Route of administration affects the efficacy of moxidectin against Ostertagiinae nematodes in farmed red deer (Cervus elaphus).

The influence of route of administration on the pharmacokinetics and efficacy of macrocyclic lactone anthelmintics has been a subject of interest due to its potential to influence the development of anthelmintic resistance. For most parasite species studied so far, oral administration results in the highest concentrations of drug in the parasites and the highest efficacy against resistant genotypes. However, a recent study in cattle measured the highest levels of ivermectin in the abomasal Ostertagia ostertagi following subcutaneous injection, but it was not possible to correlate these elevated levels with efficacy. Therefore, the current study was initiated to determine whether injectable delivery might be optimal for attaining high efficacy against this important group of parasites. Three on-farm trials were conducted to measure the efficacy of moxidectin administered by the oral, injectable, and pour-on routes against Ostertagiinae parasites in farmed red deer. Groups of rising 1-year old stags (red or red-wapiti crossbreds) in the 84-104 kg weight range were randomised on liveweight into treatment groups of 6 (1 farm) or 8 (2 farms). Animals were treated to individual liveweight with moxidectin oral (0.2 mg/kg), injectable (0.2 mg/kg), pour-on (0.5 mg/kg) or remained untreated. Twelve days later all animals were euthanised and abomasa recovered for worm count. Adult worms were counted in a 2% aliquot of abomasal washings, and adult and fourth stage larvae in a 10 % aliquot following mucosal incubation in physiological saline. In addition, blood was collected from the same 5 animals in each of the treatment groups on days 0, 1, 2, 3, 5, 7 and 12 after treatment and moxidectin levels in plasma were determined using a mass spectrometer. The number of Ostertagiinae surviving treatment was significantly different for each of the treatment groups with injectable administration being most effective, oral administration being the next most effective and pour-on administration the least effective. This applied to both adult worms and fourth stage larvae. A similar pattern was seen in the levels of moxidectin in plasma with both the peak value and area under the concentration curve being highest following injectable administration and lowest following pour-on treatment. Although undertaken in a different host species, the results support the proposition that injectable administration of macrocyclic lactone anthelmintics is likely to be optimal for efficacy against Ostertagiinae parasites and potentially useful in slowing the emergence of resistance in these parasites.

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

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

Efficacy and production benefits following use of eprinomectin extended-release injection in pastured dairy heifers.

A study was conducted in grazing dairy heifers to assess anthelmintic efficacy and production responses in dairy heifers treated with a single injection of eprinomectin in an extended-release formulation over a 123 day-period. The study was conducted on a pasture-based dairy in the Southeastern United States (North Carolina) over the summer months. Sixty crossbred dairy heifers were weighed and randomly allocated into 2 groups. One group (n = 30) was given 5% eprinomectin subcutaneously in the cervical region while the other group (n = 30) was given an equivalent volume of saline. Calves were weighed every 30 days throughout the trial for calculation of average daily gain and differences in overall weight gain. In addition, fecal samples were collected at days 0, 30, 60, 90 and 123 for worm egg count and coproculture. Both groups of cattle had similar worm egg concentrations at the start of the study. However, the control group had increasing concentrations of fecal worm eggs throughout the summer months while the heifers that received eprinomectin had minimal fecal worm eggs. The primary parasite species identified in this study were Haemonchus placei, Cooperia species and Ostertagia. The heifers that received eprinomectin gained 105 + 2.8 kg during the 123-day study period, representing an average daily gain of 0.85 kg/day compared to 78.3 + 4.1 kg (0.64 kg/day) for the control group. This represented a 33 % increase in average daily gain associated with deworming. The results of this study indicate that a single dose of extended-release eprinomectin was sufficient to control parasites through a 123-day summer grazing season and that administration of the anthelmintic had a significant impact on weight gain.

Assessing anthelmintic resistance risk in the post-genomic era: a proof-of-concept study assessing the potential for widespread benzimidazole-resistant gastrointestinal nematodes in North American cattle and bison.

As genomic research continues to improve our understanding of the genetics of anthelmintic drug resistance, the revolution in DNA sequencing technologies will provide increasing opportunities for large-scale surveillance for the emergence of drug resistance. In most countries, parasite control in cattle and bison has mainly depended on pour-on macrocyclic lactone formulations resulting in widespread ivermectin resistance. Consequently, there is an increased interest in using benzimidazole drugs which have been used comparatively little in cattle and bison in recent years. This situation, together with our understanding of benzimidazole resistance genetics, provides a practical opportunity to use deep-amplicon sequencing to assess the risk of drug resistance emergence. In this paper, we use deep-amplicon sequencing to scan for those mutations in the isotype-1 ß-tubulin gene previously associated with benzimidazole resistance in many trichostrongylid nematode species. We found that several of these mutations occur at low frequency in many cattle and bison parasite populations in North America, suggesting increased use of benzimidazole drugs in cattle has the potential to result in widespread emergence of resistance in multiple parasite species. This work illustrates a post-genomic approach to large-scale surveillance of early emergence of anthelmintic resistance in the field.

Anthelmintic resistance in goat herds-In vivo versus in vitro detection methods.

Anthelmintic resistance (AR) is a serious threat to animal health and has a major economic impact worldwide due to production and financial losses. The aim of this study was to determine the occurrence of AR on 30 goat farms in Slovakia during the pasturing seasons and to compare three widely used in vitro and in vivo methods for detecting AR in field conditions. A three-year survey was conducted during the pasturing seasons of 2014-2016. Goats on each farm were split into treated and control groups and were treated by recommended (5 mg/kg body weight) and double doses (10 mg/kg b.w.) of albendazole. Comparisons between percent reduction in a faecal egg count reduction test (FECRT) and an egg hatch test (EHT) and the presence of L3 larvae in a larval development test (LDT) using resistant concentrations of benzimidazole (BZ) were monitored after treatment. The FECRT indicated percent reductions of 69.2-86.2% for the single dose and of 36.3-45.4% for the double dose. The EHT indicated that all farms had BZ-resistant nematodes. Low (<15% hatching) and high (>15% hatching) levels of resistance were detected on 13 and 17 farms, respectively. The LDT failed to detect resistant larvae on seven farms but detected low and high levels of resistance on seven and 14 farms, respectively. The data indicate a moderate correlation between in vitro and in vivo tests for detecting BZ resistance among the 30 goat farms. The hatching detected by the EHT and the presence of L3 larvae by the LDT at resistant BZ concentrations provided reasonable identification of low levels of resistance in the parasite populations, but the use of a double dose for a treatment may underestimate the real occurrence of low levels of resistant parasites on goat farms.

P-glycoprotein-9 and macrocyclic lactone resistance status in selected strains of the ovine gastrointestinal nematode, Teladorsagia circumcincta.

The Teladorsagia circumcincta P-glycoprotein-9 (Tci-pgp-9) gene has previously been implicated in multiple-anthelmintic resistance in this parasite. Here we further characterise genetic diversity in Tci-pgp-9 and its possible role in ivermectin (IVM) and multi-drug resistance using two UK field isolates of T. circumcincta, one susceptible to anthelmintics (MTci2) and the other resistant to most available anthelmintics including IVM (MTci5). A comparison of full-length Tci-pgp-9 cDNA transcripts from the MTci2 and MTci5 isolates (∼3.8 kb in both cases) indicated that they shared 95.6% and 99.5% identity at the nucleotide and amino acid levels, respectively. Nine non-synonymous SNPs were found in the MTci5 sequences relative to their MTci2 counterparts. Twelve genomic sequence variants of the first internucleotide binding domain of Tci-pgp-9 were identified and up to 10 of these were present in some individual worms, strongly supporting previous evidence that amplification of this gene has occurred in T. circumcincta. On average, fewer distinct sequence variants of Tci-pgp-9 were present in individual worms of the MTci5 isolate than in those of the MTci2 isolate. A further reduction in the number of sequence variants was observed in individuals derived from an IVM-treated sub-population of MTci5. These findings suggest that Tci-pgp-9 was under purifying selection in the face of IVM treatment in T. circumcincta, with some sequence variants being selected against.

Gastrointestinal parasites of captive European bison Bison bonasus (L.) with a sign of reduced efficacy of Haemonchus contortus to fenbendazole.

The history of European bison Bison bonasus Linnaeus, 1758 has been stormy since its extinction in the wild after the First World War. Due to the fact that the species was restored from just 12 founders, further expansion has suffered from low genetic variability, rendering the bison vulnerable to various pathogens due to inbreeding depression. Parasites are recognised as a key biological threat to bison population. Thus, parasitological examination including monitoring of the level of anthelmintic resistance in a herd should be a routine procedure involved in management and protection of European bison. This study was conducted in a group of 27 bison kept in a European bison breeding centre in Sweden. In April 2015, a faecal egg count reduction test (FECRT) was performed in animals with ≥ 100 gastrointestinal nematode (GIN) eggs per gram faeces, to determine effectiveness of fenbendazole (FBZ) treatment. Additionally, the third stage larvae were cultured for molecular examination by a conventional PCR as well as by real-time quantitative PCR (q-PCR) for detection of the blood-sucking nematode Haemonchus contortus. Faecal sampling was conducted 1 day before and 8 days after deworming each animal. Anthelmintic treatment turned to be entirely efficient toward intestinal nematodes of genera Nematodirus and Trichuris, whereas shedding of strongylid eggs from the subfamily Ostertagiinae was reduced from 81 to 30%. Polymerase chain reaction (PCR) on cultured third-stage larvae (L3) before treatment was positive for H. contortus, Ostertagia ostertagi and Cooperia oncophora, whereas post-treatment examination revealed exclusively the DNA of H. contortus. Thus, only H. contortus was involved in post-treatment faecal egg count (FEC). FECRT showed that the reduction in strongylid FEC to FBZ in the examined bison herd was 87% (95%-confidence intervals [95% CI] = 76-93), suggesting reduced efficacy of FBZ to strongylid GIN including mainly H. contortus.

Pharmaco-parasitological evaluation of the ricobendazole plus levamisole nematodicidal combination in cattle.

The goals of the current study were to evaluate the potential pharmacokinetic (PK) interactions and the clinical efficacy occurring after the subcutaneous (s.c.) administration of ricobendazole (RBZ) and levamisole (LEV) given both separately and co-administered to calves naturally infected with susceptible gastrointestinal nematodes. The clinical efficacy was shown in two seasons, winter and spring, with predominance of different nematode populations. Groups of 15 calves were treated with RBZ alone, LEV alone and RBZ + LEV combination, and an untreated group was kept as a Control. RBZ and LEV plasma concentrations were quantified by HPLC. The clinical efficacy was determined by the faecal egg count reduction test. RBZ and LEV have similar plasma persistence, being detected in plasma over 24 hr post-treatment. No PK interactions were observed after the combined treatment, with similar PK parameters (p > .05) obtained for the single-drug and the combination-based strategy. In winter, the observed clinical efficacies were 96%, 99% and 100% for groups treated with RBZ, LEV and RBZ + LEV, respectively; however, in spring, the efficacies were 95%, 93% and 96% for the same groups. Remarkably, the combination was the only treatment that achieved 100% clinical efficacy against both Haemonchus spp and Ostertagia spp in winter; but the increased presence of Ostertagia spp. in spring (28% in untreated group) determined a tendency to reduced efficacies compared to winter time (only 10% of Ostertagia spp. in untreated group), even for the combined treatment. Overall, in a scenario where the nematode population is susceptible, the RBZ + LEV treatment may be a valid combination in cattle to delay the development of resistance, especially in winter when this combination achieved 100% of efficacy. Thus, selection of anthelmintic resistance will never occur. In fact, this is one of the greatest challenges for the whole cattle production system: to be one step ahead of anthelmintic resistance.

Mid-season targeted selective anthelmintic treatment based on flexible weight gain threshold for nematode infection control in dairy calves.

The suitability of a single mid-season targeted selective treatment (TST) for gastrointestinal nematodes control, based on flexible average daily weight gain (ADWG) thresholds, was investigated in 23 groups of first grazing season calves. In each group, animals were weighed three times: before turnout, at mid-season and at housing. Just after the first weighing, each group was divided in two homogenous sub-groups in terms of age, breed and weight, and randomly allocated to one of two sub-groups intented for two different mid-season anthelmintic treatment strategies: (1) a treatment of all calves composing the sub-group (whole-group treatment (WT)) or (2) a targeted selective weight gain-based treatment (TST) of the animals showing an individual pre-treatment ADWG inferior to the mean pre-treatment ADWG of the corresponding WT sub-group. Anthelmintic treatment (levamisole 7.5 mg/kg BW) was performed 3 to 4 months after turnout. At housing, two parasitological parameters (the anti-Ostertagia ostertagi antibody level-Ostertagia optical density ratio (ODR) and the pepsinogen level) and a clinical parameter (the breech soiling score) were assessed at individual level in each group. Then, the high exposed groups to gastrointestinal nematode (GIN) were defined as groups for which untreated animals exhibited a mean Ostertagia ODR ⩾0.7 and among these groups, the ones characterized by high abomasal damage due to Ostertagia for which untreated animals exhibited a mean pepsinogen level ⩾2.5 U Tyr were also identified. Among TST sub-groups, the treatment ADWG thresholds varied from 338 to 941 g/day and the percentage of treated animals from 28% to 75%. Pre- and post-treatment ADWG as well as parasitological and clinical parameters measured at housing were similar between TST and WT sub-groups including the 17 high exposed groups to GIN. Within these 17 groups, the treatment allowed to significantly improve post-treatment ADWG compared with untreated animals. In the six high exposed groups showing mean pepsinogen level ⩾2.5 U Tyr, the average effect of treatment on post-treatment ADWG was the highest and estimated up to 14 kg after a grazing duration of 4 months. In contrast, in six other groups showing mean Ostertagia ODR<0.7 in untreated animals, no effect of treatment was seen suggesting an absence of production losses related to a low level of GIN infection. This study highlighted the suitability of a convenient mid-season TST strategy for first grazing season calves, based on the use of flexible thresholds of ADWG, allowing similar growth compared with a whole-group treatment while keeping a GIN population in refugia.

Modelling the impacts of pasture contamination and stocking rate for the development of targeted selective treatment strategies for Ostertagia ostertagi infection in calves.

A simulation study was carried out to assess whether variation in pasture contamination or stocking rate impact upon the optimal design of targeted selective treatment (TST) strategies. Two methods of TST implementation were considered: 1) treatment of a fixed percentage of a herd according to a given phenotypic trait, or 2) treatment of individuals that exceeded a threshold value for a given phenotypic trait. Four phenotypic traits, on which to base treatment were considered: 1) average daily bodyweight gain, 2) faecal egg count, 3) plasma pepsinogen, or 4) random selection. Each implementation method (fixed percentage or threshold treatment) and determinant criteria (phenotypic trait) was assessed in terms of benefit per R (BPR), the ratio of average benefit in weight gain to change in frequency of resistance alleles R (relative to an untreated population). The impact of pasture contamination on optimal TST strategy design was investigated by setting the initial pasture contamination to 100, 200 or 500 O. ostertagi L3/kg DM herbage; stocking rate was investigated at a low (3calves/ha), conventional (5 calves/ha) or high (7 calves/ha) stocking rates. When treating a fixed percentage of the herd, treatments according to plasma pepsinogen or random selection were identified as the most beneficial (i.e. resulted in the greatest BPR) for all levels of initial pasture contamination and all stocking rates. Conversely when treatments were administered according to threshold values ADG was most beneficial, and was identified as the best TST strategy (i.e. resulted in the greatest overall BPR) for all levels of initial pasture contamination and all stocking rates.

Design and evaluation of multi-indicator profiles for targeted-selective treatment against gastrointestinal nematodes at housing in adult dairy cows.

Targeted-selective treatments against gastrointestinal nematode (GIN) in adult dairy cows require the identification of "cows to treat", i.e. cows whose milk production (MP) would increase after treatment. This study aimed at quantifying the ability of multi-indicator profiles to identify such cows. A randomized controlled clinical trial was conducted at housing in 25 French pasturing dairy herds. In each herd, treated cows received fenbendazole orally, control cows remained untreated. Daily MP was recorded and the MP variation between the pre- and post-visit periods was calculated (ΔMP) for each cow. ΔMP was modelled with control cows data (n=412) (piecewise linear mixed model). Estimated parameters were applied to treated cows data (n=414) to predict the expected ΔMP in treated cows if they had not been treated. Treated cows with an observed ΔMP (with treatment) higher than the expected ΔMP (without treatment) were labelled as "cows to treat". Herds where at least 50% of the young cows were "cows to treat" were qualified as "herds to target". To characterize such cows and herds, the available candidate indicators were (i) at the cow-level: parity, stage of lactation and production level, faecal egg count (FEC), serum pepsinogen level and anti-Ostertagia antibody level (expressed as ODR); (ii) at the herd-level: bulk tank milk (BTM) Ostertagia ODR, Time of Effective Contact (TEC, in months) with GIN infective larvae before the first calving, and percentage of positive FEC. These indicators were tested one-by-one or in combination to assess their ability to characterize "herds to target" and "cows to treat" (Chi-square tests). 115 out of 414 treated cows (27.8%) were considered as "cows to treat", and 9 out of 22 herds were qualified as "herds to target". The indicators retained to profile such cows and herds were the parity, the production level, the BTM Ostertagia ODR and the TEC. Multi-indicator profiles were much more specific than single indicator profiles, induced lower treatment rates, thereby minimizing the selection pressure on parasite populations. Particularly, to target a herd, the specificity was better with the profile "high BTM Ostertagia ODR and low-TEC" than with the BTM ODR value taken into account alone. The targeted-selective treatment of "young cows, belonging to herds with a high BTM ODR at housing and a low TEC" appeared as a pertinent solution, enabling a global approach for the control of GIN infection in which GIN control in heifers is connected to GIN control in adult cows.

A multi-country study to assess the effect of a treatment with moxidectin pour-on during the dry period on milk production in dairy cows.

A randomized clinical study was conducted in a total of 45 commercial dairy farms in France (14 farms), Germany (28 farms) and the UK (3 farms) to evaluate the effect of an anthelmintic treatment on milk yield in the subsequent lactation. A total of 1287 animals with suspected exposure to Ostertagia ostertagi were included in the study. Animals were treated during the dry period (7-77days before parturition) with moxidectin pour-on (Cydectin® 0.5% Pour-On, Zoetis; 638 animals) or left untreated (649 animals) according to a randomized block design. Animals were either heifers (n=296) or multiparous cows (n=991). The milk production was monitored at regular intervals after treatment up to 335days after lactation, and analysed using a general linear mixed model with the milk production as outcome variable and several random effects. The effect on milk yield after anthelmintic treatment over the whole subsequent lactation varied from no effect (-0.43kg/day; P=0.35) to an increase of milk yield with 2.35kg/day (P=0.01), depending on the study region and parity of the cows. Lactation curve analysis suggested that the treatment effect was mainly caused by a slower decay of the milk production in the treated animals compared to untreated animals. The present study highlights the beneficial effect of a topical treatment with moxidectin before parturition on milk yield in the subsequent lactation, as well as the importance of a careful evaluation of nematode exposure risk based on local grazing management practices to guide and target production-based anthelmintic treatment decisions.

Quantification of resistant alleles in the ß-tubulin gene of field strains of gastrointestinal nematodes and their relation with the faecal egg count reduction test.

BACKGROUND: Benzimidazole (BZ) resistance in gastrointestinal nematodes is associated with a single nucleotide polymorphism (SNP) at codons 167, 198 and 200 in the isotype 1 of beta-tubulin gene although in some species these SNPs have also been associated with resistance to macrocyclic lactones. In the present study we compared the levels of resistance in Teladorsagia circumcincta and Trichostrongylus colubriformis by means of the faecal egg reduction test (FECRT) and the percentage of resistant alleles obtained after pyrosequencing. The study was conducted in 10 naturally infected sheep flocks. Each flock was divided into three groups: i) group treated with albendazole (ABZ); ii) group treated with ivermectin (IVM); iii) untreated group. The number of eggs excreted per gram of faeces was estimated at day 0 and 14 post-treatment. RESULTS: Resistance to ABZ was observed in 12.5% (1/8) of the flocks and to IVM in 44.4% (4/9) of them. One flock was resistant to both drugs according to FECRT. Coprocultures were performed at the same dates to collect L3 for DNA extraction from pooled larvae and to determine the resistant allele frequencies by pyrosequencing analysis. In T. circumcincta, SNPs were not found at any of the three codons before treatment; after the administration of ABZ, SNPs were present only in two different flocks, one of them with a frequency of 23.8% at SNP 167, and the other 13.2% % at SNP 198. In relation to T. colubriformis, we found the SNP200 before treatment in 33.3% (3/9) of the flocks with values between 48.5 and 87.8%. After treatment with ABZ and IVM, the prevalence of this SNP increased to 75 and 100% of the flocks, with a mean frequency of 95.1% and 82.6%, respectively. CONCLUSION: The frequencies observed for SNP200 in T. colubriformis indicate that the presence of resistance is more common than revealed by the FECRT.

Condensed Tannins in the Gastrointestinal Tract of Cattle after Sainfoin (Onobrychis viciifolia) Intake and Their Possible Relationship with Anthelmintic Effects.

Condensed tannins' (CTs) fate along the digestive tract of ruminants may account for the variable efficacy of CTs against gastrointestinal nematodes. We analyzed CTs in the digesta of cattle fed sainfoin. With the acetone-butanol-HCl assay, the total CTs concentrations in the digesta were close to those in the diets (6.3 and 1.5% of DM in experiments 1 and 2, respectively); thus, CTs remained potentially largely undegraded/unabsorbed. With the thiolysis assay, CTs concentration was much higher in the abomasum (2.3% of DM; expt 1) compared with the rumen and intestines, along with higher mean size and prodelphinidins percentage, corroborating CTs efficacy reported only against Ostertagia ostertagi in the abomasum. In expt 2, the dietary levels of CTs were probably too low to demonstrate anthelmintic effects in the rumen. Overall, the level of CTs accessible to thiolysis is favored under the acidic conditions of the abomasum, which seems critical for anthelmintic activity.

Modelling the consequences of targeted selective treatment strategies on performance and emergence of anthelmintic resistance amongst grazing calves.

The development of anthelmintic resistance by helminths can be slowed by maintaining refugia on pasture or in untreated hosts. Targeted selective treatments (TST) may achieve this through the treatment only of individuals that would benefit most from anthelmintic, according to certain criteria. However TST consequences on cattle are uncertain, mainly due to difficulties of comparison between alternative strategies. We developed a mathematical model to compare: 1) the most 'beneficial' indicator for treatment selection and 2) the method of selection of calves exposed to Ostertagia ostertagi, i.e. treating a fixed percentage of the population with the lowest (or highest) indicator values versus treating individuals who exceed (or are below) a given indicator threshold. The indicators evaluated were average daily gain (ADG), faecal egg counts (FEC), plasma pepsinogen, combined FEC and plasma pepsinogen, versus random selection of individuals. Treatment success was assessed in terms of benefit per R (BPR), the ratio of average benefit in weight gain to change in frequency of resistance alleles R (relative to an untreated population). The optimal indicator in terms of BPR for fixed percentages of calves treated was plasma pepsinogen and the worst ADG; in the latter case treatment was applied to some individuals who were not in need of treatment. The reverse was found when calves were treated according to threshold criteria, with ADG being the best target indicator for treatment. This was also the most beneficial strategy overall, with a significantly higher BPR value than any other strategy, but its degree of success depended on the chosen threshold of the indicator. The study shows strong support for TST, with all strategies showing improvements on calves treated selectively, compared with whole-herd treatment at 3, 8, 13 weeks post-turnout. The developed model appeared capable of assessing the consequences of other TST strategies on calf populations.

Confirmation of ivermectin resistance in Ostertagia ostertagi in cattle in New Zealand.

Six suspected cases of ivermectin resistance in Ostertagia spp. in cattle were investigated after routine anthelmintic efficacy testing on commercial farms. On four farms a comprehensive faecal egg count reduction test (FECRT) was undertaken using oral formulations of ivermectin (0.2mg/kg), albendazole (10mg/kg) and levamisole (7.5mg/kg) while on two farms only ivermectin was tested. The proportions of Ostertagia spp. in the untreated control and post-treatment larval cultures were used to apportion egg counts to genera and determine efficacy against this genus. Isolates of Ostertagia spp. recovered from three of the farms were each used to infect 18 six month old calves. The efficacy of oral formulations of ivermectin and moxidectin, both at 0.2mg/kg, was determined against each isolate by slaughter and worm count. The efficacy of ivermectin against Ostertagia spp., based on differentiated FECRT for each of the farms varied from 0% to 88%. The efficacy of ivermectin based on worm counts in the slaughter trial varied from 13% to 75% but moxidectin was >99% effective against all isolates. In addition, in the FECRT albendazole, at a dose rate of 10mg/kg, failed to achieve 95% efficacy against Ostertagia spp. on two farms (82% and 85%). Levamisole consistently failed to achieve 95% efficacy against Ostertagia spp. which is consistent with its known lesser efficacy against this parasite. These results confirm the presence of macrocyclic lactone resistant O. ostertagi in cattle in New Zealand and the likely presence of dual resistance, to macrocyclic lactones and albendazole, in some isolates. Resistant populations of this highly pathogenic parasite are probably not uncommon in New Zealand and pose a significant threat to animal production and welfare in the future.

Efficacy of ivermectin against gastrointestinal nematodes of cattle in Denmark evaluated by different methods for analysis of faecal egg count reduction.

The efficacy of ivermectin (IVM) against gastrointestinal nematodes in Danish cattle was assessed by faecal egg count reduction test (FECRT). Six cattle farms with history of clinical parasitism and avermectin use were included. On the day of treatment (Day 0), 20 naturally infected calves per farm (total n = 120) were stratified by initial faecal egg counts (FEC) and randomly allocated to a treatment group dosed with 0.2 mg IVM kg-1 body weight s.c. (IVM; n = 10) or an untreated control group (CTL; n = 10). Individual FEC were obtained at Day 0 and Day 14 post-treatment and pooled faeces by group were cultured to isolate L3 for detection of Ostertagia ostertagi and Cooperia oncophora by qPCR. Treatment efficacies were analysed using the recommended WAAVP method and two open-source statistical procedures based on Bayesian modelling: 'eggCounts' and 'Bayescount'. A simulation study evaluated the performance of the different procedures to correctly identify FEC reduction percentages of simulated bovine FEC data representing the observed real data. In the FECRT, reduced IVM efficacy was detected in three farms by all procedures using data from treated animals only, and in one farm according to the procedures including data from treated and untreated cattle. Post-treatment, O. ostertagi and C. oncophora L3 were detected by qPCR in faeces of treated animals from one and three herds with declared reduced IVM efficacy, respectively. Based on the simulation study, all methods showed a reduced performance when FEC aggregation increased post-treatment and suggested that a treatment group of 10 animals is insufficient for the FECRT in cattle. This is the first report of reduced anthelmintic efficacy in Danish cattle and warrants the implementation of larger surveys. Advantages and caveats regarding the use of Bayesian modelling and the relevance of including untreated cattle in the FECRT are discussed.

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

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

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

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

Anthelmintic resistance in gastrointestinal nematodes of dairy cattle in the Macalister Irrigation District of Victoria.

OBJECTIVE: To report anthelmintic resistance in gastrointestinal nematode parasites of cattle on commercial dairy farms in the Macalister Irrigation District of Gippsland, Victoria. METHODS: Faecal egg count reduction tests (FECRTs) were used to assess anthelmintic resistance on 20 Macalister Irrigation District dairy farms between May 2013 and June 2014. All three currently available anthelmintic classes for cattle nematodes in Australia were tested. Faecal samples were collected 10-14 days post-treatment for individual faecal egg counts (FEC) and larval differentiation. The arithmetic mean FEC for each treatment group was compared with an untreated control post-treatment. Resistance was defined as <95% reduction in FEC, with a lower 95% confidence interval <90% when the mean FEC of the control group, differentiated by genus, was greater than 25 eggs/g. RESULTS: Anthelmintic resistance was present on all 20 dairy farms involved in this study. Resistance to doramectin in at least one species was detected on 15/20 (70%) farms, fenbendazole on 16/20 (80%) farms and levamisole on 5/20 (25%) farms. On three farms, resistance by Ostertagia ostertagi to all three anthelmintic classes was detected. CONCLUSION: This is the first report of anthelmintic-resistant O. ostertagi on Australian dairy farms. Resistance to all three available anthelmintic classes is of concern, given the high pathogenicity of this species. The study highlights the need for veterinarians and dairy farmers to be aware of the risks posed by anthelmintic resistance.