MOLECULAR CLONING AND CHARACTERIZATION OF RETROVIRUSES ASSOCIATED WITH AIDS

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The objective of this project is to obtain information pertaining to the structure and diversity of HIV with respect to its biochemical, pathogenic, and immunological variability. Heterogeneity among retroviral genomes is a distinct feature of Lentiviruses. The human immunodeficiency virus type 1 Lentivirus (HIV-1), the causative agent of acquired immunodeficiency syndrome, has been isolated from CD4-positive peripheral blood T lymphocytes (PBLs) and monocyte-derived macrophages (MDMs) of seropositive individuals. Stable infectious molecular clones of HIV-1, which generate high titers of virus capable of infecting cultured PBLs, MDMs and a variety of CD4 positive T cell leukemic lines have been extensively used to delineate the structure and function of HIV-1 encoded proteins. However, only a few molecular clones have been isolated directly from infected tissue. It would be advantageous to obtain such clones without having to passage in tissue culture and lessening the probability of picking up mutations or selecting for variants that adapt to tissue culture conditions. We were able to clone directly from lung tissue obtained from a SIV-HIV (SHIV) infected, CD4 depleted cynomolgus macaque and obtained six individual clones. A highly cytopathic and syncytia forming HIV-1 virus was isolated from the blood of a patient from Uganda and was molecularly cloned from infected human PBLs. Nine complete and infectious clones were isolated from this single isolate. Of interest is the fact that the molecularly cloned viruses were able to grow in human PBLs, chimpanzee PBLs, MT-4 cells (continuous T-cell line), H-9 cells, and MDMs. This molecular isolate is dual tropic with respect to its ability to infect different cell lines.