Probes for INS

Subtype C HIV-1 is responsible for the largest proportion of people living with HIV-1 infection. However, there is limited information about the roles of the brain and its cell types as a potential sanctuary for this subtype and how the sanctuary may be affected by the administration of anti-retroviral therapy (ART). To address this issue, we collected postmortem brain tissues from ART treated HIV-1 infected Zambian individuals who experienced complete viral suppression and those who did not. Tissues from various brain compartments were collected from each individual as frozen and formalin-fixed paraffin embedded brain specimens, for detection and quantification of HIV-1 genomes and identification of the infected cell type. Genomic DNA and RNA were extracted from frozen brain tissues. The extracted DNA and RNA were then subjected to droplet digital PCR for HIV-1 quantification. RNA/DNAscope in situ hybridization (ISH) for HIV-1 was performed on formalin-fixed paraffin embedded brain tissues in conjugation with immunohistochemistry to identify the infected cell types. Droplet digital PCR revealed that HIV-1 gag DNA and RNA were detectable in half of the cases studied regardless of ART success or failure. The presence of HIV-1 lacked specific tissue compartmentalization since detection was random among various brain tissues. When combined with immunohistochemistry, RNA/DNAscope ISH demonstrated co-localization of HIV-1 DNA with CD68 expressing cells indicative of microglia or peripheral macrophage. Our study showed that brain is a potential sanctuary for subtype C HIV-1, as HIV-1 can be detected in the brain of infected individuals irrespective of ART treatment outcome and no compartmentalization of HIV-1 to specific brain compartments was evident.

We report a case of an adolescent who presented at our emergency department with acute abdominal pain. While the initial diagnosis was acute appendicitis, a secondary and coincidental diagnosis of primary HIV-1 infection was made. Concurrent and subsequent clinical and molecular biology findings form the basis of our argument that primary HIV-1 infection was the cause of acute appendicitis in this individual.

We have recently developed the first microscopy-based strategy that enables simultaneous multiplex detection of viral RNA (vRNA), viral DNA (vDNA), and viral protein. Here, we used this approach to study the kinetics of latency reactivation in cells infected with the human immunodeficiency virus (HIV). We showed the transcription of nascent vRNA from individual latently integrated and reactivated vDNA sites appearing earlier than viral protein. We further demonstrated that this method can be used to quantitatively assess the efficacy of a variety of latency reactivating agents. Finally, this microscopy-based strategy was augmented with a flow-cytometry-based approach, enabling the detection of transcriptional reactivation of large numbers of latently infected cells. Hence, these approaches are shown to be suitable for qualitative and quantitative studies of HIV-1 latency and reactivation.