Sixteen Belgian Blue beef farms with psoroptic mange were visited during the housing period during the winters of 2016-2017, 2017-2018 and 2018-2019. Skin scrapings were collected from two to five animals per farm to confirm active P. ovis infestations.
A group of seven to thirteen animals was selected on each farm. Male or female cattle older than 4 months were used. The experimental group was separated from the rest of the herd and animals in adjacent pens were treated with an acaricide (injectable ML product or topical treatment with amitraz, twice with one-week interval) for the duration of the study, to preclude re-infestation of the study animals.
All animals were treated with a commercially available injectable ML product, i.e. ivermectin (Ivomec®, Boehringer-Ingelheim), doramectin (Dectomax®, Eli Lilly) or moxidectin (Cydectin®, Zoetis) at the recommended dose rate. On one farm, a long-acting formulation of moxidectin (Cydectin 10% LA®, Zoetis) was used. The animals were weighed on a calibrated weighing scale to determine the dose prior to treatment. In absence of a weighing scale, heart girth measurements were used to estimate the body weight . All animals were sheared prior to the first treatment administration and crusts were removed as much as possible during shearing. Fresh bedding with or without cleaning of the stable was applied at least once per week.
A treatment round consisted of two injections with a 7-10-day interval or a single injection with a long-acting ML product, as recommended by the manufacturer. Treatment was evaluated 7 days after the second administration or 14 days after administration of a long-acting ML product. If living P. ovis were found in one or more animals at a given farm, a new treatment round was started for all the animals in the experiment.
Treatment with ML products was terminated when all animals had negative P. ovis mite counts, when the cattle were turned out on pasture at the end of the housing period or if the condition of the animals warranted salvage treatment. In the latter case, animals were treated topically with amitraz (Taktic®) two times with a one-week interval at the recommended dose.
In vivo efficacy
The clinical index (CI) and mite counts (MC) were determined before the first treatment and after every treatment round. The CI is the proportion of the total body surface with clinical lesions. A CI was assigned to all animals by recording the skin lesions (on both sides of the animal) on a silhouette . At each visit, the investigator assessed the surface of all the lesions on the animal’s body in the grid and then counted the percentage of skin surface affected by lesions .
Skin scrapings from three different locations at the edge of the lesions (9 cm2/sampling location) were collected in one tube per animal for MC. If sampling threatened to influence the treatment results (in case of very small lesions), samples were taken from the predilection sites (withers, back, tail basis). Samples were stored at 10°C and analysed within 24 hours after collection to identify and count living P. ovis mites using a stereoscopic microscope (magnification 100x). If the mites were too numerous to count, the sample was weighed, 1g was taken and mites were counted as previously described. Mite numbers were then recalculated to the total mite count of the pooled sample.
In vitro sensitivity
An in vitro test was based on Brimer et al. . Sterile polystyrene Petri dishes with an internal diameter of 92 mm were filled with 20 ml sterile agar solution (Bidest with 42 g/l Columbia agar) containing 1 ml horse serum (food source) and 0, 1, 10, 100, 1000, 2000 and 5000 µg/ml ivermectin (1% Ivomec®, Boehringer-Ingelheim).
The in vitro sensitivity was determined on mites from pooled skin scrapings from each farm. Ten mites (adults and nymphs from Farm 1-10, adult mites form Farms 11-16) from each pooled sample were transferred to the centre of the Petri dish. The plates were kept at room temperature for 24 h. The mites were inspected under a stereomicroscope at 0, 2, 4, 6, 8 and 24 hours of incubation. Immobility of mites and a lack of reactions or persistent immobility within 1 minute following stimulation with a needle were considered indications of death. All concentrations were tested in triplicate, except for Farms 1 and 8. The test for Farm 1 was conducted in duplo and without the 5000 µg/ml ivermectin plates. This concentration was added afterwards for the other farms, as 50% mortality was not achieved with lower concentrations on Farm 1. Farm 8 was tested in duplo, due to a shortage of mites.
Additional timepoints and an extra parameter (‘knock-down’) were monitored for Farms 11-16. The status of the mites was recorded at 0, 5 min, 0.5, 1, 2, 4,6, 8, 24, 48, 72 and 120 hours of incubation. Knock-down was defined as the absence of spontaneous mobility, while movement was still observed after stimulation with a needle.
(see Statistical analysis in the Supplemental Files)
In vitro sensitivity
The lethal dose 50 (LD50) values for each farm population was calculated for the survival at 24 and 120 hours and knock-down dose 50 (KD50) values at 24 hours using a mixed logistic regression model with mortality and knock-down as binary response variable, drug concentration as continuous independent variable and farm as random effect.
Spearman rank correlations were calculated between nTR, RedMC, RedCI, mite free proportion and log(LD50). Regression analysis was done by fitting a linear model with nTR, mean RedMC, mean RedCI or the proportion of the animals with negative mite counts as independent variable and log(LD50) as dependent variable. All calculations were done in R (version 3.5.1., Copyright © 2018 R Foundation of Statistical Computing).