Development of a broad-spectrum monoclonal antibody-based indirect competitive enzyme-linked immunosorbent assay for the multi-residue detection of avermectins in edible animal tissues and milk.

Avermectins (AVMs) are a group of anti-parasitic agents that have been widely used in food-producing animals. To monitor the residue of the AVMs, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was developed with a simple sample preparation procedure. Conjugates of 4″-HS-IVM/AVM on three different proteins were used to raise a broad-spectrum monoclonal antibody (mAb), 6D4, that had IC50 values for avermectin, ivermectin, eprinomectin and emamectin of 7.2, 10.4, 19.8 and 20.8 µg L-1, respectively. The limit of detection and limit of quantitation of this method for AVMs in various matrix samples ranged from 0.5 to 5.4 µg L-1 and 1.0 to 10.3 µg L-1, respectively. The recoveries of the samples spiked with AVMs were in the range of 78.1-110.5% with coefficients of variation below 14%. The ic-ELISA was applied to monitor ivermectin in the milk samples, and the results showed a good correlation with that of HPLC-MS/MS.

Efficacy of pyrantel pamoate and ivermectin paste formulations against naturally acquired Oxyuris equi infections in horses.

In recent years, numerous veterinary practitioners have reported anecdotal episodes in which anthelmintic treatment did not appear to deliver the expected efficacy against equine pinworms (Oxyuris equi). Anthelmintic resistance has not been demonstrated formally in equine pinworms, so a clinical study was designed to evaluate the efficacy of paste formulations of pyrantel pamoate or ivermectin against naturally acquired infections with O. equi. Twenty-one horses (>4 months to 15 years of age) with patent, naturally acquired pinworm infections were blocked by source of origin and allocated randomly to one of three treatment groups: horses (n=7) assigned to Group 1 were treated orally with pyrantel pamoate paste at a dosage of 13.2 mg/kg (2x label dosage), Group 2 horses (n=7) were untreated controls, and horses (n=7) assigned to Group 3 were treated orally with ivermectin paste at a dosage of 200 microg/kg. Fourteen days after treatment, horses were euthanatized, necropsied, and large intestinal contents were processed for recovery of adult pinworms. In addition, duplicate 1% aliquots of intestinal contents from the cecum, ventral colon, dorsal colon, and small colon were collected, preserved, and examined for recovery and enumeration of fourth-stage larval O. equi. Anthelmintic efficacy against pinworms was evaluated by comparing the post-treatment worm counts of Groups 1 and 3 to those of control animals. Mean numbers of O. equi adults recovered postmortem were significantly decreased by both pyrantel pamoate (P=0.0366) and ivermectin (P=0.0137) treatment, with respective efficacies of 91.2% and 96.0%. In addition, both products demonstrated >99% efficacy against fourth-stage O. equi larvae. The current study demonstrated acceptable adulticidal and larvicidal efficacy of both pyrantel pamoate and ivermectin paste formulations against O. equi and did not support the existence of macrocyclic lactone or pyrimidine resistance in the pinworm populations evaluated.

Lethal and sublethal toxic effects of a test chemical (ivermectin) on the yellow dung fly (Scathophaga stercoraria) based on a standardized international ring test.

A standardized bioassay using the yellow dung fly, Scathophaga stercoraria L. (Diptera: Scathophagidae), was developed to test the lethal and sublethal toxicity of parasiticide residues in livestock dung. The repeatability of the bioassay was assessed for the parasiticide ivermectin in 13 tests performed by seven laboratories in Germany, the United Kingdom, Switzerland, and Canada. Test results had an acceptable range of heterogeneity. The calculated median effective concentration for 50% (EC50) egg-to-adult mortality was 20.9 +/- 19.1 microg ivermectin/kg dung fresh weight (FW) (mean +/- standard deviation; range, 6.33-67.5 microg/kg). Mortality was not observed below a calculated no-observable-effect concentration (NOEC) of 8.1 +/- 7.7 microg/kg FW. However, prolonged development time (and, in a subset of tests, reduced body size) was observed above a calculated NOEC of 0.8 +/- 0.8 microg/kg FW. An oviposition site choice test revealed that yellow dung fly females do not discriminate among dung of different ivermectin concentrations. Thus, the yellow dung fly is suitably sensitive, and the methods are sufficiently repeatable, to support use of this standardized bioassay by the international community in the registration of new veterinary pharmaceuticals.

Parascaris equorum in foals and in their environment on a Swedish stud farm, with notes on treatment failure of ivermectin.

Environmental contamination and the egg excretion pattern of the ascarid Parascaris equorum (Nematoda) was investigated in relation to anthelmintic treatment on a Swedish stud farm. Faecal samples from 15 foals, dewormed every 8th-week with a paste formulation of ivermectin at the standard dose rate of 0.2 mg/kg bodyweight, were collected at five sampling occasions between August and November 2006. In addition, soil samples were obtained from four paddocks used by these foals in November 2006. The number of eggs per gram (epg) was counted in both faeces and soil. Egg excretion started when the foals were 3-4 months, and reached the highest levels when they were approximately 5-month-old, and was then followed by a decline. Egg excretion seemed to be unaffected by ivermectin despite these foals were dewormed at regular intervals. In four out of five foals examined 10 days after treatment, epg actually increased. In contrast, when either fenbendazol or pyrantel embonate were used instead of ivermectin, treatments were effective. The number of eggs in soil was significantly higher in the permanent paddock compared to in the temporarily used soil paddock and in the summer paddocks.

Efficacy of a long-acting formulation of ivermectin against Psoroptes ovis (Hering, 1838) on cattle.

A study was conducted in cattle experimentally infested with Psoroptes ovis to compare the prophylactic control against P. ovis provided by a long-acting injectable formulation of ivermectin to that of a commercially available injectable formulation of doramectin. Thirty Holstein steers were used. Animals were allocated by restricted randomization based on Day 0 body weight, forming six replicates of five animals each. Within each replicate, one animal was randomly allocated to one of the following treatment groups, with ivermectin and doramectin administered subcutaneously where indicated: (1) untreated controls; (2) ivermectin long-acting injectable (LAI) 630 mcg/kg, 56 days before challenge; (3) ivermectin LAI 630 mcg/kg, 42 days before challenge; (4) ivermectin LAI 630 mcg/kg, 35 days before challenge; or (5) doramectin 200 mcg/kg, 35 days before challenge. Animals were housed in individual pens 1 week prior to treatment. All animals were experimentally infested with P. ovis mites in the area between the shoulders, on the same day. Live mites were counted in scrapings from mange lesions at 2 sites on each animal 14, 21 and 28 days after challenge. Live mites were found in 33, 67 and 83% of the untreated controls on each respective evaluation. No P. ovis mites were found in steers treated with ivermectin LAI. Those animals showed lower (P < 0.05) mite counts than untreated controls on evaluations conducted 21 and 28 days after challenge. These results indicate that the ivermectin long-acting injectable formulation prevents induced infestations by P. ovis for at least 56 days after treatment. Doramectin injectable formulation, used at 200 mcg/kg, did not have a prophylactic effect 35 days after treatment.

Improved method for determination of abamectin and ivermectin in cattle plasma.

A liquid chromatographic (LC) method was developed for determination of abamectin (ABM) and ivermectin (IVM) in cattle plasma. The sample was extracted with acetonitrile and cleaned up on an alumina column. After conversion to stable fluorescent derivative with trifluoroacetic anhydride and N-methylimidazole, the sample was analyzed by LC with fluorescence detection (Ex 365 nm and Em 475 nm). Doramectin was used as an internal standard. Recoveries ranged from 91.2 to 100.7% for IVM and from 87.0 to 98.7% for ABM, with 1-50 ng/mL fortified samples. The coefficients of variation were <10.1%. The limit of detection was 0.02 ng/mL for ABM and IVM in 1.0 mL samples.

Evaluation of efficacy of selamectin and fipronil against Ctenocephalides felis in cats.

OBJECTIVE: To evaluate efficacy of monthly administration of selamectin and fipronil against Ctenocephalides felis in cats. DESIGN: Randomized controlled trial. ANIMALS: 36 healthy cats. PROCEDURE: Cats known to be free of fleas were infested with 100 unfed adult fleas on days -28 and -21. On days 0, 30, 60, 90, and 120, sixteen cats (8 pairs/treatment group) were treated by topical administration of selamectin (6 mg/kg [2.7 mg/lb] of body weight) or fipronil (7.5 mg/kg [3.4 mg/lb]). Four control cats (2 pairs) were not treated. On day -6 and every 2 weeks after initial treatment, comb counts were performed to detect fleas. Flea counts were recorded, and fleas (< or =50) that had been removed were replaced onto the cat. On day 89, fleas were not replaced. On day 91 and every 7 days until the end of the study (day 150), cats were challenged with 20 adult fleas. Flea counts were compared between and within treatments. RESULTS: 14 days after treatment, geometric mean flea counts were reduced by 71.2% by fipronil treatment and 35.3% by selamectin treatment. Both treatments resulted in 97 to 98% reduction in flea counts on day 29 and 99.8 to 100% reduction from day 44 to the end of the study. CONCLUSIONS AND CLINICAL RELEVANCE: Selamectin is as effective as fipronil in treating infestation in cats housed for 3 months in a flea-infested environment under conditions known to support the flea life cycle and in protecting against subsequent weekly challenges with C felis for an additional 2 months.

Evaluation of efficacy of selamectin, fipronil, and imidacloprid against Ctenocephalides felis in dogs.

OBJECTIVE: To evaluate efficacy of monthly administration of selamectin, fipronil, and imidacloprid against Ctenocephalides felis in dogs. DESIGN: Randomized controlled trial. ANIMALS: 44 healthy dogs. PROCEDURE: Dogs known to be free of fleas were infested with 100 unfed adult fleas on days -28 and -21. On days 0, 30, 60, 90, and 120, dogs (12/group) were treated by topical administration of selamectin (6 mg/kg [2.7 mg/lb] of body weight), fipronil (7.5 mg/kg [3.4 mg/lb]), or imidacloprid (10 mg/kg [4.5 mg/lb]); 8 untreated dogs were used as controls. On day -6 and every 2 weeks after initial treatment, comb counts of viable adult fleas were made, and fleas (< or =50/dog) were replaced onto the dog from which they were removed. On day 89, fleas were not replaced. On day 91 and every 7 days until the end of the study, dogs were challenged with 20 adult fleas. RESULTS: 14 days after initial treatment, geometric mean flea counts were reduced by 97.5 to 99.1 % for all treatments, compared with pretreatment counts on day -6. Selamectin, fipronil, and imidacloprid reduced geometric mean flea counts by 99.7 to 100% from day 29 to the end of the study. CONCLUSIONS AND CLINICAL RELEVANCE: Selamectin is as effective as fipronil and imidacloprid in reducing C felis infestation in dogs housed for 3 months in a flea-infested environment under conditions known to support the flea life cycle, and in protecting against subsequent weekly challenges with C felis for an additional 2 months.

Factors affecting adverse event reporting during mass ivermectin treatment for onchocerciasis.

This study examines the effect of age, sex, dosing round, time of day, and distance from the nurse monitor on adverse event reporting during mass ivermectin administration at Achi, south-east Nigeria. There was a significant increase in adverse event reporting with age but no significant difference between the sexes. Both compliance and the incidence of reported adverse events were less during the second dosing round than during the first. Only three (2%) out of 144 subjects interviewed gave adverse events as a reason for non-compliance. Significantly fewer adverse events were reported as starting at night (22:00-06:00 h) than by day. The effect of distance on adverse event reporting was not statistically significant for distances up to 1 km.

Efficacy of doramectin 0.5% w/v Pour-On for control of the horn fly, Haematobia irritans.

In 1998, three groups of cattle at three locations in Lousiana were treated with Dectomax((R)) (0.5% doramectin) Pour-On and horn fly populations were monitored. Acceptable levels (less than 50 flies per side) of horn fly control were observed from 4 to 8 weeks. Differences in the length of control among the three sites were most likely affected by immigration of adult flies from untreated groups. In 1999, acceptable horn fly control was obtained for 13 weeks by the use of two treatments of doramectin Pour-On.

Efficacy of an ivermectin controlled-release capsule against some rarer nematode parasites of sheep.

A controlled trial was conducted to evaluate the efficacy of the intraruminal ivermectin controlled-release capsule (CRC) (IVOMEC Maximizer CR Capsule for Sheep, Merial Ltd.) against induced incoming third-stage larvae and established adult infections with some rarer gastrointestinal nematode parasites of sheep. Twenty-one worm-free lambs were allocated by restricted randomisation based on body weight within sex to one of the following treatments: unmedicated control, ivermectin CRC given on Day 0 prior to induced infection, and ivermectin CRC given on Day 70 after establishment of induced infection. The ivermectin CRC delivers ivermectin at a minimum dose rate of 20 microg/kg/day for 100 days. Infections were induced by daily administration of third-stage larvae for five consecutive days. Nematodes were counted on Day 84, 14 days after treatment of established infection. The treatment with the ivermectin CRC prevented the establishment of Ostertagia leptospicularis, O. ostertagi, Bunostomum trigonocephalum, Cooperia oncophora, C. punctata, C. surnabada, Nematodirus helvetianus, N. roscidus and Strongyloides papillosus by >99% as compared with the untreated controls (p < 0.01). The administration of the ivermectin CRC reduced established adult infections of O. ostertagi, B. trigonocephalum, C. oncophora, C. punctata, C. surnabada, N. roscidus and S. papillosus by >99% (p < 0.01), and reduced established adult infections of O. leptospicularis and N. helvetianus by 96.5 and 98.4% (p < 0.01), respectively.

The effect of simulated rainfall on the efficacy of doramectin pour-on against nematode parasites of cattle.

Two studies were conducted with doramectin topically administered at 500 microg/kg body weight to assess retention of therapeutic efficacy against nematode infections of cattle before, and after, simulated rainfall. In the first study, 50 heifers, with patent nematode infections, were allocated to one of five treatment groups. An untreated control group and one doramectin-treated group were not exposed to simulated rainfall. Simulated rainfall was applied at a rate of 25.4 mm of water in 35 min to three of the five groups: one group immediately before treatment, the second group 90 min after treatment, and the third group 24 h after treatment. Fecal samples were collected for determining egg counts 14 days after treatment. Percentage efficacy ranged from 97.3% to 100% in all treated calves, regardless of exposure to simulated rainfall. The second study involved 40 mixed-sex cattle that were allocated to one of four treatment groups (one saline control and three doramectin-treated groups). All cattle were confirmed to be free of nematode infections prior to oral gavage with infective larvae of Dictyocaulus viviparus, Cooperia oncophora, and Ostertagia ostertagi. Twenty-six days after infection, three groups were treated with doramectin pour-on and exposed to 20 mm of simulated rainfall over 40 min: one group 60 min before treatment, the second 20 min after treatment, and the third 40 min after treatment. Approximately two weeks after treatment, all cattle were necropsied for worm counts. In all treated groups, the percentage efficacy against O. ostertagi and D. viviparus was >99% to 100%. Percentage efficacy against Cooperia ranged from 97% to 98%. Results indicated that doramectin pour-on remains efficacious against nematodes of cattle when administered immediately before or after rainfall.

Persistent activity of doramectin injectable formulation against experimental challenge with Haemonchus placei in cattle.

Two studies were conducted in North America to evaluate the persistent activity of doramectin injectable formulation against experimental challenge with Haemonchus placei. In both studies, calves were randomly assigned to 1 of 4 treatment groups (n = 10 per group) or a larval viability group (n = 2). Calves were treated subcutaneously in the lateral midline of the neck with saline (1 ml/50 kg) on Day 0, or with doramectin (200 mg/kg = 1 ml/50 kg) on Day 0, 7, or 14. Animals used to assess larval viability did not receive any treatment. Beginning on Day 14 and continuing through Day 28, each of the 40 treated calves were given approximately 300 infective larvae of H. placei per os. The two larval viability animals received approximately 10,000 larvae as a single dose on Day 28. Approximately two weeks later, all animals were slaughtered and the abomasum from each calf processed for nematode recovery. A 2% aliquot of abomasal contents plus wash was examined for enumeration and identification of nematodes. Geometric mean H. placei counts were calculated from the log (H. placei count +1) and used to estimate percentage reduction. Overall, doramectin was > or =96.9% efficacious in reducing infection with H. placei when challenged daily 14-28 days after treatment.

Persistent efficacy of doramectin injectable against artificially induced infections with Cooperia punctata and Dictyocaulus viviparus in cattle.

Three studies were conducted to evaluate the persistent efficacy of doramectin injectable solution against experimental challenges with infective larvae of Cooperia punctata and Dictyocaulus viviparus. In each study, four groups of ten randomly-assigned calves, negative for trichostrongyle-type eggs on fecal examination, were treated subcutaneously in the midline of the neck with saline (1 ml/50 kg) on Day 0 or doramectin (200 microg/kg = 1 ml/50 kg) on Day 0, 7, or 14. Two additional calves from the same pool of animals were randomly assigned as larval-viability monitors and received no treatment. On Days 14-28, approximately 1000 and 50 infective larvae of Cooperia spp. and D. viviparus, respectively, were administered daily by gavage to each animal in Groups T1-T4. On Day 28, the two larval-viability monitor calves were inoculated in a similar manner with a single dose of approximately 30000 and 2000 larvae of Cooperia spp. and D. viviparus, respectively. Equal numbers of calves from each treatment group were killed on Days 42-45, as well as the two viability monitor animals to enumerate worm numbers. A 2% or 5% aliquot of small intestinal contents and washings were examined for worm quantification and identification, while 100% of the lung recoveries were quantified and identified. For each study and across the three studies, geometric mean worm recoveries for each treatment group were calculated from the natural log transformed data (worm count + 1) and were used to estimate percentage reduction. In the three studies, doramectin injectable solution was 97.5% efficacious against lungworms for up to 28 days and was 99.8% efficacious in reducing infection resulting from challenge with infective larvae of C. punctata for at least 28 days post-treatment.

A review of the use of a controlled-release formulation of ivermectin in the treatment and prophylaxis of Psoroptes ovis infestations in sheep.

The options for the treatment and control of sheep scab (psoroptic mange) have been increased in recent years through the introduction of the endectocides ivermectin, doramectin and moxidectin. Whilst therapeutic efficacy is good, the current injectable formulations offer limited protection against re-infestation with Psoroptes ovis. An intraruminal controlled-release formulation of ivermectin has been developed to provide therapeutic and prophylactic activity against a range of sensitive endo- and ecto-parasites of sheep for 100 days after administration. These ivermectin boluses are designed to release ivermectin at 20-40 microg/kg/day over 100 days and were developed for use in sheep of 20-90 kg bodyweight. Several controlled therapeutic and prophylactic trials against sheep scab have been conducted under a variety of protocols with such boluses in Europe and South America. The results of these studies indicate that the bolus provides 100% therapeutic efficacy against established P. ovis infestations and equivalent prophylactic efficacy against challenge infestations administered during the active life of the bolus.

Persistent efficacy of doramectin pour-on against artificially induced infections of nematodes in cattle.

Two studies were conducted to determine the persistent efficacy of doramectin pour-on against an artificial, trickle challenge of mixed nematodes in calves. In each study, 42, 4-8 months old calves were randomly assigned into four groups of 10 animals each (T1-T4), plus two larval-viability monitor animals. All animals were treated with fenbendazole (10 mg kg(-1)) 14 days prior to the start of the study to clear any existing infection. Doramectin pour-on at 500 microg kg(-1) was used on each animal in Groups T2, T3, and T4 with intervals of 1 week (Day 0, 7, and 14, respectively). Calves in Group T1 were treated with saline solution on Day 0 and at the same volumetric rate (1 ml 10 kg(-1)) as the doramectin treated animals. All treatments were applied in a single passage along the midline of the back, from the withers to the tailhead. Subsequently, trickle inoculations with infective larvae were administered to all calves for 22 consecutive days (Days 14-35). Doramectin pour-on provided > or = 91.9% efficacy against challenge with Dictyocaulus viviparus, Haemonchus spp., and Ostertagia ostertagi for up to 35 days post-treatment and against challenge with Cooperia oncophora, Cooperia punctata, and Oesophagostomum radiatum for up to 28 days post-treatment.