Faecal egg count reduction test in goats: Zooming in on the genus level.

The faecal egg count reduction test (FECRT) is the most widely used method to assess treatment efficacy against gastrointestinal nematodes (GIN). Information on genera composition of the GIN community is not available with this test and it is commonly obtained by identifying cultured third-stage larvae (L3) or through molecular assays in the post-treatment survey, but results provided are usually only qualitative or semi-quantitative. The updated WAAVP guidelines now recommend assessing anthelmintic efficacy for each GIN genus/species separately (genus-specific FECRT), but this approach is poorly employed in Europe and in goats especially. For this reason, four FECRT trials were conducted using oxfendazole and eprinomectin in two Italian goat farms. Samples were processed individually using the McMaster technique and then pooled to create two samples from faeces of 5 animals each. Pooled samples were analysed using the McMaster and cultured for seven days at 26°C to obtain L3s. The genus-specific FECRT was based on larval identification, integrating coproculture and FEC results. Larvae were identified as Haemonchus, Trichostrongylus, Teladorsagia, Oesophagostomum / Chabertia and Bunostomum. Molecular assays (a multiplex real-time PCR and two end-point PCRs) were also implemented on pooled samples to support the morphological identification. The Spearmann Rho test confirmed a high correlation between the two approaches (Rho = 0.941 and Rho = 0.914 respectively for Haemonchus and Trichostrongylus, the two most common genera). Both oxfendazole and eprinomectin were effective in one farm, while none in the other farm (FECR = 75.9% and 73.3% respectively). In the second farm, the genus-specific FECRT highlighted a different response to treatment among genera: oxfendazole lacked efficacy against both Haemonchus and Trichostrongylus spp., eprinomectin only against Haemonchus, while all other genera were susceptible to both drugs. This study brings new attention on the importance of adopting a genus-specific approach to identify and quantify differences in susceptibility to anthelmintics among genera in goats, providing support for FECRT interpretation, anthelmintic resistance evaluation and evidence-based GIN control.

An alien parasite in a changing world - Ashworthius sidemi has lost its traditional seasonal dynamics.

A non-native nematode Ashworthius sidemi has emerged in captive fallow deer in Central and Eastern Europe over the last decade. Although this parasite has been spreading in the wild outside it's native distributional range and colonising local European host species since the middle of the last century, limited information has been published on the seasonality of A. sidemi and its susceptibility to anthelmintics. To address this knowledge gap, we conducted a study to investigate seasonal dynamics of the non-native parasite in the current Central European climate conditions. We collected freshly voided faecal pellets at four-week intervals from February 2018 to February 2020 at a fallow deer reserve with a known history of A. sidemi presence. The faecal pellets obtained were pooled after each site visit (n = 25) and coprocultured to obtain the third stage larvae of trichostrongylid nematodes at monthly intervals. Total genomic DNA was extracted from the recovered larvae. Using real-time multiplex PCR, A. sidemi DNA was detected in 17 out of 25 larval samples (68% prevalence). During the monitoring period, the annual administration of ivermectin based premix (Cermix) took place in January 2018, 2019, and 2020, and additionally a mixture of rafoxanide and mebendazole (Rafendazol) was administered once in spring 2019. The probability of parasite presence was significantly influenced by the time since the drug administration (p = 0.048) and the mean temperature at the location (p = 0.013). Larval samples negative for A. sidemi were always identified shortly after the drug administration. However, rapid pasture contamination by the parasite eggs from two to three months after Cermix administration and within one month after Rafendazol administration suggest only a short-lived efficacy of both administered drugs. The abundance of A. sidemi DNA was positively affected by mean temperature (p = 0.044) and remained relatively stable throughout the monitoring period, with the highest peak in August 2018 and 2019. Pasture contamination with A. sidemi eggs occurred almost all year round, with the exception of the beginning of 2018, 2019, and 2020. These findings indicate adaptation of a non-native parasite to the current climatic conditions of the Czech Republic resulted in negligible seasonal patterns of parasite egg shedding.

Non-native Nematode Ashworthius sidemi Currently Dominates the Abomasal Parasite Community of Cervid Hosts in the Czech Republic.

Ashworthius sidemi is an abomasal nematode typical for Asiatic cervids such as sambar (Rusa unicolor) or sika deer (Cervus nippon). This non-native parasite was introduced into Europe via sika deer in the late 19th and early 20th centuries. The current dynamic spread of this parasite amongst autochthonous wild cervids occurs independently of human activities, and A. sidemi has a negative impact on the health of wild ruminants and may pose a threat to the conservation of endangered wild ungulates and to livestock. This invasive parasite has been previously detected in the Czech Republic, but more accurate information on A. sidemi is required. Only limited information is generally available on the factors influencing the spread of abomasal nematodes in wild ruminants, so more information is necessary for planning effective strategies of parasite control. We therefore conducted a survey on the abomasal nematodes in cervids in both game reserves and hunting grounds across the Czech Republic, taking into account the hosts (species, age, sex) and environmental factors (monthly average temperature). The abomasa of 104 animals belonging to five cervid species originating from various locations of the country were collected. Data on host (species, sex, and age group) and the monthly average temperature in the region were obtained for each animal. The parasitological analyses indicated that 92% of the abomasa were infected by nematodes. Ashworthius sidemi was the most prevalent (72%) and abundant (80% of the total recovered individuals) nematode species and was detected in all cervid species except white-tailed deer. The intensity of A. sidemi was highest in roe deer (Capreolus capreolus) and fallow deer (Dama dama), but A. sidemi abundance did not depend substantially on the host or environmental factors. In contrast, the abundance of nematodes from the subfamily Ostertagiinae was influenced by the host species and temperature. Parasitic load was significantly higher in roe deer and during the warmer periods of the survey. We also detected another non-native nematode species, Spiculopteragia houdemeri. The results of our study suggest that the non-native nematode A. sidemi is now widespread amongst cervid hosts in the Czech Republic, probably due to the high sensitivity of autochthonous cervids to A. sidemi infections as well as adaptation of this parasite to the current climatic conditions of this country.

Total Failure of Fenbendazole to Control Strongylid Infections in Czech Horse Operations.

The control of strongylid infections has become challenging globally for equine practitioners due to the development of anthelmintic resistance. Comprehensive information on anthelmintic resistance in the Czech Republic, however, is still lacking. This study monitored the current efficacy of fenbendazole, pyrantel embonate, ivermectin and moxidectin. Forty-eight of 71 operations met the criteria (≥6 horses with ≥200 eggs per gram), with 969 fecal egg count reduction tests performed. Anthelmintic resistance was evaluated on an operation level based on fecal egg count reduction (FECR) and the lower limit of the 95% credible interval (LLCI) using Bayesian hierarchical models. General anthelmintic efficacy across all operations was assessed by posterior FECRs and the occurrence of sub-zero efficacies. Ivermectin and moxidectin demonstrated excellent efficacy (FECR 99.8-100%; 99.4-100 LLCI) in 45 and 23 operations, respectively, pyrantel embonate demonstrated sufficient efficacy in 15 operations and resistance was suspected in seven operations (FECR 88.1-99.1%; 72.5-98.5 LLCI). Fenbendazole, however, was not effective in a single operation (FECR 19.1-77.8%; 8.1-50.1 LLCI) out of 18. Fenbendazole had the highest probability of sub-zero efficacy (29.1%), i.e., post-treatment fecal egg counts exceeded the pre-treatment counts. Our data indicate an increase in the development of anthelmintic resistance, resulting in total failure of fenbendazole and a reduced efficacy of pyrantel embonate. Introducing advanced approaches of parasite control in the Czech Republic to slow the spread of anthelmintic resistance is thus needed.

Do Live Weight, Body Condition Score, Back Muscle or Back-Fat Reserves Create the Suspicion of Goats Infected with Eimeria or Trichostrongylids?

Thirty goats of the breeds Czech Brown Shorthaired and Czech White Shorthaired and their crosses were randomly selected from a flock at a farm in the Czech Republic. All animals were monitored for one year at monthly intervals for their nutritional status (live weight, LW; body-condition score, BCS; depth of musculus longissimus thoracis et lumborum, MLTL; back-fat thickness, BT) and infection intensity with Eimeria sp. (EIM) and strongylid nematodes (STR). Regression-correlation analysis showed a possible interrelation of BCS with EIM infection. Analysis of muscle and fat reserves indicated that BT was better than MLTL in identifying EIM infection. Goat nutritional status was not significantly correlated with STR infection. A linear tendency (p = 0.092), however, was detected for the response of MLTL to STR infection. Results of this study indicated theoretical use of BCS for Eimeria identification and suggested some perspective of BCS for targeting animals infected by strongylid nematode. Validity of our results, however, was limited by number of observed animals managed under specific breeding conditions.

Does the in vitro egg hatch test predict the failure of benzimidazole treatment in Haemonchus contortus?

Considerable research has been directed towards optimising in vitro tests that can diagnose resistance in pre-parasitic stages of parasites. The objective of this study was to compare the in vivo faecal egg count reduction test (FECRT), the in vitro egg hatch test (EHT), and the molecular determination of the frequency of a codon 200 allele of ß-tubulin isotype 1 associated with benzimidazole resistance in larval stages of Haemonchus contortus obtained from infected goats. Animals were infected with composite infective doses representing 10, 20, 30, 40, 60, and 80% resistant alleles. Faecal samples for the EHT were collected on 28, 33, and 35 days post-infection. The results of the in vivo FECRT indicated that albendazole treatment reduced infections consisting of composite doses of 10, 20, 30, 40, 60, and 80% larvae of the resistant isolate by 91.3, 78.0, 63.3, 48.4, 36.5, and 41.4%, respectively. The drug concentration at which 50% of the eggs were prevented from developing hatching larvae (ED50) in the in vitro EHT varied from 0.09 ± 0.01 to 15.63 ± 12.10 µg/mL thiabendazole. The results of the in vitro EHT indicated that the test could estimate in vivo resistance well. The EHT could thus accurately estimate the in vivo efficacy of the drug and percentage of the resistance allele in the population using hatching parameters in delineation doses. This finding was also supported by comparing the FECRT data to the hatching percentages in the EHT on 30 goat farms in Slovakia with natural mixed infections of gastrointestinal parasites.

TITLE: Le test d'éclosion des œufs in vitro prédit-il l'échec du traitement au benzimidazole pour Haemonchus contortus ? ABSTRACT: Des efforts de recherche considérables ont été consacrés à l'optimisation des tests in vitro permettant de diagnostiquer la résistance des parasites aux stades préparasitaires. L'objectif de cette étude était de comparer le test de réduction du nombre d'œufs fécaux in vivo (TRNOF), le test d'éclosion des œufs in vitro (TEO) et la détermination moléculaire de la fréquence d'un allèle du codon 200 de l'isotype 1 de la ß-tubuline associé à résistance au benzimidazole au stade larvaire d'Haemonchus contortus, obtenus à partir de chèvres infectées. Les animaux ont été infectés avec des doses infectieuses composites représentant 10, 20, 30, 40, 60 et 80 % d'allèles résistants. Des échantillons de matières fécales ont été prélevés pour le TEO aux jours 28, 33 et 35 après l'infection. Les résultats de la TRNOF in vivo ont indiqué que le traitement à l'albendazole réduisait respectivement de 91,3, 78,0, 63,3, 48,4, 36,5 et 41,4 % les infections composées de doses composites de 10, 20, 30, 40, 60 et 80 % de larves de l'isolat résistant. La concentration de médicament à laquelle 50 % des œufs ont été empêchés de développer des larves qui éclosent (DE50) dans le TEO in vitro variait de 0,09 ± 0,01 à 15,63 ± 12,10 µg/mL de thiabendazole. Les résultats du TEO in vitro ont indiqué que le test pouvait correctement estimer la résistance in vivo. Le TEO a ainsi pu estimer avec précision l'efficacité in vivo du médicament et le pourcentage de l'allèle de résistance dans la population en utilisant des paramètres d'éclosion dans les doses de délimitation. Cette constatation a également été étayée par la comparaison des données du TRNOF aux pourcentages d'éclosion dans le TEO sur 30 élevages de chèvres en Slovaquie, avec des infections mixtes naturelles de parasites gastro-intestinaux.

The identification and semi-quantitative assessment of gastrointestinal nematodes in faecal samples using multiplex real-time PCR assays.

BACKGROUND: The diagnosis of gastrointestinal nematode (GIN) infections in ruminants is routinely based on morphological/morphometric analysis of parasite specimens recovered by coprological methods, followed by larval culture (LC) techniques. Such an approach is laborious, time-consuming, requires a skilled expert, and moreover suffers from certain limitations. Molecular tools are able to overcome the majority of these issues, providing accurate identification of nematode species and, therefore, may be valuable in sustainable parasite control strategies. METHODS: Two multiplex real-time polymerase chain reaction (PCR) assays for specific detection of five main and one invasive GIN species, including an internal amplification control to avoid false-negative results, were designed targeting SSU rRNA and COI genetic markers, as well as established ITS1/2 sequences. The assays were optimized for analysis of DNA extracted directly from sheep faeces and verified for Haemonchus contortus, Teladorsagia circumcincta, Trichostrongylus colubriformis, Nematodirus battus, Chabertia ovina, and Ashworthius sidemi. Semi-quantitative evaluation of infection intensity was enabled using a plasmid construct and a dilution series of sheep faeces with a known number of nematode eggs. Assays were tested on 44 individually collected faecal samples from three farms, and results were compared to those from faecal egg counts (FEC) using the concentration McMaster technique and LC. RESULTS: Multiplex real-time PCR assays showed great specificity to target nematodes. During the analysis of faecal samples, the assays proved to have higher sensitivity in strongylid-type egg detection over FEC by revealing three false-negative samples, while showing moderate agreement in evaluation of infection intensity. The multiplex assays further clarified GIN species identification compared to LC, which had confused determination of Teladorsagia spp. for Trichostrongylus spp. CONCLUSIONS: Our multiplex assays proved to be a rapid and accurate approach enabling simultaneous and reliable GIN species identification from faeces and semi-quantitative estimation of the number of eggs present. This approach increases diagnostic value and may add a high degree of precision to evaluation of anthelmintic efficacy, where it is important to identify species surviving after treatment.

Resistance of strongylid nematodes to anthelmintic drugs and driving factors at Czech goat farms.

BACKGROUND: Strongylid nematode infections may negatively affect both animal health and welfare, with deleterious consequences for livestock productivity. Many farmers in recent decades have relied on anthelmintics as the sole strategy of control, but the intensive use of these chemotherapeutics has led to the development of anthelmintic resistance (AR). Knowledge of both the efficacy of anthelmintics and factors promoting AR are essential to effectively control nematode infections, but no information on these topics for goats in the Czech Republic (CR) is available. This survey aimed to determine the occurrence of AR at Czech goat farms and to identify risk factors for the development of AR. A total of 24 herds of dairy goats across the CR were evaluated using in vitro tests for detecting AR, and a questionnaire survey was carried out to evaluate factors associated with AR. RESULTS: Resistance against benzimidazoles was confirmed at 18 (75%) farms, and the level of resistance was high in four (22%) of the affected herds based on the egg hatch test. Ivermectin-resistant nematodes were detected in 13 (54%) herds using the larval development test; Teladorsagia/Trichostrongylus and Haemonchus were the predominant types of resistant larvae. Eight (62%) of the affected herds were evaluated as highly resistant to ivermectin. Eleven (46%) of the herds were resistant to both benzimidazoles and ivermectin. This report is the first on dual AR in the CR. A univariate logistic regression analysis indicated that a high stocking rate and farmer inexperience were significantly associated with ivermectin and benzimidazole resistance, respectively. CONCLUSIONS: The results of our survey suggest that AR is widespread amongst herds of dairy goats in the CR, likely due to inappropriate practices of pasture and health management. AR may be an issue for expanding dairy-goat production in the CR in the near future unless both veterinary practitioners and farmers widely adopt strategies to prevent the development of AR.

Nematode-induced pathological lesions and alterations of mucin pattern identified in abomasa of wild ruminants.

Pathological lesions as well as mucin alterations in abomasa infected by nematodes have been thoroughly studied in livestock, but such data from wild ruminants are limited or completely lacking. Pathological data for Ashworthius sidemi, an invasive nematode are particularly rare. We necropsied the abomasa of 21 wild ruminants belonging to five cervid species and detected mixed nematode infections, dominated by A. sidemi. Samples from both gross lesions and mucous membranes without macroscopically apparent pathological alterations were subjected to standard histological procedures and histochemical staining. Histological examination found chronic abomasitis, manifested by edema, and hyperemia. Various degrees of lymphoplasmacytic infiltration were observed in all samples. Initial fibrosis (8/20, 40%) was detected in samples from both gross lesions and areas without macroscopically visible changes. Tissue from hemorrhagic lesions was superficially eroded. Generalized loss of surface polysaccharides was apparent in all samples. Only residual periodic acid-Schiff and Alcian blue (pH2.5) positivity was detected in the upper abomasal pits and in mucosal neck. This study found that nematode infections, mostly by A. sidemi, caused chronic inflammation and negatively affected abomasal mucin formation in wild ruminants.

Effect of Artemisia absinthium and Malva sylvestris on Antioxidant Parameters and Abomasal Histopathology in Lambs Experimentally Infected with Haemonchus contortus.

This study evaluated the effect of Artemisia absinthium and Malva sylvestris on antioxidant response and histopathological changes in the abomasa of the Haemonchus contortus infected lambs. Twenty-four lambs were divided into four groups: unsupplemented lambs (UNS), lambs supplemented with A. absinthium (ART), lambs supplemented with M. sylvestris (MAL), and lambs supplemented with both plants (ARTMAL). Lambs were infected orally with approximately 5000 third-stage (L3) larvae of H. contortus. The experiment was conducted for 75 d (days), all animals were then slaughtered; and the abomasal tissues were examined for antioxidant parameters and histopathology. The concentration of malondialdehyde in the abomasal mucosa was lower in ARTMAL (p < 0.05), and the total antioxidant capacity was higher in MAL (p < 0.05), than in UNS. Increased mucus production was observed in the ARTMAL. The number of mast cells in UNS and ART was significantly higher than the number in MAL (p < 0.01 and p < 0.05). Plasma cell numbers were higher in ARTMAL than the number in MAL (p < 0.05). Abomasal tissue regenerated more frequently in ARTMAL. These results represent the first report of the impact of A. absinthium and M. sylvestris on antioxidant parameters and local immune responses of abomasal mucosa of lambs infected with a GIN parasite.

Setaria cervi (Filarioidea, Onchocercidae) undressing in ungulates: altered morphology of developmental stages, their molecular detection and complete sequence cox1 gene.

This work introduces new morphological and molecular information on the filaroid nematode Setaria cervi (Rudolphi, 1819) obtained from 13 infected game ungulates out of 96 dissected. The hosts comprised the following: a single moose (Alces alces), ten red deer (Cervus elaphus) and two sika deer (Cervus nippon) originating from the western and northern regions of the Czech Republic. Based on the complete sequences of the gene encoding mitochondrial cytochrome c oxidase subunit 1 (cox1), all 20 females and four males belonged to the species S. cervi. We detected three developmental female stages (adult fertile females, juvenile L5 females and L4 female larvae) differing in size and some morphological traits as the subtle structure of peribuccal crown and shape and features of tail knob. Such differences were described in detail for the first time. The phylogenetic relationships within the family Onchocercidae have been evaluated using new information on the cox1 sequence of S. cervi (maximum likelihood method, GTR + I + G model). In accordance with the latest phylogenetic studies, the present analysis confirmed the ancient separation of the subclass Setariinae from the remaining two onchocercid lineages Dirofilariinae and Onchocerinae.

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.

TITLE: Importance croissante de la résistance aux anthelminthiques chez les ruminants européens : création et méta-analyse d'une base de données ouverte. ABSTRACT: Les helminthes sont omniprésents dans les systèmes de production de ruminants au pâturage et sont responsables de coûts et de pertes de production importants. La résistance aux anthelminthiques (RA) des parasites est maintenant répandue dans toute l'Europe, bien qu'il existe encore des lacunes dans nos connaissances dans certaines régions et certains pays. La RA est une menace majeure pour la pérennité de la production moderne de ruminants, en diminuant la productivité, en compromettant la santé et le bien-être des animaux, et en augmentant les émissions de gaz à effet de serre au travers d'une augmentation du parasitisme et des intrants agricoles. Une meilleure compréhension de l'étendue de la RA en Europe est nécessaire pour développer et préconiser des approches de lutte antiparasitaire plus durables. Une base de données intégrant des informations publiées et non publiées en Europe concernant la RA des nématodes gastro-intestinaux (NGI) et de la douve du foie (Fasciola hepatica) a été compilée par les membres de l'action européenne COST « COMBAR ¼ (« Combattre la résistance aux anthelminthiques chez les ruminants ¼) et combinée avec les données d'une précédente étude systématique concernant la RA des NGI. Au total, 197 publications sur la RA des NGI étaient disponibles pour analyse, représentant 535 études dans 22 pays et couvrant la période 1980­2020. Des signalements de RA étaient présents sur tout le continent européen et certains rapports indiquaient une forte prévalence nationale. Les estimations heuristiques pondérées par la taille de l'échantillon de la prévalence de la RA en Europe sur toute la période d'étude, stratifiées par classe d'anthelminthiques, variaient de 0 à 48 %. La prévalence régionale (nationale) estimée était très hétérogène, variant entre 0 % et 100 % selon le secteur de l'élevage et la classe d'anthelminthique, et augmentait généralement avec les efforts de recherche dans le pays. Dans les quelques pays disposant de données longitudinales adéquates, il y avait une tendance à l'augmentation de la RA au fil du temps pour toutes les classes d'anthelminthiques des NGI : les résultats agrégés chez les ovins et caprins depuis 2010 révèlent une prévalence moyenne de résistance aux benzimidazoles (BZ) de 86 %, aux lactones macrocycliques sauf moxidectine (ML) de 52 %, au lévamisole (LEV) de 48 % et à la moxidectine (MOX) de 21 %. Tous les genres principaux de NGI ont survécu au traitement dans diverses études. Chez les bovins, la prévalence de la RA variait selon les classes d'anthelminthiques de 0 à 100 % (BZ et ML), 0 à 17 % (LEV) et 0 à 73 % (MOX), et Cooperia et Ostertagia ont survécu aux traitements. Une RA suspectée chez F. hepatica a été signalée dans 21 études portant sur 6 pays. Pour les NGI, et encore plus pour F. hepatica, il y avait un biais d'échantillonnage en faveur des exploitations individuelles suspectées de RA, et l'effort de recherche était biaisé vers l'Europe occidentale et en particulier le Royaume-Uni. La capture continue des résultats futurs dans la base de données, en direct, les efforts pour éviter les biais dans le recrutement des exploitations, des tests plus précis pour la RA et une meilleure appréciation de l'importance de la RA parmi l'industrie agricole et les décideurs politiques, soutiendront des analyses plus sophistiquées des facteurs contribuant à la RA, et des stratégies efficaces pour ralentir sa propagation.

The use of high resolution melting analysis of ITS-1 for rapid differentiation of parasitic nematodes Haemonchus contortus and Ashworthius sidemi.

Among gastrointestinal nematodes, haematophagous strongylids Haemonchus contortus and Ashworthius sidemi belong to the most pathogenic parasites of both domestic and wild ruminants. Correct identification of parasitic taxa is of crucial importance in many areas of parasite research, including monitoring of occurrence, epidemiological studies, or testing of effectiveness of therapy. In this study, we identified H. contortus and A. sidemi in a broad range of ruminant hosts that occur in the Czech Republic using morphological/morphometric and molecular approaches. As an advanced molecular method, we employed qPCR followed by High Resolution Melting analysis, specifically targeting the internal transcribed spacer 1 (ITS-1) sequence to distinguish the two nematode species. We demonstrate that High Resolution Melting curves allow for taxonomic affiliation, making it a convenient, rapid, and reliable identification tool.

The response of soil nematode Caenorhabditis elegans on the sewage sludge-derived micropollutants.

Sewage sludge application to soil is of great interest, due to required organic matter and the wide spectra of nutrients it provides. However, the presence of unpredictable content of emerging contaminants may turn this valuable raw material into a hazardous substance. In this study, three selected sewage sludges derived micropollutants from different origins; that is, one each under persistent organic pollutants (POPs), pharmaceuticals and personal care products (PPCPs) were considered. The effect of each micropollutant on the feeding activity of free-living soil nematode Caenorhabditis elegans was analysed. The analysis was performed in model soil solution using a larval feeding inhibition assay. The results showed no significant effects from selected POP-2,2',4,4',5-pentabromodiphenyl either and pharmaceutical-chlortetracycline on the feeding activity of tested nematodes. On the contrary, feeding activity was inhibited by PPCP-galaxolide (HHCB) with an effective concentration of 12.2 ±â€¯2.2 mg.l-1. The calculated risk quotient for galaxolide (RQ = 0.14) demonstrated a medium ecological risk to the nematodes. Based on our findings, concentrations of micropollutants in sewage sludge treated soil pose negligible risk to feeding activity of soil nematode. However, the potential impact of musk compounds on free-living soil biota requires detailed evaluation in further research.

Natural chemotherapeutic alternatives for controlling of haemonchosis in sheep.

BACKGROUND: Parallel in vitro and in vivo experiments were designed to evaluate promising chemotherapeutic alternatives for controlling haemonchosis in ruminants. In vitro anthelmintic activities (egg hatch test - EHT; larval development test - LDT) of aqueous and methanolic herbal extracts Mix1 and Mix2 were investigated. The in vivo effects of dietary supplementation with Mix1 and Mix2 on the parasitological status, inflammatory response, antioxidant parameters and microbial community of the lambs infected experimentally with Haemonchus contortus were investigated. Lambs were divided into four groups for the in vivo study: uninfected control lambs (C), infected lambs (I), infected lambs supplemented with Mix1 (I + Mix1) and infected lambs supplemented with Mix2 (I + Mix2). The experimental period was 70 days. RESULTS: The number of eggs per gram (EPG) of feces was quantified 22, 30, 37, 44, 51, 58, 65 and 70 days post-infection, and mean abomasal worm counts were assessed 70 days post-infection. Quantitative analyses identified 57.3 and 22.2 mg/g phenolic acids, 41.5 and 29.5 mg/g flavonoids and 1.4 and 1.33 mg/g protoberberine-type alkaloids in Mix1 and Mix2, respectively. The methanolic extracts of the herbal mixtures in both in vitro tests had higher anthelmintic effects (P < 0.01) than the aqueous extracts, but the effects did not differ significantly between Mix1 and Mix2 (P > 0.05). I + Mix1 and I + Mix2 lowered mean EPGs between 44 and 70 d by 58.1 and 51.6%, respectively. The level of IgG antibodies against H. contortus increased significantly after infection in each infected group. CONCLUSION: These results represent the first monitoring of the in vitro anthelmintic effects of herbal mixtures on H. contortus. The in vivo experiment indicated that the anthelmintic effect was not sufficient for the elimination of parasites, but this herbal treatment may affect the host over a longer term, reducing the parasitic infection in the host.

Ovicidal and larvicidal activity of extracts from medicinal-plants against Haemonchus contortus.

The use of medicinal plants (MP) containing bioactive compounds is an alternative strategy to control of parasitic nematode of small ruminants Haemonchus contortus at various stages of their life cycle. The aims of this study were to determine the in vitro anthelmintic activity of both aqueous and methanolic extracts from 13 medicinal plants typical for Central Europe, and to determine quantity of selected plant secondary metabolites (PSMs) in the methanolic extracts. In vitro egg hatch test and larval development tests were conducted to determine the possible anthelmintic effects of methanolic and aqueous extracts of the roots of Althaea officinalis L., Petasites hybridus L. and Inula helenium L.; flowers of Malva sylvestris L. and Chamomilla recutita L.; leaves of Plantago lanceolata L. and Rosmarinus officinalis L.; seeds of Foeniculum vulgare Mill. and stems of Solidago virgaurea L., Fumaria officinalis L., Hyssopus officinalis L., Melisa officinalis L. and Artemisia absinthium L. on eggs and larvae of H. contortus. Ultra-performance liquid chromatography and tandem mass spectroscopy was used for quantifying six PSMs: gallic acid (GA), rutin (RU), diosmin (DI), hesperidin (HE), quercetin (QU) and kaempferol (KA). RU content of the most effective methanolic extracts was in the order: M. sylvestris (9.33 mg/g DM) > A. absinthium (6.10 mg/g DM) > C. recutita (0.42 mg/g DM). The highest concentration of QU (44.8 mg/g DM) and KA (6.59 mg/g DM) were detected in stems of F. officinalis comparing to the other evaluated plants. The most significant (p < 0.05) anthelmintic effects exhibited methanolic extracts of A. absinthium in both in vitro tests (i.e., egg hatch test and larval development test). Additionally, only two methanolic extracts of C. recutita and M. sylvestris were comparable to activity of A. absinthium using the larval development test. Wider spectrum of aqueous extracts exhibited stronger ovicidal activity in comparison to methanolic extracts. The similar trend was observed in evaluating of larvicidal activity of aqueous and methanolic plant extracts.

Long-term occurrence of Trichuris species in wild ruminants in the Czech Republic.

The aim of this study was to identify Trichuris species in wild ruminants from 32 localities in the Czech Republic using morphological and molecular methods (ITS1-5.8S RNA-ITS2 region polymorphisms). Trichurids were obtained from 176 wild ruminants (roe deer, sika deer, red deer, fallow deer and mouflons) that were culled between 2009 and 2017. Trichuris discolor is the predominant trichurid of all of the above-mentioned wild ruminants, whereas Trichuris ovis was identified less frequently in roe deer, fallow deer, sika deer and mouflons. Red deer were parasitised exclusively by T. discolor. Young hosts under 1 year of age were more intensively infected by trichurids than were adults (χ2 = 32.02, p = 0.00). Trichurid prevalence results obtained through coprological methods and those based on parasitological dissections differed significantly (χ2 = 16.26, p = 0.00). The regression analysis indicated that the eggs per gram (EPG) threshold (20 EPG) was exceeded only if the host was parasitised by more than 7 trichurid females. Full concordance between the positive results obtained by the coprological methods and those obtained via direct dissections was achieved when the number of trichurid females per host exceeded 51.

Seasonal dynamics of endoparasitic infections at an organic goat farm and the impact of detected infections on milk production.

This study evaluated patterns and species composition of parasitic infections detected over a 1-year period at an organic goat farm. As a result of coprological examination, the overall prevalence of observed strongylids (99%), coccidia of the genus Eimeria (98%), and Muellerius capillaris lungworms (93%) was calculated. The most prevalent strongylids recovered from incubated fecal samples were Haemonchus contortus (42%), genera Trichostrongylus (23%), Oesophagostomum columbianum (13%), and Teladorsagia circumcincta (11%). A maximum intensity of coccidia infection 5150 oocysts per gram, strongylids infection 9900 eggs per gram and lungworm infection 867.26 larvae per gram were detected. The various effects (including environment, host, and parasites) on milk yield, lactose, protein, and fat were evaluated using generalized linear mixed models. Milk yield (P < 0.0001), milk fat (P < 0.01), and lactose (P < 0.0001) were affected by month, i.e., these parameters were influenced by the month of the year, regardless of the individual goat. With the intensity of infection detected in our study, only protein content was affected (P < 0.01) by parasitic infection (exclusively caused by strongylids). Correlation between measurements from one individual revealed that the goat itself can substantially decrease protein content but has much less of an effect on fat, milk yield, and lactose. Based on our results, we can conclude that a low intensity of parasitic infections does not significantly affect milk yield and the qualitative parameters of milk.

Reliable molecular differentiation of Trichuris ovis and Trichuris discolor from sheep (Ovis orientalis aries) and roe deer (Capreolus capreolus) and morphological characterisation of their females: morphology does not work sufficiently.

The main aim of the study was to evaluate associations between morphological variability of Trichuris females from sheep and roe deer and their rDNA polymorphism in whipworm populations from the Czech Republic. The results introduced the use of new molecular markers based on the internal transcribed spacer (ITS)1-5.8S RNA-ITS2 region polymorphisms, as useful tools for the unambiguous differentiation of congeners Trichuris ovis and Trichuris discolor. These markers revealed both parasites in roe deer and in sheep; however, T. ovis females predominated in sheep while T. discolor females occurred mostly in roe deer. Additional analysis of ITS1-5.8 rRNA-ITS2 discovered the genetic uniformity of the analysed T. discolor but high haplotype variation of T. ovis. Simultaneously, molecularly designated female individuals of both species were categorised into four morphotypes (MT) on the basis of morphology of genital pore area. MT1 and MT4 (vulvar opening on everted vaginal appendage/on visible cuticular bulge) occurred only in T. ovis, MT2 (uneverted vagina-vulvar opening without any elevation) was identified only in T. discolor and MT3 (transient type of vulvar opening on a small swelling) was observed in both species. Statistical analysis of biometric data confirmed that morphology of vulva is not a reliable marker for the species determination. On the basis of the ITS1-5.8S RNA-ITS2 region variability, we carried out a phylogenetic analysis (maximum likelihood method, Hasegawa-Kishino-Yano model) which showed that T. ovis haplotypes from the Czech Republic and Ireland and T. discolor haplotypes from the Czech Republic, Spain, Iran and Japan are sister OTUs.

Efficacy and persistent activity of moxidectin against natural Muellerius capillaris infection in goats and pathological consequences of muelleriosis.

The effect of moxidectin against natural Muellerius capillaris infection in goats was evaluated in this study. Long-acting moxidectin at a single dose of 1 mg kg(-1) body weight was administered to an entire flock (n=10) of goats. The individual faecal larval count reduction was applied as an indicator of treatment efficacy. A significant reduction (>98%) in larval counts was observed in all surveyed animals 14 days after drug administration. Moxidectin demonstrated persistent activity in this study; the mean faecal larval count reduction was 99.1% ± 1.8 on day 77 of the treatment. Macroscopic abnormalities and histological changes in the lungs of two infected goats were evident during the post-mortem examination. The pathological consequences of M. capillaris infection were observed even three months after parasite elimination. The results of this study indicate that moxidectin is a highly effective anthelmintic agent for the control of muelleriosis in goats. This drug provides animals with fifteen weeks of protections against M. capillaris reinfection.