The Citrus tristeza virus (CTV) strains present in an area can determine the type and severity of disease produced. Using real time RT-qPCR, we screened a series of representative samples collected throughout Florida from 1964 to 2002 for CTV strain presence. We found that contrary to previous reports, the historical samples were infected with both the T30 and T36 strains, the latter often at low titer. The VT strain was rare, with a scattered distribution. We compared this to samples collected in 2014 and 2015, and found that T30 and T36 are prevalent in nearly all commercial groves; VT strain incidence has increased markedly. With changing cultural practices, such as an increase in sour orange rootstock use, tristeza disease continues to be a significant threat to the Florida citrus industry.