IN-VIVO STUDY OF GENETICALLY SIMPLIFIED BOVINE LEUKEMIA-VIRUS DERIVATIVES THAT LACK TAX AND REX

Genetically simplified derivatives of complex retroviruses that replic ate in animal models are useful tools to study the role of the complex regulatory genes in virus infection and pathogenesis and were propose d as a novel approach toward the development of vaccines against compl ex retroviruses. Previously we developed genetically simple derivative s of bovine leukemia virus (BLV) that can replicate in tissue culture independently of the BLV regulatory proteins, Tax and Rex, and the RII I and GIV open reading frames (K. Boris-Lawrie and H. M. Temin, J. Vir ol. 69:1920-1924, 1995). These derivatives are encoded on novel, hybri d retrovirus genomes that contain transcriptional control sequences of a simple retrovirus and gag-pol or env genes of the complex BLV. The first-generation simple BLV derivatives replicate as complementary vir uses (coviruses) by using separate gag-pol or env genomes, and therefo re virus spread is limited to cells that are infected with both coviru s genomes. Here we describe a second-generation simple BLV derivative that is encoded on a single hybrid genome. We show the virus to be rep lication competent by successive passage on D17 target cells and by an alysis of viral RNA and proteins in the infected cells. Furthermore, w e evaluate the immunogenicity and infectivity of the simple BLV deriva tives in a BLV animal model. Small groups of rats were injected either with virus-producing cells or with proviral DNA. Western immunoblot a nalysis revealed that antibodies against the major viral antigenic det erminants are induced in response to either method of introduction and that seroconversion is sustained in most of the rats for at least 6 m onths (the duration of the study). The magnitudes of the antiviral res ponses were similar in rats infected with the first-generation simple BLV coviruses, the second-generation replication-competent derivative, or wild-type BLV. Wild-type BLV typically infects peripheral blood mo nonuclear cells (PBMC), and the simple BLV derivatives were also found to infect PBMC as demonstrated by PCR amplification of proviral seque nces and reverse transcriptase PCR amplification of viral RNA in treat ed rats. These results establish that simple BLV derivatives lacking t ax and rex are infectious and immunogenic in rats. These viruses will be useful tools in comparative studies with BLV to evaluate the role o f tax and rex in maintenance of virus load and in disease outcome.