Technical Abstract: Viruses have evolved as combinations of genes that interact with cellular components to produce progeny virus throughout the plants. Some viral genes, particularly those that are involved in replication and assembly, tend to be relatively conserved, while other genes that have evolved for interactions with the specific host for movement and to counter host defense systems tend to be less conserved. Closteroviridae encode 1 to 5 non-conserved open reading frames (ORFs). Citrus tristeza virus (CTV), a Closterovirus, possesses non-conserved p33, p18 and p13 genes that are expendable for systemic infection of the two major laboratory hosts, Citrus macrophylla and Mexican lime. In this study, we show that the extended host range of CTV does require these non-conserved genes. The p33 ORF was required to systemically infect sour orange and lemon trees, while either the p33 or the p18 gene was sufficient for systemic infection of grapefruit trees and the p33 or the p13 gene was sufficient for systemic infection of calamondin plants. Thus, these three genes are required for systemic infection of the full host range of CTV, but different genes were specific for different hosts. Remarkably, either of two genes was sufficient for infection of some citrus hybrids. These findings suggest that CTV acquired multiple non-conserved genes (p33, p18 and p13) to extend its ability to interact with multiple hosts, thus extending its host range during the course of evolution. These results greatly extend the complexity of known virus plant interactions.