 1.75 mg/ml and 3.12 mg/ml, respectively. NanoTTO had lower activity, with 82.6% inhibition at the same concentration. Using LMIT, TTO and nanoTTO had a similar activity with 88.0% and 84.8% inhibition, respectively, at a concentration of 56 mg/ml. Terp-4-ol had a greater effect on larvae, with 85.7% inhibition at a concentration of 56 mg/ml and 82.4% at 3.5 mg/ml, demonstrating high activity at the lowest concentration tested. Therefore, the results indicate that all substances tested showed ovicidal and larvicidal activity against H. contortus. TTO, terp-4-ol and, mainly, nanoTTO may be targeted in in vivo studies, besides being a promising line of research into the control and treatment of veterinary important helminths.

Acr-23 Encodes a Monepantel-Sensitive Channel in Caenorhabditis elegans

Monepantel is a member of the recently identified class of anthelmintics known as the amino-acetonitrile derivatives (AADs). Monepantel controls all major gastro-intestinal nematodes in sheep including those that are resistant to the classical anthelmintics. Previous studies have shown that the Caenorhabditis elegans acr-23 and the Haemonchus contortus Hco-mptl-1 genes may be prominent targets of monepantel. With this discovery it became possible to investigate the mode of action of monepantel in nematodes at the molecular level. In the present study, we show that a C. elegans mutant acr-23 strain is fully rescued by expressing the wild-type acr-23 gene. Moreover, we present a new mutant allele, and characterize acr-23 alleles genetically. We also show that acr-23 is expressed in body wall muscle cells, and provide therefore a possible explanation for the paralysis caused by monepantel. Furthermore, genetic evidence suggests that the chaperone RIC-3 is required for expression of full monepantel resistance. Finally, we present reconstitution of the C. elegans ACR-23 receptor in Xenopus laevis oocytes and provide direct evidence of its modulation by monepantel. Conversely, co-injection of the chaperone RIC-3 had no impact for channel reconstitution in X. laevis oocytes. These results reinforce the involvement of the ACR-23 family in the mode of action of monepantel and advance our understanding of this new class of anthelmintics.

Integrated parasite management: Products for adoption by the Australian sheep industry

The increasing cost of production loss caused by gastrointestinal nematodes (GIN) highlights the importance of good control programs. However, the endemic nature of anthelmintic resistance also reminds of the need for nonchemical options. Both chemical and nonchemical control options need to be integrated into regional parasite management programs (IPM) with the emphasis on component options determined by factors such as major GIN species, regional climate, property size and enterprise structure. The Integrated Parasite Management of Sheep project was established to develop and demonstrate regional parasite control programs, that integrated chemical and nonchemical options, for the main sheep-producing regions of Australia. The project included research about the ecology of the main endo and ecto-parasites of sheep and a national survey of parasite control practices by sheep producers. IPM approaches developed for two contrasting regions of Australia are discussed.Barriers for the adoption of IPM programs include perceived complexity associated with a multi-component approach, time requirements and difficulty. Facilitating the industry adoption of IPM programs is discussed with relevance to the use of small group extension and involvement of the commercial sector. Perceptions of complexity of IPM may be managed by facilitating adoption of components in a step-wise process such that learning outcomes accumulate over time. Extension efforts must address the needs of industry sectors other than sheep producers and explore user pay approaches. The success of these approaches will depend on the relation of the extra profit to producers, through adoption of IPM programs, with remuneration sufficient to attract a commercial service.