Search

To provide an accessible and inexpensive method to surveil for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mutations, we developed a multiplex real-time reverse transcription-PCR (rRT-PCR) assay, the Spike single-nucleotide polymorphism (SNP) assay, to detect specific mutations in the spike receptor binding domain. A single primer pair was designed to amplify a 348-bp region of spike, and probes were initially designed to detect K417, E484K, and N501Y. The assay was evaluated using characterized variant sample pools and residual nasopharyngeal samples. Variant calls were confirmed by SARS-CoV-2 genome sequencing in a subset of samples. Subsequently, a fourth probe was designed to detect L452R. The lower limit of 95% detection was 2.46 to 2.48 log10 genome equivalents (GE)/ml for the three initial targets (∼1 to 2 GE/reaction). Among 253 residual nasopharyngeal swabs with detectable SARS-CoV-2 RNA, the Spike SNP assay was positive in 238 (94.1%) samples. All 220 samples with threshold cycle (CT) values of

Inducing latency reversal to reveal infected cells to the host immune system represents a potential strategy to cure HIV infection. In separate studies, we have previously shown that CD8+ T cells may contribute to the maintenance of viral latency and identified a novel SMAC mimetic/IAP inhibitor (AZD5582) capable of reversing HIV/SIV latency in vivo by activating the non-canonical (nc) NF-κB pathway. Here, we use AZD5582 in combination with antibody-mediated depletion of CD8α+ cells to further evaluate the role of CD8+ T cells in viral latency maintenance. Six rhesus macaques (RM) were infected with SIVmac239 and treated with ART starting at week 8 post-infection. After 84-85 weeks of ART, all animals received a single dose of the anti-CD8α depleting antibody (Ab), MT807R1 (50mg/kg, s.c.), followed by 5 weekly doses of AZD5582 (0.1 mg/kg, i.v.). Following CD8α depletion + AZD5582 combined treatment, 100% of RMs experienced on-ART viremia above 60 copies per ml of plasma. In comparator groups of ART-suppressed SIV-infected RMs treated with AZD5582 only or CD8α depletion only, on-ART viremia was experienced by 56% and 57% of the animals respectively. Furthermore, the frequency of increased viremic episodes during the treatment period was greater in the CD8α depletion + AZD5582 group as compared to other groups. Mathematical modeling of virus reactivation suggested that, in addition to viral dynamics during acute infection, CD8α depletion influenced the response to AZD5582. This work suggests that the latency reversal induced by activation of the ncNF-κB signaling pathway with AZD5582 can be enhanced by CD8α+ cell depletion.