We isolated plaque-size variants of a South American strain of yellow fever virus, and compared their ability to infect orally and be transmitted by vector Aedes aegypti mosquitoes with that of the uncloned, parental virus. We analyzed the same clonal isolates in mouse virulence experiments. No significant differences could be demonstrated in the capacities of the variants to infect and be transmitted by mosquitoes or in mouse virulence tests. The 17D vaccine virus (derived from the African Asibi strain) was, however, markedly attenuated in mosquitoes; also, a variant virus (Asibi strain) derived by continuous passage in HeLa cells (and attenuated for monkeys and mice) was markedly attenuated in mosquitoes when compared with its parent virus. The results suggest that vector competence and mouse virulence are similar in plaque-size variants of the South American strain of yellow fever virus. However, if vigorous and appropriate selection pressures are applied (as established by the derivation of the 17D vaccine and HeLa passaged viruses), attenuated variants can be discovered; their role and prevalence within a virus population in nature is unknown.