This study was conducted to evaluate the role of level infestation of the Varroa destructormite on virgin queen's acceptance, mating success and pre-oviposition period when their introduced into Carniolan honeybee, Apis mellifera colonies. According to level infestation of mite, the colonies were divided into three groups, each contained 15 colonies, low (5% ± 0.3); moderate (12% ± 0.7) and high (20% ± 0.9).          The results showed that the varroa level significant influence virgin queen's acceptance, mating success and somewhat influence their onset of oviposition. The results mentioned that, low infestation level was more accepted and reached mating success 100% than both moderate and high levels (mating success were 91.67% and 80.0%, respectively). Pre-oviposition periods were (8.9 ± 1.05; 11.1 ± 0.89 and 11.6 ± 1.45 days), recorded at low, moderate and high mite infestation levels, respectively. Beekeepers can get an economic benefit from introducing queen bees and improving introduction acceptance and mating success by control varroa mite before queen introduction.