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Abstract Background Gastrointestinal nematodes (GINs) have a major impact on sheep production, health, and welfare worldwide. Norway is no exception, but there are only a few studies on the prevalence of GINs in Norwegian sheep. The aim of this study was to investigate the current occurrence of the most important nematodes in sheep flocks in Norway. Faecal samples were collected from flocks in 2021/2022, mainly from three geographical regions in Norway, i.e., northern, eastern, and western. In each of 134 flocks included, individual samples from 10 lambs (autumn) were pooled. Third stage larvae (L3) were cultivated and harvested (Baermann method) from the pooled samples. The DNA was then extracted and further analysed using droplet digital PCR (ddPCR). This enables assessment of the proportions of the three most important nematode species/genera, i.e., H. contortus, T. circumcincta, and Trichostrongylus. The fractional abundance/relative proportion of each species/genus was assessed by performing duplex assays with universal strongyle and species/genus-specific primers and probe sets. In addition, the occurrence of Nematodirus eggs was assessed by standard faecal egg counts (i.e., McMaster method). Results Of the 134 flocks sampled, 24 were from the northern region, 31 from eastern, and 71 from western Norway. In addition, some flocks from central (n = 7), and southern (n = 1) Norway were included. Among the sampled flocks, T. circumcincta occurred most commonly (94%), followed by H. contortus (60%) and Trichostrongylus (55%), and Nematodirus (51%). In general, mixed infections were observed, with 38% and 18% of flocks infected with three or all four genera, respectively. Conclusions The results of this study indicate that GINs are widespread in Norway. Teladorsagia circumcincta seems to be present in most flocks based on this screening. Moreover, the results show that Nematodirus spp. infect lambs throughout the country, predominantly N. battus, and indicate that this nematode has become more abundant, which could lead to an increase in nematodirosis.

Reports of Ascaridia galli in laying hens in Europe have increased since the ban on conventional battery cages in 2012. As this parasite is transmitted directly via the faecal-oral route by parasite eggs containing a larva, it is reasonable to assume that the escalating problem is related to the increased exposure now occurring in modern welfare-friendly cage-free housing systems. On many farms, A. galli reappears in subsequent flocks, even though the birds have no access to the outdoors, biosecurity is high and empty houses are cleaned and disinfected during downtime. Since the egg production cycle lasts only ≈80 weeks and recombinant antigen production for helminth vaccines has not yet been solved, the development of a vaccine seems to be an unrealistic option. Therefore, disrupting the life cycle of the parasite by other means, including the strategic use of dewormers, appears to be the key to controlling infection. Of concern is that only one class of anthelmintics is licenced for poultry in Europe and that are usually administered indiscriminately through the birds' drinking water and often too late when the parasite is already established. If current calendar-based parasite control strategies are not changed, there is a risk that resistance to anthelmintics may develop, as has already been demonstrated with nematodes in livestock. We insist that treatments can be more effective and the risk of developing drug resistance can be mitigated if we invest in a better understanding of A. galli responses to more prudent and judicious use of anthelmintics. This review identifies knowledge gaps and highlights aspects of sustainable parasite control that require further research to support commercial egg producers.

Lungworms of the genus Dictyocaulus are causative agents of parasitic bronchitis in domestic and wild ungulates. This study investigates the distribution, morphology and genetic diversity of D. cervi and a new lungworm species, Dictyocaulus skrjabini n. sp. infecting red deer Cervus elaphus, fallow deer Dama dama and moose Alces alces in Poland and Sweden. The study was conducted on 167 red deer from Poland and on the DNA of lungworms derived from 7 fallow deer, 4 red deer and 2 moose collected in Sweden. The prevalence of D. cervi and D. skrjabini n. sp. in dissected red deer in Poland was 31.1% and 7.2%, respectively. Moreover, D. skrjabini n. sp. was confirmed molecularly in 7 isolates of fallow deer lungworms and 1 isolate of red deer lungworms from Sweden. Dictyocaulus skrjabini n. sp. was established based on combination of their distinct molecular and morphological features; these included the length of cephalic vesicle, buccal capsule (BC), buccal capsule wall (BCW), distance from anterior extremity to the nerve ring, the width of head, oesophagus, cephalic vesicle, BC and BCW, as well as the dimensions of reproductive organs of male and female. Additionally, molecular analyses revealed 0.9% nucleotide sequence divergence for 1,605 bp SSU rDNA, and 16.5–17.3% nucleotide sequence divergence for 642 bp mitochondrial cytB between D. skrjabini n. sp. and D. cervi, respectively, and 18.7–19% between D. skrjabini n. sp. and D. eckerti, which translates into 18.2–18.7% amino acid sequence divergence between D. skrjabini n. sp. and both lungworms.

The aim of this study was to investigate how the activity of cattle under natural grazing conditions is related to their individual parasite contribution (IPC). Potentially, the individuals contributing the most to the contamination of the pasture with gastrointestinal nematodes (GIN) could then be identified and selectively treated based on sensor data thresholds. A total of 58 steers of the dairy breeds Swedish Red (SR, n = 19) and Swedish Holstein (SH, n = 39) were used for sensor-based data collection that lasted for a total of 10 weeks from 4 May to 13 July 2022. All steers were inoculated with a priming dose of nematode larvae and weighed in conjunction with the pasture release. The animals were then divided into four experimental groups and treated with ivermectin (IVM PO, Boehringer Ingelheim, 0.5 mg/kg BW) at different intervals to obtain an exposure contrast (Group A was left untreated, Group B/IVM PO-4w was treated after 4 weeks, Group C/IVM PO-8w was treated after 8 weeks, and Group D/IVM PO-4&8w was treated after 4 weeks and then after 8 weeks). The steers were weighed on four further occasions, during which faecal samples were also taken for parasite testing. Activity data were collected using leg-mounted IceQube sensors and body weight data and faecal samples were collected bi-weekly. The new threshold metric (IPC) was proposed for individuals with different faecal egg count (FEC) levels [NO (no contribution)

Parasitic sheep nematodes, among which Haemonchus contortus is often considered to be the most clinically important, exact a significant toll on the animals, not least because of their capacity to evolve drug resistance. Despite decades of research, our understanding of the mechanism of resistance to compounds such as levamisole is fairly limited, which therefore constrains our ability to develop sensitive and efficient molecular diagnostic tools for rapid and accurate resistance detection in field settings. Herein, we investigated the presence and frequency of the newly reported, levamisole-resistance-associated, mutation, yielding a S168T substitution in exon 4 of hco-acr-8, in six different phenotypically described isolates (three susceptible and three resistant), three Swedish field isolates and eight larvae culture samples, the latter two of which originated on farms where levamisole showed complete parasite elimination. For this purpose, we created both an allele-specific and droplet digital PCR approaches and found the mutated allele to be present only in the Kokstad isolate, whereas the other five as well as both the Swedish isolates and larvae cultures displayed only the non-mutated, serine-encoding, allele. While the finding of only the non-mutated allele in the phenotypically susceptible and Swedish isolate and larvae culture samples seemed sensible, we speculate that for the other two phenotypically resistant isolates, different (perhaps secondary) variants are responsible for conferring the resistance to levamisole phenotype, given the polygenic nature of levamisole resistance. All in all, despite the limited number of samples tested here, the mutation causing the S168T substitution in hco-acr-8 represents a plausible levamisole resistance-associated variant in, at least, some isolates of H. contortus.

Abstract The use of polymerase chain reaction (PCR)-based diagnostic approaches has steadily increased in the field of parasitology in recent decades. The most recent large-scale technological modification of the PCR formula, also known as third-generation PCR, came in the form of digital PCR (dPCR). Currently, the most common form of dPCR on the market is digital droplet PCR (ddPCR). Unlike quantitative real-time PCR (qPCR), the digital format allows for highly sensitive, absolute quantification of nucleic acid targets and does not require external standards to be included in the developed assays. Dividing each sample into thousands of compartments and using statistical models also eliminates the need for technical replicates. With unprecedented sensitivity and enforcement of binary endpoint reactions, ddPCR not only allows the use of tiny sample volumes (especially important when working with limited amounts of DNA) but also minimises the impact of variations in amplification efficiency and the presence of inhibitors. As ddPCR is characterised by excellent features such as high throughput, sensitivity and robust quantification, it is widely used as a diagnostic tool in clinical microbiology. Due to recent advances, both the theoretical background and the practical, current applications related to the quantification of nucleic acids of eukaryotic parasites need to be updated. In this review, we present the basics of this technology (particularly useful for new users) and consolidate recent advances in the field with a focus on applications to the study of helminths and protozoan parasites. Graphical Abstract

Abstract The diagnostics of ruminant parasites remains one of the cornerstones for parasite control best practices. Field veterinarians have several techniques at their disposal (fecal egg count, coproculture, FAMACHA®, plasma pepsinogen, ELISA-Ostertagia, ELISA-Fasciola, Baermann and ELISA-Lungworm) for the identification and/or quantification of gastrointestinal nematodes, lungworms and liver fluke infecting small ruminants and cattle. Each of these diagnostic tools has its own strengths and weaknesses and is more appropriate for a specific production operation and/or age of the animal (young and adults). This review focuses on the usability and interpretation of the results of these diagnostic tools. The most advanced technical information on sampling, storage, advantages and limitations of each tool for different types of production operations and animal categories is provided. Graphical abstract

Abstract Background Wildlife hosts may serve as reservoirs for strongyles, which can be transmitted to domestic livestock. Therefore, studies evaluating nemabiome compositions in wildlife ruminants are of great use in assessing the possibility of transmission of important nematode pathogens to domestic sheep in Sweden. Methods First, fecal samples were collected from roe deer (n = 125), fallow deer (n = 106), red deer (n = 18) and mouflon (n = 13) in south central Sweden during the hunting season in 2019. Second, after fecal examination samples were cultured and the larvae were harvested, followed by DNA extractions. Third, all samples were barcoded and processed for sequence analysis on the PacBio platform. Finally, bioinformatic sequence analysis was conducted with DADA2, while species diversity and richness, as well as interactions between the different hosts, were calculated and analyzed in R. Results Nematode ITS2 sequences were found in 225 of 262 (86%) samples. In total, 31 taxa were identified, among which 26 (86%) to the species level. These were found in different combinations, among which 24 (77%) occurred in roe deer, 19 (61%) in fallow deer, 20 (65%) in red deer and 10 (32%) in mouflon. Five of the species found are known to be associated with livestock (Chabertia ovina, Haemonchus contortus, Oesophagostomum venulosum, Teladorsagia circumcincta and Trichostrongylus axei). However, in the present study the relative abundance and prevalence of most of these species were low. The most striking exception was T. axei, which was relatively abundant in all wildlife hosts. Mostly a wide range of wildlife specific nematodes such as Ostertagia leptospicularis and Spiculopteragia spp. were identified including the invasive nematode Spiculopteragia houdemeri, which was found for the first time in red deer, fallow deer, and mouflon in Sweden. The difference in the number of shared species between mouflon and all cervids (n = 6) was less than among all three cervids (n = 8). Conclusion In this study, we investigated the community structure of parasitic intestinal nematodes in four wildlife hosts, and we found that the majority of the parasite species identified were wildlife specific. We also found a new, potentially invasive species not reported before. After comparing the nemabiome of the wildlife hosts in this study with a previous study in sheep from the same geographical region, we conclude that the horizontal transmission potential appears to be relatively low. Still, cross-infections of nematodes between game and sheep cannot be completely ignored. Graphical Abstract

Abstract Background Next-generation sequencing (NGS) has provided an alternative strategy to study the composition of nematode communities with increased resolution and sensitivity. However, the handling and processing of gigabytes worth of amplicon sequence data produced by an NGS platform is still a major hurdle, limiting the use and adoption of faster and more convenient analysis software. Methods In total 32 paired, fecal samples from Swedish sheep flocks were cultured and the larvae subsequently harvested subjected to internal transcribed spacer 2 (ITS2) amplicon sequencing using the PacBio platform. Samples were analyzed with three different bioinformatic pipelines, i.e. the DADA2, Mothur and SCATA pipelines, to determine species composition and richness. Results For the the major species tested in this study (Haemonchus contortus, Teladorsagia circumcinta and Trichostrongylus colubriformis) neither relative abundances nor species diversity differed significantly between the three pipelines, effectively showing that all three analysis pipelines, although different in their approaches, yield nearly identical outcomes. In addition, the samples analyzed here had especially high frequencies of H. contortus (90–95% across the three pipelines) both before and after sample treatment, followed by T. circumcinta (3.5–4%). This shows that H. contortus is the parasite of primary importance in contemporary Swedish sheep farms struggling with anthelmintic resistance. Finally, although on average a significant reduction in egg counts was achieved post-treatment, no significant shifts in major species relative frequencies occurred, indicating highly rigid community structures at sheep farms where anthelmintic resistance has been reported. Conclusions The findings presented here further contribute to the development and application of NGS technology to study nemabiome compositions in sheep, in addition to expanding our understanding about the most recent changes in parasite species abundances from Swedish sheep farms struggling with anthelmintic resistance. Graphical Abstract

Haemonchus contortus is a pathogenic gastrointestinal nematode of small ruminants and, in part due to its capacity to develop resistance to drugs, contributes to significant losses in the animal production sector worldwide. Despite decades of research, comparatively little is known about the specific mechanism(s) driving resistance to drugs such as ivermectin in this species. Here we describe a genome-wide approach to detect evidence of selection by ivermectin treatment in a field population of H. contortus from Sweden, using parasites sampled from the same animals before and seven days after ivermectin exposure followed by whole-genome sequencing. Despite an 89% reduction in parasites recovered after treatment measured by the fecal egg count reduction test, the surviving population was highly genetically similar to the population before treatment, suggesting that resistance has likely evolved over time and that resistance alleles are present on diverse haplotypes. Pairwise gene and SNP frequency comparisons indicated the highest degree of differentiation was found at the terminal end of chromosome 4, whereas the most striking difference in nucleotide diversity was observed in a region on chromosome 5 previously reported to harbor a major quantitative trait locus involved in ivermectin resistance. These data provide novel insight into the genome-wide effect of ivermectin selection in a field population as well as confirm the importance of the previously established quantitative trait locus in the development of resistance to ivermectin.

Abstract Background Haemonchus contortus is one of the most pathogenic gastrointestinal nematodes of small ruminants. The current diagnostic approach for the detection of this species relies on coproscopic methods, which both have low sensitivity and are time consuming. Methods employing detection through DNA amplification, such as droplet digital polymerase chain reaction (ddPCR), offer an advantageous approach to the diagnosis of H. contortus. However, DNA extraction protocols need to be constantly updated for the optimal retrieval of diagnostically usable template. Here, we describe the evaluation of three genomic DNA extraction kits for the detection and quantification of H. contortus ITS2 amplicon DNA from faecal samples, using droplet digital PCR. Results DNA samples, extracted from faecal material with the Nucleospin DNA Stool kit, produced the highest amounts of ITS2 amplicon copies and had the lowest coefficient of variation across different dilutions and sample types (fresh or frozen) out of the tested kits (Nucleospin DNA Stool, E.Z.N.A.® Stool DNA Kit and QIAamp Fast DNA Stool Mini Kit). Furthermore, the protocol of this kit has the fewest number of steps and the price of DNA extraction per sample is reasonable (2.77 €). Conclusions The Nucleospin DNA Stool kit is an attractive option for the detection and quantification of H. contortus DNA in faecal samples of small ruminants in a diagnostic setting.

Benzimidazole (BZ) drugs are frequently used to treat infections with the equine ascarid Parascaris univalens due to increasing resistance to macrocyclic lactones and pyrantel. Benzimidazole resistance is rare in ascarids in contrast to strongyle parasites where this resistance is widespread. In strongyles, single nucleotide polymorphisms (SNPs) at codons 167, 198 and 200 in a β-tubulin gene have been correlated to BZ resistance, but little is known about the β-tubulin genes and their possible involvement in BZ resistance in P. univalens and other ascarids. Previously two β-tubulin genes have been identified in P. univalens. In this study, we present five additional β-tubulin genes as well as the phylogenetic relationship of all seven genes to β-tubulins of other clade III and V nematodes. In addition, the efficacy of fenbendazole for treatment of P. univalens on a Swedish stud farm was studied in 2019 and 2020 using faecal egg count reduction test. Reductions varied from 73% to 88%, indicating the presence of a resistant P. univalens population on the farm. The emergence of BZ resistance emphasizes the need for development of molecular markers for rapid and more sensitive detection of resistant populations. We therefore investigated whether possible SNPs at positions 167, 198 or 200 in any of the β-tubulin genes could be used to distinguish between resistant and susceptible P. univalens populations. Amplicon sequencing covering the mutation sites 167, 198 and 200 in all seven β-tubulin genes revealed an absence of SNPs in both resistant and susceptible populations, suggesting that the mechanism behind BZ resistance in ascarids is different from that in strongyle nematodes and the search for a molecular marker for BZ resistance in P. univalens needs to continue.

Abstract 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. Graphical Abstract

The O. ostertagi-Ab ELISA assay is widely used as a diagnostic tool for monitoring gastrointestinal (GI) nematodes using milk samples from adult dairy cows. This assay is potentially also useful to analyse serum samples from first-season grazing (FSG) calves, providing a more cost-effective and robust diagnostic technique than the current serum pepsinogen assay. However, a comprehensive evaluation of its use in serum samples from FSG calves has not yet been conducted. In this study, we first reviewed the available scientific literature in which the O. ostertagi-Ab ELISA was applied to serum samples from FSG calves. Then, a field study was conducted to compare results from the O. ostertagi-Ab ELISA assay with a serum pepsinogen assay on a set of 230 serum samples from 11 commercial dairy herds (seven in Belgium and four in Sweden). The literature review showed an increase in mean antibody levels, expressed as optical density ratio (ODR) values, from O. ostertagi antibody levels measured during the late grazing season and weight gain, while the other two studies found no correlation between the two variables. Our field studies showed a good correlation between O. ostertagi antibody levels and the results from the pepsinogen assay. Both indicators were negatively related to average daily weight gain in the Belgian herds, but not in the Swedish herds. Overall, the results suggest that the O. ostertagi-Ab ELISA test can be a useful tool in FSG calves and could replace the use of the serum pepsinogen assay at the end of the grazing season for general monitoring purposes.

Gastrointestinal nematodes in grazing livestock are ubiquitous and can cause severe damage, leading to substantial losses in agricultural yields. It is undeniable that the integrated use of anthelmintics is often an essential component of successful intensive livestock management. However, anthelmintic resistance has been a major challenge for several decades, especially in pasture-based lamb production. Measures are therefore needed to reduce the risk and prevent further spread. In many countries with more extensive lamb production and pronounced resistance problems than in Sweden, the importance of keeping parasites in refugia is emphasised. To ensure that treatment is necessary, the Swedish model is based on deworming certain groups of ewes based on the parasitological results of a faecal examination and then releasing them with their lambs to safe pastures. This is intended to reduce the risk of infection, which ultimately reduces the number of subsequent treatments. Whether this preventive strategy in turn means an increased risk of resistance is debatable. In this review, we explain the importance of parasites in refugia and how they can help delay the development of resistance to anthelmintics. We also discuss how likely it is that our model contributes to an increase in resistance risk and whether there is reason to question whether it is a sustainable strategy in the long term.

Abstract Gastrointestinal nematodes (GIN) in small ruminants result in production losses, and consequently economic losses, and are an animal welfare problem in most countries in the Nordic-Baltic region. Intensive use of anthelmintics to control helminth infections has led to anthelmintic resistance (AR), which has become a major issue in many European countries. Several studies have been performed in countries in the Nordic-Baltic region (e.g. Denmark, Sweden, Norway and Lithuania) showing increasing/emerging levels of AR. The aim of this paper is to provide an overview of the problem of AR on sheep and goat farms in the Nordic-Baltic region. This region has a limited number of registered anthelmintics. However, researchers in this area have discovered some surprising findings, such as ivermectin (IVM) resistance on a farm that had never used IVM. In Sweden there is evidence of macrocyclic lactone (ML)-resistant Haemonchus contortus being introduced with sheep imported from the Netherlands. As elsewhere in the world, the livestock trade appears to be contributing to the spread of AR in the region and isolated cases of multidrug-resistant cases have also been reported. This is surprising given that the frequency of treatments here is much lower than in other countries where sheep production is economically more important. The prevailing nematodes are Haemonchus, Teledorsagia and Trichostrongylus, while on some farms Haemonchus is dominant and clinical haemonchosis has increasingly been observed in recent decades. The reasons for this are unclear, but are probably related to this parasite’s propensity to rapidly develop drug resistance and a general lack of awareness of the problem, possibly in combination with global warming and the increased livestock trade within the EU. In addition, domestic interactions through contacts with wildlife ruminants, alpacas may also be a contributing factor for transmission of AR.

The nematode Haemonchus contortus is one of the most prevalent and pathogenic parasites in small ruminants. Although usually controlled using anthelmintics, the development of drug resistance by the parasite has become a major issue in livestock production. While the molecular detection of benzimidazole resistance in H. contortus is well developed, the molecular tools and protocols are far less advanced for the detection of levamisole resistance. The hco-acr-8 gene encodes a critical acetylcholine susceptible subunit that confers levamisole-sensitivity to the receptor. Here, we report the development of a droplet digital PCR assay as a molecular tool to detect a 63 bp deletion in the hco-acr-8 that has been previously associated with levamisole resistance. Sanger sequencing of single adult H. contortus yielded 56 high-quality consensus sequences surrounding the region containing the deletion. Based on the sequencing data, new primers and probes were designed and validated with a novel droplet digital PCR assay for the quantification of the deletion containing “resistant” allele in genomic DNA samples. Single adult worms from six phenotypically described isolates (n = 60) and from two Swedish sheep farms (n = 30) where levamisole was effective were tested. Even though a significant difference in genotype frequencies between the resistant and susceptible reference isolates was found (p = 0.01), the homozygous “resistant” genotype was observed to be abundantly present in both the susceptible isolates as well as in some Swedish H. contortus samples. Furthermore, field larval culture samples, collected pre- (n = 7) and post- (n = 6) levamisole treatment on seven Swedish sheep farms where levamisole was fully efficacious according to Fecal Egg Count Reduction Test results, were tested to evaluate the frequency of the “resistant” allele in each. Frequencies of the deletion ranged from 35 to 80% in the pre-treatment samples, whereas no amplifiable H. contortus genomic DNA was detected in the post-treatment samples. Together, these data reveal relatively high frequencies of the 63 bp deletion in the hco-acr-8 both on individual H. contortus and field larval culture scales, and cast doubt on the utility of the deletion in the hco-acr-8 as a molecular marker for levamisole resistance detection on sheep farms.

Abstract Background A novel way to study the species composition and diversity of nematode parasites in livestock is to perform deep sequencing on composite samples containing a mixture of different species. Herein we describe for the first time the nematode community structures (nemabiomes) inhabiting Swedish sheep and how these are/were affected by host age and recent anthelmintic treatments. Methods A total of 158 fecal samples were collected (n = 35 in 2007 and n = 123 in 2013–2016) and cultured from groups of sheep on 61 commercial farms in the south-central part of the country where most animals are grazed. Among the samples, 2 × 44 (56%) were paired collections from the same groups pre- and post-treatment with anthelmintics such as macrocyclic lactones, benzimidazoles or levamisole. Samples were analyzed for their nemabiome using the PacBio platform followed by bioinformatic sequence analysis with SCATA. Species richness and diversity were calculated and analyzed in R. Results Nematode ITS2 sequences were found in all larval culture samples except two, even though the fecal egg counts were below the McMaster threshold in 20 samples. Sequencing yielded, on average, 1008 sequences per sample. In total, 16 operational taxonomical units (OTU), all with ≥ 98 % identity to sequences in the NCBI database, were recognized. The OTUs found represented nematode species of which ten are commonly associated with sheep. Multiple species were identified in all pre-anthelmintic treatment larval culture samples. No effects on nematode diversity were found in relation to host age. On the other hand, recent anthelmintic treatment lowered species richness, especially after use of ivermectin and albendazole. Interestingly, despite zero egg counts after use of levamisole, these samples still contained nematode DNA and especially H. contortus. Conclusions Our findings provide evidence that nemabiome analysis combined with diversity index analysis provides an objective methodology in the study of the efficacy of anthelmintic treatment as both high and low abundant species were detected.

Abstract Background Internal parasites are common in pigs worldwide and may induce clinical disease or subclinical infections with negative effects such as poor weight gain and reduced welfare, which in turn affect productivity. Effective parasite control to reduce the negative impact of parasitic infections demands a combination of antiparasitic drugs as well as various hygiene and biosecurity practices. The aim of this study was to obtain information on current management practices and parasite control routines used on Swedish pig farms using an online questionnaire. Results Antiparasitic drugs were used on 69% of the farms routinely and were mainly administered to sows just prior to farrowing. Less than 5% of the herds conducted faecal analysis for parasites. Batchwise, age segregated rearing was common and overall, it was practiced for piglets, growers, and fatteners on 88, 80 and 75% of the farms, respectively. Large and medium sized farms appeared to apply stricter hygiene and biosecurity measures to the growing pigs compared to small farms. Dry sows were mainly housed in groups on deep litter straw beds and cleaning, as well as disinfection, between each group was less common compared to what was practiced for growing pigs. Outdoor access was rare and only occurred on organic and small farms. Most of the farms, 54, 74 and 82% of small, medium, and large sized herds respectively, reported to have less than 5% white spot lesions, caused by migrating A. suum larvae, registered at slaughter. Conclusion Several risk factors for parasite infections, such as bedding material, group housing and solid floors, are mandatory requirements by national law. However, it was evident from this study that although strategic hygiene and biosecurity practices appeared common, they were not practiced in all herds and less so for dry sows. Antiparasitic drugs were used frequently and mainly through routine prophylactic treatments without prior testing for parasites. A holistic approach is necessary when designing efficient parasite control programs, and it is essential that management factors and routine monitoring of parasites are given attention. This to achieve efficient parasite control and reduce the risk of unnecessary use of antiparasitic drugs.

Although the Dictyocaulus lungworm, the agent of dictyocaulosis, is one of parasitological threats to European bison, its systematic position remains unclear. The aim of the present study was to evaluate the morphological features of the lungworm and the pathological lesions it induces, and to analyse mitochondrial (mt) genetic markers for systematic and molecular epidemiological studies. The morphological findings indicate that Dictyocaulus lungworms of European bison can be distinguished from those of cattle on the basis of differences in buccal capsule wall length, total body length, and spicules length in males, all of which were significantly longer in those of European bison. Nucleotide diversity calculated from pairwise sequence alignments of partial cytochrome c oxidase subunit 1 (cox1), cytochrome B (cytB) and NADH dehydrogenase subunit 5 (nad5) of specimens from cattle and European bison varied from 1.7% for nad5, 2.1% for cytB, to 3.7% for cox1 gene. Thus, among the lungworms of European bison and cattle, nad5 and cytB were the most conserved proteins, whereas cox1 was the most diverse. The mt cytB marker gene may be a suitable candidate for distinguishing between the two genotypes, as nad5 demonstrated the greatest within-genus sequence variation. The lung tissue of infected European bison manifests signs of verminous pneumonia characterized by interstitial pneumonia, bronchitis and bronchiolitis. Therefore, it appears that European bison and cattle are infected with slightly diverged, morphologically-different, genotypes of D. viviparus, indicating they belong to two separate worm populations. We propose, therefore, that the lungworm of European bison should be classified as D. viviparus subsp. bisontis.

Abstract Background Parascaris univalens is a pathogenic parasite of foals and yearlings worldwide. In recent years, Parascaris spp. worms have developed resistance to several of the commonly used anthelmintics, though currently the mechanisms behind this development are unknown. The aim of this study was to investigate the transcriptional responses in adult P. univalens worms after in vitro exposure to different concentrations of three anthelmintic drugs, focusing on drug targets and drug metabolising pathways. Methods Adult worms were collected from the intestines of two foals at slaughter. The foals were naturally infected and had never been treated with anthelmintics. Worms were incubated in cell culture media containing different concentrations of either ivermectin (10−9 M, 10−11 M, 10−13 M), pyrantel citrate (10−6 M, 10−8 M, 10−10 M), thiabendazole (10−5 M, 10−7 M, 10−9 M) or without anthelmintics (control) at 37 °C for 24 h. After incubation, the viability of the worms was assessed and RNA extracted from the anterior region of 36 worms and sequenced on an Illumina NovaSeq 6000 system. Results All worms were alive at the end of the incubation but showed varying degrees of viability depending on the drug and concentration used. Differential expression (Padj

At its inception, the COVID-19 pandemic was described as something inherently new, capable of crossing and erasing the economic, racial, gendered, and religious divides that stratify societies around the world. However, the ongoing pandemic is not new or egalitarian, but fuelled by, and fuelling, crises already under way on a global scale. In this article we examine on the one hand the relationship between the pandemic and still-active formations of racialised and gendered power, and on the other the pandemic's inextricability from a dispersed and uneven planetary emergency. As the environmental historian Jason W. Moore notes, this emergency disproportionately affects ‘women, people of colour and (neo)colonial populations’ (2019: 54), and the effects of COVID-19 are similarly unevenly allocated.

The occurrence of endoparasites in Swedish adult dogs (n = 303) was investigated between January and October 2014. Included dogs had to be clinically healthy, older than 1 year and untreated with anthelmintics or endectocides for at least 3 months prior to sampling. They were grouped according to age, category of dog and time since last antiparasitic treatment. Samples were analyzed by flotation to detect parasitic eggs and cysts/oocysts. Among these, 129 (43%) dogs were also analyzed with the Baermann-technique to detect cardiopulmonary larval stages. Parasite dispersal stages were found in 24 (7.9%, CI 95% 4.9–10.1) of the dogs at flotation, while no dog shed cardiopulmonary larval stages. Giardia sp. cysts were observed in 2.6% (n = 8) of dogs examined, cysts of Sarcocystis spp. were observed in 0.6% (n = 2), oocysts of Cystosisopora ohioensis were found in one dog (0.3%). Eggs of Toxocara canis (2.3%, n = 7), Uncinaria stenocephala (1.3%, n = 4) and Trichuris vulpis (0.3%, one dog) were found. None of the dogs were diagnosed with more than one species. Although the occurrence of endoparasites was above the average in dogs ≤ 2 years of age (11.5%), nematodes were more common in older dogs ≥4 years (77.0%). Although the occurrence was lower in working/exhibition dogs (5.9%) than in companion dogs (8.4%) and hunting-dogs (8.6%), these differences were not significant. However, dogs exposed to prey according to the owner had a statistically significant higher prevalence than other dogs (20.5 vs. 5.7%). The Odds Ratio (OR) was 4.0 (CI 95%, 1.58–10.11) for dogs having access to prey, 2.4 (CI 95%, 0.37–8.06) for dogs staying at day-care, and 2 (CI 95%, 0.96–5.96) for bitches. Furthermore, a significant association was observed between infection with nematodes and exposures to prey (p = 0.006). As a reference, data on the endoparasites in canine fecal samples submitted to the National Veterinary Institute (SVA, Uppsala) during 2014 are presented. Overall, this study shows a low occurrence of endoparasites among dogs in Sweden. Any risk-assessment on zoonotic parasites as well as deworming recommendations will take advantage from these updated figures.

Sickness behaviour has been suggested as an applicable indicator for monitoring disease. Deviating feeding behaviour and activity can provide information about animals’ health and welfare status. Recent advances in sensor technology enable monitoring of such behaviours and could potentially be utilized as an indicator of gastrointestinal nematode (GIN) infections. This study investigated activity and rumination patterns in first-season grazing steers exposed to subclinical infection levels of the GIN Ostertagia ostertagi and Cooperia oncophora. At turnout, animals were allocated to one of four experimental groups and were faced with “high” (H1, n = 15; H2, n = 17) or “low” (L1, n = 17; L2, n = 11) levels of parasite exposure by grazing in similar enclosures contaminated with overwintering third stage (L3) GIN larvae. Animals in H1 and H2 (HP) received a 1:1 mix of approximately 10,000 O. ostertagi and C. oncophora L3 at turnout; whereas the animals in L1 and L2 (LP) were treated monthly with ivermectin. Activity and rumination patterns were monitored by fitting animals with leg- (IceQube) and neck-mounted (Heatime) sensors. BW was recorded every fortnight, whereas faecal and blood samples were examined every four weeks for nematode faecal egg count and serum pepsinogen concentrations (SPCs). There was an interaction effect of exposure level and period (P

Eimeria spp. and Clostridium perfringens (CP) are pathogens associated with coccidiosis and necrotic enteritis (NE) in broiler chickens. In this study we evaluated the effect of anticoccidial vaccination on intestinal health in clinically healthy organic Ross 308 chickens. On each of two farms, one unvaccinated flock (A1 and B1) was compared to one vaccinated flock (A2 and B2) until ten weeks of age (WOA). Faecal oocysts were counted weekly, and species were identified by PCR (ITS-1 gene). Lesion scoring, CP quantification and PCR targeting the CP NetB toxin gene were performed at three, four, and six WOA and chickens were weighed. Necropsies were performed on randomly selected chickens to identify coccidiosis/NE. Oocyst shedding peaked at three WOA in all flocks. Later oocyst shedding (E. tenella/E. maxima) in unvaccinated flocks at 5–7 WOA coincided with coccidiosis/NE. Although results differed somewhat between farms, vaccination was associated with lower intestinal lesion scores, reduced caecal CP counts, lower proportions of netB-positive CP, lower body weight at three–four WOA, and similar or slightly increased body weight at six WOA. In conclusion, the intestinal health of organic broilers can benefit from anticoccidial vaccination when oocyst exposure levels are high.

Helminth infections, mainly by gastrointestinal nematodes (GIN), are one of the main concerns for animal health, welfare and productivity in grazing ruminant livestock worldwide. The use of a sensitive, precise, accurate, low-cost, and easy-to-perform copromicroscopic technique is of pivotal importance to perform reliable fecal egg count (FEC) and fecal egg count reduction test (FECRT), in order to determine the need of anthelmintic treatment, but also anthelmintic efficacy or resistance. This approach is fundamental to a correct and efficient control of GIN. Unfortunately, in worldwide ruminant farm practice, repeated anthelmintic treatments are carried out, without prior diagnosis of infection, contributing to the spread of Anthelmintic Resistance (AR). Tackling this phenomenon, improving mainly the GIN diagnosis and AR status in farm animals, is a priority of the European COST Action “COMBAR—COMBatting Anthelmintic Resistance in Ruminants” and of the STAR-IDAZ International Research Consortium on Animal Health. One of the specific objectives of the COMBAR Working Group 1 (WG1) is to conduct an European market analysis of new diagnostics and develop a business plan for commercial test introduction, leveraging technical know-how of participants. Since the Mini-FLOTAC in combination with the Fill-FLOTAC may be considered a good candidate for a standardized FEC and FECRT in the laboratory, as well as directly in the field, the aim of this study was to conduct SWOT (Strength—Weaknesses—Opportunities—Threats) and PESTEL (Political, Economic, Social, Technological, Environmental, and Legal) analyses of these tools in 20 European countries involved in the COMBAR WG1, in order to identify the opportunities, barriers, and challenges that might affect the Mini-FLOTAC and Fill-FLOTAC commercialization in Europe.

Anthelmintic resistance in gastrointestinal nematode (GIN) parasites of grazing ruminants is on the rise in countries across the world. Haemonchus contortus is one of most frequently encountered drug-resistant GINs in small ruminants. This blood-sucking abomasal nematode contributes to massive treatment costs and poses a serious threat to farm animal health. To prevent the establishment of resistant strains of this parasite, up-to-date molecular techniques need to be proposed which would allow for quick, cheap and accurate identification of individuals infected with resistant worms. The effort has been made in the previous decade, with the development of the pyrosequencing method to detect resistance-predicting alleles. Here we propose a novel droplet digital PCR (ddPCR) assay for rapid and precise identification of H. contortus strains as being resistant or susceptible to benzimidazole drugs based on the presence or absence of the most common resistance-conferring mutation F200Y (TAC) in the β tubulin isotype 1 gene. The newly developed ddPCR assay was first optimized and validated utilizing DNA templates from single-worm samples, which were previously sequenced using the next generation PacBio RSII Sequencing (NGS) platform. Subsequent NGS results for faecal larval cultures were then used as a reference to compare the obtained values for fractional abundances of the resistance-determining mutant allele between ddPCR and NGS techniques in each sample. Both methods managed to produce highly similar results and ddPCR proved to be a reliable tool which, when utilized at full capacity, can be used to create a powerful mutation detection and quantification assay. Keywords: Strongyles, ddPCR, NGS-Sequencing, Benzimidazoles, Anthelmintic resistance

Resistance to ivermectin (IVM) in the nematode Haemonchus contortus in small ruminants is an increasing problem throughout the world. Access to molecular diagnostics will allow early detection of IVM resistance, which in turn can limit the spread of resistant isolates. One candidate gene which has recently been suggested as a marker for IVM resistance is that for dye-filling protein (dyf-7). In this study, we critically investigated the suitability of A141G and G153T single nucleotide polymorphisms (SNP) of dyf-7 as a marker in larval cultures collected from sheep farms in Sweden, involving several isolates for which resistance status had been characterised by the faecal egg count reduction test (FECRT). Initially, we designed dyf-7 primers from a worldwide collection of adult Haemonchus contortus DNA. With the sequence data, we created a haplotype network. We then optimised and used the same sets of primers and probes in a droplet digital PCR (ddPCR) assay for precise quantification of dyf-7 allele frequencies in pre- and post-anthelmintic treatment faecal larval cultures. The fractional abundance (FA) of the mutant SNP was within the range 7.8 and 31%. However, the FA was generally stable in samples collected from the same farms, even though they were obtained on different occasions up to 25 months apart. There was also no indication that the level of IVM resistance as measured by the faecal egg count reduction test was higher on farms with high FA. Furthermore, by comparing FA in samples from the same farms pre- and post-IVM treatment, we found no evidence of a correlation between dyf-7 and level of IVM resistance. Based on these results, dyf-7 is not a suitable marker for field testing of IVM resistance in H. contortus. Keywords: Amphids, Macrocyclic lactones, Haemonchosis, FECRT, qPCR, Molecular target

The presence of subcutaneous nodular onchocercosis was investigated at slaughter of 151 red deer (Cervus elaphus) (107 juveniles and 44 adults) between October–December 2015. The prevalence of subcutaneous nodules was 56%. Nodules were located in the lumbar region of the back in 96% of the cases, and 38% of the infected red deer had additional parasitic nodules in other body locations, such as rump, thorax, forelimbs and neck. The number of nodules per deer was 1–10 in two-thirds of the affected animals, and only 2% had more than 50 nodules. Significant differences in prevalence of nodule presence were found between adult (81%) and juvenile red deer less than two years of age (45%) (p = 0.0001). Species confirmation and identification was done on 14 randomly selected nodules taken from the lumbar region of 14 deer of different geographical origin, by sequencing the mitochondrial 12S, 16S and nad5 gene fragments. The sequences were more or less identical (12S = 99–100%; 16S = 93–100%; nad5 = 92–100%) to previously published sequences for Onchocerca flexuosa. The high prevalence of nodular onchocercosis caused by O. flexuosa in red deer in Sweden shows that the investigated areas in the southern third of Sweden have favorable vector (simuliids and ceratopogonids) conditions and suitable environment for the maintenance of the parasite. To our knowledge, this is the first systematic study of O. flexuosa in red deer in Sweden. Keywords: Onchocerca flexuosa, Cervid, DNA genotyping, Filarioidea, Onchocercidae, Vector-borne helminthes

Abstract Background According to targeted treatment (TT), the whole flock is dewormed based on knowledge of the risk, or parameters that quantify the mean level of infection, whereas according to targeted selective treatment (TST), only individual animals within the grazing group are treated, based on parasitological, production and/or morbidity parameters. The aim of this study was to compare two different treatment protocols on sheep farms in Lithuania. The study was conducted from 15 April to 31 October 2014 on three sheep farms. On the TT (the whole flock) and T(S)T (with FECs ≥ 300, respectively) farms all adult animals were treated orally with fenbendazole irrespective of EPG counts before the grazing season. The second treatment was applied with injectable ivermectin on both farms. However, on the TT farm all sheep were also treated on 2nd August regardless of their EPG counts, while on the T(S)T farm only those animals with an EPG ≥ 300 were treated on 1 July using a threshold of ≥ 300 EPG. No treatments were administered on the control farm (n = 1) during the study. Results Spring treatment of ewes significantly reduced nematode faecal egg counts (FEC) both on the TT and T(S)T farms, with the benefit of lowering pasture contamination with infective L3 stage larvae at the start of grazing season, while it remained significantly higher on the control farm. The positive effect of the spring treatment of ewes was reflected by increased body weight gains (BWG) in lambs in the first half of the grazing season. Following the second treatment, the weight gains in lambs on the T(S)T farm were higher compared to lambs on the TT farm, while BWG in the control lambs started to decrease. The difference was also substantiated by the body condition scores (BCS) and dag scores (DS) of lambs, which were highest on the T(S)T farm compared with those on the control and TT farms. Conclusions The results of this study show that both treatment strategies were useful in reducing clinical effects (BCS and DS) of gastrointestinal nematode parasitism and increasing the performance in lambs. Furthermore, on the T(S)T farm some of animals were left in refugia, helping to slow down the development of anthelmintic resistance (AR) in future.

The presence ofTaenia solium DNA from eggs in soils around the households in four Tanzanian villages in Kongwa district were analysed in relation to seasonal fluctuations and infection risk implications. A total of 192 pooled soil samples from five sampling points per household were examined by droplet digital Polymerase Chain Reaction (ddPCR) from 96 pig-keeping households both during the dry and rainy seasons. The pooled samples were first processed by a flotation-double sieving technique, followed by screening for worm DNA employing universal primers targeting the mitochondrial cytochrome c oxidase subunit I (cox1) gene of human taeniid species and some other helminths. All DNA positive samples were later confirmed by a specific ddPCR probe assay targeting the mitochondrial cox1 gene of T. solium. A total of 17.2% (n = 33) samples were positive with the universal ddPCR, whereas T. solium DNA was confirmed by the specific ddPCR only in 3.1% (n = 3) of the surveyed households. The detection of T. solium DNA in this study spells out a low risk of exposure to T. solium eggs from contaminated household soil. Based on our results, ddPCR seems to be a promising technology for screening T. solium eggs in soil.

The aim of this study was to investigate exposure to porcine cysticercosis (PC) and associated risk factors in the Kongwa District, eastern-central Tanzania. For the first time a cross-sectional investigation of the seroprevalence in pigs using a commercial genus specific cysticercosis enzyme linked immunosorbent assay (apDia Ag-ELISA) was undertaken in eastern-central Tanzania. Moreover, the identity of suspected T. solium cysts from pigs in the study area were confirmed by sequencing parasites’ mitochondrial cox1 gene. Structured questionnaires and direct observations were used to investigate risk factors associated with parasite transmission. A total of 102 pig-keeping households were surveyed during the dry season between July and August 2017 and 126 households in the rainy season between March and April 2018. Of the 447 examined pigs, 77 (17%, 95% C.I. 14%–20%) tested positive in the ELISA. Seroprevalence was higher in pigs examined during the rainy (21%, 95% C.I. 16%–26%) than dry (12%, 95% C.I. 7%–17%) season (p = 0.019). Eight cyst-positive-pigs were confirmed to be infected with T. solium by sequencing. Risk factors associated with PC seropositivity included origin of piglets or pigs (OR = 0.27, 95% C.I. 0.13–0.42, p = 0.001), socioeconomic factors and pig production system (OR = 0.22, 95% C.I. 0.07–0.37, p = 0.005) and sanitation and hygiene practices (OR = 0.19, 95% C.I. 0.04–0.34, p = 0.014). This study has recorded a high Taenia spp. seroprevalence in pigs in Kongwa suggesting the presence of people in the community carrying the adult parasite, Taenia solium. Our findings also suggest risk of infection by T. solium to people in urban centres and cities consuming pigs from rural areas in Kongwa. The high seroprevalence in Kongwa calls for further studies on taeniasis and cysticercosis in the human population in order to determine suitable control strategies. Keywords: Taenia solium, Seroprevalence, Porcine cysticercosis, Antigen, Kongwa, Tanzania

Anthelmintic resistance (AR) to Fasciola hepatica is emerging worldwide. Recently, AR to the adulticide compound albendazole (ABZ) was shown in Argentina and Spain. In Sweden, ABZ treatment failure against F. hepatica was first reported in sheep in 2012. The present study tested the efficacy of ABZ and triclabendazole (TCBZ) in sheep naturally infected with F. hepatica using a combination of three different diagnostic methods: faecal egg counts (FEC), coproantigen ELISA (cELISA) and Fasciola egg hatch test (FEHT). Two deworming trials, in November 2014 and January 2015, were performed on two sheep farms (farms A and B) in south-western Sweden. Except ABZ in November, treatment with ABZ or TCBZ achieved sufficient efficacy (97–100%) against adult F. hepatica on farm A. In contrast, ABZ treatment failed in the sheep flock on farm B, despite low initial faecal egg output. On farm B, ABZ efficacy based on FEC was 67% (95% CI: 35–84) and four of eight ewes tested were coproantigen-positive 21 days post-treatment. Ovicidal activity of ABZ against Fasciola eggs in isolates from both farms and one additional bovine isolate were tested by FEHT to exclude the presence of juvenile flukes and other factors such as dosing failure and poor quality of drug product. Irrespective of drug trial, data from FEHT showed significantly lower ovicidal activity of ABZ for the ovine farm B isolate than for the isolate from farm A. This confirms that the low efficacy of ABZ in sheep flock B was associated with ABZ resistance. Overall, the usefulness of three complementary methods for detection of ABZ resistance in the field was demonstrated.

Echinococcus multilocularis is a zoonotic tapeworm with a sylvatic lifecycle and an expanding range in Europe. Monitoring efforts following its first identification in 2011 in Sweden have focused on the parasite's definitive host, the red fox (Vulpes vulpes). However, identifying rodent intermediate hosts is important to recognize opportunities for parasite transmission. During 2013–2015, livers from a total of 1566 rodents from four regions in Sweden were examined for E. multilocularis metacestode lesions. Species identity of suspect parasite lesions was confirmed by PCR and sequencing. E. multilocularis positive lesions >6 mm in diameter were also examined histologically. One Microtus agrestis out of 187 (0.5%, 95%CI: 0–2.9%), 8/439 (1.8%, 95%CI: 0.8–3.6%) Arvicola amphibius, 0/655 (0%, 95%CI: 0–0.6%) Myodes glareolus, and 0/285 (0%, 95%CI: 0–1.3%) Apodemus spp. contained E. multilocularis metacestode lesions. Presence of protoscoleces was confirmed in the infected M. agrestis and in three of eight infected A. amphibius. Six of the nine positive rodents were captured from the same field. This is the first report of E. multilocularis in intermediate hosts in Sweden. The cluster of positive rodents in one field shows that local parasite prevalence can be high in Sweden despite overall low national prevalence in foxes (

This article investigates the non-fiction book Black Hawk Down (1999) by Mark Bowden, Black Hawk Down the movie (2001) directed by Ridley Scott, and the computer game Delta Force: Black Hawk Down (2003). The article suggests that while the movie and the game must be studied as adaptations of the first text, the tools developed by adaptation studies, and that are typically used to study the transfer of narratives from one media form to another, do not suffice to fully describe the ways in which these narratives change between iterations. To provide a more complete account of these adaptations, the article therefore also considers the shifting political climate of the 9/11 era, the expectations from different audiences and industries, and, in particular, the role that what James Der Derian has termed the Military-Industrial-Media-Entertainment Network (MIME-Net) plays in the production of narrative. The article thus investigates how a specific political climate and MIME-Net help to produce certain adaptations. Based on this investigation, the article argues that MIME-Net plays a very important role in the adaptation of the Black Hawk Down story by directing attention away from historical specificity and nuance, towards the spectacle of war. Thus, in Black Hawk Down the movie and in Delta Force: Black Hawk Down, authenticity is understood as residing in the spectacular rendering of carnage rather than in historical facts. The article concludes that scholarly investigations of the adaptation of military narratives should combine traditional adaptation studies tools with theory and method that highlight the role that politics and complexes such as MIME-Net play within the culture industry.

Control of Fasciola hepatica infection in livestock is based on annual treatment using flukicides such as triclabendazole, albendazole and closantel. However, triclabendazole resistant F. hepatica populations are emerging worldwide and resistance is emerging to albendazole, whereas it has until now never been described for closantel. In Sweden, a topical formulation containing a combination of closantel and ivermectin (Closamectin Pour On) has been registered for use in cattle only since 2011. This study evaluated the efficacy of closantel against F. hepatica in naturally infected beef cattle using both coproantigen and faecal egg count reduction tests. Faecal egg counts (FEC) and coproantigen ELISA examinations were conducted in February 2014 in three beef cattle herds (A, B, C) in south-western Sweden. On each farm, 10 F. hepatica coproantigen-positive and F. hepatica egg-positive animals were allocated after 12–16 weeks of housing into groups and treated topically with a minimum of 20 mg closantel per kg body weight. Faecal samples were collected from selected animals on 0, 7 and 21 day post-treatment (PT). Based on FEC, closantel efficacy 21 days PT was 72% (95% CI: 65–77%) and 97% (95% CI: 95–98%) on farms A and B, respectively. No FEC reduction at all was observed on farm C. In total, 4, 1 and 6 animals remained coproantigen-positive at 21 days PT on farms A, B and C, respectively. Closantel treatment failure was confirmed on two of the farms. As the animals were housed 12–16 weeks before treatment and thereafter during the entire study, failure due to the presence of juvenile flukes was excluded. Although the cause of closantel failure currently remains unclear, development of resistance or/and absorption failure of topical administration should be considered. To our knowledge, this is the first report of closantel treatment failure against F. hepatica in cattle.

Abstract Background A major constraint for the effective control and management of helminth parasites is the lack of rapid, high-throughput, routine diagnostic tests to assess the health status of individual animals and herds and to identify the parasite species responsible for these helminthoses. The capability of a multiplex platform for the simultaneous detection of three pasture associated parasite species was evaluated and compared to existing ELISAs. Methods The recombinant antigens 14.2 kDa ES protein for Cooperia oncophora, major sperm protein for Dictyocaulus viviparus and Cathepsin L1 for Fasciola hepatica were recombinantly expressed either in Escherichia coli or Pichia pastoris. Antigens were covalently coupled onto magnetic beads. Optimal concentrations for coupling were determined following the examination of serum samples collected from experimentally mono-infected animals, before and after their infection with the target species. Absence of cross-reactivity was further determined with sera from calves mono-infected with Haemonchus contortus, Ostertagia ostertagi and Trichostrongylus colubriformis. Examination of negative serum samples was characterised by low median fluorescence intensity (MFI). Results Establishment of the optimal serum dilution of 1:200 was achieved for all three bead sets. Receiver Operating Characteristic analyses were performed to obtain cut-off MFI values for each parasite separately. Sensitivity and specificity at the chosen cut-off values were close to, or 100 % for all bead sets. Examination of serum samples collected on different days post infection from different animals showed a high reproducibility of the assays. Serum samples were additionally examined with two already established ELISAs, an in-house ELISA using the recombinant MSP as an antigen and a DRG ELISA using Cathepsin L1 for liver fluke. The results between the assays were compared and kappa tests revealed an overall good agreement. Conclusions A versatile bead-based assay using fluorescence detection (xMAP® technology) was developed to simultaneously detect antibodies against C. oncophora, D. viviparus and F. hepatica in cattle serum samples. This platform provides rapid, high-throughput results and is highly sensitive and specific in comparison to existing serological as well as coproscopical diagnostic techniques.

A harmonized sampling approach in combination with spatial modelling is required to update current knowledge of fasciolosis in dairy cattle in Europe. Within the scope of the EU project GLOWORM, samples from 3,359 randomly selected farms in 849 municipalities in Belgium, Germany, Ireland, Poland and Sweden were collected and their infection status assessed using an indirect bulk tank milk (BTM) enzyme-linked immunosorbent assay (ELISA). Dairy farms were considered exposed when the optical density ratio (ODR) exceeded the 0.3 cut-off. Two ensemble-modelling techniques, Random Forests (RF) and Boosted Regression Trees (BRT), were used to obtain the spatial distribution of the probability of exposure to Fasciola hepatica using remotely sensed environmental variables (1-km spatial resolution) and interpolated values from meteorological stations as predictors. The median ODRs amounted to 0.31, 0.12, 0.54, 0.25 and 0.44 for Belgium, Germany, Ireland, Poland and southern Sweden, respectively. Using the 0.3 threshold, 571 municipalities were categorized as positive and 429 as negative. RF was seen as capable of predicting the spatial distribution of exposure with an area under the receiver operation characteristic (ROC) curve (AUC) of 0.83 (0.96 for BRT). Both models identified rainfall and temperature as the most important factors for probability of exposure. Areas of high and low exposure were identified by both models, with BRT better at discriminating between low-probability and high-probability exposure; this model may therefore be more useful in practise. Given a harmonized sampling strategy, it should be possible to generate robust spatial models for fasciolosis in dairy cattle in Europe to be used as input for temporal models and for the detection of deviations in baseline probability. Further research is required for model output in areas outside the eco-climatic range investigated.

The geographic distribution of Fasciola hepatica infection in relation to management routines was studied in Swedish dairy herds by testing for F. hepatica antibodies with the enzyme-linked immunosorbent assay (ELISA). In addition, all farmers were sent a questionnaire asking for information about type of production, management routines and historical record of F. hepatica at slaughter. A total of 176 farmers (41%) responded to the questionnaire. A total of 426 bulk tank milk (BTM) samples were randomly selected from the period September to October 2012 representing approximately 10% of all herds in Sweden. The overall seroprevalence was 25% (n = 107; 95% confidence interval = 21-29%) with a concentration of herds located in south-western Sweden. Among the seropositive herds, 31 (29%) had antibody levels indicating production loss. There were no significant differences in seropositivity between organic and conventional herds or due to pasture management routines. The length of grazing period, which increased the risk for heifers, was found to be the most influential factor. A discrepancy was noted between reported F. hepatica presence at meat inspection and herds that were seropositive based on BTM-ELISA results. Although the largest proportion of seropositive BTM samples (80%) came from herds where liver fluke presence had been observed at meat inspection after slaughter, seropositive BTM samples were also diagnosed in five herds (17%) with no remarks at slaughter. In conclusion, F. hepatica is a common parasite in Swedish dairy herds and the month of heifer turn-out and the grazing period length were the most influential factors observed.

We compared four methods for identification of Haemonchus contortus eggs. With increased trade in animals within and between countries and continents, it has become important to correctly identify H. contortus eggs in fecal samples. To validate the outcome of diagnostic tests, sheep feces (n = 38) were collected from naturally infected flocks in Sweden. Subsamples were analyzed with (a) McMaster egg counting; (b) differential counting of eggs after staining with peanut agglutinin (PNA); (c) detection of DNA following amplification by real-time quantitative polymerase chain reaction (qPCR); and (d) loop-mediated isothermal amplification (LAMP). Differences between similar tests (microscopic and molecular) and SD (±SD) were analyzed with Bland–Altman plots and Spearman rank correlation. Strongylid egg counts ranged from 200 to 12,100 eggs per gram (epg) (mean epg ± SD = 1,278 ± 2,049). Microscopy showed presence of H. contortus eggs in 27 (73%) unstained samples and in 28 (76%) samples stained with PNA, whereas 29 samples (78%) tested positive in LAMP and 34 (91%) in qPCR analysis. The cycle threshold (Ct) values with LAMP ranged between 13 and 38 (mean ± SD = 21 ± 7), and those in qPCR between 25 and 49 (mean ± SD = 33 ± 6). In the LAMP and qPCR analyses, seven (19%) and three (8%) samples, respectively, had a cycle threshold (Ct) >35, whereas no reactions were observed in eight (22%) and three (8%) samples, respectively. There was good agreement between the diagnostic tests based on microscopic examination and DNA detection, although the molecular tests were more sensitive. The bias between the microscopy methods (−4.2 ± 11) was smaller than for the molecular tests (−9.8 ± 10). The observed ranking in terms of test sensitivity was: McMaster counting by conventional microscopy

Abstract Background The development of anthelmintic resistance (AR) to macrocyclic lactones in the equine roundworm Parascaris equorum has resulted in benzimidazoles now being the most widely used substance to control Parascaris infections. However, over-reliance on one drug class is a risk factor for the development of AR. Consequently, benzimidazole resistance is widespread in several veterinary parasites, where it is associated with single nucleotide polymorphisms (SNPs) in drug targets encoded by the β-tubulin genes. The importance of these SNPs varies between different parasitic nematodes, but it has been hypothesised that they occur, at low allele frequencies, even in unselected populations. This study investigated whether these SNPs exist in the P. equorum population and tested the hypothesis that BZ resistance can develop from pre-existing SNPs in codons 167, 198 and 200 of the β-tubulin isotype 1 and 2 genes, reported to be associated with AR in strongylids. The efficacy of the oral paste formula fenbendazole on 11 farms in Sweden was also assessed. Methods Two isotype-specific primer pairs were designed, one on either side of the codon 167 and one on either side of codons 198 and 200. A pool of 100 000 larvae was sequenced using deep amplicon sequencing by Illumina HiSeq. Faecal egg count reduction test was used to assess the efficacy of fenbendazole. Results No SNPs were observed in codons 167, 198 or 200 of the β-tubulin isotype 1 or 2 genes of P. equorum, even though 100 000 larvae were sequenced. Faecal egg count reduction testing of fenbendazole showed that this anthelmintic was still 100% effective, meaning that the likelihood of finding high allele frequency of SNPs associated with benzimidazoles resistance in P. equorum was low. Unexpectedly, the allele frequencies observed in single worms were comparable to those in pooled samples. Conclusions We concluded that fenbendazole does not exert selection pressure on the β-tubulin genes of isotypes 1 and 2 in P. equorum. The fact that no pre-existing SNPs were found in codons 167, 198 and 200 in P. equorum also illustrates the difficulties in generalising about AR mechanisms between different taxonomic groups of nematodes.

Abstract Background Anthelmintic resistance (AR) is an increasing problem for the ruminant livestock sector worldwide. However, the extent of the problem is still relatively unknown, especially for parasitic nematodes of cattle. The effect of ivermectin (IVM) (Ivomec inj.®, Merial) was investigated in Swedish isolates of gastrointestinal nematode (GIN) populations showing signs of AR in the field to further characterise the AR status by a range of in vivo and in vitro methods. Methods Three groups, each of 11 calves, were infected with an equal mixture of third stage larvae (L3) of Cooperia oncophora and Ostertagia ostertagi. Group A was inoculated with an IVM-susceptible laboratory isolate and groups B and C with isolates originating from ‘resistant’ cattle farms. Faecal egg counts (FEC) were monitored from 0 to 45 days post infection (d.p.i.), and L3 were harvested continuously for larval migration inhibition testing (LMIT) and species-specific PCR (ITS2). At 31 d.p.i., one calf from each group was necropsied and adult worms were recovered pre-treatment. At 35 d.p.i., calves from all groups were injected with IVM at the recommended dose (0.2 mg/kg bodyweight). At 45 d.p.i., another two animals from each group were sacrificed and established gastrointestinal worms were collected and counted. Results A few animals in all three groups were still excreting eggs (50-150 per g faeces) 10 days post IVM injection. However, there was no significant difference in the FEC reductions in groups A (95%; 95% CI 81-99), B (98%; 92-100) and C (99%; 97-100) between 35 and 44 d.p.i. Furthermore, LMIT showed no significant difference between the three groups. Approximately 100 adult O. ostertagi were found in the abomasum of one calf (group B), whereas low to moderate numbers (400-12 200) of C. oncophora remained in the small intestine of the calves in all three groups at 45 d.p.i. PCR on L3 harvested from faecal samples up to 10 days post treatment showed a ratio of 100% C. oncophora in the calves inoculated with isolates A and B, whereas C also had 8% O. ostertagi. Conclusions Overall, this experiment showed that the animals were successfully treated according to the Faecal egg count reduction test (FECRT) standard (≥ 95% reduction). However, several adult worms of the dose-limiting species C. oncophora demonstrably survived the IVM treatment.

Infections with parasitic helminths (nematodes and trematodes) represent a significant economic and welfare burden to the global ruminant livestock industry. The increasing prevalence of anthelmintic resistance means that current control programmes are costly and unsustainable in the long term. Recent changes in the epidemiology, seasonality and geographic distribution of helminth infections have been attributed to climate change. However, other changes in environment (e.g., land use) and in livestock farming, such as intensification and altered management practices, will also have an impact on helminth infections. Sustainable control of helminth infections in a changing world requires detailed knowledge of these interactions. In particular, there is a need to devise new, sustainable strategies for the effective control of ruminant helminthoses in the face of global change. In this paper, we consider the impact of helminth infections in grazing ruminants, taking a European perspective, and identify scientific and applied priorities to mitigate these impacts. These include the development and deployment of efficient, high-throughput diagnostic tests to support targeted intervention, modelling of geographic and seasonal trends in infection, more thorough economic data and analysis of the impact of helminth infections and greater translation and involvement of end-users in devising and disseminating best practices. Complex changes in helminth epidemiology will require innovative solutions. By developing and using new technologies and models, the use of anthelmintics can be optimised to limit the development and spread of drug resistance and to reduce the overall economic impact of helminth infections. This will be essential to the continued productivity and profitability of livestock farming in Europe and its contribution to regional and global food security.