Epidemiological and clinical insights from SARS-CoV-2 RT-PCR crossing threshold values, France, January to November 2020.

BackgroundThe COVID-19 pandemic has led to an unprecedented daily use of RT-PCR tests. These tests are interpreted qualitatively for diagnosis, and the relevance of the test result intensity, i.e. the number of quantification cycles (Cq), is debated because of strong potential biases.AimWe explored the possibility to use Cq values from SARS-CoV-2 screening tests to better understand the spread of an epidemic and to better understand the biology of the infection.MethodsWe used linear regression models to analyse a large database of 793,479 Cq values from tests performed on more than 2 million samples between 21 January and 30 November 2020, i.e. the first two pandemic waves. We performed time series analysis using autoregressive integrated moving average (ARIMA) models to estimate whether Cq data information improves short-term predictions of epidemiological dynamics.ResultsAlthough we found that the Cq values varied depending on the testing laboratory or the assay used, we detected strong significant trends associated with patient age, number of days after symptoms onset or the state of the epidemic (the temporal reproduction number) at the time of the test. Furthermore, knowing the quartiles of the Cq distribution greatly reduced the error in predicting the temporal reproduction number of the COVID-19 epidemic.ConclusionOur results suggest that Cq values of screening tests performed in the general population generate testable hypotheses and help improve short-term predictions for epidemic surveillance.

Ultrasensitive Detection of p24 in Plasma Samples from People with Primary and Chronic HIV-1 Infection.

HIV-1 Gag p24 has long been identified as an informative biomarker of HIV replication, disease progression, and therapeutic efficacy, but the lower sensitivity of immunoassays in comparison to molecular tests and the interference with antibodies in chronic HIV infection limit its application for clinical monitoring. The development of ultrasensitive protein detection technologies may help in overcoming these limitations. Here, we evaluated whether immune complex dissociation combined with ultrasensitive digital enzyme-linked immunosorbent assay (ELISA) single-molecule array (Simoa) technology could be used to quantify p24 in plasma samples from people with HIV-1 infection. We found that, among different immune complex dissociation methods, only acid-mediated dissociation was compatible with ultrasensitive p24 quantification by digital ELISA, strongly enhancing p24 detection at different stages of HIV-1 infection. We show that ultrasensitive p24 levels correlated positively with plasma HIV RNA and HIV DNA and negatively with CD4-positive (CD4+) T cells in the samples from people with primary and chronic HIV-1 infection. In addition, p24 levels also correlated with plasma D-dimers and interferon alpha (IFN-α) levels. p24 levels sharply decreased to undetectable levels after initiation of combined antiretroviral treatment (cART). However, we identified a group of people who, 48 weeks after cART initiation, had detectable p24 levels despite most having undetectable viral loads. These people had different virological and immunological baseline characteristics compared with people who had undetectable p24 after cART. These results demonstrate that ultrasensitive p24 analysis provides an efficient and robust means to monitor p24 antigen in plasma samples from people with HIV-1 infection, including during antiretroviral treatment, and may provide complementary information to other commonly used biomarkers. IMPORTANCE The introduction of combined antiretroviral treatment has transformed HIV-1 infection into a manageable condition. In this context, there is a need for additional biomarkers to monitor HIV-1 residual disease or the outcome of new interventions, such as in the case of HIV cure strategies. The p24 antigen has a long half-life outside viral particles, and it is, therefore, a very promising marker to monitor episodes of viral replication or transient activation of the viral reservoir. However, the formation of immune complexes with anti-p24 antibodies makes its quantification difficult beyond acute HIV-1 infection. We show here that, upon immune complex dissociation, new technologies allow the ultrasensitive p24 quantification in plasma samples throughout HIV-1 infection at levels close to those of viral RNA and DNA determinations. Our results further indicate that ultrasensitive p24 quantification may have added value when used in combination with other classic clinical biomarkers.

Impact of Raltegravir or Efavirenz on Cell-Associated Human Immunodeficiency Virus-1 (HIV-1) Deoxyribonucleic Acid and Systemic Inflammation in HIV-1/Tuberculosis Coinfected Adults Initiating Antiretroviral Therapy.

BACKGROUND: In view of the fast viremia decline obtained with integrase inhibitors, we studied the respective effects of initiating efavirenz (EFV) or raltegravir (RAL)-based antiretroviral therapy (ART) regimens on human immunodeficiency virus (HIV)-1 deoxyribonucleic acid (DNA) levels and inflammation biomarkers in the highly inflammatory setting of advanced HIV-1 disease with tuberculosis (TB) coinfection. METHODS: We followed cell-associated HIV-1 DNA, high-sensitivity C-reactive protein (hsCRP), interleukin 6 (IL-6), soluble CD14 and D-Dimer levels for 48 weeks after ART initiation in the participants to the ANRS12-180 REFLATE-TB study. This phase II open-label randomized study included ART-naive people with HIV and TB treated with rifampicin to receive RAL 400 mg twice daily (RAL400), RAL 800 mg twice daily (RAL800) or EFV 600 mg QD with tenofovir and lamivudine. RESULTS: In 146 participants, the median (interquartile range [IQR]) week (W)0 HIV-1 DNA level was 4.7 (IQR, 4.3-5.1) log10 copies/106 CD4+, and the reduction by W48 was -0.8 log10 copies/106 CD4+ on EFV, -0.9 on RAL400, and -1.0 on RAL800 (P = .74). Baseline median (IQR) hsCRP, IL-6, sCD14, and D-Dimer levels were 6.9 (IQR, 3.3-15.6) mg/L, 7.3 (IQR, 3.5-12.3) pg/mL, 3221 (IQR, 2383-4130) ng/mL, and 975 (IQR, 535-1970) ng/mL. All biomarker levels decreased over the study: the overall W0-W48 mean (95% confidence interval) fold-change on ART was 0.37 (IQR, 0.28-0.48) for hsCRP, 0.42 (IQR, 0.35-0.51) for IL-6, 0.51 (IQR, 0.47-0.56) for sCD14, and 0.39 (IQR, 0.32-0.47) for D-Dimers. There were no differences in biomarker reduction across treatment arms. CONCLUSIONS: In participants with HIV and TB, EFV, RAL400, or RAL800 effectively and equally reduced inflammation and HIV-1 DNA levels.

Anti-gp41 antibody levels reflect HIV viral suppression and cellular reservoir in long-term antiretroviral-treated trial participants.

BACKGROUND: A major challenge to HIV cure strategies is the quantification of persistent reactivation-prone virus in people living with HIV. OBJECTIVES: To determine whether anti-gp41 antibody levels correlate with viral suppression and HIV-1 DNA levels in patients on ART. METHODS: Participants with plasma HIV-1 RNA below 50 copies/mL for >12 months were included from three ANRS cohorts (COPANA, MONOI and APROCO). Antibody levels to gp41 were measured by a low-sensitivity enzyme-linked immunoassay. Correlations with patient and virus characteristics, plasma HIV-1 RNA load (standard and ultrasensitive tests) and cell-associated HIV-1 DNA were assessed. RESULTS: Median age was 41 years and 77.5% of the 683 participants were men. Median CD4+ T cell count was 582 cells/mm3 and median viral suppression duration was 6.6 years (IQR 2.0-9.5). The overall median anti-gp41 antibody titre was 1.3 (IQR 0.6-1.9); median HIV-1 DNA level was 2.6 (IQR 2.1-3.0) log10 copies/106 leucocytes; and HIV-1 RNA was undetectable in 56% of samples. A lower titre of anti-gp41 antibodies correlated with male gender, longer viral suppression and lower HIV-1 DNA burden. Sustained undetectable HIV-1 RNA was associated with lower anti-gp41 levels [median 1.1 (IQR 0.5-1.6) versus 1.4 (IQR 0.7-1.9), P = 0.009]. CONCLUSIONS: Anti-gp41 levels decreased with the duration of antiviral suppression on ART. Lower titres were associated with lower HIV-1 DNA levels and longer duration of viral suppression, reflecting minimal antigen stimulation. Anti-gp41 antibody titration may be a useful biomarker reflecting long-term HIV-1 suppression on ART.

Reconstitution of HIV-1 reservoir following high-dose chemotherapy/autologous stem cell transplantation for lymphoma.

OBJECTIVES: Autologous stem cell transplantation following high-dose chemotherapy (HDC/ASCT) is the prime model to study the impact of HDC in HIV-1-infected participants. We analyzed the impact of HDC/ASCT on the resurgent reservoir composition and origin. DESIGN: We included retrospectively a homogenous group of HIV-1-infected patients treated for high-risk lymphoma in a reference center with similar chemotherapy regimens. METHODS: Thirteen participants treated with HDC/ASCT from 2012 to 2015 were included. A median seven longitudinal blood samples per participant were available. Total HIV-1 DNA levels in peripheral blood mononuclear cells (PBMCs) were quantified by quantitative PCR. In six participants with sustained viral suppression, the highly variable C2V3 viral region was subjected to next-generation sequencing. Maximum-likelihood phylogeny trees were generated from the reconstructed viral haplotypes. Lymphocyte subsets were studied by flow cytometry. RESULTS: PBMC-associated HIV-1 DNA levels were stable over time. Viral diversity decreased along lymphoma treatment, but increased promptly back to prechemotherapy numbers after HDC/ASCT. Blood viral populations from all time-points were intermingled in phylogeny trees: the resurgent reservoir was similar to pre-HDC circulating proviruses. Memory subsets were the main contributor to the early restoration of the CD4+ T-cell pool, with a delayed increase in naïve cell counts. CONCLUSIONS: The characterization of HIV-1 reservoir in blood revealed a fast and consistent replenishment from memory CD4+ T cells after HDC/ASCT. As HDC/ASCT is increasingly involved in HIV cure trials with gene-modified hematopoietic stem cells, the management of infected T cells in HIV-positive autologous transplants will be critical.

Ongoing Clinical Trials of Human Immunodeficiency Virus Latency-Reversing and Immunomodulatory Agents.

In chronic human immunodeficiency virus (HIV)-1 infection, long-lived latently infected cells are the major barrier to virus eradication and functional cure. Several therapeutic strategies to perturb, eliminate, and/or control this reservoir are now being pursued in the clinic. These strategies include latency reversal agents (LRAs) designed to reactivate HIV-1 ribonucleic acid transcription and virus production and a variety of immune-modifying drugs designed to reverse latency, block homeostatic proliferation, and replenish the viral reservoir, eliminate virus-producing cells, and/or control HIV replication after cessation of antiretroviral therapy. This review provides a summary of ongoing clinical trials of HIV LRAs and immunomodulatory molecules, and it highlights challenges in the comparison and interpretation of the expected trial results.