In silico chemical library screening and experimental validation of novel compounds with potential varroacide activities.

The mite Varroa destructor is an ectoparasite and has been identified as a major cause of worldwide honey bee colony losses. The use of yearly treatments for the control of varroosis is the most common answer to prevent collapses of honey bee colonies due to the mite. However, the number of effective acaricides is small and the mite tends to become resistant to these few active molecules. In this study, we have been looking for a new original varroacide treatment inhibiting selectively Varroa destructor AChE (vdAChE) with respect to Apis mellifera AChE (amAChE). To do this an original drug design methodology was used applying virtual screening of the CERMN chemolibrary, starting from a vdAChE homology sequence model. By combining the in silico screening with in vitro experiments, two promising compounds were found. In vitro tests of AChE inhibition for both species have confirmed good selectivity toward the mite vdAChE. Moreover, an in vivo protocol was performed and highlighted a varroacide activity without acute consequences on honey bee survival. The two compounds discovered have the potential to become new drug leads for the development of new treatments against the mite varroa. The method described here clearly shows the potential of a drug-design approach to develop new solutions to safeguard honey bee health.

Lethal and sublethal effects of imidacloprid, after chronic exposure, on the insect model Drosophila melanogaster.

Neonicotinoids are subjected to vigilance because of environmental contaminations and deleterious effects on bees. Imidacloprid (IMI) is one of the most representative insecticides of this family. At chronic exposure, concentration-effect relationships are non linear. An insect model should allow a better description of this toxicity. We compared the lethal concentration 50% (LC50) of IMI for a Drosophila-field strain, after acute and chronic exposure. Relative to the acute LC50, the chronic LC50 was lowered by a factor of 29 for males (1.3 mM/45 µM), 52 for larvae (157 µM/3 µM) and more than 172 for females (>3.1 mM/18 µM). Chronic exposure also revealed significant lethal and sublethal effects, at concentrations 3-5 orders of magnitude lower than the chronic LC50. Mean mortalities reached 28% (at 3.91 nM) and 27% (at 39.1 nM) for females and males, respectively. Fecundity decreased of 16% at 1.96 nM. Mating increased of 30% at 0.391 nM. The LOEC (lowest observed effect concentration: 0.391 nM) was 46 000 times lower than the chronic LC50 for males; it was 115 000 times lower than the chronic LC50 for females. This study illuminates effects that neonicotinoids can induce at very low concentrations. This is of particular interest for nontarget insects and for insect dependent species.

Lethal and sub-lethal effects of thymol on honeybee (Apis mellifera) larvae reared in vitro.

BACKGROUND: Thymol offers an attractive alternative to synthetic chemicals to keep Varroa under control. However, thymol accumulates in bee products and is suspected of having adverse effects on colonies and especially on larvae. In this study, we investigated the effects of acute and chronic exposure to thymol on larvae reared in vitro with contaminated food and compared results to the theoretical larval exposure based on the amount of pollen and honey consumed by larvae during their development. RESULTS: The laboratory assays reveal that, first, the 48 h-LD50 of thymol introduced into larval food is 0.044 mg larva(-1) . Second, the 6 day-LC50 is 700 mg kg(-1) food. A significant decrease of larval survival and mass occurred from 500 mg thymol kg(-1) food (P < 0.0001). Finally, vitellogenin expression, which reached a maximum at the fifth instar larvae, is delayed for individuals exposed to 50 mg thymol kg(-1) food (P < 0.0006). That is 10 times higher than the theoretical level of exposure. CONCLUSION: Based on the level of thymol residue found in honey and pollen, these results suggest that the contamination of food by thymol represents no notable risk for the early-developing larvae.