Ostertagia ostertagi Mediates Early Host Immune Responses via Macrophage and Toll-Like Receptor Pathways.

Ostertagia ostertagi is an abomasal parasite with significant economic impact on the cattle industry. Early host immune responses are poorly understood. Here, we examined time course expression of Toll-like receptors (TLRs) in peripheral blood mononuclear cells (PBMC) during infection where PBMC macrophages (Mϕ) generated both pro- and anti-inflammatory responses when incubated with excretory/secretory products (ESP) from fourth-stage larvae (OoESP-L4) or adult worms (OoESP-Ad). First, changes in cell morphology clearly showed that both OoESP-L4 and OoESP-Ad activated PBMC-Mϕ in vitro, resulting in suppressed CD40 and increased CD80 expression. Expression of mRNAs for TLR1, -4, -5, and -7 peaked 7 days postinfection (dpi) (early L4), decreased by 19 dpi (postemergent L4 and adults) and then increased at 27 dpi (late adults). The proinflammatory cytokine tumor necrosis factor alpha (TNF-α) (transcript and protein) increased in the presence of OoESP-Ad, and the anti-inflammatory cytokine interleukin 10 (IL-10) (protein) decreased in the presence of OoESP-L4 or OoESP-Ad; however, IL-10 mRNA was upregulated, and IL-6 (protein) was downregulated by OoESP-L4. When PBMC-Mϕ were treated with ligands for TLR4 or TLR5 in combination with OoESP-Ad, the transcripts for TNF-α, IL-1, IL-6, and IL-10 were significantly downregulated relative to treatment with TLR4 and TLR5 ligands only. However, the effects of TLR2 ligand and OoESP-Ad were additive, but only at the lower concentration. We propose that O. ostertagi L4 and adult worms utilize competing strategies via TLRs and Mϕ to confuse the immune system, which allows the worm to evade the host innate responses.

Prevalence, economic assessment, and risk factors of gastrointestinal nematodes infecting herds in tropical, dry and temperate climate regions in Mexico.

This study investigated exposure to gastrointestinal (GI) nematodes on dairy cattle farms by antibody level determination in bulk tank milk (BTM) samples and its influence on production to detect the risk factors for infection in different climate regions in three states of Mexico. From January to April 2017, BTM samples were collected from 1058 dairy cattle herds and used to establish three Köppen climate classes (tropical, dry and temperate) and states of Mexico. A questionnaire on farm management was applied. The overall herd prevalence of parasites was 67.20%. The highest percentage of positive herds was detected in Veracruz state (78.45%). In addition, the highest prevalence among the climate regions was found in the tropical climate (78.59%). In general, production losses were approximately 1.37-1.78 kg of milk/cow per day. The annual costs of milk production losses per farm were estimated for three different climate regions, ranging between $5541.49 and 6982.50 US$, and those in the three states varied between 5974.10 and 8660.06 US$. The costs for anthelmintic treatments for young stock and adult cows ranged between 57.51 and 192.75 US$, respectively, among the three climate regions and between 46.02 and 189.49 US$, respectively, among the three states. The overall annual costs of milk yield loss per cow were estimated to be 150.74 US$ for the climate regions and 190.54 US$ for the three states of Mexico, followed by the treatment costs for young stock (4.02 US$) and adult cows (3.99 US$). The results suggest that the economic losses due to GI nematodes in Mexican dairy herds are approximately 248 million US$ per annum. Four final models were built based on multivariate logistic regression for potential statistical association from the ELISA results using climatic/environmental and management factors so that each model used different risk factors that were significantly associated with helminth infections in dairy herds.

A longitudinal study of gastrointestinal parasites in English dairy farms. Practices and factors associated with first lactation heifer exposure to Ostertagia ostertagi on pasture.

The gastrointestinal nematode Ostertagia ostertagi is an important cause of lost production, health, and welfare in cattle. Detailed records were obtained over a 5-yr period (2010-2015) by questionnaires and qualitative interviews to investigate the practices adopted by dairy farmers to control cattle helminth infections and the factors associated with heifer exposure to O. ostertagi on pasture. In total, 1,454 heifers' individual milk samples were collected over a 1-yr period (2014-2015) in 43 dairy farms in England and tested for O. ostertagi antibody by ELISA. Multilevel linear regression models were used to investigate the association between individual milk optical density ratio (ODR) against O. ostertagi and heifer management from birth to time of sampling. Farm and heifer median ODR against O. ostertagi were 0.98 (interquartile range = 0.76-1.02) and 0.64 (interquartile range = 0.42-0.84), respectively. The majority of heifers (88%) received an anthelmintic treatment before sampling in this study. After controlling for the effect of anthelmintic treatments, heifer individual milk ODR against O. ostertagi significantly increased with high stocking rate at first grazing and co-grazing with adult cows before calving. Conversely, heifer individual milk ODR against O. ostertagi significantly decreased when heifers had co-grazed with sheep and pasture grass had frequently been mowed. Overall, these results provide evidence to support targeting grazing management toward limiting the use of anthelmintics in dairy young stock to enable sustainable control of cattle helminth infections in England. However, to be accepted and adopted by farmers, these best practices would need to take into account farmers' perspectives and contextual challenges.

Bulk tank milk prevalence and production losses, spatial analysis, and predictive risk mapping of Ostertagia ostertagi infections in Mexican cattle herds.

This study investigated the prevalence, production losses, spatial clustering, and predictive risk mapping in different climate zones in five states of Mexico. The bulk tank milk samples obtained between January and April 2015 were analyzed for antibodies against Ostertagia ostertagi using the Svanovir ELISA. A total of 1204 farm owners or managers answered the questionnaire. The overall herd prevalence and mean optical density ratio (ODR) of parasite were 61.96% and 0.55, respectively. Overall, the production loss was approximately 0.542 kg of milk per parasited cow per day (mean ODR = 0.92, 142 farms, 11.79%). The spatial disease cluster analysis using SatScan software indicated that two high-risk clusters were observed. In the multivariable analysis, three models were tested for potential association with the ELISA results supported by climatic, environmental, and management factors. The final logistic regression model based on both climatic/environmental and management variables included the factors rainfall, elevation, land surface temperature (LST) day, and parasite control program that were significantly associated with an increased risk of infection. Geostatistical kriging was applied to generate a risk map for the presence of parasite in dairy cattle herds in Mexico. The results indicate that climatic and meteorological factors had a higher potential impact on the spatial distribution of O. ostertagi than the management factors.

Characterization of Ostertagia ostertagi annexin-like proteins at different developmental stages.

Ostertagiosis remains an economically important parasitic disease in cattle in the temperate regions of the world. Repeated exposures to Ostertagia ostertagi in calves cause significant pathology in the abomasum but elicit little protective immunity. The larvae use the host's gastric glands as a niche for development, where the parasite completes its parasitic stages, while in the gastric glands, the larvae must down-regulate the host inflammatory immune responses. Annexin (ANX) A1, commonly found in most eukaryotes, is heavily involved in controlling anti-inflammatory responses by binding receptors on leukocytes. We hypothesized, therefore, that parasite proteins of the ANX family may be involved in host-parasite interactions during ostertagiosis. BLASTN search with the bovine ANXA1 identified two families of Oos-ANX like proteins (Oos-ANXL), each of which was highly conserved at the genetic level and identical at the amino acid sequence level. Oos-ANXL-1 is encoded by two transcripts and Oos-ANXL-2 by 20 transcripts. The present study characterized one Oos-ANXL, representing the most abundant Oos-ANXL, which was further defined as Oost-ANXL-2.1. Oos-ANXL-2.1 with a coding sequence of 519 bp was PCR-amplified, cloned, and expressed. Oos-ANXL-2.1 was immunolocalized to both L3 and adult, but not L4. The staining appeared to be associated with the gut and hypodermis in L3, but it was specifically localized to the hypodermis in adult worms. Western blots detected three protein bands in parasite lysates using anti-recombinant Oos-ANXL-2.1 antibody. Integrated optical density for each of the 3 Oos-ANXL-2s or the total Oos-ANXL-2s detected by Western blots (P < 0.05) was higher in adult worms than in L3 or L4. The results indicate that the production of Oos-ANXL-2s is developmentally regulated and most abundant in the adult worm. This rather large family of proteins could be a potential vaccine target against O. ostertagi infection and warrants further investigation.

Impact of chemical structure of flavanol monomers and condensed tannins on in vitro anthelmintic activity against bovine nematodes.

Plants containing condensed tannins (CT) may have potential to control gastrointestinal nematodes (GIN) of cattle. The aim was to investigate the anthelmintic activities of four flavan-3-ols, two galloyl derivatives and 14 purified CT fractions, and to define which structural features of CT determine the anti-parasitic effects against the main cattle nematodes. We used in vitro tests targeting L1 larvae (feeding inhibition assay) and adults (motility assay) of Ostertagia ostertagi and Cooperia oncophora. In the larval feeding inhibition assay, O. ostertagi L1 were significantly more susceptible to all CT fractions than C. oncophora L1. The mean degree of polymerization of CT (i.e. average size) was the most important structural parameter: large CT reduced larval feeding more than small CT. The flavan-3-ols of prodelphinidin (PD)-type tannins had a stronger negative influence on parasite activity than the stereochemistry, i.e. cis- vs trans-configurations, or the presence of a gallate group. In contrast, for C. oncophora high reductions in the motility of larvae and adult worms were strongly related with a higher percentage of PDs within the CT fractions while there was no effect of size. Overall, the size and the percentage of PDs within CT seemed to be the most important parameters that influence anti-parasitic activity.

Anthelmintic effects of forage chicory (Cichorium intybus) against gastrointestinal nematode parasites in experimentally infected cattle.

Two experiments studied the effects of dietary chicory against gastrointestinal nematodes in cattle. In Experiment (Exp.) 1, stabled calves were fed chicory silage (CHI1; n = 9) or ryegrass/clover hay (CTL1; n = 6) with balanced protein/energy intakes between groups. After 16 days, all calves received 10 000 Ostertagia ostertagi and 66 000 Cooperia oncophora third-stage larvae (L3) [day (D) 0 post-infection (p.i.)]. In Exp. 2, calves were assigned to pure chicory (CHI2; n=10) or ryegrass/clover (CTL2; n = 10) pastures. After 7 days, animals received 20 000 O. ostertagi L3/calf (D0 p.i.) and were moved regularly preventing pasture-borne infections. Due to poor regrowth of the chicory pasture, CHI2 was supplemented with chicory silage. At D40 p.i. (Exp. 1) and D35 p.i. (Exp. 2) calves were slaughtered for worm recovery. In Exp.1, fecal egg counts (FEC) were similar between groups. However, O. ostertagi counts were significantly reduced in CHI1 by 60% (geometric mean; P < 0·01), whereas C. oncophora burdens were unaffected (P = 0·12). In Exp. 2, FEC were markedly lowered in CHI2 from D22 p.i onwards (P < 0·01). Ostertagia ostertagi adult burdens were significantly reduced in CHI2 by 66% (P < 0·001). Sesquiterpene lactones were identified only in chicory (fresh/silage). Chicory shows promise as an anti-Ostertagia feed for cattle and further studies should investigate its on-farm use.

A stochastic model to investigate the effects of control strategies on calves exposed to Ostertagia ostertagi.

Predicting the effectiveness of parasite control strategies requires accounting for the responses of individual hosts and the epidemiology of parasite supra- and infra-populations. The first objective was to develop a stochastic model that predicted the parasitological interactions within a group of first season grazing calves challenged by Ostertagia ostertagi, by considering phenotypic variation amongst the calves and variation in parasite infra-population. Model behaviour was assessed using variations in parasite supra-population and calf stocking rate. The model showed the initial pasture infection level to have little impact on parasitological output traits, such as worm burdens and FEC, or overall performance of calves, whereas increasing stocking rate had a disproportionately large effect on both parasitological and performance traits. Model predictions were compared with published data taken from experiments on common control strategies, such as reducing stocking rates, the 'dose and move' strategy and strategic treatment with anthelmintic at specific times. Model predictions showed in most cases reasonable agreement with observations, supporting model robustness. The stochastic model developed is flexible, with the potential to predict the consequences of other nematode control strategies, such as targeted selective treatments on groups of grazing calves.

[A comparative analysis of various antigenic proteins found in Haemonchus contortus--a review].

Many innovative researches on the development and introduction of recombinant vaccines against many economically important parasites were carried out in the 20th century. Research continues to hold promise with the development of immunological and molecular approaches for control of these parasites and in this regard it has already been seen that blood-sucking parasites such as Haemonchus contortus and Ostertagia ostertagi are susceptible to control by vaccines containing "novel" or "concealed" antigens. Haemonchus contortus is primarily pathogenic to sheep and its blood-feeding behaviour causes effects ranging from mild anaemia to mortality in young animals. Current means of control which are dependent on repeated treatment with anthelmintics are responsible for the increasing drug resistance of this parasite. Together with the growing concern of residual chemicals in the environment and food chain, this has led to attempts to better understand the biology of the parasite with an aim to develop alternate means of control, including the development of molecular vaccines. More problematic and also important is the formulation and delivery strategy to induce expulsion of this parasite, using vaccines containing recombinant "conventional" antigens. Tremendous progress has been made in the last decade in identifying several antigens from Haemonchus contortus which in their native form stimulate useful levels of protective immunity. Vaccines have been developed against H. contortus using 'novel' gut antigens from the parasite, but variable responsiveness of the host sheep has resulted in varying degrees of protection which are stimulated by these vaccines. Computer models have also been used to simulate vaccine efficacy in worm control and have yielded good results. This review will try to summarise the protective efficacy and also the molecular properties of principal candidate antigens which are expressed by this parasite. The review will try to cover the aspirations, current success, limitations and problems faced by researchers in the control of this economically important parasite.

A stochastic frontier approach to study the relationship between gastrointestinal nematode infections and technical efficiency of dairy farms.

The impact of gastrointestinal (GI) nematode infections in dairy farming has traditionally been assessed using partial productivity indicators. But such approaches ignore the impact of infection on the performance of the whole farm. In this study, efficiency analysis was used to study the association of the GI nematode Ostertagia ostertagi on the technical efficiency of dairy farms. Five years of accountancy data were linked to GI nematode infection data gained from a longitudinal parasitic monitoring campaign. The level of exposure to GI nematodes was based on bulk-tank milk ELISA tests, which measure the antibodies to O. ostertagi and was expressed as an optical density ratio (ODR). Two unbalanced data panels were created for the period 2006 to 2010. The first data panel contained 198 observations from the Belgian Farm Accountancy Data Network (Brussels, Belgium) and the second contained 622 observations from the Boerenbond Flemish farmers' union (Leuven, Belgium) accountancy system (Tiber Farm Accounting System). We used the stochastic frontier analysis approach and defined inefficiency effect models specified with the Cobb-Douglas and transcendental logarithmic (Translog) functional form. To assess the efficiency scores, milk production was considered as the main output variable. Six input variables were used: concentrates, roughage, pasture, number of dairy cows, animal health costs, and labor. The ODR of each individual farm served as an explanatory variable of inefficiency. An increase in the level of exposure to GI nematodes was associated with a decrease in technical efficiency. Exposure to GI nematodes constrains the productivity of pasture, health, and labor but does not cause inefficiency in the use of concentrates, roughage, and dairy cows. Lowering the level of infection in the interquartile range (0.271 ODR) was associated with an average milk production increase of 27, 19, and 9L/cow per year for Farm Accountancy Data Network farms and 63, 49, and 23L/cow per year for Tiber Farm Accounting System farms in the low- (0-90), medium- (90-95), and high- (95-99) efficiency score groups, respectively. The potential milk increase associated with reducing the level of infection was higher for highly efficient farms (6.7% of the total possible milk increase when becoming fully technically efficient) than for less efficient farms (3.8% of the total possible milk increase when becoming fully technically efficient).

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.

Structure of Ostertagia ostertagi ASP-1: insights into disulfide-mediated cyclization and dimerization.

The cysteine-rich secretory/antigen 5/pathogenesis-related 1 (CAP) protein superfamily is composed of a functionally diverse group of members that are found in both eukaryotes and prokaryotes. The excretome/secretome of numerous helminths (parasitic nematodes) contains abundant amounts of CAP members termed activation-associated secreted proteins (ASPs). Although ASPs are necessary for the parasitic life cycle in the host, the current lack of structural and functional information limits both understanding of their actual role in host-parasite interactions and the development of new routes in controlling parasitic infections and diseases. Alleviating this knowledge gap, a 1.85 Å resolution structure of recombinantly produced Oo-ASP-1 from Ostertagia ostertagi, which is one of the most prevalent gastrointestinal parasites in cattle worldwide, was solved. Overall, Oo-ASP-1 displays the common hallmark architecture shared by all CAP-superfamily members, including the N-terminal CAP and C-terminal cysteine-rich domains, but it also reveals a number of highly peculiar features. In agreement with studies of the natively produced protein, the crystal structure shows that Oo-ASP-1 forms a stable dimer that has been found to be primarily maintained via an intermolecular disulfide bridge, hence the small interaction surface of only 306.8 Å(2). Moreover, unlike any other ASP described to date, an additional intramolecular disulfide bridge links the N- and C-termini of each monomer, thereby yielding a quasi-cyclic molecule. Taken together, the insights presented here form an initial step towards a better understanding of the actual biological role(s) that this ASP plays in host-parasite interactions. The structure is also essential to help to define the key regions of the protein suitable for development of ASP-based vaccines, which would enable the current issues surrounding anthelmintic resistance in the treatment of parasitic infections and diseases to be circumvented.

Novel epidemiological model of gastrointestinal nematode infection to assess grazing cattle resilience by integrating host growth, parasite, grass and environmental dynamics.

Gastrointestinal nematode (GIN) infections are ubiquitous and often cause morbidity and reduced performance in livestock. Emerging anthelmintic resistance and increasing change in climate patterns require evaluation of alternatives to traditional treatment and management practices. Mathematical models of parasite transmission between hosts and the environment have contributed towards the design of appropriate control strategies in ruminants, but have yet to account for relationships between climate, infection pressure, immunity, resources, and growth. Here, we develop a new epidemiological model of GIN transmission in a herd of grazing cattle, including host tolerance (body weight and feed intake), parasite burden and acquisition of immunity, together with weather-dependent development of parasite free-living stages, and the influence of grass availability on parasite transmission. Dynamic host, parasite and environmental factors drive a variable rate of transmission. Using literature sources, the model was parametrised for Ostertagia ostertagi, the prevailing pathogenic GIN in grazing cattle populations in temperate climates. Model outputs were validated on published empirical studies from first season grazing cattle in northern Europe. These results show satisfactory qualitative and quantitative performance of the model; they also indicate the model may approximate the dynamics of grazing systems under co-infection by O. ostertagi and Cooperia oncophora, a second GIN species common in cattle. In addition, model behaviour was explored under illustrative anthelmintic treatment strategies, considering impacts on parasitological and performance variables. The model has potential for extension to explore altered infection dynamics as a result of management and climate change, and to optimise treatment strategies accordingly. As the first known mechanistic model to combine parasitic and free-living stages of GIN with host feed-intake and growth, it is well suited to predict complex system responses under non-stationary conditions. We discuss the implications, limitations and extensions of the model, and its potential to assist in the development of sustainable parasite control strategies.

Development of Bovine Gastric Organoids as a Novel In Vitro Model to Study Host-Parasite Interactions in Gastrointestinal Nematode Infections.

Gastro-intestinal nematode (GIN) parasites are a major cause of production losses in grazing cattle, primarily through reduced growth rates in young animals. Control of these parasites relies heavily on anthelmintic drugs; however, with growing reports of resistance to currently available anthelmintics, alternative methods of control are required. A major hurdle in this work has been the lack of physiologically relevant in vitro infection models that has made studying precise interactions between the host and the GINs difficult. Such mechanistic insights into the infection process will be valuable for the development of novel targets for drugs, vaccines, or other interventions. Here we created bovine gastric epithelial organoids from abomasal gastric tissue and studied their application as in vitro models for understanding host invasion by GIN parasites. Transcriptomic analysis of gastric organoids across multiple passages and the corresponding abomasal tissue showed conserved expression of tissue-specific genes across samples, demonstrating that the organoids are representative of bovine gastric tissue from which they were derived. We also show that self-renewing and self-organising three-dimensional organoids can also be serially passaged, cryopreserved, and resuscitated. Using Ostertagia ostertagi, the most pathogenic gastric parasite in cattle in temperate regions, we show that cattle gastric organoids are biologically relevant models for studying GIN invasion in the bovine abomasum. Within 24 h of exposure, exsheathed larvae rapidly and repeatedly infiltrated the lumen of the organoids. Prior to invasion by the parasites, the abomasal organoids rapidly expanded, developing a 'ballooning' phenotype. Ballooning of the organoids could also be induced in response to exposure to parasite excretory/secretory products. In summary, we demonstrate the power of using abomasal organoids as a physiologically relevant in vitro model system to study interactions of O. ostertagi and other GIN with bovine gastrointestinal epithelium.

Tracking gastrointestinal nematode risk on cattle farms through pasture contamination mapping.

Gastrointestinal nematode (GIN) parasites in grazing cattle are a major cause of production loss and their control is increasingly difficult due to anthelmintic resistance and climate change. Rotational grazing can support control and decrease reliance on chemical intervention, but is often complex due to the need to track grazing periods and infection levels, and the effect of weather on larval availability. In this paper, a simulation model was developed to predict the availability of infective larvae of the bovine GIN, Ostertagia ostertagi, at the level of individual pastures. The model was applied within a complex rotational grazing system and successfully reproduced observed variation in larval density between fields and over time. Four groups of cattle in their second grazing season (n = 44) were followed throughout the temperate grazing season with regular assessment of GIN faecal egg counts, which were dominated by O. ostertagi, animal weight and recording of field rotations. Each group of cattle was rotationally grazed on six group-specific fields throughout the 2019 grazing season. Maps and calendars were produced to illustrate the change in pasture infectivity (density of L3 on herbage) across the 24 separate grazing fields. Simulations predicted differences in pasture contamination levels in relation to the timing of grazing and the return period. A proportion of L3 was predicted to persist on herbage over winter, declining to similar intensities across fields before the start of the following grazing season, irrespective of contamination levels in the previous year. Model predictions showed good agreement with pasture larval counts. The model also simulated differences in seasonal pasture infectivity under rotational grazing in systems that differed in temperature and rainfall profiles. Further application could support individual farm decisions on evasive grazing and refugia management, and improved regional evaluation of optimal grazing strategies for parasite control. The integration of weather and livestock movement is inherent to the model, and facilitates consideration of climate change adaptation through improved disease control.

Gastrointestinal nematode and lungworm infections in organic dairy calves reared with nurse cows during their first grazing season in western France.

The rearing system of dairy calves with nurse cows has been developing since 2010 in organic farms in western France. This system allows cow-calf contact until a weaning age close to the natural weaning for cattle and is characterized by an early turnout for calves at around one month of age with their nurse cows and a first grazing season with mixed grazing of calves and adults at a ratio of 2-4 calves per nurse cow. The objectives of this study were to assess the gastrointestinal (GIN) and lungworm infections in such reared calves and their variability during the first grazing season. Faecal egg count (FEC), pepsinogen (PEP) concentration and Ostertagia ELISA optical density ratio (ODR) were determined in calves (n = 497) at housing in 33 groups from 24 farms in 2018, and in calves (n = 405) and nurse cows (n = 199) throughout the 2019 grazing season in 41 groups from 20 farms. For lungworm infection, information was obtained during 2019 through the recording of coughing episodes along the grazing season and the Dictyocaulus ELISA ODR determination at housing both in calves and nurses. Results indicated that the level of GIN infection was overall low for calves during the first grazing season with PEP and Ostertagia ODR group-average values ranging from 0.97 to 1.6 U Tyr and 0.23 to 0.71 ODR respectively. No anthelmintic treatment being given in any group of calves. Ostertagia ODR values increased with the duration of the grazing season (>240 d) and with the ratio calves/nurse (>2). GIN parameters for nurses remained fairly stable during the grazing season with mean FEC, PEP and Ostertagia ODR group-average values of 13 epg, 2.28 U Tyr and 0.81 ODR, respectively. Antibodies against lungworms were detected in 3-62 % of calves depending on the duration of grazing, but only 6% of calves showed a coughing episode. The dilution effect due to the mixed grazing of resistant (nurse cows) and susceptible (calves) animals associated with predominant milk diet of calves during the first months of grazing in combination with protective grazing management allow calves to be turned out at an early age without using anthelmintic treatments. Further studies are needed to assess the GIN infection dynamics during the second grazing season in weaned heifers.

Molecular and phenotypic characterisation of fenbendazole resistance in a field-derived isolate of Ostertagia ostertagi.

The prevalence of anthelmintic resistance in the bovine nematode Cooperia oncophora has been well documented globally but lack of efficacy against the more pathogenic nematode species Ostertagia ostertagi is less common. The sensitivity of an O. ostertagi isolate to the benzimidazole class of anthelmintic was investigated using classical parasitological techniques following apparent clinical failure of controlled release fenbendazole capsule administration in first season grazers at pasture. A controlled efficacy test (CET) was conducted in conjunction with sequencing of the ß-tubulin isotype 1 gene of larvae pre- and post-fenbendazole administration. Twelve helminth-naïve calves were infected experimentally with 20,000 third stage larvae; six received oral fenbendazole (7.5 mg/kg bodyweight) 28 days post infection. Total abomasal nematode burdens were compared between treatment and control groups to determine efficacy. Fenbendazole resistance in O. ostertagi was confirmed with a total treatment failure in reducing worm burden: efficacy of 0%. Sequence analysis of the ß-tubulin isotype-1 gene from forty-five infective larvae from both control and treated groups was performed. The three commonest single nucleotide polymorphisms (SNPs) associated with benzimidazole resistance, namely F167Y, E198A and F200Y, were examined. The predominant resistance-associated SNPs were F200Y (78 % control and 79 % treated groups) and F167Y (remaining genotypes) and emphasises the importance of these SNPs in clinical disease in this isolate. The development of diagnostic molecular tools based on a characterised field-derived isolate of benzimidazole-resistant Ostertagia will enable future prevalence surveys to be undertaken to assess the possible risk posed by resistance in this economically important species.

Gastrointestinal nematodes and Fasciola hepatica in Norwegian cattle herds: a questionnaire to investigate farmers' perceptions and control strategies.

BACKGROUND: Pasture management influences the prevalence and impact of the pasture parasites (PP) in cattle herds, which cause production-limiting disease worldwide. Evaluating farmer management strategies is vital when considering sustainable PP control practices. The aim of this questionnaire-based study was to describe the pasture management and control strategies regarding PP in Norwegian beef cattle (BC) and dairy cattle (DC) production systems with a focus on gastrointestinal nematodes (GIN) and Fasciola hepatica. RESULTS: A total of 745 responses from BC (return rate 20.5%) and 1347 responses from DC farmers (30.7%) were included. The mean total pasture time for DC was 4.2 months for first-season grazers and 4.3 months for second-season grazers and cows, while the corresponding finding in BC was 5.4 months. Home pasture was used for most of the pasture period, particularly for first-season grazer dairy heifers (81%), which were also commonly grazed on the same pasture every year (79%). For most farmers it was necessary for grazing areas to be used for cattle for more than one season (77% of BC farmers and 89% of DC farmers). However, changing the pasture during the season was common in both DC (67%) and BC (70%) herds. The majority of DC farmers (60%) stated that they did not consider that they had a problem with PP. Of the remaining 40%, few respondents could specify whether their herds had a problem due to infection by GIN (11%) or liver flukes (12%). Treatment for GIN was performed by 52% of DC and 34% of BC farmers. Diagnostic faecal samples were collected upon suspicion of parasitic disease by 5% of DC and 16% of BC farmers. Veterinarians were stated as a central source of information about parasite management and treatment. CONCLUSIONS: Potential risks for exposure to PP were identified, such as use of the same pasture every year for first-season grazers and frequent use of home pasture. The perception of problems related to PP appeared low. Regular anthelmintic treatment without concurrent use of diagnostic faecal samples seems to be common practice.

Effect of Mixed Invasions of Hypoderma bovis and Ostertagia ostertagi in Cattle on Milk Yield and Contents in Polish Dairy Farms.

Wide distribution of ecto- and endoparasites in cattle is a serious problem in the sustainability of a farm, due to the negative impact on animals' health and productivity. The aim of this study was to determine the presence of antibodies against Ostertagia and Hypoderma in udder milk samples and the comparison of milk yield and content of the basic components of milk in ELISA-positive and ELISA-negative cows. Milk samples were collected from 148 lactating cows from 3 herds. Antibody detection was performed using specific ELISAs for Ostertagia ostertagi and Hypoderma bovis. Milk yield and content of protein, fat, and dry matter were studied in samples from each individual cow 11 times per year at 4 week intervals. The extensiveness of dual parasitic invasions in individual herds, estimated on the basis of udder milk testing with the ELISA test, varied and amounted to 3.22%, 11.36%, and 4.76% in the three studied herds, respectively. No antibodies were found in 61.2%, 22.7%, and 57.1% of the milk samples from the cows in each herd. The milk yield of ELISA-positive cows was significantly lower in comparison to the efficiency of ELISA-negative cows and amounted to 294 kg and even to 3672 kg of milk per year, per cow. No significant differences were found between the fat and protein contents of milk between ELISA-positive and -negative cows for both parasites.

Repeated, drug-truncated infections with Ostertagia ostertagi elicit strong humoral and cell-mediated immune responses and confer partial protection in cattle.

Bovine ostertagiasis causes significant production losses to the cattle industry. Protective immunity induced by natural infection is slow to develop and anthelmintic resistance is rapidly developing. There is a need to advance alternatives for control of gastrointestinal nematode parasites. The present study investigated the effects of repeated, drug-truncated infections (rDTI) on development of protective immunity and attenuation of a challenge infection by O. ostertagi. Helminth-free calves were randomly assigned to either a rDTI or a control group (n = 5). The rDTI group received daily oral infections of 5000 Ostertagia L3 for 5 consecutive days, then were drug-treated on 14 and 15 days post infection (dpi), to attenuate O. ostertagi at the late fourth larval (L4) through young adult stages. DTI was repeated 3 weeks after the drug treatment. A total of 5 DTIs were administered to the DTI-treated animals. Non-DTI-treated, control animals received tap water as infection control. All animals were drug-treated at the same time. Animals were challenge-infected 4 weeks following the final round of rDTI. The results show that eggs per gram of feces (EPG) in the rDTI group were significantly reduced (P < 0.05) from 21 to 39 dpi, with an overall reduction in cumulative EPG. The control group exhibited reduced (P = 0.0564) average weight gains when compared to those of the rDTI group during weeks 4-5 post infection, a period coinciding with peak EPG output of control animals. Antigen-specific IgG, IgE and IgA responses were detected after the 2nd DTI, and stronger antibody recall responses were elicited by challenge infection. High levels of antigen-specific peripheral blood mononuclear cell (PBMC)/T cell proliferation to whole worm and excretory-secretory (ES) antigens were detected in rDTI-treated animals. These data indicate that partial protective immunity against ostertagiasis, involving cell-mediated and humoral responses, can be attained by rDTI which allowed for maximal antigen exposure from staggered parasitic developmental stages. The data suggest that rDTI can be used as a model to study host-parasite interactions and identify parasite antigens responsible for eliciting host protective immune responses.