The soybean aphid (Aphis glycines) was a poor vector (0.83% transmission) when the aphids were allowed overnight acquisition feed on Soybean mosaic virus (SMV)-infected soybean leaves. However, A. glycines was shown to be a very efficient vector (34.72% transmission) when individual aphids were allowed a 1-min acquisition probe on the same infected leaves used for the feeding treatment. Similar results were obtained with Myzus persicae and tobacco in transmission experiments of the potyviruses Tobacco etch virus (feeding: 1.36%; probing: 45.5%) and Tobacco vein mottling virus (feeding: 2.0%; probing: 47.5%). A reverse transcription-polymerase chain reaction (RT-PCR) assay was developed to detect SMV in single soybean aphids using a pair of primers designed to amplify a 469-bp PCR fragment in the coding region of SMV coat protein. In contrast to the low transmission rate obtained with the soybean aphids that acquired virus through overnight feeding, RT-PCR detected SMV in 100% of these aphids. Interestingly, the rate of SMV detection by RT-PCR in aphids that were allowed a 1-min acquisition probe (31.67%) coincided with percent transmission (34.72%). The practical application of RT-PCR in detecting nonpersistently transmitted viruses and its implications for virus epidemiology are discussed.