Amplification and detection of lentiviral DNA inside cells

Abstract

Visna virus and human immunodeficiency virus are prototypes of animal and human lentiviruses, respectively, that persist and are disseminated despite the host immune response because cells in the tissues and the bloodstream harbor viral genomes in a covert state. To facilitate identification of these latently infected cells, the polymerase chain reaction has been adapted to amplify viral DNA in fixed cells for detection by in situ hybridization. By using a multiple primer set that generates DNA segments with overlapping cohesive termini, visna virus DNA can be amplified, retained, and detected in infected cells with sensitivities that exceed those of existing methods by more than 2 orders of magnitude. This advance in single-cell technology should prove useful in diagnosing and gaining insight into the pathogenesis of viral infections and provide new opportunities to look for viruses in chronic diseases of unknown etiology.