## 4 Discussion  
Varroa mite is a major cause of overwintering honey bee (_A. mellifera_) colony losses across the globe [36]. In recent years, several surveys were conducted to analyze beekeepers' treatment practices and overwintering [36; 37; 38; 39; 40]. One of the findings of the above-mentioned studies was that the worst scenario concerning the overwintering ability was a combination of weakness of honey bee populations, low food reserves and high Varroa infestation levels [37]. In our study, we administered VarroMed(r) in several apiaries with different climatic conditions. The average temperatures near the experimental apiaries in Croatia, Italy, Slovenia and Belgium show a versatile range (30.2-12.4 \({}^{\circ}\)C) only in summer/autumn treatment period in 2018. Our results show that the efficacy of VarroMed(r) treatment was above 70% and not highly variable, regardless of the presence of honey bee brood and environmental temperature, despite some relevant differences between minimum and maximum efficacy values in some sites, as shown in Tables 2 and 5. High efficacies of other oxalic acid-based treatments (application by trickling or sublimation) were reported by [PERSON] et al. [8]. Colonies were treated in a broodless period in summer time (using brood interruption techniques, such as queen caging or brood removal) and the efficacy ranged from 48 to 89% mite removal. The protocols of the treatments against _V. destructor_ may require quite a long time (over 40 days) to reach a high acaricide efficacy, especially in broodign colonies with high levels of infestation [31]. The field trials were carried out in medium infested colonies and the treatment was, therefore, appropriate. In the experimental apiaries, the efficacy ranged from 71.2 to 89.3% in summer/autumn treatment. [PERSON] and [PERSON] [41] reported even higher acaricide efficacy (91.5%) in summer, after three consecutive applications of the oxalic acid based complementary feed HiveClean(r) (former product, comparable with VarroMed(r)). In winter treatments reported in this paper, the overall acaricide efficacies ranged from 71.8 to 95.6%. According to the report of the European Medicines Agency [42], the efficacy of winter treatment with VarroMed(r) was 88%. The level of efficacy of VarroMed(r) treatment in summer/autumn and winter was found sufficient, which can be supported by the average number of mite fall after queen caging and follow-up treatment. Figure 8: Boxplot of adult honey bee coverage in all groups recorded during the winter trial and compared to the beginning of the treatment (%). The horizontal line in boxes shows the median. The box shows the first and third quartile. Whiskers extend to 1.5 \(\times\) interquartile range (IQR). Additionally, the pluses show mean values. Another critical aspect to the use of mixture of organic acids (oxalic and formic acid) and additional substances against Varroa mites that should be considered is a possible toxicity to honey bees and colonies. The use of oxalic acid by trickling, evaporation and spraying, honey bee tolerability and efficacy has been reviewed by [PERSON] and [PERSON] [12]. In general, the application of oxalic acid in a formulation with sugar syrup increases the efficacy against Varroa mites. However, there are some negative effects of oxalic acid on queen health [43; 44] and on worker bees' digestive system [45]. Formic acid operates through the inhibition of mitochondrial energetic metabolism of Varroa mites binding the cytochrome C oxidase enzyme [46], as well as through a significant neuroexcitation process [47]. It also affects honey bee colony, as it reduces the longevity of worker bees [48] and affects brood survival [49]. There is a report on an increased number of dead bees in front of the hive, queen rejection and decrease of honey yield during treatment [50]. Considering the above-mentioned toxicity aspects, we evaluated the effects on the strength of the colonies after VarroMed(r) treatments.