Technical Abstract: Citrus tristeza virus (CTV) is the most destructive viral pathogen of citrus and has been an important concern for the citrus industry in the Dominican Republic. Earlier studies documented widespread distribution of mild isolates of the T30 genotype, which caused no disease in the infected trees, and a low incidence of isolates of the VT and T3 genotypes, which were associated with economically damaging decline and stem pitting symptoms. In light of the dramatic increase in the number of severely diseased citrus trees throughout the country over the last decade suggesting that field populations of CTV have changed, we examined the CTV pathosystem in the Dominican Republic to assess the dynamics of virus populations. In this work, we characterized the molecular composition of 163 CTV isolates from different citrus-growing regions. Our data demonstrate a dramatic change in CTV populations, with the VT genotype now widely disseminated throughout the different regions and with the apparent ingress of two new virus genotypes, T36 and RB. Remarkably, these viruses were able to superinfect trees that appeared to be infected with other genotypes of CTV prior to their invasion. Multiple infections of trees resulted in development of complex virus populations composed of different genotypes.