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Background: In dogs, infections with Giardia duodenalis are mainly caused by assemblages C and D, but also by the potentially zoonotic assemblages A and B. The aims of this study were to assess differences in assemblages (i) between dogs living mainly in close proximity to humans (synanthropic dogs) versus dogs living mainly among other dogs, (ii) between samples of dogs with or without loose stool, and (iii) related to the amount of cysts shedding., Methods: One hundred eighty-nine qPCR Giardia positive fecal samples of dogs originating from four groups (household, sheltered, hunting, and dogs for which a veterinarian sent a fecal sample to a diagnostic laboratory) were used for genotyping. For this, multi-locus genotyping of beta-giardin, triose phosphate isomerase, and glutamate dehydrogenase and genotyping of SSU rDNA gene fragments were performed. Fecal consistency was scored (loose or non-loose stool), and cysts per gram of feces were determined with qPCR., Results: Assemblage D was the most prevalent in all groups, followed by the other canid assemblage C. Also, mixed C/D was common. In two (synanthropic) household dogs, the potentially zoonotic assemblage AI was present. Although occurrence of assemblage AI in household dogs was not significantly different from dogs living among other dogs (sheltered and hunting dogs), it was significantly higher compared to dogs for which a sample was sent to a diagnostic laboratory. Dogs with assemblage D shed significantly more cysts than dogs with other assemblages (except for mixed C/D results) or dogs in which no assemblage could be determined. None of the assemblages was significantly associated with loose stool., Conclusion: Not only do dogs mainly shed the canid Giardia duodenalis assemblages D and/or C, the numbers of cysts per gram for the canid assemblage D were also higher than for the potential zoonotic assemblage AI. Based on the assemblages shed by dogs, the risk to public health posed by dogs is estimated to be low, even though the dogs that shed AI were synanthropic household dogs. Loose stool in infected dogs was not associated with any particular Giardia assemblage.

We report a European wide assessment of the economic burden of gastrointestinal nematodes, Fasciola hepatica (common liver fluke) and Dictyocaulus viviparus (bovine lungworm) infections to the ruminant livestock industry. The economic impact of these parasitic helminth infections was estimated by a deterministic spreadsheet model as a function of the proportion of the ruminant population exposed to grazing, the infection frequency and intensity, the effect of the infection on animal productivity and mortality and anthelmintic treatment costs. In addition, we estimated the costs of anthelmintic resistant nematode infections and collected information on public research budgets addressing helminth infections in ruminant livestock. The epidemiologic and economic input data were collected from international databases and via expert opinion of the Working Group members of the European Co-operation in Science and Technology (COST) action COMbatting Anthelmintic Resistance in ruminants (COMBAR). In order to reflect the effects of uncertainty in the input data, low and high cost estimates were obtained by varying uncertain input data arbitrarily in both directions by 20 %. The combined annual cost [low estimate-high estimate] of the three helminth infections in 18 participating countries was estimated at € 1.8 billion [€ 1.0-2.7 billion]. Eighty-one percent of this cost was due to lost production and 19 % was attributed to treatment costs. The cost of gastrointestinal nematode infections with resistance against macrocyclic lactones was estimated to be € 38 million [€ 11-87 million] annually. The annual estimated costs of helminth infections per sector were € 941 million [€ 488 - 1442 million] in dairy cattle, € 423 million [€ 205-663 million] in beef cattle, € 151million [€ 90-213 million] in dairy sheep, € 206 million [€ 132-248 million] in meat sheep and € 86 million [€ 67-107 million] in dairy goats. Important data gaps were present in all phases of the calculations which lead to large uncertainties around the estimates. Accessibility of more granular animal population datasets at EU level, deeper knowledge of the effects of infection on production, levels of infection and livestock grazing exposure across Europe would make the largest contribution to improved burden assessments. The known current public investment in research on helminth control was 0.15 % of the estimated annual costs for the considered parasitic diseases. Our data suggest that the costs of enzootic helminth infections which usually occur at high prevalence annually in ruminants, are similar or higher than reported costs of epizootic diseases. Our data can support decision making in research and policy to mitigate the negative impacts of helminth infections and anthelmintic resistance in Europe, and provide a baseline against which to measure future changes., (Copyright © 2020 Elsevier B.V. All rights reserved.)

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

Background: The aim of this study was to assess potential associations between Giardia duodenalis infection in dogs, as determined by three diagnostic tests, and dog's group of origin, fecal consistency, age, sex, neuter status, and co-infections with other gastrointestinal parasites., Methods: Fecal samples from 1291 dogs from four groups (household, shelter, hunting and clinical dogs) were tested with qPCR, rapid enzyme immunochromatographic assay (IDEXX SNAP ® Giardia), and direct immunofluorescence (DFA, Merifluor) for presence of G. duodenalis. Moreover, fecal samples were tested with centrifugation sedimentation flotation (CSF) coproscopical analysis for presence of gastrointestinal parasites. Associations were expressed as odds ratios (ORs)., Results: Several significant associations were found, of which a few were consistent for all three tests and Giardia positivity in general (positive with at least one of these tests). Dogs older than one year were significantly less likely to test positive for Giardia than younger dogs. Group-housed dogs, especially hunting dogs, were significantly more likely to test positive for Giardia compared to household and clinical dogs. A consistently significant association with Trichuris appeared to be driven by the high prevalence in hunting dogs. Although there was no significant association between loose stool and Giardia infection in the overall population, household dogs were significantly more likely to test Giardia-positive when having loose stool. Overall, Giardia-positive dogs with loose stool shed significantly more cysts, both determined semi-quantitatively with CSF and quantitatively by qPCR, than positive dogs with no loose stool. When other gastrointestinal parasites were present, significantly fewer cysts were detected with CSF, but this was not confirmed with qPCR., Conclusion: Giardia is the most common gastrointestinal parasite in Dutch dogs, except for hunting dogs, in which Trichuris and strongyle-type eggs (hookworms) prevailed. Giardia infection was not significantly associated with loose stool, except for household dogs. Young dogs and group-housed dogs were significantly more often Giardia-positive. These associations were consistent across diagnostic tests. Young dogs, clinical dogs and dogs with loose stool shed Giardia cysts in the highest numbers. If another gastrointestinal parasite was present lower numbers of cysts were observed by microscope (CSF), but not with a molecular method (qPCR).

An elicitation exercise was conducted to collect and identify pressing questions concerning the study of helminths in livestock, to help guide research priorities. Questions were invited from the research community in an inclusive way. Of 385 questions submitted, 100 were chosen by online vote, with priority given to open questions in important areas that are specific enough to permit investigation within a focused project or programme of research. The final list of questions was divided into ten themes. We present the questions and set them briefly in the context of the current state of knowledge. Although subjective, the results provide a snapshot of current concerns and perceived priorities in the field of livestock helminthology, and we hope that they will stimulate ongoing or new research efforts., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

In a survey involving 34 sheep flocks spread over the Netherlands anthelmintic resistance (AR), based on a fecal egg count reduction (FECR) test, was determined for six different products. The study was conducted in ewes shortly after lambing during spring 2015. A FECR of less than 90%, indicating presence of AR against one or more nematode genera producing strongylid eggs, was found in 22 of 30 (73.3%) flocks against oxfendazole, 18 of 23 (78.3%) flocks against ivermectin, 15 of 32 (46.9%) flocks against moxidectin, and 2 of 26 (7.7%) flocks against monepantel. No AR was observed against levamisole. If oxfendazole resistance was observed, Haemonchus contortus was involved in 90.5% of the cases. If resistance against ivermectin, moxidectin or monepantel was observed, it invariably involved H. contortus. In the majority of cases resistance was also observed for Teladorsagia circumcincta and/or Trichostrongylus spp, between which no distinction was made in this study. Based on FECR 9 of 15 (60.0%) flocks showed resistance against closantel, which was mainly due to closantel not being effective against most other nematode species than H. contortus. However, in 44.4% of flocks showing reduced FECR it did involve H. contortus as well. Multi-drug resistance (excluding closantel) was found in 16 flocks, of which 8 showed resistance against 2 products, 7 against 3 products and 1 flock showed resistance against 4 products. If resistance against 3 or 4 products was present, there invariably was resistance against both ivermectin and moxidectin. Overall, of the 22 flocks in which both macrocyclic lactones (ML) were tested, 4 (18.2%) showed no resistance against both products, 9 (40.9%) showed resistance against ivermectin only, and 9 (40.9%) showed resistance against both MLs. It is concluded that AR is widespread in sheep in the Netherlands and involves products from all major anthelmintic classes, with possibly the exception of levamisole. It appears that the macrocyclic lactones have lost much of their efficacy against sheep nematodes over the last decade., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: To accurately diagnose giardiosis in dogs, knowledge of diagnostic test characteristics and expected prevalence are required. The aim of this work was to estimate test characteristics (sensitivity and specificity) of four commonly used diagnostic tests for detection of Giardia duodenalis in dogs., Methods: Fecal samples from 573 dogs originating from four populations (household dogs, shelter dogs, hunting dogs and clinical dogs) were examined with centrifugation sedimentation flotation (CSF) coproscopical analysis, direct immunofluorescence assay (DFA, Merifluor Cryptosporidium/Giardia®), a rapid enzyme immunochromatographic assay (IDEXX SNAP Giardia®) and qPCR (SSU rDNA) for presence of G. duodenalis. Bayesian latent class analysis was used to determine test performance characteristics and to estimate G. duodenalis prevalence of each of the four dog populations., Results: All tests were highly specific. IDEXX SNAP Giardia® showed the highest specificity (99.6%) and qPCR the lowest (85.6%). The sensitivities were much more variable, with qPCR showing the highest (97.0%) and CSF the lowest (48.2%) sensitivity. DFA was more sensitive than IDEXX SNAP Giardia®, but slightly less specific. Prevalences of G. duodenalis differed substantially between populations, with the hunting dogs showing the highest G. duodenalis prevalence (64.9%) and the household dogs the lowest (7.9%)., Conclusions: This study identifies qPCR as a valuable screening tool because of its high sensitivity, whereas methods using microscopy for cyst identification or cyst wall detection should be used in situations where high specificity is required. G. duodenalis is a prevalent gastro-intestinal parasite in Dutch dogs, especially in dogs living in groups (hunting and shelter dogs) and clinical dogs.

The Faculty of Veterinary Medicine at Utrecht University has seen three major curriculum changes, in 1995, 2001 and 2007. The last change was made because of the European change to a Bachelor-Master system. Almost each time teaching hours tagged for veterinary parasitology have been reduced to currently a minimum of between 46 and 51.5 h, which is much less than the WAAVP-recommended minimum of 70-90 h. This results in a challenge to maintain a qualitatively adequate veterinary parasitology program in a curriculum. Following a brief historic account of previous curricula and implemented curriculum changes, experiences, limitations and opportunities are discussed, including the potential of introducing new teaching materials based, for example, on digital technologies and gaming., (Copyright © 2018 The Author(s). Published by Elsevier B.V. All rights reserved.)

The purpose of the study was to gain knowledge about the prevalence and identity of rumen flukes (RF) in cattle and sheep in the Netherlands. Routine faecal examinations of diagnostic submissions between May 2009 and September 2014 showed a mean annual herd or flock RF prevalence of 15.8% for cattle and 8.0% for sheep. Prevalence in cattle was higher after 2012 than before, which may reflect a change in detection method as well as an increase in true prevalence. During November and December 2014, an abattoir survey was conducted to allow for scoring of rumen fluke burden and to obtain specimens for molecular species characterization. Over 8 visits to 5 abattoirs in areas deemed to pose a high risk for trematode infection, 116 cows and 41 sheep from 27 herds and 10 flocks were examined. Prevalence of RF was higher in beef cattle than in dairy cattle and higher in cattle than in sheep. Median fluke burden was >100 specimens per animal for most positive animals. Using a semi-quantitative RF density score as a gold standard, sensitivity and specificity of a modified quantitative Dorsman egg counting method were estimated at 82.6% and 83.3%, respectively. Of 14 collected adult rumen flukes, twelve (8 bovine and 4 ovine specimens) were identified as Calicophoron daubneyi. The other two, of bovine origin, were identified as Paramphistomum leydeni, which was unexpected as in other European countries all recently collected rumen flukes in both cattle and sheep were identified as C. daubneyi. The findings implicate that multiple rumen fluke species, intermediate host species and transmission cycles may play a role in rumen fluke infections in the Netherlands., (Copyright © 2017. Published by Elsevier B.V.)

The prevalence of and risk factors for shedding Toxocara eggs in cats older than 6 months were determined by examining 670 faecal samples collected in 4 cross-sectional studies in the Netherlands. Additionally, cat owners provided information on their attitude towards routine deworming. Samples were examined using the centrifugal sedimentation flotation method. Overall Toxocara prevalence was 7.2 %. Multivariable logistic regression analysis revealed that young age and living in rural areas were significant risk factors for shedding Toxocara eggs. Moreover, the more time a cat was allowed to roam outdoors, the higher was its risk to shed Toxocara as compared to cats with no outdoor access at all. For 199 cats (81.6 % of cats subjected to a deworming regimen) owners provided the reason for treatment. The main reason for routine deworming (80.4 %) concerned the cat's health and only 10.6 % of the cats were treated for public health reasons. Moreover, the generally advocated four-times-a-year deworming advice was applied on only 24.5 % of cats. We concluded that free roaming is a key factor in the acquisition of patent Toxocara infections leading to the environmental contamination with Toxocara eggs. Additionally, the knowledge of cat owners is still insufficient to expect them to make sound decisions on routine deworming.

Gastrointestinal (GI) nematode infections are considered among one of the toughest challenges sheep farmers face worldwide. Control still is largely based on the use of anthelmintics, but anthelmintic resistance is becoming rampant. To facilitate implementation of alternative nematode control strategies and to reduce anthelmintic usage, the purpose of this study was twofold: (i) to gain insight in common practices, knowledge gaps and perceptions of farmers regarding nematode control, and (ii) to provide foci of attention for improving parasite control practices and transfer of knowledge within the sheep husbandry. An internet-based questionnaire was made available to all sheep farmers pertaining to the year 2013, resulting in 450 entered questionnaires for analysis. The two most important nematodes mentioned, were Haemonchus contortus and, to a lesser extent, Nematodirus battus. Of all respondents, 25.6% said they did not have any worm problems. Of these, almost a third did notice clinical signs that can be related to worm infections and about three quarters did use anthelmintics. Overall, clinical symptoms mentioned by farmers matched the worm species they identified as the cause of problems. Ewes and lambs were treated up to 6 times in 2013. On average, ewes were treated 1.53 and lambs 2.05 times. Farmers who treated their ewes more often, also treated their lambs more often (P<0.001). Both ewes and lambs were frequently treated based on fixed moments such as around lambing, at weaning and before mating, rather than based on faecal egg counts. Treatments based on faecal egg counts were practiced, but on a minority of the farms (32.7%). The majority of the farms (75.6%) did not leave 2-5% of the sheep within a flock untreated. About 74% of farmers keep newly purchased animals quarantined for at least 10days, but some (13.4%) leave quarantined animals untreated nor check faecal egg counts. Of farmers who do treat their quarantined animals, just 12.6% check the efficacy of the treatment. Slightly over 40% of the respondents said they did not experience bottlenecks in parasite control. Yet, over half of these said having problems with worm infections, over half did see clinical signs related to worm infections and over three quarters used anthelmintics. Within the group of farmers experiencing difficulties in parasite control, the most often mentioned bottleneck concerned pasture management (75.8%). When asking farmers for solutions, 90% of all respondents indicated they are willing to adjust their pasture management. Farmers are also interested in other methods to reduce the risk of worm infections, such as possibilities to enhance the immune system of sheep in general (71%), to increase specific genetic resistance to worms and to apply anti-parasite forages, both about 40%. Results of this study gave the following potential foci of attention: (1) making complex scientific knowledge more accessible to farmers through simple tools and applicable in the daily farming process; (2) changing the mindset of farmers about their current worm control practices, i.e. breaking long-standing habits such as treating ewes and lambs at fixed moments rather than based on actual worm infection monitoring data; (3) demonstrating effective pasture rotation schemes on specific farms and using these in extension work; (4) making farmers more aware that checking anthelmintic efficacy is important; (5) improving quarantine procedures; (6) creating a wider array of applicable alternative control measures from which individual farmers can choose what fits them most; and finally, (7) improving mutual understanding among farmers, veterinary practitioners and parasitologists alike., (Copyright © 2016 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: To reduce environmental contamination with Toxocara canis eggs, the current general advice is to deworm all dogs older than six months on average four times a year. However, only a small proportion of non-juvenile household dogs actually shed T. canis eggs, and some dogs shed eggs more frequently than others. The identification of these frequent shedders and the associated risk factors is an important cornerstone for constructing evidence-based deworming regimens. The purpose of this study is to identify risk factors associated with recurrence of periods of shedding Toxocara eggs in a cohort of household dogs older than six months., Methods: We performed a prospective study (July 2011 to October 2014) on shedding Toxocara eggs in a cohort of 938 household dogs older than six months from all over the Netherlands. The median follow-up time was 14 months. Monthly, owners sent faecal samples of their dogs for Toxocara testing and completed a questionnaire. Dogs were dewormed only after diagnosis of a patent infection (PI). Survival analysis was used to assess factors influencing the time to first diagnosed PIs (FPI) and the time to recurrent PIs (RPI)., Results: The overall prevalence of PIs was 4.5 %, resulting in an estimated average incidence of 0.54 PIs/dog/year. No PI was diagnosed in 67.9 % of the dogs, 17.5 % of the dogs went through only one PI and 14.6 % had > 1 PI. Prevalence of PIs always peaked during wintertime. Increased hazards for first diagnosed PIs were associated with coprophagy, geophagy, walking off-leash for ≥ 80 % of walking time, reported worms in the faeces, feeding a commercial diet and suffering from urologic or respiratory conditions. Median time to reinfection was nine months. Factors associated with increased hazards for recurrent PIs were taking corticosteroids, changing dog's main purpose, and proxies for veterinary care-seeking behaviours., Conclusions: We concluded that targeted anthelmintic treatments in household dogs may be feasible as PIs tend to (re)occur in specific periods and in groups of dogs at high risk. Moreover, recurrent PIs appear to be influenced more by factors related to impaired immunity than environmental exposure to Toxocara eggs.

For the control of cyathostomins in horses, the macrocyclic lactones (MLs), moxidectin (MOX) and ivermectin (IVM) are the most commonly used anthelmintics. However, reduced activity, observed as shortening of the egg reappearance period (ERP) has been described. Shortening of the ERP may be caused by a decreased susceptibility of immature worms for MLs. Alternatively, immature worms may develop faster into egg producing adults as a result of repeated ML treatments. The species composition of the larval cultures obtained shortly after ML and pyrantel (PYR) treatment can confirm the hypothesis of decreased ML susceptibility, as this is often class-specific, whereas faster development would also occur after treatment with anthelmintics with a different mode of action. From 3 farms with a known history of shortened ERP, 8 horses per farm were selected and divided into 2 groups. The MOX-PYR-MOX group was treated twice with MOX (day 0 and 126) and once with PYR (day 84) and the IVM-PYR-IVM group was treated twice with IVM (day 0 and 98) and once with PYR (day 56). Cultured infective larvae (L3s) were counted and differentiated with the reverse line blot on pooled samples. Per cyathostomin species, the number of larvae per gram was calculated. The efficacy of all ML treatments was 100% and a shortened ERP was found on all 3 farms. The species composition of the larval cultures after ML treatment did not differ significantly from that after PYR treatment in the IVM-PYR-IVM group, but it did differ in the MOX-PYR-MOX group. The larval cultures obtained after MOX treatment consisted mostly of Cylicocyclus nassatus, while after PYR treatment Cylicostephanus longibursatus was the most abundant species. In the cultures from 42days after MOX treatment 6 cyathostomin species from 3 genera were found on the farm with the lowest activity (farm 1), while on the farm with the highest activity (farm 3) only 3 species from one genus were found in the same number of examined L3s. The high numbers of L3s of Cylicocyclus species 42days after MOX treatment and the low numbers 42days after PYR treatment can be explained by reduced susceptibility of the immature worms to MOX, but not by a faster development. In conclusion, shortening of the ERP following MOX treatment is most likely a process in which an increasing number of immature worms from an increasing number of species is becoming less susceptible to the active compound., (Copyright © 2016 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Environmental contamination with Toxocara eggs is considered the main source of human toxocariasis. The contribution of different groups of hosts to this contamination is largely unknown. Current deworming advices focus mainly on dogs. However, controversy exists about blind deworming regimens for >6-month-old dogs, as most of them do not actually shed Toxocara eggs. We aim to estimate the contribution of different non-juvenile hosts to the environmental Toxocara egg contamination and to assess the effects of different Toxocara-reducing interventions for dogs., Methods: A stochastic model was developed to quantify the relative contribution to the environmental contamination with Toxocara eggs of household dogs, household cats, stray cats, and foxes, all older than 6 months in areas with varying urbanization degrees. The model was built upon an existing model developed by Morgan et al. (2013). We used both original and published data on host density, prevalence and intensity of infection, coprophagic behaviour, faeces disposal by owners, and cats' outdoor access. Scenario analyses were performed to assess the expected reduction in dogs' egg output according to different deworming regimens and faeces clean-up compliances. Estimates referred to the Netherlands, a country free of stray dogs., Results: Household dogs accounted for 39% of the overall egg output of >6-month-old hosts in the Netherlands, followed by stray cats (27%), household cats (19%), and foxes (15%). In urban areas, egg output was dominated by stray cats (81%). Intervention scenarios revealed that only with a high compliance (90%) to the four times a year deworming advice, dogs' contribution would drop from 39 to 28%. Alternatively, when 50% of owners would always remove their dogs' faeces, dogs' contribution would drop to 20%., Conclusion: Among final hosts of Toxocara older than 6 months, dogs are the main contributors to the environmental egg contamination, though cats in total (i.e. both owned and stray) transcend this contribution. A higher than expected compliance to deworming advice is necessary to reduce dogs' egg output meaningfully. Actions focusing solely on household dogs and cats are unlikely to sufficiently reduce environmental contamination with eggs, as stray cats and foxes are also important contributors.

The prevalence of gastrointestinal parasites and risk factors for shedding of Toxocara eggs were determined for 916 Dutch household dogs older than 6 months. Additionally, the owners answered a questionnaire about their dogs and their attitude towards routine deworming was assessed. Faecal samples were examined using the centrifugal sedimentation flotation method. The overall prevalence of dogs shedding Toxocara eggs was 4.6 %. Multivariable logistic regression analysis revealed that the risk for 1-7-year-old dogs to shed Toxocara eggs was significantly lower (OR 0.38) than that of 6-12-month-old dogs. Compared to dogs walking ≤20 % of the time off-leash, those ranging freely 50-80 % and 80-100 % of the time had a significantly higher risk (OR 10.49 and 13.52, respectively) of shedding Toxocara eggs. Other risk factors were coprophagy (OR 2.44) and recently being kenneled (OR 2.76). Although the applied deworming frequency was not significantly associated with shedding Toxocara eggs, there was a trend towards no shedding in dogs under strict supervision that were dewormed 3-4 times a year. Most dog owners (68 %) recognized 'dog's health' as the main reason for deworming. Only 16 % of dogs were dewormed four times a year. It was concluded that the prevalence of Toxocara egg-shedding household dogs is almost unchanged over recent years and that the knowledge of owners is insufficient to expect sound decisions on routine deworming.

The normal time interval between treatment of horses with a macrocyclic lactone (ML) and reappearance of strongyle eggs in the feces, or 'Egg Reappearance Period (ERP)', is at its shortest 8 weeks for ivermectin (IVM) and 12 weeks for moxidectin (MOX). Nowadays, it is not uncommon to find shorter ERPs, potentially indicating the beginning of the development of drug resistance. Whether all cyathostomin species contribute equally to a shortened ERP is not known. In the present study a Reverse Line Blot (RLB) on individual infective larvae was used to compare species composition before and after ML-treatment in horses with either a normal or a shortened ERP. Species composition was determined for a total of 21 horses on 4 farms and was based on differentiation of approximately 40 larvae per horse per sampling day. After IVM treatment the shortest ERP was found to be 28-42 days. After MOX treatment the shortest ERP was found to be 42-56 days. The RLB showed that early egg shedding was dominated by species of the genus Cylicocyclus (Cyc.) (Cyc. ashworthi, Cyc. insigne, Cyc. leptostomum and Cyc. nassatus). Before treatment, species composition was generally more diverse with species from several other genera also present. Interestingly, on the farm with the overall shortest ERP and where all horses showed a shortened ERP, more species/genera were present during early egg excretion than on other farms. Results suggest that cyathostomin species within the genus Cylicocyclus contribute to a shortened ERP more than species from other genera. However, if shortening of ERP overall becomes more pronounced and widespread within a farm, species from other genera will also be present during early egg shedding., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Many dogs display coprophagic behaviour. Helminth eggs can passively pass the dog's digestive tract and this may result in a false positive diagnosis of infection with gastrointestinal helminth parasites. For a period of one year, faecal samples of dogs were examined monthly using the Centrifugal Sedimentation Flotation (CSF) technique with a sugar flotation solution (s.g. 1.27-1.30 g/cm(3)). If a sample tested positive for canine helminth eggs, the owner was asked to submit another sample after preventing the dog from eating faeces for 3 days. If the second sample again tested positive for the same type of helminth egg, the dog was considered to have a patent infection. If the second sample tested negative, the first sample was considered a false positive due to coprophagy. The focus of this study was on dogs shedding Toxocara eggs. At the first examination, 246 samples (out of 308 samples testing positive for canine-specific helminth eggs) tested positive for Toxocara spp. Of these, 120 (49%) tested negative at the second examination. Coprophagic behaviour was recognized by 261 of the 564 owners that answered the accompanying questionnaire. This concerned 391 dogs. Coproscopical examination also provided proof of coprophagy (e.g. oocysts of Eimeria spp. or non-dog typical helminth eggs) in dogs belonging to owners that did not report coprophagic behaviour in their dogs. Results indicate that coprophagy in dogs may result in an overestimation of the prevalence of patent helminth infections and that dogs may serve as a transport host for helminth eggs., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Lungworm antibody ELISAs developed in Germany (DE) and The Netherlands (NL) were compared using four sets of serum (S) and bulk-tank milk (BTM) samples from adult dairy cows. The samples originated from 37 farms with or without a suspected clinical lungworm infection during August-October 2010 (Dataset 1), from cows excreting lungworm larvae or not during August-October 2010 (n=59) or May-June 2011 (n=164) (Dataset 2), from 305 farms in a national survey during October 2010 (Dataset 3), and 14 zero-grazing farms during February-April 2011 (Dataset 4). During August-October 2010, covering the second half of the grazing season, the NL-S and NL-BTM ELISA outperformed the DE-S and DE-BTM ELISAs in terms of sensitivity. For at least the NL-S and DE-S ELISA the opposite was found during May-June 2011, covering the end of the winter housing period and the early grazing season. Of the 305 farms in the survey 62.6% were found positive with the NL-BTM ELISA, whereas 2.6% was found positive with the DE-BTM ELISA. ODR values for the zero-grazing farms indicated that a cut-off value of 30% for the DE-BTM ELISA might be more appropriate than the currently used 41%. Results suggest that the NL ELISAs also respond to lungworm antigens other than Major Sperm Protein as used in the DE ELISAs. It is concluded that the generally higher sensitivity of the NL-BTM ELISA makes it better suited for large-scale prevalence studies and herd health monitoring programmes than the DE-BTM ELISA, positivity of which is more associated with the presence of clinical lungworm infection. Reducing the cut-off value of the DE-BTM ELISA from the original 49.3% to the current 41% or the possibly more appropriate 30% increased its sensitivity for detecting lungworm infections, but did not lead to similar sensitivity estimates as found for the NL-BTM ELISA., (Copyright © 2013 Elsevier B.V. All rights reserved.)

A study was conducted to evaluate the occurrence of resistance against, in particular, ivermectin in cyathostomins in the Netherlands. Seventy horse farms were visited between October 2007 and November 2009. In initial screening, faecal samples were collected 2 weeks after deworming with either ivermectin, moxidectin or doramectin. Pooled faecal samples from a maximum of 10 horses were examined for worm eggs using a modified McMaster technique and for worm larvae after faecal larval cultures. In total 931 horses were involved. On 15 of 70 farms eggs and/or larvae were found. On 8 of these 15 farms a FECRT with ivermectin was performed on 43 horses. Efficacy of ivermectin against cyathostomins of 93% was found in one animal on one farm. Additionally, the strategies and efforts of the horse owners to control cyathostomins, as well as risk factors for the development of macrocyclic lactone resistance were evaluated with a questionnaire. Strikingly, many responders indicated that the control of cyathostomins in horses is achieved through very frequent deworming. Fourteen percent of these owners deworm seven times per year or more. On 34% of the 70 farms treatment was repeated within the Egg Reappearance Period of a product., (Copyright © 2011 Elsevier B.V. All rights reserved.)

On two dairy farms it was attempted to eradicate lungworm, Dictyocaulus viviparus, by means of a single mass-treatment of all cattle that had been grazed the previous year(s), before turnout in the spring. Both farms experienced two years of lungworm outbreaks in the adult dairy herd prior to this study. Following confirmation that both herds contained lungworm carriers, all animals older than approximately 6 months were treated with eprinomectin in March 2007. One week after treatment none of the animals were shedding lungworm larvae. Subsequently, animals were pastured according to normal farm routine. From August to November all first-calving heifers were coprologically and serologically monitored for lungworm infection. During 2007 both farms remained lungworm-negative and did not report any clinical sign indicative for a lungworm infection. The following year, on one of the farms replacements grazing on cow pastures, started showing signs of parasitic bronchitis which was serologically confirmed. The other herd remained free of parasitic bronchitis until at least the fourth year after the mass treatment, although some coughing was noticed in 2008 among first-lactation heifers. It was concluded that a single mass-treatment before the grazing season may be useful to break a series of annual lungworm outbreaks. However, it is not a secure method to prevent parasitic bronchitis for more than one year., (Copyright © 2011 Elsevier B.V. All rights reserved.)

To test the value of a recently developed bulk-tank milk (BTM) ELISA for diagnosing (sub)clinical Dictyocaulus viviparus infection in lactating dairy herds under field conditions, bulk milk samples were collected from farms with or without clinical symptoms suspected to be caused by lungworm infection. Results of the BTM ELISA were compared against individual examinations for lungworm larvae in faeces and lungworm antibodies in serum from up to 20 heifers (parity 1) and up to 20 cows (parity ≥ 2) on the same farms. This also allowed, for the first time, to examine the value of individual faecal and serological examinations in the diagnosis of (sub)clinical lungworm infections. In total, 33 farms participated. Of these, 16 reported clinical symptoms possibly related to lungworm infection (defined as a suspected positive clinical status or CS(+)) and 17 reported having no such symptoms (CS(-)). In total, 503 heifers and 649 cows were sampled. Of all faeces samples positive for lungworm larvae, 94 were from heifers (18.9% of all heifers) and 75 from cows (11.7% of all cows) (P<0.001). Of all sera positive for lungworm antibodies, 130 were from heifers (26.1% of all heifers) and 113 from cows (17.5% of all cows) (P<0.001). Of the CS(-) farms 41% had at least one heifer or cow shedding larvae and 71% had at least one seropositive heifer or cow. Of the CS(+) farms this was 81% and 94%, respectively. There were only 4 farms, all CS(-), where none of the animals were found shedding larvae and all animals tested seronegative. This implies that on 76% of the CS(-) farms lungworm infection circulated unnoticed. On all CS(+) farms the suspicion that lungworm caused the respiratory symptoms was confirmed by the individual faecal and serological examinations, whereas the BTM ELISA confirmed presence of lungworm on half of the CS(+) farms. The latter in particular occurred on farms with the more severe outbreaks. Overall, of 32 available BTM samples 10 tested positive (8 of 15 CS(+) and 2 of 17 CS(-) farms). For diagnosing suspected lungworm disease it was concluded that testing a BTM sample might suffice in case of moderate to severe outbreaks. However, in case of a mild outbreak with just a few animals coughing, examining individual animals has to be preferred over testing a BTM sample. The likelihood to detect lungworm infection is higher if heifers are sampled compared to cows. Sensitivity of the BTM ELISA was 35.7% if the presence of at least one seropositive and/or one larvae shedding animal in the herd was used to define lungworm positive farms. On average, at least 30% of the herd had to be seropositive before the BTM ELISA was found positive for lungworm antibodies. Results indicate that the BTM ELISA in its current form does not appear to be suitable for surveys on the prevalence of lungworm presence on farms. However, this BTM ELISA might be used in large-scale surveys to detect, for instance, annual changes in percentage positive farms, as long as it is recognized that positivity is more closely related to incidence of lungworm disease than to prevalence of lungworm infection., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Two lungworm outbreaks in dairy herds were investigated in order to estimate the resulting economic costs. On the two farms, with 110 and 95 cows, total costs were estimated at €159 and €167 per cow, respectively. Overall, milk production reduced by 15 to 20 per cent during the outbreaks. Five cows died on one farm, while on the other farm seven cows died as a result of the lungworm outbreak. On one farm, 51.7 per cent of the total costs was due to reduced milk production and 33.1 per cent was due to disposal of dead animals. On the other farm, it was 36.3 and 50.9 per cent, respectively. The remaining 13 to 15 per cent of the total costs were due to extra inseminations, laboratory diagnosis and treatments. The history and development of the outbreaks are described. One lesson from these outbreaks is that recognising that potentially lungworm-naïve animals are to be introduced into the adult herd allows for timely measures (for example, vaccination) to prevent a lungworm outbreak.

Cyathostomins are considered to be the primary helminth pathogen of horses and macrocyclic lactones (ML) are the most frequently used anthelmintics. Therefore, ML resistance is a serious threat for the control of these parasites. In the present study ivermectin resistant cyathostomin L3 were in vitro selected, using a reiterative larval migration inhibition assay (rLMIA) and differentiated by reverse line blot (RLB). Larvae were obtained from two populations, one from a never treated, free-roaming horse population in the nature reserve Oostvaardersplassen (OVP) and the other from regularly ivermectin-treated ponies of Utrecht University (UU). In the rLMIA the proportion of larvae that migrated increased with each passage, demonstrating that the applied procedure indeed selects for larvae the least susceptible for ivermectin. This was further supported by the fact that glutamate addition to this procedure reversed the selection effect, which also suggests that glutamate-gated chloride channels (GluCls) play a role in the ivermectin resistance of the selected L3. In both populations the predominant species were Cyathostomum catinatum, Cylicostephanus longibursatus and Cylicocyclus nassatus. After in vitro selection in the rLMIA in the presence of ivermectin the predominant species became C. catinatum in both larval populations, while C. nassatus disappeared in the never treated OVP larval population but not in the regularly ivermectin-exposed UU population. It is concluded that the rLMIA and RLB can be used to study anthelmintic resistance in cyathostomin populations and to study differences and changes in species composition between populations with different anthelmintic exposure histories., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Prevalence of Tritrichomonas foetus among Dutch cats The aim of this study was to determine the prevalence of and risk factors for, Tritrichomonas foetus among cats in the Netherlands. A total of 154 faecal samples were collected from three groups of cats: cats with diarrhoea (n=53), cattery cats (n=47), and healthy pet cats (n=54). Faecal samples were examined with a T. foetus specific real-time PCR. All PCR-positive samples were run on gel electrophoresis for definitive diagnosis. The prevalence of T. foetus was 2% among cats with diarrhoea and 4% among cattery cats; T. foetus was not prevalent among pet cats (none of the samples tested positive). Questionnaires had been distributed to cat and cattery owners to determine risk factors for T. foetus, but the low prevalence precluded statistical analysis of the questionnaire results.

Until recently, autochthonous infections with Angiostrongylus vasorum were not thought to occur in The Netherlands. However, in 2007 four dogs were diagnosed with angiostrongylosis. Three of them had never left the country. Subsequently, faecal samples from 485 dogs living in the same region where the positive dogs had been diagnosed in 2007 were collected through veterinarians, hunting associations and kennels between January and April 2008. A. vasorum larvae were found in faecal samples of four dogs. Three of those had never been outside the country. Though only few samples were found positive, A. vasorum is now considered autochthonous in The Netherlands.

Background: Parasitic gastroenteritis caused by nematodes is only second to mastitis in terms of health costs to dairy farmers in developed countries. Sustainable control strategies complementing anthelmintics are desired, including selective breeding for enhanced resistance., Results and Conclusion: To quantify and characterize the genetic contribution to variation in resistance to gastro-intestinal parasites, we measured the heritability of faecal egg and larval counts in the Dutch Holstein-Friesian dairy cattle population. The heritability of faecal egg counts ranged from 7 to 21% and was generally higher than for larval counts. We performed a whole genome scan in 12 paternal half-daughter groups for a total of 768 cows, corresponding to the approximately 10% most and least infected daughters within each family (selective genotyping). Two genome-wide significant QTL were identified in an across-family analysis, respectively on chromosomes 9 and 19, coinciding with previous findings in orthologous chromosomal regions in sheep. We identified six more suggestive QTL by within-family analysis. An additional 73 informative SNPs were genotyped on chromosome 19 and the ensuing high density map used in a variance component approach to simultaneously exploit linkage and linkage disequilibrium in an initial inconclusive attempt to refine the QTL map position.

A study was performed on two horse farms to evaluate the use of age-clustered pooled faecal samples for monitoring worm control in horses. In total 109 horses, 57 on farm A and 52 on farm B, were monitored at weekly intervals between 6 and 14 weeks after ivermectin treatment. This was performed through pooled faecal samples of pools of up to 10 horses of the groups 'yearlings' (both farms), '2-year-old' (two pools in farm A), '3-year-old' (farm A) and adult horses (four pools on farm A and five pools on farm B), which were compared with the mean individual faecal egg counts of the same pools. A very high correlation between the faecal egg counts in pooled samples and the mean faecal egg counts was seen and also between the faecal egg counts in pooled samples and larval counts from pooled faecal larval cultures. Faecal egg counts increased more rapidly in yearlings and 2-year-old horses than in older horses. This implied that in these groups of young animals faecal egg counts of more than 200 EPG were reached at or just after the egg reappearance period (ERP) of 8 weeks that is usually indicated for ivermectin. This probably means that, certainly under intensive conditions, repeated treatment at this ERP is warranted in these young animals, with or without monitoring through faecal examination. A different situation is seen in adult animals. Based on the mean faecal egg counts on both farms and on the results of pooled samples in farm A, using 100 EPG as threshold, no justification for treatment was seen throughout the experimental period. However, on farm B values of 100 EPG were seen at 9 and 11, 13 and 14 and 14 weeks after ivermectin treatment in pools 10, 12 and 13, respectively. This coincided with the presence of one or two horses with egg counts above 200 EPG. The conclusion is that random pooled faecal samples of 10 adult horses from a larger herd, starting at the ERP and repeating it at, for instance, 4-week intervals, could be used for decisions on worm control. However, there would be a certain risk for underestimating pasture contamination through missing high-egg excreters. An alternative use of pooled samples would be as a cheap first screening to detect which adult horses really contribute to pasture contamination with worm eggs on a farm. All horses should be sampled and subsequently animals from 'positive' pools can be reexamined individually.

Parasite N-glycans may play an important role in helminth infections. As antibodies from Dictyocaulus viviparus-infected calves strongly react with N-glycans, we investigated the characteristics of the major immunodominant glycoprotein (GP300) of this parasite. Probing of worm extracts with various lectins demonstrated unique binding of GP300 to wheat germ agglutinin. Analysis of lectin-purified GP300 revealed that the glycan was substituted with phosphorylcholine and reacted with the phosphorylcholine-specific antibody TEPC-15. Competitive enzyme-linked immunosorbent assay with GP300-coated plates and GP300-specific immunoglobulin G (IgG) in conjunction with free phosphorylcholine or TEPC-15 demonstrated that antibodies from infected calves recognized phosphorylcholine on GP300. Additional assays showed that these antibodies cross-reacted with the phosphorylcholine moiety present on platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine), a proinflammatory mediator of the host. Heavily infected calves contained high levels of serum GP300-specific IgG1 but low levels of IgA and IgG2 and showed a reduced influx of eosinophils in the lungs, all consistent with a neutralization of PAF activity. In conclusion, we demonstrated that D. viviparus infection elicits GP300-specific antibodies that cross-react with PAF and may neutralize PAF function, thus limiting the development of a protective response as well as parasite-induced host pathology.

Calves with naturally acquired Dictyocaulus viviparus infection mount an effective immune response. In the search for protection-inducing antigens, we found that several D. viviparus third-stage larval (L3) and adult ES products carry N-glycans. Deglycosylation of the worm antigens using PNGase F resulted in reduced IgA, IgE, IgG1 and IgG2 (but not IgM) reactivities in sera of primary infected animals, suggesting that the carbohydrate moieties contained immunodominant epitopes. Challenge infection resulted in increased specific serum antibody levels against ES and L3 in the re-infected and challenge control groups. Testing of sera by enzyme-linked immunosorbent assay (ELISA) demonstrated a significant increase in IgG1 and IgE (but not IgA or IgG2) reactivity against the deglycosylated antigens in the re-infected group compared with the challenge control group. Sera from calves vaccinated with irradiated larvae showed a strong anti-N-glycan response, but no booster response against the protein backbone after challenge infection, consistent with the absence of a memory response. Together, our results suggest that D. viviparus proteins carry immunodominant N-glycan moieties that elicit a strong but short-lived immune response during infection and after vaccination, whereas the protein backbones effectively induce a memory response which results in a long-lasting, potentially protective immune response in re-infected, but not in vaccinated calves.

Large-scale parasite quantification is required for improving our understanding of the epidemiology and genetics of host-parasite interactions. We describe a protocol that uses a low-density salt solution for flotation and centrifugation of nematode eggs. Subsequently, sucrose flotation and precipitation are used to obtain clear egg preparations. Most traditional quantification protocols such as the McMaster technique are unsuited for the standardized processing of large numbers of samples and the analysis of large amounts of feces per sample. Consequently, they are suited only for small-scale surveys. Our protocol, which can be used to analyze up to 6 g of feces, results in clear egg preparations that are concentrated in wells of a microtiter plate and that are suited for digital recording and automated counting. Starting from a fecal suspension in the first flotation solution to a digital recording requires approximately 40 min per 24 samples.

Two experiments were performed in 2002 and 2003 to evaluate the effect of biological control of gastrointestinal nematodes in sheep through the daily feeding of 500,000 chlamydospores of Duddingtonia flagrans/kg bodyweight to lactating ewes during the first 9 weeks with their young lambs on pasture. In both experiments four groups of eight ewes and their April-borne lambs were used. They were turned out on four separate plots (plots A) at the beginning of May, moved to similar separate plots after 3 (plots B) and 6 weeks (plots C), respectively, and weaning occurred after 9 weeks. In both experiments, two groups were fed spores daily while the two other groups served as controls. The effect of D. flagrans application was evaluated through faecal egg counts of ewes and lambs, the yield of faecal cultures in ewes, pasture larval counts and worm counts of lambs and tracer lambs. The results demonstrated no effect of D. flagrans application during the first 5 (2002) or 4 (2003) weeks. Subsequently, fungus application strongly reduced the yield in faecal cultures of the ewes. This was, however, not reflected in the pasture larval counts, but lower worm burdens were observed in tracer lambs of 'treated' plots C in 2002 than on those of 'control' plots. In 2003 worm burdens in 'treated' lambs returned to plots B were lower than those of 'control' lambs and a tendency for the same was observed for plots C. However, in all groups, lambs and tracer lambs developed severe haemonchosis.

In 2004, an experiment was carried out to evaluate the effect of biological control through feeding spores of Duddingtonia flagrans on parasitic gastroenteritis in lambs, kept under an evasive grazing system. In total 66 lambs were used. Forty naturally infected 3-month old ram lambs were weaned in mid June, and divided into four groups of 10 lambs. On 21 June, G1-G4 were moved to four separate virtually clean plots, they were moved after 4 and 8 weeks to similar plots, and housed after 12 weeks to be necropsied 16 days later. The other 26 lambs had been raised helminth-free, and were used as pairs of tracer lambs. All but one of these pairs, were grazed during the last 2 weeks on each plot. The remaining pair (TA) was grazed during the last 2 weeks on pasture (30 August to 13 September) on the plot that had been grazed by G3 between 19 July and 16 August, to study inhibited development in Haemonchus contortus. All lambs were fed 200 g of concentrates daily throughout the whole period, and those of G1 and G2 were also fed 500,000 spores of D. flagrans/kg bodyweight daily. The faecal cultures demonstrated a high reduction in yield as a result of fungal application. However, no differences between groups were seen in weight gain, faecal egg counts, pasture larval counts, worm counts and tracer worm counts. H. contortus was the dominant species, and it is obvious that the moves at 4-week intervals prevented the development of severe haemonchosis. This is in particular demonstrated by the much higher worm counts in the two TA tracer lambs grazed. Nevertheless, increases to high faecal egg counts 3 weeks after the first and second moves, indicated acquisition of infection before these moves and at least subclinical haemonchosis. This was supported with the worm counts of lambs and tracer lambs. A higher proportion of inhibited early L4 than in other tracers and than in the permanent lambs were found in the pair of TA tracer lambs. This indicates that moves to new pastures in late summer and autumn delays the onset of inhibition.

Due to excessive and inappropriate use of dewormers anthelmintic resistance has developed as a significant problem in horse parasites in the Netherlands. Since it is unlikely that new classes of anthelmintics against horse nematodes will be introduced in the near future, it is important to use the present drugs wisely. Veterinarians should advice horse owners about worm control programs with a more targeted approach. The number of anthelmintic treatments should be reduced and, through selective anthelmintic treatments, further development of anthelmintic resistance should be delayed. Preferably, horses with a low faecal egg count should not be treated at all to ascertain a reduction of the selection pressure for anthelmintic resistance. The propensity for low faecal egg counts is hereditary. This implies that mature horses with consistent low egg counts can be detected by faecal examination and that it is not necessary to repeat faecal examination each time in these animals. New horses on the farm should always be dewormed on arrival and should be introduced only after the efficacy of treatment has been determined. Anthelmintic resistance can also be introduced with the arrival of a new animal that is infected with drug-resistant parasites.

Resistance of Teladorsagia circumcincta in goats to ivermectin In 2005, suspected ivermectin resistance of gastrointestinal nematodes was investigated in 17 goats on a farm in the Netherlands, using a faecal egg count reduction test combined with faecal larval cultures. Eight goats were treated orally with 0.2 mg/ kg ivermectin on 1 August 2005 and the other nine goats served as untreated controls. Faeces were collected from all goats on 1 August and on 12 August. Faecal egg counts were reduced by 80.6% in goats receiving ivermectin. On the basis of faecal larval cultures and the egg counts, ivermectin was found to be 100% effective against Haemonchus contortus, 15.0% effective against Teladorsagia circumcincta, and 93.3% effective against Trichostrongylus spp. The effect of levamisole and oxfendazole on gastrointestinal nematodes was also evaluated. Nine goats were treated subcutaneously with 5 mg/kg levamisole and seven goats orally with 4.5 mg/kg oxfendazole on 12 September On the basis of faecal egg counts performed on 12 and 22 September, levamisole reduced the total egg count by 95.4% and oxfendazole reduced the egg count by 85.8%. Differentiation of pooled faecal larval cultures revealed that levamisole was 99.9% effective against H. contortus, 81.9% effective against T. circumcincta, and 99.5% against Trichostrongylus spp. Oxfendazole was 75.5% effective against H. contortus, 33.2% effective against T. circumcincta, and 100% effective against Trichostrongylus spp. On 28 October the nine goats that were still present on the farm were orally treated with 0.4 mg/kg ivermectin. Faecal egg counts and differentiation of pooled larval cultures of faecal samples collected on 28 October and 9 November, respectively, revealed that total egg counts were reduced by 93.1%. Ivermectin was 49.1% effective against T. circumcincta but 100% effective against H. contortus and Trichostrongylus spp. Thus H. contortus appears to be resistant to oxfendazole and T. circumcinta appears to be resistant to oxfendazole and ivermectin. This is the first documented case of ivermectin resistance of a gastrointestinal nematode of small ruminants in the Netherlands.

During 2003 a grazing study was performed at Utrecht University to evaluate evasive grazing and application of Duddingtonia flagrans for the control of parasitic gastroenteritis in sheep. However, the summer of 2003 (June-August) was unusually warm and dry. As a result the patterns of gastrointestinal nematode infections deviated from those observed in more average years. The proportion of eggs that developed to infective larvae was far lower than normal in July-August. On the other hand, survival of larvae that had developed before the middle of July was not affected compared to other years. In fact, severe haemonchosis was observed in (tracer) lambs grazed at the end of July on pastures that had been contaminated from 26-05 to 16-06 and from 16-06 to 07-07. Moreover, tracer lambs grazing in September on some of these plots still acquired large Haemonchus contortus burdens. Over 60% of H. contortus that had established before the middle of July appeared to be able to survive until October in the virtual absence of re-infection. That may have consequences for the application of evasive grazing as a control option in that suppression of adult burdens might still be necessary, through a limited use of anthelmintics or through alternative deworming strategies.

Between 1999 and 2003 studies were done at Utrecht University on the population dynamics of gastrointestinal nematodes and the potential to use evasive grazing for the control of nematode infections in small ruminants. On most pastures grazed in May and June it took at least 3 weeks before high pasture infectivity levels for Haemonchus contortus developed while it took at least 2 weeks on pastures grazed in July, August and September. Occasionally, larvae emerged earlier on pasture when high temperatures coincided with rain. Pasture infectivity levels for H. contortus subsequently increased and the highest levels were found between 5 and 9 weeks on pastures grazed in May-June and between 3 and 9 weeks on pastures grazed in July, August and September. Pasture infectivity usually had decreased again to low levels after approximately 3 months. Larvae of the other trichostrongylids emerged later on pasture. However, patterns of Teladorsagia circumcincta, Trichostrongylus spp. and Cooperia curticei were basically similar to H. contortus. Strongyloides papillosus larvae emerge within 2 weeks on pasture and survival is short. On the other hand Nematodirus larvae took a lot longer to emerge on pasture than the other trichostrongylids. The results imply that only on a small proportion of farms (mixed dairy cattle/sheep farms; some organic dairy goat farms) evasive grazing can be recommended as only control measure for parasitic gastroenteritis. However, the present data offer possibilities to combine evasive grazing with other methods in an integrated control scheme.

Control of parasitic gastroenteritis in small ruminants is threatened by the worldwide growing problem of anthelmintic resistance. Therefore, alternative approaches for worm control are imperative. Of utmost importance is to slow down selection pressure for anthelmintic resistance by using alternative control measures combined with minimal anthelmintic usage. New control options for the Netherlands are discussed.

A study of the safety of a vaccine against lungworm was carried out with pregnant and lactating heifers from three dairy herds with a previous history of lungworm outbreaks in adult cows. Half of the heifers were vaccinated while the other half were not. A slight temporary cough following the vaccination was only observed in one herd. No adverse effects on pregnancy or milk production were seen. All heifers were serologically and coprologically examined before the first, before and after the second immunization, 3 months after introduction to pasture and at the end of the grazing season. Serological and faecal examination of the dairy cows before introduction into pasture confirmed the presence of at least one Dictyocaulus viviparus carrier in each herd. Lungworm infection occurred in all herds during the grazing season, most prominently in the herd with the highest number of heifers. In this herd, mild coughing associated with the lungworm infection was noticed, especially in the non vaccinated heifers. No other signs or symptoms were observed. It is concluded that a vaccine against D. viviparus can be used safely in heifers, before they are introduced into the adult herd, and that this vaccine can be used as a preventive measure against lungworm outbreaks in adult cattle.

We investigated the effect of bovine anamnestic immunity on a range of parasitological variables. To this end, calves were primed with a single oral dose of 30000 or 100000 infective larvae (L3) of Cooperia oncophora, drenched with anthelmintic, maintained worm free in the following 2.5 months and subsequently re-infected with 100000 L3. Parasitological profiles of low, intermediate, and high responders were compared. The reduction in establishment of the worms was shown by a lower worm burden and increased percentage of fourth-stage (L4) larvae. Worm length and fecundity were similarly reduced by both priming doses but, the speed by which the effect occurred differed between animals primed with 30000 or 100000 L3. The difference in establishment between the responder types demonstrates that the ability of intermediate responders to mount a more effective and faster immune response compared to low responders is sustained after secondary infection.

We used factor and path analyses to identify the causal paths or relationships between variables affecting the demand and use by subsistence farmers of the various services offered by government-employed veterinary livestock technicians (VLTs) in Zimbabwe in 1996. We examined whether the farmer had implemented the VLT-recommended livestock-management procedures. Various factors that have been reported or theorized as important in the demand for veterinary services by subsistence farmers are described. The important factors positively affecting whether farmers had sought veterinary assistance in the last 24 months included familiarity of the farmer with the VLT, if situated closer to the VLT's base, and if the farmer received further training in agriculture (either through courses or from attendance at demonstrations on animal husbandry conducted by the VLT). Factors affecting whether farmers implemented the recommended management practices that had a monetary cost included familiarity with the VLT, proximity to the VLT's premises, farmer literacy, club membership, and farmer's wealth. There was a difference in the paths generated for preventive measures that had a monetary cost versus those measures that involved only a labour or effort costs. For the former practices, the VLT played an important role informing farmers by extension and demonstrations, and farmer's membership of a club increased their performance.

We investigated whether the generation of protective memory humoral immunity in Cooperia oncophora infected calves occurs in a dose-dependent way and whether it depends on the animal responder types. To this end, serum and mucus antibody responses were measured in animals primary-infected with 30000 or 100000 L3, treated with anthelmintics and subsequently challenged with 100000 L3. A detailed phenotypic and functional analysis of B cells was done in animals infected once or twice with 100,000 L3. Based on the similarity in parasitological variables of animals primed with 30000 or 100000 L3, we concluded that with these doses priming conferred protection in a dose-independent way. Upon challenge significant increases in Cooperia-specific serum and mucus IgG1 and IgA and total serum IgE titres were induced in primed animals in a dose-independent way. In contrast, intermediate and low responders differed in the onset of the production of Cooperia-specific serum IgG1. Furthermore, not only the onset but also the level of total serum IgE significantly differed between intermediate and low responders. Phenotypic and functional analysis of B lymphocytes revealed that (i). priming induced the generation of memory B cells which upon challenge readily differentiated into antibody secreting cells; (ii). sensitised B cells were more efficiently recruited to the intestinal effector sites; (iii). based on the expression of CD62L and CD86 two distinct B cell subpopulation could be differentiated. CD62L(+)CD86(-) B cells that were likely lymphocytes not yet activated and with an enhanced recirculation capacity, and CD62L(-)CD86(+) B cells that were activated B cells with a reduced recirculation ability; and finally (iv). the increased expression of CD86 and subsequent correlations with parameters of the T helper 2 immune response induced by C. oncophora, suggested that CD86- interactions are involved in the generation of protective immunity against Cooperia.