Zoonotic transmission of Teladorsagia circumcincta and Trichostrongylus species in Guilan province, northern Iran: molecular and morphological characterizations.

BACKGROUND: Parasitic trichostrongyloid nematodes have a worldwide distribution in ruminants and frequently have been reported from humans in Middle and Far East, particularly in rural communities with poor personal hygiene and close cohabitation with herbivorous animals. Different species of the genus Trichostrongylus are the most common trichostrongyloids in humans in endemic areas. Also, Ostertagia species are gastrointestinal nematodes that mainly infect cattle, sheep and goats and in rare occasion humans. The aim of the present study was to identify the trichostrongyloid nematodes obtained from a familial infection in Guilan province, northern Iran, using morphological and molecular criteria. METHODS: After anthelmintic treatment, all fecal materials of the patients were collected up to 48 h and male adult worms were isolated. Morphological identification of the adult worms was performed using valid nematode keys. Genomic DNA was extracted from one male worm of each species. PCR amplification of ITS2-rDNA region was carried out, and products were sequenced. Phylogenetic analysis of the nucleotide sequence data was performed using MEGA 6.0 software. RESULTS: Adult worms expelled from the patients were identified as T. colubriformis, T. vitrinus and Teladorsagia circumcincta based on morphological characteristics of the males. Phylogenetic analysis illustrated that each species obtained in current study was placed together with reference sequences submitted to GenBank database. CONCLUSIONS: The finding of current study confirms the zoonotic aspect of Trichostrongylus species and T. circumcincta in inhabitants of Guilan province. The occurrence of natural human infection by T. circumcincta is reported for the first time in Iran and the second time in the world.

Genomic introgression mapping of field-derived multiple-anthelmintic resistance in Teladorsagia circumcincta.

Preventive chemotherapy has long been practiced against nematode parasites of livestock, leading to widespread drug resistance, and is increasingly being adopted for eradication of human parasitic nematodes even though it is similarly likely to lead to drug resistance. Given that the genetic architecture of resistance is poorly understood for any nematode, we have analyzed multidrug resistant Teladorsagia circumcincta, a major parasite of sheep, as a model for analysis of resistance selection. We introgressed a field-derived multiresistant genotype into a partially inbred susceptible genetic background (through repeated backcrossing and drug selection) and performed genome-wide scans in the backcross progeny and drug-selected F2 populations to identify the major genes responsible for the multidrug resistance. We identified variation linking candidate resistance genes to each drug class. Putative mechanisms included target site polymorphism, changes in likely regulatory regions and copy number variation in efflux transporters. This work elucidates the genetic architecture of multiple anthelmintic resistance in a parasitic nematode for the first time and establishes a framework for future studies of anthelmintic resistance in nematode parasites of humans.

Effects of selected Palestinian plants on the in vitro exsheathment of the third stage larvae of gastrointestinal nematodes.

BACKGROUND: Gastrointestinal parasites are one of the main restrictions to small ruminant production. Their pathological importance is primarily related to the major production losses, in quantity or quality, induced by the direct action of worms. Control of these parasites is based exclusively on the frequent use of anthelmintic drugs. However, the resistance to anthelmintics in worm populations after commercialisation of chemical drugs is now widespread. Therefore, there is a need to find new natural resources to ensure sustainable and effective treatment and control of these parasites. The aim of this study was to evaluate the anthelmintic activity, as minimum inhibitory concentration (IC 50 mg/mL), of different plant extracts using larval exsheathment inhibition assay using a two-species but steady population of parasitic nematodes (ca. 20% Teladorsagia circumcinta and 80% Trichostrongylus colubriformis). RESULTS: The study showed that the ethanolic extracts of 22 out of the 48 plant extracts, obtained from 46 plant species, have an inhibitory effect >50% (at concentrations of 100 mg/mL) on the third stage larvae (L3) of the nematodes exhibited the strongest inhibition activity (94%) with IC 50 of 0.02 mg/mL, where other members of the Rhamnaceae family have shown to possess strong anthelmintic activity (70-89%). CONCLUSIONS: Plant extracts are potential rich resources of anthelmintics to combat helminthic diseases. Our results suggest that extracts from Rhamnus elaternus, Epilobium hirsutum, Leucaena leucocephala and Rhamnus palaestinus have promising anthelmintic activity, with potential applications in animal therapeutics and feed.

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.

Modelling the short- and long-term impacts of drenching frequency and targeted selective treatment on the performance of grazing lambs and the emergence of anthelmintic resistance.

Refugia-based treatment strategies aim to prolong anthelmintic efficacy by maintaining a parasite population unexposed to anthelmintics. Targeted selective treatment (TST) achieves this by treating only animals that will benefit most from treatment, using a determinant criterion (DC). We developed a mathematical model to compare various traits proposed as DC, and investigate impacts of TST and drenching frequency on sheep performance and anthelmintic resistance. Short term, decreasing the proportion of animals drenched reduced benefits of anthelmintic treatment, assessed by empty body weight (EBW), but decreased the rate of anthelmintic resistance development; each consecutive drenching had a reduced impact on average EBW and an increased impact on the rate of anthelmintic resistance emergences. The optimal DC was fecal egg count, maintaining the highest average EBW when reducing the proportion of animals drenched. Long-term, reducing the proportion of animals drenched had little impact on total weight gain benefits, across animals and years, whilst reducing drenching frequency increased it. Decreasing the frequency and proportion of animals drenched were both predicted to increase the duration of anthelmintic efficacy but reduce the total number of drenches administered before resistance was observed. TST and frequency of drenching may lead to different benefits in the short versus long term.

Development and application of a delayed-release anthelmintic intra-ruminal bolus system for experimental manipulation of nematode worm burdens.

In order to quantify the impact of parasites on host population dynamics, experimental manipulations that perturb the parasite-host relationship are needed but, logistically, this is difficult for wild hosts. Here, we describe the use of a delayed-release anthelmintic delivery system that can be administered when the hosts can be captured and its activity delayed until a more appropriate period in the host-parasite cycle. Our model system is Svalbard reindeer infected with a nematode parasite, Marshallagia marshalli, which appears to accumulate during the Arctic winter. To determine the extent to which this occurs and the effect on host fitness, reindeer need to be treated with anthelmintics in late autumn but they can only be caught and handled in April. To solve this problem, we devised an intra-ruminal capsule that releases the anthelmintic from up to 6 months after being administered. The capsule was trialed in cannulated sheep and red deer to determine optimum capsule orifice size and release rates. Capsules were estimated to release placebo for 100-153 days followed by abamectin for 22-34 days. To test the efficacy of treatment in reindeer, capsules were administered in April and retrieved in October. All capsules had fully released the anthelmintic and treated reindeer had significantly lower worm burdens than controls. Thus, success of this system allows repeated treatment over several years to test the effect of winter parasitism on host fitness.

Changes in protein expression in the sheep abomasum following trickle infection with Teladorsagia circumcincta.

Continual low-level exposure of sheep to the helminth Teladorsagia circumcincta elicits a temporary protective immunity, where factors in the immune abomasal mucosa prevent penetration of infective larvae, but which is essentially lost within 6 weeks of cessation of parasite challenge. Here, a proteomic approach was used to identify proteins that are differentially regulated in immune compared to naïve sheep, as potential key mediators of immunity. Six naïve sheep and 12 sheep trickle-infected with T. circumcincta were treated with anthelmintic, and the naïve (control) and 6 immune sheep were killed 7 days later. The remaining 6 sheep (immune waning) were killed 42 days after anthelmintic treatment. Abomasal tissue samples were subjected to 2D-gel electrophoresis and densitometric analysis. Selected spots (n=73) were identified by peptide mass fingerprinting and confirmatory Western blotting was carried out for 10 proteins. Spots selectively up-regulated in immune versus control, but not immune waning versus control sheep, included galectin-15 and thioredoxin, which were confirmed by Western blotting. In immune sheep, serum albumin was significantly down-regulated and albumin proteolytic cleavage fragments were increased compared to controls. Unexpectedly, albumin mRNA was relatively highly expressed in control mucosa, down-regulated in immune, and was immunolocalized to mucus-producing epithelial cells. Thus we have identified differential expression of a number of proteins following T. circumcincta trickle infection that may play a role in host protection and inhibition of parasite establishment.

Teladorsagia circumcincta beta tubulin: the presence of the E198L polymorphism on its own is associated with benzimidazole resistance.

BACKGROUND: Benzimidazole resistance is associated with isotype-1 ß-tubulin gene F200Y, E198A and F167Y SNPs. In this study, the recently described polymorphism E198L was reported and analysed in Teladorsagia circumcincta. METHODS: The benzimidazole phenotypic resistance was measured by the faecal egg count reduction test (FECRT) and the egg hatch test (EHT) using a discriminating dose (DD) in 39 sheep flocks. Around 1000 larvae collected before and after treatment were used for DNA extraction. The resistant species identified in all flocks was T. circumcincta. The resistance alleles frequencies were measured for F200Y and E198A. A 371-bp fragment of the isotype-1 ß-tubulin gene was analysed, including the three codons of interest, and a new pyrosequencing assay was designed for testing E198L. RESULTS: The percentage of resistant flocks was 35% by FECRT or 26% by EHT; however, F200Y and E198A SNPs were absent in T. circumcincta. The amplification of a 371-bp fragment confirmed the absence of F167Y and F200Y in 6 resistant flocks. Regarding codon 198, all samples after treatment carried a leucine (CTA). A pyrosequencing assay analysed the allele frequencies for the first two bases at codon 198 independently, G/C and A/T. The correlation between C and T frequencies was almost 1 (r = 0.929, P < 0.0001) and the mean value of both was calculated to measure the leucine frequency; this value ranged between 10.4-80.7% before treatment, and 82.3-92.8% after treatment. High and similar correlations were reported between the genotypic variables (C frequency, T frequency or mean of both frequencies) and phenotypic resistance (r > 0.720, P < 0.0001), although negatively associated with the FECRT and positively with the EHT. According to multivariate linear regression analysis, the T frequency was the most significant variable influencing the phenotypic resistance (FECRT or EHT; P < 0.0001). In the EHT, 67.1% of the phenotypic variability is associated with the T frequency but in the FECRT only 33.4%; therefore, the EHT using a DD seems to detect the genotypic resistance more accurately than the FECRT. CONCLUSIONS: The E198L polymorphism can confer BZ resistance on its own in T. circumcincta.

Ellagitannins Inhibit the Exsheathment of Haemonchus contortus and Trichostrongylus colubriformis Larvae: The Efficiency Increases Together with the Molecular Size.

Worldwide, parasitic gastrointestinal nematodes continue to threaten animal health, welfare, and production in outdoor breeding systems of small ruminants. For more than 50 years, the control of these parasitic worms has relied on the use of commercial synthetic anthelmintics. However, anthelmintic resistance in worm populations is nowadays widespread and requires novel solutions. The use of tannin-rich plants has been suggested as an alternative to synthetic anthelmintics to control gastrointestinal nematodes. The majority of previous studies have focused on the activity of proanthocyanidins (syn condensed tannins), and less is known about ellagitannins. In this study, the effects of 30 structurally unique ellagitannins on the exsheathment of third-stage infective larvae were examined on Haemonchus contortus and Trichostrongylus colubriformis by the in vitro larval exsheathment inhibition assay. Ellagitannins were found to be promising natural anthelmintics as they showed direct inhibition on larval exsheathment for both nematode species. In general, ellagitannins were more efficient at inhibiting the exsheathment of H. contortus larvae than those of T. colubriformis. The efficiency of inhibition increased as the degree of oligomerization or the molecular weight of the ellagitannin increased. Otherwise, we found no other structural features of ellagitannins that significantly affected the anthelmintic activity on the third-stage infective larvae. The effective concentrations were physiologically relevant and should be achievable in the gastrointestinal tract also in in vivo conditions.

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.

Honest sexual signalling mediated by parasite and testosterone effects on oxidative balance.

Extravagant ornaments evolved to advertise their bearers' quality, the honesty of the signal being ensured by the cost paid to produce or maintain it. The oxidation handicap hypothesis (OHH) proposes that a main cost of testosterone-dependent ornamentation is oxidative stress, a condition whereby the production of reactive oxygen and nitrogen species (ROS/RNS) overwhelms the capacity of antioxidant defences. ROS/RNS are unstable, very reactive by-products of normal metabolic processes that can cause extensive damage to key biomolecules (cellular proteins, lipids and DNA). Oxidative stress has been implicated in the aetiology of many diseases and could link ornamentation and genetic variation in fitness-related traits. We tested the OHH in a free-living bird, the red grouse. We show that elevated testosterone enhanced ornamentation and increased circulating antioxidant levels, but caused oxidative damage. Males with smaller ornaments suffered more oxidative damage than those with larger ornaments when forced to increase testosterone levels, consistent with a handicap mechanism. Parasites depleted antioxidant defences, caused oxidative damage and reduced ornament expression. Oxidative damage extent and the ability of males to increase antioxidant defences also explained the impacts of testosterone and parasites on ornamentation within treatment groups. Because oxidative stress is intimately linked to immune function, parasite resistance and fitness, it provides a reliable currency in the trade-off between individual health and ornamentation. The costs induced by oxidative stress can apply to a wide range of signals, which are testosterone-dependent or coloured by pigments with antioxidant properties.

Associations between nematode larval challenge and gastrointestinal tract size that affect carcass productivity in sheep.

Effects of gastrointestinal parasitism on sheep productivity are usually described using live weight change, however carcass productivity is more accurately described using dressing percentage (carcass weight as a proportion of live weight). This experiment had a 2x2x2 factorial design whereby 10-month-old Merino wethers were fed lucerne (Medicago sativa) diets (fresh lucerne or lucerne chaff) with 2 levels of carboxymethycellulose (CMC) inclusion (0% or 8% CMC) and nematode larval challenge (no larval challenge or 10,000 Teladorsagia circumcincta and 10,000 Trichostrongylus colubriformis per week). Sheep were weighed and euthanased 50 or 51 days after larval challenge and CMC supplementation commenced. Weight of the carcass (hot standard carcass weight) and gastrointestinal organs (full and empty) were recorded and expressed as a proportion of live weight. Larval challenged sheep had a worm egg count (mean+/-standard error) of 173+/-38 eggs per gram of faeces and total worm count of 30,237+/-2013 at slaughter. Larval challenged sheep had 1.3% lower dressing percentage (p=0.048), and 2% heavier full (p=0.007) and 1.2% heavier empty gastrointestinal tracts (p=0.012) compared to unchallenged sheep. There was no effect of CMC inclusion or lucerne type (fresh or chaff) on gastrointestinal tract weight or dressing percentage. Larval challenged sheep had 1.1% heavier full (p<0.001) and 0.6% heavier empty (p<0.001) small intestines, and 0.6% heavier full (p=0.005) and 0.3% heavier empty (p=0.026) large intestines compared to unchallenged sheep. Use of live weight change or other measures based on live weight (e.g. feed conversion efficiency) to assess the impact of nematode challenge in sheep may underestimate carcass productivity losses associated with larval challenge in sheep even at moderate levels of larval intake and without overt clinical signs of parasitism. Measurement of carcass weight and/or lean meat yield may better reflect the true economic effects of parasitism in sheep.

Effect of fasting sheep for a short period on the efficacy and safety of monepantel.

Eighteen, six- to seven-month-old lambs were infected experimentally with larvae of Haemonchus contortus, Teladorsagia circumcincta, Trichostrongylus axei, Trichostrongylus colubriformis, Cooperia curticei and Nematodirus spathiger, and allocated to three equal groups. The infections were timed to ensure that fourth-stage larvae were present when groups 1 and 2 were treated orally with monepantel. Group 1 was not fed for 24 hours before the treatment, group 2 was fed two hours before the treatment and group 3 was fed at the same time as group 2 but not treated with monepantel. All the sheep had access to water. Worm burdens were determined 15 days after the treatments. Fasting or feeding had no statistically significant effects on the efficacy of the monepantel solution against the nematodes, and the period of fasting had no adverse effects.

Long-acting moxidectin for the control of trichostrongylid infections of sheep in south-eastern Australia.

OBJECTIVE: To compare the effectiveness of a single long-acting injection of moxidectin, given in either summer or winter, with a two-summer ('standard') treatment strategy for controlling gastrointestinal nematodes in spring-lambing Merino ewes. DESIGN: A replicated study over 2 years in the high winter rainfall environment of western Victoria. METHODS: Measures of worm infection were worm egg counts of ewes and total worm counts of tracer lambs. Measures of production were body weight, proportion of pregnant ewes and number of weaned lambs. RESULTS: The egg counts of ewes given the standard strategy followed a characteristic pattern, rising to 300 to 600 eggs per gram before the second summer treatment in February. During this time, ewes given long-acting moxidectin in December had zero or negligible counts. There were no consistent differences between tracer worm counts or production measures for the two groups treated in summer, but when data from both years were pooled, the total egg output from December to May was significantly lower for ewes treated with long-acting moxidectin in December. Ewes not treated during summer had lower body weights, and higher worm egg counts and tracer worm counts, showing that this was not a suitable strategy at that time of the year. However, following treatment with long-acting moxidectin in winter, ewes had rapid compensatory weight gain and lower egg output than ewes treated in summer. CONCLUSION: A single injection of long-acting moxidectin in December is an effective treatment strategy for Merino ewes lambing in spring.

Genotypic characterisation of monepantel resistance in historical and newly derived field strains of Teladorsagia circumcincta.

Recent reports of monepantel (MPTL) resistance in UK field isolates of Teladorsagia circumcincta has highlighted the need for a better understanding of the mechanism of MPTL-resistance in order to preserve its anthelmintic efficacy in this economically important species. Nine discrete populations of T. circumcincta were genotypically characterised; three MPTL-susceptible isolates, three experimentally selected MPTL-resistant strains and three field derived populations. Full-length Tci-mptl-1 gene sequences were generated and comparisons between the MPTL-susceptible isolates, MPTL-resistant strains and one field isolate, showed that different putative MPTL-resistance conferring mutations were present in different resistant isolates. Truncated forms of the Tci-mptl-1 gene were also observed. The genetic variability of individual larvae, within and between populations, was examined using microsatellite analyses at 10 'neutral' loci (presumed to be unaffected by MPTL). Results confirmed that there was little background genetic variation between the populations, global FST <0.038. Polymorphisms present in exons 7 and 8 of Tci-mptl-1 enabled genotyping of individual larvae. A reduction in the number of genotypes was observed in all MPTL-resistant strains compared to the MPTL-susceptible strains that they were derived from, suggesting there was purifying selection at Tci-mptl-1 as a result of MPTL-treatment. The potential link between benzimidazole (BZ)-resistance and MPTL-resistance was examined by screening individual larvae for the presence of three SNPs associated with BZ-resistance in the ß-tubulin isotype-1 gene. The majority of larvae were BZ-susceptible homozygotes at positions 167 and 198. Increased heterozygosity at position 200 was observed in the MPTL-resistant strains compared to their respective MPTL-susceptible population. There was no decrease in the occurrence of BZ-resistant genotypes in larvae from each population. These differences, in light of the purifying selection at this locus in all MPTL-resistant isolates, suggests that Tci-mptl-1 confers MPTL-resistance in T. circumcincta, as in Haemonchus contortus, but that different mutations in Tci-mptl-1 can confer resistance in different populations.

Field evaluation for anthelmintic-resistant ovine gastrointestinal nematodes by in vitro and in vivo assays.

A coprological survey to analyze the presence of flock resistance to benzimidazoles (BZ) and macrocyclic lactones (ML) was performed in sheep under field conditions. Fecal samples were collected from 2,625 sheep in 72 commercial farms from Galicia (NW Spain). The in vitro (FECRT, fecal egg count reduction test) and in vivo (EHA, egg hatch assay, and LFIA, larval feeding inhibition assay) tests were used to assess the efficacy of these anthelmintics. Coprocultures were also developed to obtain knowledge on the main genera of trichostrongylid nematoda prior to, and after, the administration of the anthelmintics. By using the FECRT, BZ resistance was observed in 13 (18%) flocks, whereas ML resistance was only detected in 2 (3%) farms. The number of resistant flocks to BZ was 21 (29%) by using the EHA and 7 (10%) by means of the LFIA. None of the flocks used in this study showed simultaneous resistance to both employed anthelmintics. The results from the in vitro and in vivo tests revealed that 92% of the flocks FECRT resistant to BZ were also resistant with the EHA. The LFIA confirmed all the farms resistant to ML by using the in vivo test. After the administration of BZ, nematode larvae belonging to Teladorsagia circumcincta (32.2%), Trichostrongylus spp. (29%), Nematodirus spp. (6.5%), and Chabertia ovis (3.2%) were identified. In the flocks receiving ML, only T. circumcincta was identified (57%). We recommend the use of in vitro tests because they are more efficient. As the use of macrocyclic lactones is increasing in this region, further investigation is needed for detecting resistance to the anthelmintic family compounds by the LFIA.

Establishment of Cooperia oncophora in calves.

The establishment rate of Cooperia oncophora related to host age and previous infection was investigated in young calves. Calves of similar age were kept on a feed pad and allocated into multiple groups, based on their age and weight. Two groups (each n = 16) received trickle infections with an ivermectin-susceptible C. oncophora isolate of 2000 or 10,000 infective stage larvae per week while another group (n = 16) was kept as an uninfected control. At intervals over a period of 11 months, two animals from each group were challenged with 15,000 infective stage larvae of an ivermectin-resistant isolate, 25 days later orally treated with ivermectin and 5 days after that slaughtered for worm counts. On three occasions additional calves (n = 2), subjected to the high trickle infection rate, received an ivermectin treatment to remove the existing worm burden, prior to challenge as above. Further calves (n = 4) of similar age were introduced at the beginning and the end of the experiment to determine the effect of larval age on establishment rate. The establishment in the two trickle infection groups declined to <10% within the first three months, which was significantly different from the control group. In the animals receiving the high trickle infection, but an anthelmintic treatment before challenge the establishment rate was not significantly different from the controls. Over the duration of the experiment establishment in the control group declined from 53% to <20%, which was similar to the decrease recorded at the beginning and the end of the experiment in the animals to determine the effect of larval age. The findings indicate that an existing C. oncophora burden had a strong effect on the establishment of incoming larvae in the trickle infected groups, but this was not observed if the existing burden was removed before the final challenge. The decline in establishment rate in the control group was attributed to the age of the larvae and not the age of the calves per se.

Combination deworming for the control of double-resistant cyathostomin parasites - short and long term consequences.

Equine cyathostomin are pervasive gastrointestinal parasites with wide-spread resistance to the benzimidazole and tetrahydropyrimidine drug classes worldwide. Combination deworming has been proposed as a more sustainable parasite control strategy. Simulation studies have found combination deworming to be effective in controlling drug resistant ovine trichostrongylid parasites. One equine study demonstrated an additive effect of a combination of oxibendazole and pyrantel pamoate against cyathostomins. However, this is the only equine study evaluating combination therapy, and the effects of repeated combination treatments administered over time remain unknown. The purpose of this study was to observe the efficacy of repeated oxibendazole/pyrantel pamoate combination therapy administered over one year against a cyathostomin population with resistance to benzimidazole and pyrantel products. Fecal egg counts were determined for the entire herd (N = 21) at the day of anthelmintic treatment and at two-week intervals for eight weeks post treatment. Starting efficacies of oxibendazole (OBZ, 10 mg/kg) and pyrantel pamoate (PYR, 6.6 mg base/kg) were 66.7% and 63.3%, respectively. Hereafter, the herd was treated four times with an oxibendazole/pyrantel pamoate combination, eight weeks apart, followed by repeating the single active treatments before concluding the study. While the first combination treatment exhibited an additive effect of the two active ingredients, this efficacy was not sustained over the course of the study. Mean fecal egg count reduction (FECR) was significantly greater for the first combination treatment (76.6%) than the second (42.6%, p = 0.0454), third (41.6%, p = 0.0318), and fourth (40.7%, p = 0.0372) combination treatments. The final single active mean FECRs were 42.3% for oxibendazole, and 42.7% for pyrantel pamoate. These efficacies were not significantly different from the initial single active efficacies (OBZ, p = 0.4421; PYR, p = 0.8361). These results suggest that combination therapy against double resistant equine cyathostomin populations is not sustainable, when using actives with markedly decreased starting efficacies.

Effect of Gliricidia sepium leaves intake on larval establishment of Cooperia punctata in calves and bio-guided fractionation of bioactive molecules.

The aims of this study were: 1) to assess the anthelmintic effect of Gliricidia sepium on the establishment of C. punctata third-stage larvae (L3) in calves, and 2) to isolate and to elucidate an anti-exsheathment phytochemical from the plant offered during the trial. Twelve ¾ Holstein × Zebu calves were divided in two experimental groups: control (T1) and treatment (T2) (n = 6). After adaptation, each calf was infected with an oral dose of 400 C. punctata L3/Kg LW. Basal diet consisted of Digitaria decumbens hay (6.27% CP) and commercial concentrate (12% CP). In addition, during the experimental period T2 received fresh G. sepium leaves (26.88% CP) ad libitum. On day 9 post-infection, three calves per treatment were randomly selected for slaughter, and worm counts were performed. Larval establishment rates obtained were 13.44 ±â€¯0.13% and 3.1 ±â€¯1.42% for T1 and T2, respectively (P < .05). The reduction of larval establishment was 76.9%. The total length of worms recovered from the animals was also affected by the intake of G. sepium (P < .05). Phytochemicals present in G. sepium leaves offered to calves were isolated through silica gel columns and elucidated through Magnetic Nuclear Resonance (1H and 13C). Bio-guided isolation procedures lead to the elucidation of Oxytroside (Kaempferol 3-O-rhamnopyranosyl-(1 → 6)-ß-D-glucopyranoside-7-O-rhamnopyranoside), which fully inhibited the C. punctata exsheathment process (2400 µg mL-1). Gliricidia sepium represents an alternative to prevent severe C. punctata infections by reducing larval establishment in cattle.

The effect on liveweight gain of using anthelmintics with incomplete efficacy against resistant Cooperia oncophora in cattle.

A replicated field trial was conducted to measure the effect on liveweight gain of failing to adequately control anthelmintic resistant populations of Cooperia oncophora and to determine whether populations, and hence production losses, increased with time. Eight mobs of 10 Friesian-Hereford calves were run on independent farmlets from January to December, over each of two years. All mobs were routinely treated with a pour-on formulation of eprinomectin every six weeks, which controlled parasites other than Cooperia. Four mobs also received six weekly treatments with an oral levamisole plus albendazole combination anthelmintic to control Cooperia. Liveweights, condition scores, faecal egg counts and larval numbers on pasture were measured throughout. In the first year animals treated with eprinomectin alone were 12.9 kg lighter in November than those treated with eprinomectin plus albendazole and levamisole, however, in the second year there was no difference between the treatment groups. The data, therefore, support the view that while C. oncophora is less pathogenic than other cattle parasite species it can still cause production losses when present in sufficient numbers. In the first year of the study, parasite load, as measured by faecal nematode egg count and larval numbers on herbage, tended to be higher and calf growth rates lower than in the second year. In both years, counts of infective larvae on herbage declined over winter-spring to be at low levels before mid-summer. This suggests that the carry-over of infection from one crop of calves to the next was relatively small and hence that the level of challenge to the young calves at the start of each year was largely due to the effectiveness of the quarantine treatments administered when the animals arrived on the trial site. Low survival of larvae on pasture between grazing seasons, resulting in small larval populations on pasture when drenching programmes start each summer, might help to explain the widespread development of anthelmintic resistance in this parasite under New Zealand grazing systems.