Virological characterization of HIV-1 RNA elements detected exclusively through the LTR region by the dual-target Aptima HIV-1 Quant Dx assay in a subset of positive patients

Background: In a restricted subset of HIV patients with suppressed viral load (i.e., pol-undetected HIV-RNA), the Aptima HIV-1 Quant Dx Assay (Aptima), a dual-target (pol and LTR) and dual-probe test for viral load (VL) monitoring, can detect HIV-RNA exclusively through amplification of the LTR region.Objectives: To analyze the virological characteristics of the HIV-RNA elements detected only through LTR amplification (LTR-e). Study design: LTR-e isolated from plasma and peripheral blood mononuclear cells (PBMC) were evaluated for their ability to trigger productive infections. Viral pellets morphology and ultrastructural characteristics of PBMC from LTR-e patients were examined by electron microscopy. Plasma LTR-e underwent Sanger sequencing. Exosomes were examined with Aptima for LTR-e content.Results: In-vitro, LTR-e could not infect PBMC, induce cytopathic effects, or cause syncytia, even at high VL (e.g., >10,000 copies/mL). Under the electron microscope, plasma pellets and PBMC from patients with LTR-e showed atypical vesicles. Sanger sequencing of LTR-e yielded no results. Moreover, in plasma samples, LTR-e were associated with cell debris, never with exosomes.Conclusions: Differently from other dual-target but single-probe assays, Aptima unveils VL based only on LTR amplification in some HIV patients. Here, we show that LTR-e represent partial/incomplete/non-canonical transcripts unable to trigger productive infection or transmit HIV-1 infection. The recognition of VL based only on LTR-e in infected individuals is crucial as it allows to avoid inappropriate decisions in the clinical management of HIV patients, such as retesting of VL and switching of ART. Physicians and HIV-RNA dual-target assay manufacturers should consider the important implications of not recognizing this singular type of VL.

Automated summary

In a subset of HIV patients, the Aptima HIV-1 Quant Dx Assay (Aptima) can detect viral load exclusively through amplification of the LTR region. The viral RNA detected through LTR amplification does not have the ability to infect or transmit HIV-1. The recognition of viral load based on LTR amplification is crucial for the clinical management of HIV patients.