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Deep convection in the Labrador Sea supplies large amounts of anthropogenic carbon to the ocean's interior. We use measurements of all four measurable CO 2 system parameters made along AR7W (across Labrador Sea) between 2013 and 2015 to assess the internal consistency of the carbonate system , including, as appropriate, conversion to in situ temperature (T) and pressure (P). The best agreement between measured and calculated values was obtained through combination of T,P-dependent (pH or pCO2 ) and non-dependent (TA or DIC) parameters. Use of the dissociation constants of Mehrbach et al. (1973) as refit by Dickson and Millero (1987) and Lueker et al. (2000) yielded the best internal consistency irrespective of the input parameters used. A Monte Carlo simulation demonstrated that the propagated uncertainty ( i.e. combined standard uncertainty) of calculated parameters of the carbonate system is (a) always larger than the analytical precision of the measurements themselves; (b) strongly dependent on the choice of input parameters and uncertainties; (c) less dependent on choice of the specific set of constants. For calculation of other parameters of the carbonate system from TA and DIC measurements made throughout the Labrador Sea time-series, the estimated combined standard uncertainty of calculated pCO2 and pH based on the Monte Carlo simulation is ~13 μatm and ~0.012 pH units respectively, with accuracy relative to laboratory-based measurement estimated to be between −3 and − 13 μatm and 0.002 and 0.007 pH units. Internal consistency especially at in situ temperature and pressure conditions is important for rapidly developing sensor-based monitoring programs in the region, including measurement of pH and/or pCO2 from gliders, profiling floats and moorings. We highlight uncertainty associated with the large pressure effect on pH and pCO2, and recommend a study of carbonate system internal consistency under deep ocean conditions that addresses pressure effects on calculations.

Anthropogenic climate change profoundly alters the ocean’s environmental conditions, which, in turn, impact marine ecosystems. Some of these changes are happening fast and may be difficult to reverse. The identification and monitoring of such changes, which also includes tipping points, is an ongoing and emerging research effort. Prevention of negative impacts requires mitigation efforts based on feasible research-based pathways. Climate-induced tipping points are traditionally associated with singular catastrophic events (relative to natural variations) of dramatic negative impact. High-probability high-impact ocean tipping points due to warming, ocean acidification, and deoxygenation may be more fragmented both regionally and in time but add up to global dimensions. These tipping points in combination with gradual changes need to be addressed as seriously as singular catastrophic events in order to prevent the cumulative and often compounding negative societal and Earth system impacts.There are no data underlying this work.

Transport of freshwater from the Labrador Shelf into the interior Labrador Sea has the potential to impact deep convection via its influence on the salinity of surface waters. To examine this transport, the authors deployed two underwater gliders on a mission to traverse the continental shelf break multiple times between 5 July and 22 August 2014, the period when Arctic meltwater has historically peaked in transport down the Labrador Shelf. The field campaign yielded a unique dataset of temperature, salinity, and oxygen across the shelf break to a depth of 1000 m at unprecedented spatial resolution. Two mechanisms of cross-shelf transport were examined: Ekman transport and transport due to mesoscale eddies. Ekman transport is quantified using satellite wind stress and near-surface hydrographic properties, and eddy-induced transport is scaled using a parameterized eddy diffusivity and thickness gradients of layers of uniform potential density, as well as the tracer gradients along those isopycnals. Both the Ekman and eddy terms transport high-oxygen and low-salinity water from the shelf to the Labrador Sea during the field campaign. The influence of the eddy-driven oxygen flux from the shelf to the Labrador Sea on oxygen budgets depends strongly on the size of the region over which this eddy flux converges. The deduced offshore transport of freshwater (4 ± 6 mSv; 1 mSv = 103 m3 s−1) from both Ekman and eddy mechanisms, which is likely at a seasonal maximum during this summertime survey, represents about 3% of the annual-mean freshwater flowing through Hudson and Davis Straits but may be an important component of the total freshwater budget of the interior Labrador Sea.

The control of equid gastrointestinal nematodes in developed countries, in particular the cyathostomins, is threatened by high levels of anthelmintic resistance. In recent years, there has been increasing interest in the evaluation of traditional ‘ethnoveterinary’ medicines as alternatives to chemical anthelmintics. The cysteine proteinases (CPs), a group of enzymes derived from fruits such as papaya (Carica papaya), pineapple (Ananas comosus) and figs (Ficus spp.), have shown good efficacy against adult stages of a range of parasitic nematodes, in vitro and in vivo. The efficacy of CPs against cyathostomins remains to be explored. In this study, the efficacy of a crude preparation of CPs, papaya latex supernatant (PLS), against the free-living stages of cyathostomins was evaluated using two in vitro tests, the egg hatch test (EHT) and the larval migration inhibition test (LMIT). It was demonstrated that PLS had a potent effect in the EHT, with EC-50 values in the range of 0.12–0.22 μM. At concentrations above 6.25 μM the eggs did not develop, below this concentration the L1 developed but they lost integrity of the cuticle upon hatching. These effects were inhibited by pre-incubation of PLS with the CP inhibitor L-trans-epoxysuccinyl- l -leucylamido-(4-guanidino butane) (E64), indicating that CPs were responsible for the anti-parasitic activity. A dose-dependent inhibition of migration of third stage larvae (L3) in the LMIT was demonstrated at higher concentrations of PLS, with EC-50 values in the range of 67.35–106.31 μM. Incubation of PLS with E64 prior to use in the LMIT did not reverse the anti-migratory effect, suggesting that CPs were not responsible for the reduced migration of cyathostomin L3 and that PLS also contains an additional active compound. This is the first report of PLS and/or CPs showing activity against the free-living stages of a parasitic helminth. In addition, it suggests that cyathostomins are highly sensitive to the effects of CPs and further evaluation of their efficacy against parasitic stages and in vivo are strongly indicated.

This is the second of two papers on observational timeseries of top of ocean heat capture. The first reports hourly and daily meridional central tropical Pacific top 3 m timeseries showing high Southern Hemisphere evaporation (2.67 m yr−1) and Northern Hemisphere trapped heat (12 MJ m−2 day−1). We suggested that wind drift/geostrophic stratified gyre circulation transported warm water to the Arctic and led to three phases of Arctic basal ice melt and fluxes of brackish nutrient-rich waters to north Atlantic on centennial timescales. Here we examine daily top metre 1904–2012 timeseries at Isle of Man west coast ~54° N for evidence of tropical and polar surface waters. We compare these to Central England (CET) daily land-air temperatures and to Arctic floating ice heat content and extent. We find three phases of ocean surface heating consistent with basal icemelt buffering greenhouse gas warming until a regime shift post-1986 led to the modern surface temperature rise of ~1 °C in 20 yr. Three phases were: warming +0.018 °C yr−1 from 1904–1939, slight cooling −0.002 °C yr−11940–86 and strong warming +0.037 °C yr−1 1986–2012. For the same periods CET land-air showed: warming +0.015 °C yr−1, slight cooling −0.004 °C yr−1, about half SST warming at +0.018 °C yr−1. The mid-century cooling and a 1959/1963 hot/cold event is consistent with sunspot/solar radiation maximum 1923–2008 leading to record volumes of Arctic ice meltwater and runoff that peaked in 1962/3 British Isles extreme cold winter. The warming Arctic resulted in wind regime and surface water regime shifts post 1986. This coincides with the onset of rapid Arctic annual ice melt. Continued heat imbalance is likely to lead to tidewater glacier basal icemelt and future sealevel rise after remaining relatively stable over 4000 yr. Our work needs confirmation by further fieldwork concentrating on the dynamics and thermodynamics of ocean top 3 m that controls the 93 % anthropogenic global warming in the oceans. This may be done most cost-effectively through focussed multidisciplinary scientific research adaptively managed and funded.

Cyathostomins are the most important gastrointestinal nematode infecting equids. Their effective control is currently under threat due to widespread resistance to the broad spectrum anthelmintics licenced for use in equids. In response to similar resistance issues in other helminths, there has been increasing interest in alternative control strategies, such as bioactive plant compounds derived from traditional ethnoveterinary treatments. This study used an evidence-based approach to evaluate the potential use of plant extracts from the UK and Ethiopia to treat cyathostomins. Plants were shortlisted based on findings from a literature review and additionally, in Ethiopia, the results of a participatory rural appraisal (PRA) in the Oromia region of the country. Systematic selection criteria were applied to both groups to identify five Ethiopian and four UK plants for in vitro screening. These included Acacia nilotica (L.) Delile, Cucumis prophetarum L., Rumex abyssinicus Jacq., Vernonia amygdalina Delile. and Withania somnifera (L.) Dunal from Ethiopia and Allium sativum L. (garlic), Artemisia absinthium L., Chenopodium album L. and Zingiber officinale Roscoe. (ginger) from the UK. Plant material was collected, dried and milled prior to hydro-alcoholic extraction. Crude extracts were dissolved in distilled water (dH2O) and dimethyl sulfoxide (DMSO), serially diluted and screened for anthelmintic activity in the larval migration inhibition test (LMIT) and the egg hatch test (EHT). Repeated measures ANOVA was used to identify extracts that had a significant effect on larval migration and/or egg hatch, compared to non-treated controls. The median effective concentration (EC-50) for each extract was calculated using PROBIT analysis. Of the Ethiopian extracts A. nilotica, R. abyssinicus and C. prophetarum showed significant anthelmintic activity. Their lowest EC-50 values were 0.18 (confidence interval (CI): 0.1-0.3), 1.1 (CI 0.2-2.2) and 1.1 (CI 0.9-1.4)mg/ml, respectively. All four UK extracts, A. sativum, C. album, Z. officinale and A. absinthium, showed significant anthelmintic activity. Their lowest EC-50 values were 1.1 (CI 0.9-1.3), 2.3 (CI 1.9-2.7) and 0.3 (CI 0.2-0.4)mg/ml, respectively. Extract of A. absinthium had a relatively low efficacy and the data did not accurately fit a PROBIT model for the dose response relationship, thus an EC-50 value was not calculated. Differences in efficacy for each extract were noted, dependent on the assay and solvent used, highlighting the need for a systematic approach to the evaluation of bioactive plant compounds. This study has identified bioactive plant extracts from the UK and Ethiopia which have potential as anthelmintic forages or feed supplements in equids.

Discrepancies between historical sea surface temperature (SST) datasets have been partly ascribed to use of different adjustments to account for variable measurement methods. Until recently, adjustments had only been applied to bucket temperatures from the late 19th and early 20th centuries, with the aim of correcting their supposed coolness relative to engine cooling water intake temperatures. In the UK Met Office Hadley Centre SST 3 dataset (HadSST3), adjustments have been applied over its full duration to observations from buckets, buoys and engine intakes. Here we investigate uncertainties in the accuracy of such adjustments by direct field comparison of historical and modern methods of shipboard SST measurement. We compare wood, canvas and rubber bucket temperatures to 3 m seawater intake temperature along a central tropical Pacific transect conducted in May and June 2008. We find no average difference between the temperatures obtained with the different bucket types in our short measurement period (∼1 min). Previous field, lab and model experiments have found sizeable temperature change of seawater samples in buckets of smaller volume under longer exposure times. We do, however, report the presence of strong near-surface temperature gradients day and night, indicating that intake and bucket measurements cannot be assumed equivalent in this region. We thus suggest bucket and buoy measurements be considered distinct from intake measurements due to differences in sampling depth. As such, we argue for exclusion of intake temperatures from historical SST datasets and suggest this would likely reduce the need for poorly field-tested bucket adjustments. We also call for improvement in the general quality of intake temperatures from Voluntary Observing Ships. Using a physical model we demonstrate that warming of intake seawater by hot engine room air is an unlikely cause of overly warm intake temperatures. We suggest that reliable correction for such warm errors is not possible since they are largely of unknown origin and can be offset by real near-surface temperature gradients.

Equine gastrointestinal nematodes are ubiquitous; in horses that graze contaminated pasture and that are not treated appropriately, large numbers of worms can accumulate, which can lead to serious clinical disease. Nematode control has traditionally followed interval treatment regimens, which involve regular anthelmintic administration to all horses based on the strongyle egg reappearance periods of each drug, usually defined around the time of licensing. Interval treatment programmes have resulted in substantial reductions in large strongyle disease, but have made major contributions to the development of anthelmintic resistance, particularly in cyathostomins. Cyathostomin resistance to 2 of the 3 available anthelmintic classes is widespread, and resistance to both classes in single populations is not uncommon. Reduced efficacy of the most commonly used macrocyclic lactone anthelmintics, as measured by shortened egg reappearance periods after treatment, is emerging in cyathostomins. Macrocyclic lactone resistance is also now commonly reported in Parascaris equorum on stud farms. Faecal worm egg counts (FWEC) are increasingly being used as part of targeted approaches to parasite control, whereby only those horses with moderate to high FWEC within a group are treated with an anthelmintic. The objective of this approach is to reduce environmental contamination, while leaving a proportion of the worm population in some horses unexposed to selection pressure for anthelmintic resistance. This article reviews recent findings in equine parasitology research that will underpin guidelines for control, with a particular focus on how to optimise the value of FWEC methodologies and anthelmintic efficacy analyses.

†‡ § # Joint last authors. Summary Reasons for performing study: Anthelmintic resistance in equine gastrointestinal nematodes is a threat to equine health and welfare. Detailed knowledge of anthelmintic use and parasite control methods is a prerequisite to identification of potential risk factors for resistance. Objectives: To identify parasite control practices employed by equine owners in Scotland and investigate management factors associated with anthelmintic resistance. Study design: Questionnaire study of equine parasite control in Scotland. Methods: Questionnaires were available electronically, distributed at a conference and mailed to clients. Key areas explored included general background, grazing management, anthelmintic treatment practices and use of diagnostic tests. Results: A total of 193 responses detailing information on parasite control programmes of 993 equids were analysed. Moxidectin (MOX) and ivermectin or related combination products were the most commonly administered anthelmintics in the preceding 12 months. Treatments licensed for use against cyathostomin encysted larvae and tapeworms were administered by 80% and 90% of respondents, respectively. This was often achieved through indiscriminate use of MOX and MOX-praziquantel products. Faecal egg count (FEC) analysis had been performed on 62% of yards and regular use of FECs reduced annual anthelmintic treatment frequency. Veterinarians had the greatest influence on control practices. While 40% of respondents believed that they practised targeted dosing, this was not associated with delaying treatment beyond the egg reappearance period of the anthelmintic used. Conclusions: Responses indicated increasing veterinary involvement and use of FECs. The majority of respondents administered anthelmintics licensed against cyathostomin encysted larvae and tapeworms. However, responses suggested that owners did not understand the definition of ‘targeted’ dosing regimens. Potential relevance: The high frequency of MOX use represents a potential risk factor for macrocyclic lactone resistance. As veterinarians were the most influential factor in anthelmintic choice, awareness of macrocyclic lactone resistance and potential risk factors for its development and spread should be incorporated into client advice.

Sea Surface Temperature (SST) measurements have been obtained from a variety of different platforms, instruments and depths over the post-industrial period. Today most measurements come from ships, moored and drifting buoys and satellites. Shipboard methods include temperature measurement of seawater sampled by bucket and in engine cooling water intakes. Engine intake temperatures are generally thought to average a few tenths of a °C warmer than simultaneous bucket temperatures. Here I review SST measurement methods, studies comparing shipboard methods by field experiment and adjustments applied to SST datasets to account for variable methods. In opposition to contemporary thinking, I find average bucket-intake temperature differences reported from field studies inconclusive. Non-zero average differences often have associated standard deviations that are several times larger than the averages themselves. Further, average differences have been found to vary widely between ships and between cruises on the same ship. The cause of non-zero average differences is typically unclear given the general absence of additional temperature observations to those from buckets and engine intakes. Shipboard measurements appear of variable quality, highly dependent upon the accuracy and precision of the thermometer used and the care of the observer where manually read. Methods are generally poorly documented, with written instructions not necessarily reflecting actual practices of merchant mariners. Measurements cannot be expected to be of high quality where obtained by untrained sailors using thermometers of low accuracy and precision.

Summary Reasons for performing study Cyathostomins comprise 50 helminth species, considered the most problematic equine endoparasites. Three classes of anthelmintic are currently licensed for their control, namely the benzimidazoles (fenbendazole), tetrahydropyrimidines (pyrantel) and macrocyclic lactones (ivermectin and moxidectin). Anthelmintic resistance in cyathostomins is common. With no new classes expected in the near future, it is essential to determine the efficacy of the available anthelmintics to inform future control programmes. Objectives To determine the efficacy of all 3 anthelmintic classes against strongyles in equids on livery yards in east and central Scotland. Study design Anthelmintic efficacy testing using the faecal egg count reduction test (FECRT). Methods FECRTs were performed on equids with initial strongyle faecal egg counts (FECs) of ≥50 eggs per gram. Efficacy was determined by comparing pretreatment (Day 0) and 14 days post treatment (Day 14) FECs. The following chemicals were tested: fenbendazole, pyrantel, ivermectin and moxidectin. Group arithmetic mean FECR of >90% for fenbendazole and pyrantel, and >95% for ivermectin and moxidectin, represented efficacy, whereas lower mean FECR indicated potential resistance. Results A total of 447 FECRTs were performed on 15 yards, as follows (the numbers in parentheses represent the number of yards each anthelmintic was tested on): 55 equids (7 yards) fenbendazole, 111 (8 yards) pyrantel, 163 (13 yards) ivermectin and 118 (10 yards) moxidectin. Fenbendazole resistance was documented on all yards (range of mean FECR, 15.8–83.4%), whereas pyrantel (90.4–99.6%), ivermectin (99.5–100%) and moxidectin (99.4–100%) treatment had acceptable efficacy. Conclusions Reduced efficacy of fenbendazole was widespread, whereas >90% efficacy was found after pyrantel, and >95% efficacy after ivermectin and moxidectin. Overall, efficacies were higher than reported previously in Europe and the USA, potentially reflecting differences in management and anthelmintic use on the yards surveyed. Potential relevance The use of fenbendazole for strongyle control in Scotland should be questioned. Targeted use of pyrantel should be encouraged to reduce reliance on macrocyclic lactones. Further work to correlate management practices with the presence of anthelmintic resistance is warranted. The Summary is available in Chinese – see Supporting information.

Summary Roundworms and flatworms that affect donkeys can cause disease. All common helminth parasites that affect horses also infect donkeys, so animals that co-graze can act as a source of infection for either species. Of the gastrointestinal nematodes, those belonging to the cyathostomin (small strongyle) group are the most problematic in UK donkeys. Most grazing animals are exposed to these parasites and some animals will be infected all of their lives. Control is threatened by anthelmintic resistance: resistance to all 3 available anthelmintic classes has now been recorded in UK donkeys. The lungworm, Dictyocaulus arnfieldi, is also problematical, particularly when donkeys co-graze with horses. Mature horses are not permissive hosts to the full life cycle of this parasite, but develop clinical signs on infection. In contrast, donkeys are permissive hosts without displaying overt clinical signs and act as a source of infection to co-grazing horses. Donkeys are also susceptible to the fluke, Fasciola hepatica. This flatworm can be transmitted, via snails and the environment, from ruminants. As with cyathostomins, anthelmintic resistance is increasing in fluke populations in the UK. A number of the anthelmintic products available for horses do not have a licence for use in donkeys, and this complicates the design of parasite control programmes. As no new equine anthelmintic classes appear to be near market, it is important that the efficacy of currently effective drugs is maintained. It is important that strategies are used that attempt to preserve anthelmintic efficacy. These strategies should be based on the concept that the proportion of worms in a population not exposed to anthelmintic at each treatment act as a source of ‘refugia’. The latter is an important factor in the rate at which resistance develops. Thus, it is imperative that parasite control programmes take into account the need to balance therapy to control helminth-associated disease with the requirement to preserve anthelmintic effectiveness.

Summary Reasons for performing study: Improved education of veterinarians and equine owners/managers is essential in implementing parasite control strategies that are less reliant on chemicals. Methods: This questionnaire study, conducted on 61 UK Thoroughbred (TB) establishments during 2009 and 2010, was designed to obtain an understanding of current helminth control practices on studs. To our knowledge, this is the first occasion that statements obtained from TB studs via questionnaire have been supported by statistical analysis. Results: Despite many respondents indicating high levels of concern regarding anthelmintic resistance, 56% of these establishments that received visiting equines co-grazed these animals with permanent stock and 25% of respondents), indicates that many stud owners/managers are not aware of the strong risk factors for the development of anthelmintic resistance. Few studs had conducted faecal egg count (FEC) analysis in the past and only 22% indicated that they considered this form of analysis beneficial in determining anthelmintic choice. Conclusions and potential relevance: The challenge now is to convince stud owners/managers to deviate from their current practices to control strategies that are more likely to preserve anthelmintic efficacy. Veterinarians need to get more involved in implementing these control strategies, with better emphasis placed on the role of diagnostic tests in facilitating targeted treatments and in investigating anthelmintic sensitivity in the associated nematode populations.

Summary Horses worldwide are exposed to a complex mixture of intestinal parasitic helminths. When burdens are high, these parasites can seriously compromise health and welfare. Some helminth species have an extremely high prevalence and are difficult to control, not least because there is a limited understanding of their most basic biology. Furthermore, levels of resistance to some of the commonly used anthelmintics are widespread and increasing. The cyathostomins are the most common nematode species affecting equids worldwide. Within this group of parasites are more than 50 different species. Until recent research activities, little was known about the contribution that individual species make to clinical disease, parasite epidemiology and anthelmintic resistance. This review describes some of the recent research advances in the understanding of cyathostomins in these areas. As part of the research effort, molecular tools were developed to facilitate identification of the non-parasitic stages of cyathostomins. These tools have proved invaluable in the investigation of the relative contributions that individual species make to the pathology and epidemiology of mixed infections. At the more applied level, research has also progressed in the development of a diagnostic test that will allow numbers of cyathostomin encysted larvae to be estimated. This test utilises cyathostomin-specific serum antibody responses as markers of infection. As anthelmintic resistance will be the major constraint on parasite control in future, researchers are actively investigating mechanisms of drug resistance and how to improve the detection of resistance in the field. Recent developments in these areas are also outlined.

Summary Like many other complex human disorders of unknown aetiology, autoimmune-mediated type 1 diabetes may ultimately be controlled via a therapeutic approach that combines multiple agents, each with differing modes of action. The numerous advantages of such a strategy include the ability to minimize toxicities and realize synergies to enhance and prolong efficacy. The recognition that combinations might offer far-reaching benefits, at a time when few single agents have yet proved themselves in well-powered trials, represents a significant challenge to our ability to conceive and implement rational treatment designs. As a first step in this process, the Immune Tolerance Network, in collaboration with the Juvenile Diabetes Research Foundation, convened a Type 1 Diabetes Combination Therapy Assessment Group, the recommendations of which are discussed in this Perspective paper.

A wide range of proteins belonging to the SCP/TAPS “family” has been described for various eukaryotic organisms, including plants and animals (vertebrates and invertebrates, such as helminths). Although SCP/TAPS proteins have been proposed to play key roles in a number of fundamental biological processes, such as host–pathogen interactions and defence mechanisms, there is a paucity of information on their genetic relationships, structures and functions, and there is no standardised nomenclature for these proteins. A detailed analysis of the relationships of members of the SCP/TAPS family of proteins, based on key protein signatures, could provide a foundation for investigating these areas. In this article, we review the current state of knowledge of key SCP/TAPS proteins of eukaryotes, with an emphasis on those from parasitic helminths, and undertake a comprehensive, systematic phylogenetic analysis of currently available full-length protein sequence data (considering characteristic protein signatures or motifs) to infer relationships and provide a framework (based on statistical support) for the naming of these proteins. This framework is intended to guide genomic and molecular biological explorations of key SCP/TAPS molecules associated with infectious diseases of plants and animals. In particular, fundamental investigations of these molecules in parasites and the integration of structural and functional data could lead to new and innovative approaches for the control of parasitic diseases, with important biotechnological outcomes.

Cyathostomins are recognised as the primary parasitic pathogens of horses. Despite the use of benzimidazole (BZ) anthelmintics in horses for more than 40 years and widespread drug resistance in the field, the mechanisms of resistance to this drug class in cyathostomins are not fully understood. The results presented here constitute a detailed comparison of beta tubulin gene mutations and mRNA transcript levels in populations of BZ-susceptible (BZ-S) and -resistant (BZ-R) cyathostomins. Full-length cDNA sequences were generated from individual parasites of four (n=24) and two (n=19) cyathostomin species for isotypes 1 and 2, respectively. Levels of intra- and inter-specific nucleotide sequence variation were comparable with previous findings and single amino acid substitutions were observed at several locations. On comparison of BZ-S and BZ-R parasites, differences were consistently observed at only two sites, codons 167 and 200 of the beta tubulin isotype 1 gene. Four populations of parasites were genotyped at these two loci by pyrosequencing; one that was fenbendazole (FBZ)-sensitive (FBZ-rS), two that were FBZ-resistant (FBZ-R1 and -R2) and one that was oxibendazole-resistant (OBZ-R), as previously assessed by faecal egg count reduction tests. This analysis revealed statistically significant differences between FBZ-rS and FBZ-R populations at both loci and this was highly significant for codon 167. For the OBZ-R population, the only significant difference compared with the FBZ-rS population was observed at codon 200. These observations suggest that mutations at codons 167 and 200 are important in BZ resistance and raise the possibility that selection at different loci may occur in FBZ- and OBZ-resistant parasites. Multiple parasites (n=158) were genotyped for both codons 167 and 200, the majority of which showed homozygous 'resistant' mutations at one locus only and none showed homozygous 'resistant' genotypes at both loci. No significant differences in mRNA levels of beta tubulin isotypes 1 and 2 were observed between the FBZ-rS and FBZ-R1 populations.

Suppression subtractive hybridisation was used to enrich genes expressed in a stage-specific manner in infective, exsheathed L3s (xL3) versus early L4s of the ovine nematode, Teladorsagia circumcincta prior to gene expression profiling by microarray. The 769 cDNA sequences obtained from the xL3-enriched library contained 361 unique sequences, with 292 expressed sequence tags (ESTs) being represented once ("singletons") and 69 sequences which were represented more than once (overlapping and non-overlapping "contigs"). The L4-enriched EST dataset contained 472 unique sequences, with 314 singletons and 158 contigs. Of these 833 sequences, 85% of the xL3 sequences and 86% of the L4 sequences exhibited homology to known genes or ESTs derived from other species of nematode. Quantitative differential expression (P0.05) was demonstrated for 563 (68%) of the ESTs by microarray. Within the L3-specific dataset, more than 30% of the transcripts represented the enzyme, guanosine-5'-triphosphate (GTP)-cyclohydrolase, which is the first and rate-limiting enzyme of the tetrahydrobiopterin synthesis pathway and may be involved in critical elements of larval development. In L4s, proteolytic enzymes were highly up-regulated, as were collagens and a number of previously characterised secretory proteins, reflecting the rapid growth of these larvae in abomasal glands. Nucleotide sequence data reported in this paper are available in the EMBL, GenBank and DDJB databases under accession numbers AM 743198-AM 744942.

The global increase in anthelmintic resistant nematodes of ruminants, together with consumer concerns about chemicals in food, necessitates the development of alternative methods of control for these pathogens. Subunit recombinant vaccines are ideally placed to fill this gap. Indeed, they are probably the only valid option for the long-term control of ruminant parasitic nematodes given the increasing ubiquity of multidrug resistance in a range of worm species across the world. The development of a subunit multicellular parasite vaccine to the point of practical application would be a groundbreaking step in the control of these important endemic infections of livestock. This review summarizes the current status of subunit vaccine development for a number of important gastrointestinal nematodes of cattle and sheep, with a focus on the limitations and problems encountered thus far, and suggestions as to how these hurdles might be overcome.

SUMMARY Groups of yearling sheep, which had been trickle infected with Teladorsagia circumcincta for 8 weeks and then drenched, were challenged with 50 000 T. circumcincta larvae together with groups of worm-free controls. Fewer parasites and a greater proportion of early fourth stage larvae were recovered from previously infected sheep compared to controls. Worm loss and arrested development were evident by 5 days after challenge whereas growth retardation of developing worms was observed by day 10. In the previously infected sheep a secondary IgA response was observed in the efferent gastric lymph from 5 days post-infection. Western blot analysis showed the lymph IgA to be predominantly dimeric and nonsecretory in nature and that the somatic antigens recognized were predominantly in the 100‐250 kDa range. The concentration of IgA in lymph was always higher than in blood and in the previously infected sheep increased fivefold 8 days post-challenge in contrast to blood where IgA levels were unchanged. The timing of the response suggested that it occurred too late to have been the cause of worm loss or arrested development, though it may have retarded the growth of developing parasites .

Tumour necrosis factor alpha (TNF-alpha) is a pro-inflammatory cytokine that plays a crucial role in the regulation of inflammatory and immune responses. In all vertebrate species the genes encoding TNF-alpha are located within the major histocompatability complex. In the horse TNF-alpha has been ascribed a role in a variety of important disease processes. Previously two single nucleotide polymorphisms (SNPs) have been reported within the 5' un-translated region of the equine TNF-alpha gene. We have examined the equine TNF-alpha promoter region further for additional SNPs by analysing DNA from 131 horses (Equus caballus), 19 donkeys (E. asinus), 2 Grant's zebras (E. burchellii boehmi) and one onager (E. hemionus). Two further SNPs were identified at nucleotide positions 24 (T/G) and 452 (T/C) relative to the first nucleotide of the 522 bp polymerase chain reaction product. A sequence variant at position 51 was observed between equidae. SNaPSHOTtrade mark genotyping assays for these and the two previously reported SNPs were performed on 457 horses comprising seven different breeds and 23 donkeys to determine the gene frequencies. SNP frequencies varied considerably between different horse breeds and also between the equine species. In total, nine different TNF-alpha promoter SNP haplotypes and their frequencies were established amongst the various equidae examined, with some haplotypes being found only in horses and others only in donkeys or zebras. The haplotype frequencies observed varied greatly between different horse breeds. Such haplotypes may relate to levels of TNF-alpha production and disease susceptibility and further investigation is required to identify associations between particular haplotypes and altered risk of disease.

Teladorsagia circumcincta is a common, pathogenic abomasal nematode of sheep. In order to improve disease control in parasite isolates resistant to several anthelmintics, alternative methods must be sought. Sheep develop acquired immunity to T. circumcincta so vaccination is a valid option for control. For this reason, we are investigating parasite excretory/secretory products for molecules, which have potential to invoke protective immunity against T. circumcincta. Here, we describe experiments in which we identified a novel, immunogenic cathepsin F secreted by L4 T. circumcincta. This protease, initially identified by mass spectrometry analysis, is the most abundant molecule in excretory/secretory products released in vitro by T. circumcincta harvested at 5, 6 or 9 days p.i. and is a target of specific, local IgA responses in sheep which are immune to challenge infection. The full-length cDNA encoding this secreted protease was isolated. Sequence and phylogenetic analyses indicated that the protease (designated T. circumcincta cathepsin F-1, Tci-CF-1) belongs to the cathepsin F class and exhibits greatest identity (>60%) to expressed sequence tags present in the Ostertagia ostertagi and Haemonchus contortus expressed sequence tag databases. Tci-CF-1 also displays high identity to hypothetical proteins identified in the genomes of Caenorhabditis elegans and Caenorhabditis briggsae, both proteins having been described as cathepsin F enzymes. Specific inhibitor binding assay of larval excretory/secretory products confirmed the classification of this excretory/secretory component as a cathepsin F. Reverse transcription-PCR studies indicated that Tci-cf-1 is developmentally regulated and is particular to the host parasitic stages of T. circumcincta. The abundance, immunogenicity and temporal expression pattern of Tci-CF-1 make this a potential vaccine candidate for teladorsagiosis.

This study describes the isolation and characterisation of beta tubulin isotype 2 cDNA sequences from two common species of cyathostomin, Cylicocyclus nassatus and Cyathostomum catinatum. The full-length cDNAs for these species were 1709 and 1753 bp in length, respectively, including 1350 bp of sequence inferred to encode 450 amino acids of peptide sequence. They had greatest identity with previously characterised isotype 2 sequences from Trichostrongylus colubriformis, Cooperia oncophora and Haemonchus contortus (96% for C. nassatus and 95% for C. catinatum), and grouped together with these sequences in phylogenetic analysis. Both cyathostomin beta tubulin isotype 2 sequences contained the isotype-specific carboxyl terminal region described previously in other nematode species. Alignment with beta tubulin isotype 1 proteins from other trichostrongyloids, indicated 95 and 94% identity for the isotype 2 sequences of C. nassatus and C. catinatum, respectively. This comparison revealed 14 isotype-specific amino acid substitutions. Also, 2605 bp of beta tubulin isotype 2 genomic DNA sequence were isolated from C. nassatus. Comparison with the previously published isotype 1 gene of C. nassatus indicated differences in genomic organisation between the two isotypes. Reverse transcriptase (RT)-polymerase chain reaction analysis revealed constitutive temporal expression of beta tubulin isotype 1, whilst isotype 2 appeared to be developmentally expressed, with transcripts detected only in RNA derived from adult parasites.

Proteomics has come to the forefront in the post-genomic era. The ability to compare and identify proteins expressed in a particular cell type under specific physiological or pathological states requires a range of technologies, including separation of complex protein or peptide mixtures, densitometry-based or isotope-coded methods for comparison of multiple proteomes, and mass spectrometric methods for identification of individual low abundance proteins. Although an emergent technology, thus far, proteomics has provided new perspectives on many problems in biomedical science. In parasitology, proteomics has been used to answer specific biological questions relating to survival and development, and also to identify candidates for vaccines. Here, we describe an ongoing research programme in which proteomics is being used to identify potential vaccine candidates for the bovine lungworm, Dictyocaulus viviparus. This work is focusing on antibody responses to the adult parasite excretory/secretory (ES) products, with selection of candidate antigens based on differential screening with serum from immune versus non-immune animals to simplify the proteome and the ensuing analytical challenges. Thus far, we have identified seven candidate proteins using this strategy. Of these, one protein showed significant identity to a previously cloned gene from D. viviparus, whilst the other six proteins have shown no significant identities. Isolation of further peptide sequences is now warranted to facilitate cloning of the genes encoding these antigens.

Equine clinical larval cyathostominosis is caused by simultaneous mass emergence of previously inhibited larvae from the mucosa of the colon. Clinical signs include diarrhoea, colic, weight loss and malaise, and in up to 50% of cases, the disease results in death. Cyathostominae spend a large part of their life cycle as larval stages in the intestinal mucosa. Definitive diagnosis is difficult due to the lack of diagnostic methods for pre-patent infection. In the present study, the enzyme-linked immunosorbent assay (ELISA) was used to investigate isotype responses to larval cyathostominae somatic antigen. Measurement of anti-larval IgG(T) responses appeared to have the most immunodiagnostic potential. An increase in IgG(T) response was detected to crude larval antigen by 5 weeks post-infection (PI) in individual infected ponies. Subsequently, IgG(T) responses to larval and adult somatic extracts were examined by Western blotting using sera from experimentally-infected horses and helminth-naive animals (n=6). Two antigen complexes, designated A and B, in larval somatic antigen were recognised specifically by the infected animals by 7 weeks PI. Sera taken from 23 endemically-infected animals, whose cyathostominae burdens had been enumerated, were also used to identify putative diagnostic antigens. Eighteen horses had positive mucosal worm burdens (range 723-3,595,725) and all but two of these animals had serum IgG(T) antibody specific to either complex. Moreover, IgG(T) responses specific to antigen complexes A and B were absent in all five parasite negative horses that were tested. Serum IgG(T) responses to either of the two complexes were identified in five clinical cases tested. IgG(T) responses to adult antigen somatic extracts were more heterogeneous, with no clear pattern between experimentally-infected ponies and helminth-free controls. The results indicate that increases in serum IgG(T) to mucosal larvae occur in the pre-patent period and that two antigenic complexes within somatic preparations of these stages have immunodiagnostic potential.

International audience; Benzimidazole resistance is common amongst many ovine trichostrongylid nematodes species globally. Although anthelmintics have been used for over half a century in some areas of the world for the control of Nematodirus battus, resistance has never been detected. Veterinary investigations conducted in 2010 demonstrated reduced efficacy in a flock that had been treated previously with fenbendazole (FBZ), suggesting probable resistance in N. battus. Infective larvae (L3; designated MNba2) were generated from the original material to conduct a controlled efficacy test (CET). Faecal egg counts showed an average of 37% reduction in the FBZ treated group 7 days post treatment compared to the untreated lambs. Average worm burden results showed no reduction after FBZ treatment compared to the untreated group (3850 and 3850 worms respectively). A molecular assay to assess the frequency of the commonly associated single nucleotide polymorphisms (SNP) in the β-tubulin isotype 1 gene, F200Y and E198A, was developed. Larval genotypes were predominantly homozygous resistant at codon 200 SNP, ranging from 56%-83% and remained stable at 70% for adult worm populations taken from treated and control lambs in the CET. Only susceptible genotypes were found at codon 198. The allele frequency for F200Y ranged between 80-83% in adult worms taken from the CET from treated and control lambs. The results confirmed initial findings and demonstrated the first report of FBZ resistance in N. battus whilst providing evidence that the P200 point mutation in the β-tubulin isotype 1 gene is a potential mechanism of resistance in the species.

This collection of articles provides an account of six presentations delivered at the 19th International Conference of the World Association for the Advancement of Veterinary Parasitology(WAAVP) (held in New Orleans, Louisiana, USA, from 10 to 14 August 2003) in a symposium session on Molecular Systematics and Diagnosis, organised and chaired by R.B. Gasser and D.S. Zarlenga. The focus was on recent advances in molecular tools for specific and genotypic identification,diagnosis, systematics and population genetics, with special emphasis on investigations of parasitic nematodes and protists.

Here, we report evaluation of five oligoprobes designed from intergenic spacer (IGS) region sequences for identification of cyathostomin species. Oligoprobes were designed for identification of Cylicocyclus ashworthi, Cylicocyclus nassatus, Cylicostephanus longibursatus, Cylicostephanus goldi and a fifth probe designed to identify all members of this tribe. PCR amplification of IGS DNA from 16 cyathostomin species allowed sequence comparison and identification of four putative species-specific probes. Southern blotting of amplified products from 16 species showed that all probes were species-specific. The fifth probe recognised all 16 cyathostomin species but did not bind to members of the genus Strongylus. Furthermore, these probes were used to identify individual infective L3, eggs and L4 indicating that they will be invaluable to furthering the study of the epidemiology and pathogenesis of these important equine nematodes.

The frequent use of anthelmintics in horses has contributed to the development of widespread anthelmintic resistance in cyathostomins (Matthews 2008, 2011), a group of parasitic nematodes comprising around 50 species. Cyathostomins are highly prevalent in grazing equids globally and have great pathogenic potential due to their capacity to cause life-threatening colitis when present in high numbers (Love and others 1999). Most cyathostomin infections are well tolerated; however, mass emergence of encysted larvae from the large intestinal wall can result in larval cyathostominosis, characterised by diarrhoea, rapid weight loss and oedema, which can be fatal in up to 50 per cent of cases (Love and others 1999). Currently, three classes of broad spectrum anthelmintics are licensed for use against adult cyathostomin worms in the UK; fenbendazole (FBZ; a benzimidazole, BZ), pyrantel embonate (PYR; a tetrahydropyrimidine, THP) and the macrocyclic lactones (ML), ivermectin (IVM) and moxidectin (MOX). BZ resistance is ubiquitous, (Kaplan and others 2004, Osterman Lind and others 2007, Traversa and others 2012, Stratford and others 2013a) and reduced sensitivity of cyathostomins to PYR is common in certain geographic areas (Kaplan and others 2004, Comer and others 2006, Osterman Lind and others, 2007, Traversa and others 2009). Single cyathostomin populations have been identified that exhibit both FBZ and PYR resistance (Kaplan 2004, Traversa and others, 2009). A reduction in the standard strongyle egg reappearance period following ML administration has been reported, and is generally accepted as an early indicator of resistance …

SUMMARYFew studies have described the combined effect of age, gender, management and control programmes on helminth prevalence and egg shedding in grazing equines. Here, fecal samples collected from 1221 Thoroughbred horses, residing at 22 studs in the UK, were analysed. The distribution of strongyle eggs amongst individuals in relation to age, gender and management practices was investigated. Fecal worm egg counts (FWECs), described as the number of eggs per gramme (epg) of feces, were determined using a modification of the salt flotation method. The FWEC prevalence (mean%) of strongyles,Parascaris equorum, tapeworm spp. andStrongyloides westeriwas 56, 9, 4 and 8%, respectively. Strongyle,P. equorum, tapeworm spp. andS. westeriinfections were detected on 22 (100%), 11 (50%), 9 (41%) and 8 (36%) of studs, respectively. Within all age and gender categories, strongyle FWECs were highly over-dispersed (arithmetic mean = 95 epg, aggregation parameterk=0·111) amongst horses. Animal age, last anthelmintic type administered and management practices (for example, group rotation on grazing) most strongly influenced strongyle prevalence and level of egg shedding (P

Anthelmintic resistance in equine gastrointestinal nematodes is a threat to equine health and welfare. Detailed knowledge of anthelmintic use and parasite control methods is a prerequisite to identification of potential risk factors for resistance.To identify parasite control practices employed by equine owners in Scotland and investigate management factors associated with anthelmintic resistance.Questionnaire study of equine parasite control in Scotland.Questionnaires were available electronically, distributed at a conference and mailed to clients. Key areas explored included general background, grazing management, anthelmintic treatment practices and use of diagnostic tests.A total of 193 responses detailing information on parasite control programmes of 993 equids were analysed. Moxidectin (MOX) and ivermectin or related combination products were the most commonly administered anthelmintics in the preceding 12 months. Treatments licensed for use against cyathostomin encysted larvae and tapeworms were administered by 80% and 90% of respondents, respectively. This was often achieved through indiscriminate use of MOX and MOX-praziquantel products. Faecal egg count (FEC) analysis had been performed on 62% of yards and regular use of FECs reduced annual anthelmintic treatment frequency. Veterinarians had the greatest influence on control practices. While 40% of respondents believed that they practised targeted dosing, this was not associated with delaying treatment beyond the egg reappearance period of the anthelmintic used.Responses indicated increasing veterinary involvement and use of FECs. The majority of respondents administered anthelmintics licensed against cyathostomin encysted larvae and tapeworms. However, responses suggested that owners did not understand the definition of 'targeted' dosing regimens.The high frequency of MOX use represents a potential risk factor for macrocyclic lactone resistance. As veterinarians were the most influential factor in anthelmintic choice, awareness of macrocyclic lactone resistance and potential risk factors for its development and spread should be incorporated into client advice.

Background Gastrointestinal nematode infections constitute a threat to the health and welfare of donkeys worldwide. Their primary means of control is via anthelmintic treatments; however, use of these drugs has constraints in developing countries, including cost, limited availability, access to cheaper generic forms of variable quality and potential anthelmintic resistance. As an alternative, bioactive plants have been proposed as an option to treat and control gastrointestinal helminths in donkeys. This study aimed to use participatory methodology to explore donkey owner knowledge, attitudes and beliefs relating to the use of plant-based treatments for gastrointestinal parasites of donkeys in Ethiopia. Results In focus groups, 22/29 groups stated they knew of plants used for the treatment of gastrointestinal parasites in donkeys. All groups volunteered plants that were used in cattle and/or small ruminants. In total, 21 plants were named by participants. ‘Koso’ (Hagenia abyssinica) ‘Grawa’ (Vernonia amygdalina) and a mixed roots and leaves preparation were the most frequently named plant preparations. ‘Enkoko’ (Embelia shimperi) and ‘a mixture of roots and leaves’ were ranked highly for effectiveness in donkeys. However, ‘Grawa’ and ‘Koso’ were the highest ranked when taking into account both the rank position and the number of groups ranking the plant. Thematic analysis of participants’ current attitudes and beliefs surrounding traditional plant-based remedies for gastrointestinal parasites revealed that anthelmintics obtained from clinics were generally favoured due to their ease of administration and perceived higher effectiveness. There was doubt surrounding the effectiveness of some plant-based treatments, but there were also perceived advantages including their low cost, ease of cultivation and availability. However, plant-based treatments were considered a “past trend” and people favoured “modern” medicine, particularly among the younger generation. Conclusions There was extensive knowledge of plant-based treatments for gastrointestinal parasites in livestock in Ethiopia. In donkeys, Koso (Hagenia abyssinica), Grawa (Vernonia amygdalina), Enkoko (Embelia shimperi) and ‘mixed roots and leaves’ were the most frequently named and/or highest ranked plants with reported efficacy against gastrointestinal parasites. Further in vitro and in vivo investigation of these plants is now required to determine viable alternatives for the treatment and control of gastrointestinal parasites in Ethiopia.

Cyathostomins comprise 50 helminth species, considered the most problematic equine endoparasites. Three classes of anthelmintic are currently licensed for their control, namely the benzimidazoles (fenbendazole), tetrahydropyrimidines (pyrantel) and macrocyclic lactones (ivermectin and moxidectin). Anthelmintic resistance in cyathostomins is common. With no new classes expected in the near future, it is essential to determine the efficacy of the available anthelmintics to inform future control programmes.To determine the efficacy of all 3 anthelmintic classes against strongyles in equids on livery yards in east and central Scotland.Anthelmintic efficacy testing using the faecal egg count reduction test (FECRT).FECRTs were performed on equids with initial strongyle faecal egg counts (FECs) of ≥50 eggs per gram. Efficacy was determined by comparing pretreatment (Day 0) and 14 days post treatment (Day 14) FECs. The following chemicals were tested: fenbendazole, pyrantel, ivermectin and moxidectin. Group arithmetic mean FECR of90% for fenbendazole and pyrantel, and95% for ivermectin and moxidectin, represented efficacy, whereas lower mean FECR indicated potential resistance.A total of 447 FECRTs were performed on 15 yards, as follows (the numbers in parentheses represent the number of yards each anthelmintic was tested on): 55 equids (7 yards) fenbendazole, 111 (8 yards) pyrantel, 163 (13 yards) ivermectin and 118 (10 yards) moxidectin. Fenbendazole resistance was documented on all yards (range of mean FECR, 15.8-83.4%), whereas pyrantel (90.4-99.6%), ivermectin (99.5-100%) and moxidectin (99.4-100%) treatment had acceptable efficacy.Reduced efficacy of fenbendazole was widespread, whereas90% efficacy was found after pyrantel, and95% efficacy after ivermectin and moxidectin. Overall, efficacies were higher than reported previously in Europe and the USA, potentially reflecting differences in management and anthelmintic use on the yards surveyed.The use of fenbendazole for strongyle control in Scotland should be questioned. Targeted use of pyrantel should be encouraged to reduce reliance on macrocyclic lactones. Further work to correlate management practices with the presence of anthelmintic resistance is warranted.

International audience; The anthelmintic sensitivity of two field-derived isolates (designated FI001 and FI004) of cattle nematodes from beef farms in Scotland were investigated in a controlled efficacy test (CET). Efficacies of ivermectin pour-on (IVM-PO), IVM injectable (IVM-INJ) and moxidectin pour-on (MOX-PO) formulations were assessed. In each group, five helminth-naive calves were infected experimentally with 50,000 third stage larvae from either isolate and administered with anthelmintic at the manufacturers' recommended dose rate 28 days later. For each isolate, nematode burdens were compared between treatment and control groups to determine efficacy. Nematode species composition, based on data derived from the untreated control groups' burden estimations, were 39 and 14% Cooperia oncophora and 61 and 86% Ostertagia ostertagi for isolates FI001 and FI004, respectively. Macrocyclic lactone resistance in C. oncophora was confirmed for both FI001 and FI004 isolates. Efficacies (as determined by nematode burden analysis) of 4,21 and 31% for FI001, and 10, 1 and 74% for FI004, were obtained for IVM-INJ, IVM-PO and MOX-PO, respectively. Efficacy based on faecal egg count reduction at seven days post anthelmintic administration were 8, 99 and 100% for FI001, and 37, 20 and 100% for FI004 for IVM-INJ, IVM-PO and MOX-PO, respectively. In summary, this study details two macrocyclic lactone resistant isolates of C. oncophora obtained from cattle from two distinct geographical locales in the UK.

Improved education of veterinarians and equine owners/managers is essential in implementing parasite control strategies that are less reliant on chemicals.This questionnaire study, conducted on 61 UK Thoroughbred (TB) establishments during 2009 and 2010, was designed to obtain an understanding of current helminth control practices on studs. To our knowledge, this is the first occasion that statements obtained from TB studs via questionnaire have been supported by statistical analysis.Despite many respondents indicating high levels of concern regarding anthelmintic resistance, 56% of these establishments that received visiting equines co-grazed these animals with permanent stock and74% administered anthelmintics prior to integration. In the 12 months preceding the study, most respondents administered frequent macrocyclic lactone (ML) treatments, with none appearing to leave any animals in groups untreated at each administration. Indiscriminate whole group treatments with MLs and movement of animals to 'clean grazing' post treatment (reported by25% of respondents), indicates that many stud owners/managers are not aware of the strong risk factors for the development of anthelmintic resistance. Few studs had conducted faecal egg count (FEC) analysis in the past and only 22% indicated that they considered this form of analysis beneficial in determining anthelmintic choice.The challenge now is to convince stud owners/managers to deviate from their current practices to control strategies that are more likely to preserve anthelmintic efficacy. Veterinarians need to get more involved in implementing these control strategies, with better emphasis placed on the role of diagnostic tests in facilitating targeted treatments and in investigating anthelmintic sensitivity in the associated nematode populations.

Cyathostomins represent a potential cause of equine morbidity and have become the main focus of endoparasite control in managed horses. All grazing horses are at risk of infection with cyathostomins; therefore, the application of appropriate management measures is essential. Anthelmintics currently comprise the main method of control for equine nematodes and the ready availability of these products in some countries has resulted in their use becoming dissociated from veterinary involvement. This is concerning given the levels of anthelmintic resistance that have been recorded in cyathostomin populations. It is important that veterinarians re-establish control over the implementation of parasite control programmes, a major objective of which should be the preservation of anthelmintic efficacy. This article details the principles of cyathostomin control in horses with particular reference to anthelmintic resistance, and the use and interpretation of diagnostic tests for detecting cyathostomins and identifying anthelmintic resistance.

Infections with parasitic nematodes are of significant welfare and economic importance worldwide, and because of the emergence of anthelmintic resistance, this has lead to alternative methods of parasite control being required. Vaccination offers a feasible alternative control, and the majority of research has focused on the production of recombinant versions of native antigens previously identified as protective in vaccinated animals. Attempts at the production of protective recombinant subunit vaccines have been hindered, however, as these antigens have invariably failed to replicate the same level of protective immune response as seen with the native versions. It has been proposed that these failures are owing to the fact that the recombinant proteins do not contain the appropriate post-translational modifications to retain the protective capacity of the native molecules. In this review, we discuss a novel approach to vaccine antigen identification through the application of random peptide phage-display libraries and their use to identify peptide sequences that potentially mimic the structure(s) of antigenic epitopes. This area of research is still relatively novel with respect to parasites, and the current state of the art will be discussed here.

A cDNA representing the gene Teladorsagia circumcincta apyrase-1 (Tci-apy-1) was isolated, by PCR, from a T. circumcincta fourth-stage larval (L4) cDNA library. The closest orthologue of this gene is a Ca(2+)-dependent apyrase from Ostertagia ostertagi, with 92% amino acid identity across all 339 residues. Tci-apy-1 is transcribed in a stage-specific manner, the transcript being predominant in L4, detectable in the adult cDNA, but absent from eggs and infective third-stage larvae (L3). The protein, Tci-APY-1, was detected by immunoblotting in extracts of L4 nematodes and was present in excretory/secretory products from the same developmental stage. A recombinant version of Tci-APY-1 was expressed in bacteria as an active enzyme that hydrolysed nucleoside triphosphate substrates with a preference of ATP over other nucleoside triphosphates. Recombinant Tci-APY-1 hydrolysed ATP and ADP but not AMP. Apyrase activity was divalent cation-dependent, with no hydrolysis in the presence of Mg(2+), but activation in the presence of Ca(2+). Recombinant Tci-APY-1 was bound by IgG present in serum and both IgG and IgA present in abomasal mucus from trickle-infected, immune sheep but not in material derived from lambs exposed to a single infection. The potential immunomodulatory roles of this Tci-APY-1 are discussed in relation to purinergic signalling.

Abnormal neutrophil responses have been observed in periodontitis patients, including hyper-reactivity in terms of production of reactive oxygen species (ROS) following exposure to the key quorum-sensing plaque bacterium, Fusobacterium nucleatum. This study was designed to characterize the transcriptional response of neutrophils to F. nucleatum.Peripheral blood neutrophils were exposed to F. nucleatum, and gene expression was analysed using high-throughput transcriptomics.Microarray technology demonstrated differential expression of 208 genes (163 increased and 43 decreased relative to control genes), which identified regulation of several ontological classes, including signal transduction (13%), transcription regulation (7%) and ROS response (14%). Individual gene expression analysis of selected transcripts, including CSF, CXCL3, FOS, HMOX1, HSP40, SOD2, NFKB2 and GP91, in individual and pooled RNA samples from control and F. nucleatum-exposed neutrophils corroborated microarray data. Analysis of ROS generation, combined with transcript analysis, in response to a panel of proinflammatory stimuli (F. nucleatum, Porphyromonas gingivalis, Escherichia coli lipopolysaccharide and opsonized Staphylococcus aureus) identified significant differences in ROS and transcript regulatory control. Further analyses of neutrophils from periodontitis patients and periodontally healthy control subjects stimulated with F. nucleatum indicated significant differential induction of several ROS response-related transcripts.These data demonstrate that neutrophils are transcriptionally active in response to the periodontal pathogen F. nucleatum and that these changes in gene expression are likely to affect neutrophil function. The differential response of neutrophils to a range of stimuli combined with data demonstrating differences between patient and control neutrophils indicate the importance of this cell and its interaction with the local tissue environment in the pathogenesis of periodontitis.

Sixty-seven invited participants involved in the development, evaluation, and use of therapies for acid-peptic disorders participated in a meeting to discuss the scientific basis for healing actions of ulcer drugs and the prospects for future developments (“Realities of Mucosal Protection in the Upper Gastrointestinal Tract”, Lausanne, Switzerland, November 8–10, 1987). Eighty-one key statements were prepared and subsequently analyzed on the basis of a voting system. Of the 45 statements that dealt with existing therapies, only 3 statements showed positive consensus (agreement of two-thirds or more of voters) about mechanisms of ulcer healing. Participants agreed that both (1) hydrogen/potassium adenosine triphosphatase inhibitors and (2) histamine H2 antagonists healed ulcers solely by acid inhibition, and (3) that sucralfate works by topical action. Substantial uncertainty about the mechanisms by which bismuth compounds and antacids heal ulcers was noted as well as their wide range of effects. The mechanism of ulcer healing by prostaglandins and the clinical relevance of antiulcer effects of drugs demonstrated in acute studies with animals were also controversial. There was greater agreement among participants about unexplored drug effects that might produce ulcer healing. Of the 36 such mechanisms surveyed, the most support went to therapies aimed at enhancement of mucosal blood flow, epithelial restitution, and mucosal alkaline secretion or inhibition of luminal pepsin activity. The diversity of opinions among participants suggests a high level of empiricism in the development of ulcer healing drugs apart from those that inhibit acid secretion. This empiricism probably arises from inadequate understanding of processes of mucosal injury and repair.