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In this study, we investigated the effect of queen caging on honey bee colonies’ post-treatment development and the optimal timing of method application on honey production during the main summer nectar flow. We conducted the study in nine apiaries (N = 9) across six Mediterranean countries, with a total of 178 colonies. The colonies were divided into three test groups: QC1, QC2, and C. The QC1 group involved queens caged for a total of 28 days before the expected harvesting day. In the QC2 group, queens were caged for 28 days, but only 14 days before the expected harvesting day. The C group consisted of queens that were not caged, and the colonies received common local treatments. In both the QC1 and QC2 groups, the colonies were treated with a 4.2% oxalic acid (OA) solution by trickling after the queen release. Our findings revealed no significant adverse effects (p > 0.05) on colony strength at the end of the study resulting from queen caging. However, significantly lower amounts of honey were extracted from the QC1 group compared to both the QC2 group (p = 0.001) and the C group (p = 0.009). Although there were no initial differences in Varroa destructor infestation between the groups, ten weeks later, a significantly higher infestation was detected in the C group compared to both the QC1 group (p < 0.01) and the QC2 group (p = 0.003). Overall, our study demonstrates that queen caging, in combination with the use of OA, is an effective treatment for controlling V. destructor. However, the timing of caging plays a crucial role in honey production outcomes.