Equine infectious anemia virus (EIAV) is a lentivirus that causes lifelong persistent infection in horses. In vivo, EIAV typically progresses in four disease stages. The initial acute stage of infection leads to a chronic stage of recurring fever cycles. After the host mounts a broadly acting immune response, EIAV enters an inapparent stage of clinical quiescence, from which it may later recrudesce. To identify genetic changes that affect immune escape during each disease stage, viral RNA was isolated from sequential sera samples collected from a horse experimentally infected with EIAV. The 3' end of the viral genome was amplified using single genome amplification (SGA), and individual amplicons were sequenced. Genotypes representative of each disease stage were identified using a phylogenetic clustering analysis developed by modeling each subpopulation as an unobserved ancestral sequence and a cloud of descendents. The SU (surface envelope glycoprotein) of representative genotypes was used to generate pseudotyped reporter viruses, which were tested to determine how genetic variation alters neutralization phenotypes and/or infectivity. The representative sequences showed a continuous selection of antigenic escape variants. Successive variants became increasingly resistant to broadly neutralizing antibodies; however, this did not appear to incur a cost to viral infectivity. Identifying genetic changes that contribute to immune escape and viral persistence can aid in the creation of effective vaccines against animal and human lentiviruses, including HIV-1.