Equine infectious anemia virus (EIAV) is an equine lentivirus responsible for a persistent infection manifested by recurring episodes of fever, anemia and edema. Each episode of clinical disease is associated with the emergence of a unique antigenic variant of EIAV. During phase I, this project will attempt to develop an infectious molecular clone of EIAV from genomic RNA extracted from the plasma of a pony acutely infected with the wild-type Wyoming strain of EIAV. We will determine the entire nucleotide sequence of this infectious clone using the dideoxy, chain terminating method of sequencing. Comparison of the nucleotide sequence of this infectious, virulent clone with previously developed avirulent, noninfectious clones will help define some molecular determinants of infectivity. In addition, combined in situ hybridization and immunocytochemical studies using monoclonal antibodies to equine monocytes/macrophages and EIAV will be performed. This will allow identification of cellular sites of viral replication and persistence during acute disease and during the chronic, asymptomatic stage of infection. During Phase II, ponies will be infected with the infectious, virulent clone of wild-type Wyoming strain of EIAV. Virus isolates from sequential febrile episodes will be analyzed by oligonucleotide mapping to determine sites and types of nucleotide mutations occurring in vivo. Molecular determinants of infectivity and virulence will be further investigated using serial allelic replacements of nucleotide segments of noninfectious avirulent clones with comparable segments from our infectious clones. Taken together, the proposed experiments should provide much-needed information on mechanisms of lentivirus persistence and in vivo antigenic variation following an initial homogeneous infection. These same mechanisms may be integral in the pathogenesis of human lentivirus infections, including the acquired immune deficiency syndrome (AIDS) associated with human immunodeficiency virus 1.