SARS-CoV-2 infects blood monocytes to activate NLRP3 and AIM2 inflammasomes, pyroptosis and cytokine release

SARS-CoV-2 causes acute respiratory distress that can progress to multiorgan failure and death in some patients. Although severe COVID-19 disease is linked to exuberant inflammation, how SARS-CoV-2 triggers inflammation is not understood. Monocytes are sentinel blood cells that sense invasive infection to form inflammasomes that activate caspase-1 and gasdermin D (GSDMD) pores, leading to inflammatory death (pyroptosis) and processing and release of IL-1 family cytokines, potent inflammatory mediators. Here we show that ~10% of blood monocytes in COVID-19 patients are dying and infected with SARS-CoV-2. Monocyte infection, which depends on antiviral antibodies, activates NLRP3 and AIM2 inflammasomes, caspase-1 and GSDMD cleavage and relocalization. Signs of pyroptosis (IL-1 family cytokines, LDH) in the plasma correlate with development of severe disease. Moreover, expression quantitative trait loci (eQTLs) linked to higherGSDMDexpression increase the risk of severe COVID-19 disease (odds ratio, 1.3, p<0.005). These findings taken together suggest that antibody-mediated SARS-CoV-2 infection of monocytes triggers inflammation that contributes to severe COVID-19 disease pathogenesis.One sentence summaryAntibody-mediated SARS-CoV-2 infection of monocytes activates inflammation and cytokine release.

Infrequent HIV Infection of Circulating Monocytes during Antiretroviral Therapy

ABSTRACT Whereas human immunodeficiency virus (HIV) persists in tissue macrophages during antiretroviral therapy (ART), the role of circulating monocytes as HIV reservoirs remains controversial. Three magnetic bead selection methods and flow cytometry cell sorting were compared for their capacity to yield pure CD14+ monocyte populations. Cell sorting by flow cytometry provided the purest population of monocytes (median CD4+ T-cell contamination, 0.06%), and the levels of CD4+ T-cell contamination were positively correlated with the levels of integrated HIV DNA in the monocyte populations. Using cell sorting by flow cytometry, we assessed longitudinally the infection of monocytes and other cell subsets in a cohort of 29 Thai HIV-infected individuals. Low levels of HIV DNA were detected in a minority of monocyte fractions obtained before and after 1 year of ART (27% and 33%, respectively), whereas HIV DNA was readily detected in CD4+ T cells from all samples. Additional samples (2 to 5 years of ART) were obtained from 5 individuals in whom monocyte infection was previously detected. Whereas CD4+ T cells were infected at high levels at all time points, monocyte infection was inconsistent and absent in at least one longitudinal sample from 4/5 individuals. Our results indicate that infection of monocytes is infrequent and highlight the importance of using flow cytometry cell sorting to minimize contamination by CD4+ T cells. IMPORTANCE The role of circulating monocytes as persistent HIV reservoirs during ART is still controversial. Several studies have reported persistent infection of monocytes in virally suppressed individuals; however, others failed to detect HIV in this subset. These discrepancies are likely explained by the diversity of the methods used to isolate monocytes and to detect HIV infection. In this study, we show that only flow cytometry cell sorting yields a highly pure population of monocytes largely devoid of CD4 contaminants. Using this approach in a longitudinal cohort of HIV-infected individuals before and during ART, we demonstrate that HIV is rarely found in monocytes from untreated and treated HIV-infected individuals. This study highlights the importance of using methods that yield highly pure populations of cells as flow cytometry cell sorting to minimize and control for CD4+ T-cell contamination.

Monocyte Infection Dynamics Shape HIV-1 Phyloanatomy in the Peripheral Blood

Human immunodeficiency virus (HIV) RNA and DNA have been isolated from patient monocytes, an immune cell population that is quite different in several aspects from the canonical T-cell viral target. Because monocytes are migratory and resilient to both natural and synthetic antiviral defenses, knowledge of the contribution of monocyte infection to ongoing viral evolution and spread \textit{in vivo} is of significant interest for drug development and treatment strategies. Using single viral genome sequencing from different peripheral blood compartments and phyloanatomic statistical inference, we demonstrate that productively infected monocytes follow an evolutionary trajectory that is distinct from peripheral T cells during multiple stages of disease progression. Gene flow and selection analysis reveal plasticity in the source of monocyte infection and in the region of the HIV envelope glycoprotein that experiences selection pressure across individuals. The findings, thus, point to a potential reservoir showing a range of infection and transmission dynamics, for which the current universal, T cell-targeted treatment strategies would be inadequate.

Pharmacodynamic Evaluation of the Intracellular Activity of Antibiotics towards Pseudomonas aeruginosa PAO1 in a Model of THP-1 Human Monocytes

ABSTRACTPseudomonas aeruginosainvades epithelial and phagocytic cells, which may play an important role in the persistence of infection. We have developed a 24-h model of THP-1 monocyte infection withP. aeruginosaPAO1 in which bacteria are seen multiplying in vacuoles by electron microscopy. The model has been used to quantitatively assess antibiotic activity against intracellular and extracellular bacteria by using a pharmacodynamic approach (concentration-dependent experiments over a wide range of extracellular concentrations to calculate bacteriostatic concentrations [Cs] and maximal relative efficacies [Emax]; Hill-Langmuir equation). Using 16 antipseudomonal antibiotics (three aminoglycosides, nine β-lactams, three fluoroquinolones, and colistin), dose-response curves were found to be undistinguishable for antibiotics of the same pharmacological class if data were expressed as a function of the corresponding MICs. Extracellularly, all of the antibiotics reached a bacteriostatic effect at their MIC, and theirEmaxexceeded the limit of detection (−4.5 log10CFU compared to the initial inoculum). Intracellularly,Csvalues remained unchanged for β-lactams, fluoroquinolones, and colistin but were approximately 10 times higher for aminoglycosides, whereasEmaxvalues were markedly reduced (less negative), reaching −3 log10CFU for fluoroquinolones and only −1 to −1.5 log10CFU for all other antibiotics. The decrease in intracellular aminoglycoside potency (higherCs) can be ascribed to the acid pH to which bacteria are exposed in vacuoles. The decrease in theEmaxmay reflect a reversible alteration of bacterial responsiveness to antibiotics in the intracellular milieu. The model may prove useful for comparison of antipseudomonal antibiotics to reduce the risk of persistence or relapse of pseudomonal infections.