An attempt to replace an ivermectin-resistant Cooperia spp. population by a susceptible one on grazing pastures based on epidemiological principles and refugia management.

The maintenance of anthelmintic-susceptible parasite refugia to delay the onset of anthelmintic resistance is an almost impossible effort in many grazing livestock production countries given that current refugia consist of already resistant parasites. Rather, efforts could be focused on replacing the resistant parasite refugia by susceptible parasite ones and implementing sustainable parasite control measures from then on. To this purpose, a trial was conducted to attempt to establish a new population of ivermectin-susceptible Cooperia sp. on a beef cattle farm with proven problems of ivermectin-resistant Cooperia. During two consecutive years, 82 (Year 1) and 100 (Year 2) recently weaned and parasite-free heifers were inoculated with 40,000 or 30,000 susceptible Cooperia L3, respectively, at a time when levels of resistant parasite refugia were normally low. The animals were subsequently allowed to graze on the problem pastures during autumn until the end of spring. Levels of parasitism in the animals and on pasture were monitored monthly and animals were treated with levamisole when needed. The combination of parasitological monitoring and local epidemiological knowledge was essential to determine when treatments were to be administered. No clinical signs of gastrointestinal parasitosis in the herd were observed throughout the study and unnecessary treatments were avoided. Faecal egg counts reduction tests (FECRT) and controlled efficacy tests (CET) employing worm counts were carried out at different times throughout the study to determine the clinical efficacy (FECRT) and the absolute efficacy (CET) of ivermectin, respectively. The clinical efficacy of ivermectin increased from an initial 73% to 99.4%, while the absolute efficacy increased from 54.1% to 87.5% after just two animal production cycles. The switch from a resistant parasite population to a susceptible one requires knowledge of parasitological epidemiology, especially in relation to seasonal variations of parasite populations in both the host and in refugia.

Observations on the free-living stages of cattle gastrointestinal nematodes.

A 4-year study on the free-living stages of cattle gastrointestinal nematodes was conducted to determine (a) the development time from egg to infective larvae (L3) inside the faecal pats, (b) the pasture infectivity levels over time, and (c) the survival of L3 on pasture. Naturally infected calves were allowed to contaminate 16 plots on monthly basis. Weekly monitoring of eggs per gram of faeces (epg) values and faecal cultures from these animals provided data for the contamination patterns and the relative nematode population composition. At the same time, faecal pats were shaped and deposited monthly onto herbage and sampled weekly to determine the development time from egg to L3. Herbage samples were collected fortnightly over a 16-month period after deposition to evaluate the pasture larval infectivity and survival of L3 over time. The development time from egg to L3 was 1-2 weeks in summer, 3-5 weeks in autumn, 4-6 weeks in winter, and 1-4 weeks in spring. The levels of contamination and pasture infectivity showed a clear seasonality during autumn-winter and spring, whilst a high mortality of larvae on pasture occurred in summer. Ostertagia spp., Cooperia spp. and Trichostrongylus spp. were predominant and a survival of L3 on pasture over a 1-year period was recorded in this study.

Environmental impact of ivermectin excreted by cattle treated in autumn on dung fauna and degradation of faeces on pasture.

The effect of ivermectin excreted in faeces of treated cattle on dung fauna and dung degradation on pasture during autumn was evaluated. Two groups of calves were used. One group was treated subcutaneously with ivermectin while the other remained as untreated control. Faeces deposited on 1, 3, 7, 14 and 21 days post-treatment (dpt) were removed on 1, 3, 7, 14, 21, 30 and 60 days post-deposition (dpd) and were used to determine the concentration of ivermectin and the percentage of organic matter and for the collection of colonising organisms. Samples from 1 and 3 dpt contained the highest drug concentration and percentage of organic matter compared to the control group (p<0.05). Faeces from the treated group showed lesser abundance and diversity of arthropods (p<0.05) than the control group. A reduction in numbers and diversity of dung fauna in faecal samples from treated animals was most remarkable at 1, 3 and 7 dpt, coinciding with the highest concentration of ivermectin and organic matter percentage.

Arrested development of Ostertagia ostertagi: effect of the exposure of infective larvae to natural spring conditions of the humid pampa (Argentina).

The aim of this study was to determine the effect of environmental conditions and the time of exposure to the conditions required for Ostertagia ostertagi to become inhibited in development at the early fourth larval stage in the host. Two comparable experiments were conducted from September to January, experiment I in 1997-1998 and experiment II in 1999-2000. Twenty-thousand third-stage larvae (L3), freshly obtained from coprocultures, were spread in different parasite-free grass plots at the beginning of September, October and November in each experiment and exposed to environmental conditions throughout spring and early summer. Duplicate plots for each exposure period were grazed for 3 days by two dewormed tracer calves after 1, 2, 3, 4 weeks of exposure during the corresponding month, and the remaining plots were grazed for 3 days at monthly intervals until the end of the experimental period. For each month in both experiments, control animals were inoculated orally with 20,000 L3 newly recovered from coprocultures (week 0 animals; infection controls). The control and tracer calves were sacrificed and their parasite burdens analysed. The time required to obtain greater than 50% inhibited larvae (IeL4) in the tracer animals during September and October was 3 weeks, whereas during November around 60% of the parasites were inhibited after one week of exposure. During the period tested, greater than 50% inhibition was found in concurrence with a photoperiod ranging between 13 and 14 h. The highest proportion of IeL4 (75% average) in the animals was found concomitant with a 14 h 43 min photoperiod. A high correlation between the percentage of inhibition and day length was established (0.870 p < 0.001 and 0.815 p < 0.001 for experiment I and II, respectively). In both years, the capacity for developmental arrest was lost by the end of December, when the photoperiod begins to decrease, suggesting either a disappearance of the induction stimulus, or that an excess of the stimulus could block the mechanism of inhibition. The induction time was extended 2 weeks in all months tested when the coprocultures were maintained in the dark (experiment II), suggesting that accumulation of the light stimulus contributes to shortening of the induction time. The data presented here would suggest that photoperiod is a key environmental factor for the induction of hypobiosis.

Resistance of Cooperia to ivermectin treatments in grazing cattle of the Humid Pampa, Argentina.

A field experiment and a controlled efficacy trial were conducted to demonstrate resistance of cattle trichostrongylid nematodes to endectocidal compounds in grazing cattle on the Humid Pampa, Argentina. Male Polled Hereford calves, aged 9-11 months old, with a history of frequent treatments with compounds of the avermectin/milbemycin class, were used for the trial. The field experiment involved six groups of 10 animals each, which were subcutaneously treated with either ivermectin (IVM), long-acting ivermectin (LA-IVM), moxidectin (MXD) or doramectin (DRM) at a dosage of 0.2mg/kg BW. A fifth group was treated orally with fenbendazole at a dosage of 5mg/kg BW; the sixth group of calves served as non-treated control. Faecal samples were collected from each animal on the day of treatment and at 14 days after treatment. Nematode egg counts were performed to estimate the faecal egg count reduction test (FECRT). The FECRT showed that reductions were 65% (IVM), -20% (LA-IVM) and 85% (DRM). Egg counts from calves treated with MXD or fenbendazole were reduced by 95 and 100%, respectively. For the controlled efficacy trial, six animals with the highest egg counts from the IVM, LA-IVM and non-treated control groups, were necropsied on day 18 after treatment to determine numbers and nematode species in the abomasum and small intestine. The results indicated that efficacy against Cooperia oncophora was 62.7 and 48% for IVM and LA-IVM, respectively. Neither of the IVM formulations showed efficacy against Trichostrongylus spp. in the small intestine. However, the absence of Trichostrongylus spp. in some animals of both treated and control groups precluded a valid assessment of efficacy or resistance. It was concluded that resistance of trichostrongylid nematodes to the avermectin/milbemycin class of compounds in grazing cattle of the Humid Pampa, Argentina, may be strongly established on farms where cattle are treated at frequent intervals throughout the year.

Hypobiosis induction alters the protein profile of Ostertagia ostertagi (Nematoda: Trichostrongylidae).

The appearance of variations in the protein profile of Ostertagia ostertagi (Stiles, 1892) infective larvae (L3), which were induced by hypobiosis triggering factors, was evaluated by means of SDS-PAGE and densitometric analysis. Area integration analyses of their protein profiles was carried out between 66 and 77 kDa. Important quantitative variations were identified in the protein levels of the induced larvae, where a 5.25 fold increase compared to the control was observed. Two 75.4 and 70 kDa protein bands were found which exceeded the control profile by 4.5 and 44 fold, respectively. This fact suggests that the changes brought about in the process of hypobiosis induction are restricted. This work demonstrates changes at a molecular level corresponding with biological changes induced by conditions causing O. ostertagi hypobiosis.

Study on the inductive factors of hypobiosis of Ostertagia ostertagi in cattle.

Two experiments were carried out to determine the causes producing the Ostertagia ostertagi hypobiosis phenomenon in cattle. In the first experiment, the effect of time on third-stage larvae in the environment was studied during a 2-year period. Three experimental paddocks contaminated with O. ostertagi eggs at different times of the year were used, and the levels of hypobiosis were recorded by using 'indicator' and 'tracer' calves. The results suggest that time as such is not a hypobiosis-inductive factor. The second experiment was conducted under laboratory conditions, where the effects of temperature and light on infective larvae were studied. Infective larvae were subjected to different conditions of temperature and light during 6 weeks, and then inoculated to parasite-naive calves, which were slaughtered after 4 weeks. Percentages of hypobiotic larvae in these calves varied from 3.5 to 94.8%, depending on the different storage conditions the larvae underwent before inoculation. Results suggest that increasing temperature and increasing time of light exposure simulating spring conditions would be the factors which act upon third-stage larvae inducing them to a later hypobiotic stage in the host.