The COVID-19 pandemic has caused at least 780 million cases globally. While available treatments and vaccines have reduced the mortality rate, spread and evolution of the virus are ongoing processes. Despite extensive research, the long-term impact of SARS-CoV-2 infection is still poorly understood and requires further investigation. Routine analysis provides limited access to the tissues of patients, necessitating alternative approaches to investigate viral dissemination in the organism. We address this issue by implementing a whole-body in vivo imaging strategy to longitudinally assess the biodistribution of SARS-CoV-2. We demonstrate in a COVID-19 non-human primate model that a single injection of radiolabeled [89Zr]COVA1-27-DFO human monoclonal antibody targeting a preserved epitope of the SARS-CoV-2 spike protein allows longitudinal tracking of the virus by positron emission tomography with computed tomography (PET/CT). Convalescent animals exhibit a persistent [89Zr]COVA1-27-DFO PET signal in the lungs, as well as in the brain, three months following infection. This imaging approach also allows viral detection in various organs, including the airways and kidneys, of exposed animals during the acute infection phase. Overall, the technology we developed offers a comprehensive assessment of SARS-CoV-2 distribution in vivo and provides a promising approach for the non-invasive study of long-COVID pathophysiology.

Abstract Background The fight against COVID-19 requires mass vaccination strategies, and vaccines inducing durable cross-protective responses are still needed. Inactivated vaccines have proven lasting efficacy against many pathogens and good safety records. They contain multiple protein antigens that may improve response breadth and can be easily adapted every year to maintain preparedness for future seasonally emerging variants. Methods The vaccine dose was determined using ELISA and pseudoviral particle-based neutralization assay in the mice. The immunogenicity was assessed in the non-human primates with multiplex ELISA, neutralization assays, ELISpot and intracellular staining. The efficacy was demonstrated by viral quantification in fluids using RT-qPCR and respiratory tissue lesions evaluation. Results Here we report the immunogenicity and efficacy of VLA2001 in animal models. VLA2001 formulated with alum and the TLR9 agonist CpG 1018™ adjuvant generate a Th1-biased immune response and serum neutralizing antibodies in female BALB/c mice. In male cynomolgus macaques, two injections of VLA2001 are sufficient to induce specific and polyfunctional CD4+ T cell responses, predominantly Th1-biased, and high levels of antibodies neutralizing SARS-CoV-2 infection in cell culture. These antibodies also inhibit the binding of the Spike protein to human ACE2 receptor of several variants of concern most resistant to neutralization. After exposure to a high dose of homologous SARS-CoV-2, vaccinated groups exhibit significant levels of protection from viral replication in the upper and lower respiratory tracts and from lung tissue inflammation. Conclusions We demonstrate that the VLA2001 adjuvanted vaccine is immunogenic both in mouse and NHP models and prevent cynomolgus macaques from the viruses responsible of COVID-19.

Few therapeutic options are available to treat COVID-19. The KEAP1/NRF2 pathway, the major redox-responsive pathway, has emerged as a potential therapeutic target for COVID-19 as it regulates redox homeostasis and inflammation that are altered during SARS-CoV-2 infection. Here, we characterized the effects of NRF2-agonist Sulfodyne®, a stabilized natural Sulforaphane, in cellular and animal models of SARS-CoV-2 infection. In pulmonary or colonic epithelial cell lines, Sulfodyne® elicited a more efficient inhibition of SARS-CoV-2 replication than NRF2-agonists DMF and CDDO. This antiviral activity was not dependent on NRF2 but was associated with the regulation of several metabolic pathways, including the inhibition of ER stress and mTOR signaling, which are activated during SARS-CoV-2 infection. Sulfodyne® also decreased SARS-CoV-2 mediated inflammatory responses by inhibiting the delayed induction of IFNB1 and type I IFN-stimulated genes in infected epithelial cell lines and by reducing the activation of human by-stander monocytes recruited after SARS-CoV-2 infection. In K18-hACE2 mice infected with SARS-CoV-2, Sulfodyne® treatment reduced both early lung viral load and disease severity by fine-tuning IFN-beta levels. Altogether, these results provide evidence for multiple mechanisms that underlie the antiviral and anti-inflammatory activities of Sulfodyne® and pinpoint Sulfodyne® as a potent therapeutic agent against pathogenic effects of SARS-CoV-2 infection.

Background: Salivary gland epithelial cells (SGEC) play a pathogenic role in Sjögren’s disease (Sjo), mainly through their interaction with immune cells. Actual 2D culture models of SGEC do not recapitulate the complexity of cellular interactions within salivary glands and lack epithelial readouts, highlighting the necessity to develop new relevant models. Objectives: To develop and characterize differentiated organoids (DO) of SGEC derived from minor salivary gland biopsies (MSGB) of Sjo patients and controls. Methods: We included Sjo patients fulfilling the ACR/EULAR 2016 criteria and controls with sicca syndrome without autoimmune disease. MSGB were firstly dissociated enzymatically, encapsulated in extracellular matrix (ECM), and then cultured in Growth Expansion Medium (GEM) containing growth factors, until organoids formation. Organoids were passaged every 7-12 days for self-renewal and cultured in a Differentiation Medium (DM) to develop DO. Proliferation was measured by cell counting and SGEC markers were investigated by RT-qPCR, immunohistochemistry and immunofluorescence. The calcium influx was assessed by fluorescence spectroscopy. Results: We included 7 Sjo and 13 controls. Organoids were forming and differentiating in both Sjo and controls (Figure 1A-B). The mean culture duration was respectively 3.7 ± 1.5 and 2.7 ± 1.1 months in Sjo and controls. The self-renewal capacity of organoids for long-term culture was comparable between the two groups (Figure 1C). We then characterized DO and found that they highly express the ductal markers CK7/CK18 and to a lesser degree, the acinar markers AQP5 and α-amylase in both groups (Figure 1D-E). DO also expressed BAFF, CXCL10 and IL-7 when exposed to PolyIC and IFNα supporting a capacity of response to inflammation (Figure 1F). Exposure of DO to pilocarpine induced an increase of calcium levels, suggesting a capacity to secrete saliva in response to a cholinergic stimulation. Of note, the effect was reduced in Sjo-derived DO compared to control-derived DO (Figure 1G). Last, we performed preliminary experiments to develop immuno-organoids (IOSG) by adding PBMC to the DO (Figure 1H). Additional experiments are currently underway to develop and characterize IOSG. Conclusion: We established a culture system of SG organoids from MSGB from Sjo patients and controls with long term expansion, maturation markers expression and functionality. Additional investigations are ongoing to further characterize the organoids and to develop IOSG that will be used to study the crosstalk between epithelial cells and immune cells and to test the effect of drugs on this crosstalk. REFERENCES: NIL. Acknowledgements: NIL. Disclosure of Interests: None declared.Figure 1

The monkeypox virus (MPXV) outbreak of 2022 caused a human disease with unusual epidemiological and clinical features, notably an increase in human-to-human transmission through sexual contact, predominantly among men who have sex with men (MSM). This evolution underscores the need to reassess prevention and control strategies in the context of a sexually transmitted disease. Here, we show that rectal challenge of cynomolgus macaques with a 2022 clade IIb MPXV isolate mimics sexual transmission, leading to rectal infection, with systemic and male genital tract dissemination and seminal fluid shedding. Vaccination with modified-vaccinia Ankara (MVA) protected the macaques from subsequent rectal MPXV challenge. However, MVA failed to prevent the disease when administered four days post-exposure to MPXV. These findings have a critical impact on outbreak management and highlight the importance of reevaluating MVA post-exposure prophylaxis protocols.

Early pregnancy Zika virus (ZIKV) infection is associated with major brain damage in fetuses, leading to microcephaly in 0.6–5.0

The coronavirus disease 2019 (COVID-19) due to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has shown that, except vaccination, few therapeutics options for its treatment or prevention are available. Among the pathways that can be targeted for COVID-19 treatment, the Keap1/Nrf2 pathway seems of high interest as it regulates redox homeostasis and inflammation that are altered during SARS-CoV-2 infection. Here, we use three potent activators of the Keap1/Nrf2 pathway and showed that Sulfodyne®, a stabilized natural Sulforaphane preparation with optimal bioavailability, had the highest antiviral activity in pulmonary or colonic epithelial cell lines even when added late after SARS-CoV-2 infection. This antiviral activity was not dependent on NRF2 activity but associated with action on ER stress and mTOR signaling that are activated during SARS-CoV-2 infection. Sulfodyne® also decreased the inflammatory response of epithelial cell lines infected by SARS-CoV-2 independently of SARS-CoV-2 replication and reduced the activation of human monocytes that are recruited after infection of epithelial cells by SARS-CoV-2. Administration of Sulfodyne® had little effects on SARS-CoV-2 replication in mice and hamsters infected with SARS-CoV-2 but significantly reduced weight loss and disease severity. Altogether, these results pinpoint the natural compound Sulfodyne® as a potent therapeutic agent of COVID-19 symptomatology.Author Summary Accumulating evidence shows that oxidative stress coupled with the systemic inflammation contribute to COVID-19 pathogenesis. As the Keap1/Nrf2 pathway is the major regulator of redox homeostasis and promotes resolution of inflammation and as lung biopsies from COVID-19 patients showed a decreased NRF2 target gene signature, pharmacological agents that are known to activate NRF2 are good candidates for COVID-19 treatment. We show herein that Sulfodyne®, an NRF2 activator that consists in a stabilized Sulforaphane preparation with optimal bioavailability, impairs SARS-CoV-2 replication in colonic or pulmonary epithelial cells. We show that this antiviral activity of Sulfodyne® is not dependent of NRF2 activation, characterize the pathways associated with the Sulfodyne® antiviral activity and show that Sulfodyne® displays multiple actions that result in a decrease of the inflammation associated with SARS-CoV-2 infection. Finally, we show that Sulfodyne® decreases the pathogenesis of mice or hamster infected with SARS-CoV-2. Overall, this study provides mechanistic explanations of the action of Sulfodyne® during SARS-CoV-2 infection and suggests that Sulfodyne® is a potential therapeutic agent of COVID-19 pathogenesis.### Competing Interest StatementThe authors have declared no competing interest.

Abstract The fight against COVID-19 requires mass vaccination strategies, and vaccines inducing durable cross-protective responses are still needed. Inactivated vaccines have proven lasting efficacy against many pathogens and good safety records. They contain multiple protein antigens that may improve response breadth and can be easily adapted every year to maintain preparedness for future seasonally emerging variants. Here we report the immunogenicity and efficacy of VLA2001 in animal models, the first inactivated whole virus COVID-19 vaccine that has received standard marketing authorization by the European Medicines Agency. VLA2001 formulated with alum and the TLR9 agonist CpG 1018™ adjuvant generated a Th1-biased immune response and serum neutralizing antibodies in BALB/c mice. In non-human primates, two injections of VLA2001 were sufficient to induce specific and polyfunctional T cell responses, predominantly Th1-biased, and high levels of antibodies neutralizing SARS-CoV-2 infection in cell culture. These antibodies also inhibited the binding of the Spike protein to human ACE2 receptor of several variants of concern most resistant to neutralization. After exposure to a high dose of SARS-CoV-2, all vaccinated groups of cynomolgus macaques exhibited significant levels of protection from viral replication in the upper and lower respiratory tracts and from lung tissue inflammation as compared to controls.

Abstract The fight against COVID-19 requires mass vaccination strategies, and vaccines inducing durable cross-protective responses are still needed. Inactivated vaccines have proven lasting efficacy against many pathogens and good safety records. They contain multiple protein antigens that may improve response breadth and can be easily adapted every year to maintain preparedness for future seasonally emerging variants. Here we report the immunogenicity and efficacy of VLA2001 in animal models, the first inactivated whole virus COVID-19 vaccine that has received standard marketing authorization by the European Medicines Agency. VLA2001 formulated with alum and the TLR9 agonist CpG 1018™ adjuvant generated a Th1-biased immune response and serum neutralizing antibodies in BALB/c mice. In non-human primates, two injections of VLA2001 were sufficient to induce specific and polyfunctional T cell responses, predominantly Th1-biased, and high levels of antibodies neutralizing SARS-CoV-2 infection in cell culture. These antibodies also inhibited the binding of the Spike protein to human ACE2 receptor of several variants of concern most resistant to neutralization. After exposure to a high dose of SARS-CoV-2, all vaccinated groups of cynomolgus macaques exhibited significant levels of protection from viral replication in the upper and lower respiratory tracts and from lung tissue inflammation as compared to controls.

Bat sarbecovirus BANAL‐236 is highly related to SARS‐CoV‐2 and infects human cells, albeit lacking the furin cleavage site in its spike protein. BANAL‐236 replicates efficiently and pauci‐symptomatically in humanized mice and in macaques, where its tropism is enteric, strongly differing from that of SARS‐CoV‐2. BANAL‐236 infection leads to protection against superinfection by a virulent strain. We find no evidence of antibodies recognizing bat sarbecoviruses in populations in close contact with bats in which the virus was identified, indicating that such spillover infections, if they occur, are rare. Six passages in humanized mice or in human intestinal cells, mimicking putative early spillover events, select adaptive mutations without appearance of a furin cleavage site and no change in virulence. Therefore, acquisition of a furin site in the spike protein is likely a pre‐spillover event that did not occur upon replication of a SARS‐CoV‐2‐like bat virus in humans or other animals. Other hypotheses regarding the origin of the SARS‐CoV‐2 should therefore be evaluated, including the presence of sarbecoviruses carrying a spike with a furin cleavage site in bats.

Bat sarbecovirus BANAL-236 is highly related to SARS-CoV-2 and infects human cells, albeit lacking the furin cleavage site in its spike protein. To inform on the origin of SARS-CoV-2, we evaluated the clinical, epidemiological and evolutionary consequences of a potential BANAL-236 spillover into humans using animal models. The virus replicates efficiently and pauci-symptomatically in humanized mice and in macaques, where its tropism is enteric, strongly differing from that of SARS-CoV-2. BANAL-236 infection leads to protection against superinfection by a more virulent strain like Wuhan SARS-CoV-2. Yet we found no evidence of antibodies recognizing bat sarbecoviruses in populations highly exposed to bats, indicating that such infections, if they occur, are rare. Six passages in mice or in human intestinal cells, mimicking putative early spillover events, selected adaptive mutations without appearance of a furin cleavage site and not change in virulence. We thus conclude that the hypothesis of the SARS-CoV-2 pandemic being preceded by silent circulation in humans of BANAL-236-like strains leading to the acquisition of a furin cleavage site is unlikely. Our studies suggest that a specific search for a furin cleavage site in sarbecoviruses in the wild should be pursued to understand the origin of the SARS-CoV-2 pandemics.

Older age is a major risk factor for severe coronavirus disease (COVID-19) (1). Understanding the biological mechanisms linking age to the pathogenesis of COVID-19 is essential for developing preventive and therapeutic strategies. We hypothesized that cell senescence, a basic aging process that plays a pivotal role in health deterioration and diseases, particularly those targeting the lung (2), is involved in the pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–induced lung disease, including the development of long-lasting lung alterations. Senescent cells exhibit a stable proliferation arrest and acquire a specific senescence-associated secretory phenotype characterized by the release of inflammatory cytokines, immune modulators, proteases, profibrotic factors, and various effectors that can alter tissue organization and function (3). Senescence is triggered by a myriad of stressors that promote a DNA damage response leading to p53-dependent upregulation of the CDK inhibitor p21 and/or expression of p16, which is used as a reliable marker of senescent cells. Cell senescence is pivotal in age-associated lung diseases, notably lung emphysema, fibrosis, and chronic obstructive pulmonary disease (2, 4–6). In recent studies, SARS-CoV2 Spike protein-1 was shown to exacerbate the senescence-associated secretory phenotype of human senescent cells, thereby contributing to the exuberant inflammatory response seen in severe COVID-19. Targeting senescent cells using senolytic drugs reduced mortality in old mice infected with a mouse b-coronavirus (7). To further evaluate potential links between SARS-CoV-2 infection and cell senescence, we analyzed publicly available single-cell RNA sequencing data sets obtained using BAL fluid (BALF) cells from patients with moderate or severe-to-critical COVID-19 (8). We also monitored lung cell senescence in SARS-CoV-2–infected macaques, which constitute a relevant model for studying human COVID-19 (9). First, we extracted data from publicly available, BALF cell, single-cell RNA sequencing data sets from patients with moderate or severe-to-critical COVID-19 versus healthy control subjects to analyze senescence-related genes (8). In BALFs collected 10–16 days after symptom onset, mRNA of the senescence marker CDKN2A encoding p16 was mainly detected in epithelial cells, macrophages, and T cells, with higher levels in epithelial cells from patients with severe-to-critical disease compared with control subjects (Figure 1A). Expression of the senescence markers CDKN2A, CDKN1A (encoding p21), uPAR (urokinase plasminogen activator surface receptor), CXCL8, IGFBP3, andGDF15was significantly increased in epithelial ciliated and club cells from patients with severe COVID-19 compared with those with moderate disease and with healthy control subjects, suggesting that lung cell senescence induction coincided with virus detection (Figure 1B). Of note, patients with severe COVID-19 were older than those with moderate disease, whereas age was comparable between patients with moderate disease and healthy control subjects (Figure 1). In single-cell data sets from another study (10), which compared same-age patients with mild versus critical disease (see Fig. E1 in the data supplement), variations were similar, although CDKN1A and CDKN2Awere less affected than in the first data set. To further assess the extent of SARS-CoV-2–induced lung cell senescence and the fate of senescent lung cells over time, we investigated macaques at 4 and 30 dpi, or in other words, at the viral load peak and at the first negative airway sample qRT-PCR, respectively (9). Immunohistochemical studies of lung sections at 4 dpi revealed SARS-CoV-2 antigen–stained cells, including lung endothelial cells (ECs) and parenchymal cells, as well as numerous p16and p21-immunofluorescence–stained cells predominating at sites of alveolar damage (Figure 2A). Cells positive for p16 were also positive for SARS-CoV-2 Spike protein-1 at 4 dpi, indicating that senescent lung cells were infected with the virus. SARS-CoV-2 antigen–stained cells were rarer at 30 dpi, whereas massive accumulation of p16and p21-positive cells throughout the lung indicated persistence of senescent lung cells after virus clearance (Figure 2A). Cells stained for p16 were also stained for the DNA damage markers g-H2AX protein and p53-binding protein 1 at both 4 and 30 dpi (Figure 2A). Interestingly, the lungs at 30 dpi no longer exhibited the consolidated parenchymal areas seen at 4 dpi but showed extensive lung parenchyma remodeling, with thickening of the alveolar and pulmonary vessel walls and abundant extracellular matrix deposits as assessed by collagen staining (Figure 2B and Figure E2). These advanced lesions were accompanied by massive accumulation of p16and p21-positive cells, most of which were alveolar type II cells and ECs, as shown by double immunofluorescence staining for p16 and mucin 1 and for vonWillebrand factor, respectively (Figure 2B). Of note, most ECs stained for p16 in many lung vessels, notably those occluded by thrombi and showing intraluminal vonWillebrand factor and fibrin staining. Collectively, our data constitute the first evidence of temporal and topographic relations between senescent cell accumulation and pulmonary lesions induced by SARS-CoV-2. Cell senescence is usually viewed as a response to chronic stressors that severely impedes healthy aging and promotes agerelated noncommunicable diseases (11). Here, BALF cells from patients with severe COVID-19 expressed high levels of senescent This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0. For commercial usage and reprints, please e-mail Diane Gern (dgern@thoracic.org).

The COVID-19 pandemic has exemplified that rigorous evaluation in large animal models is key for translation from promising in vitro results to successful clinical implementation. Among the drugs that have been largely tested in clinical trials but failed so far to bring clear evidence of clinical efficacy is favipiravir, a nucleoside analogue with large spectrum activity against several RNA viruses in vitro and in small animal models. Here, we evaluate the antiviral activity of favipiravir against Zika or SARS-CoV-2 virus in cynomolgus macaques. In both models, high doses of favipiravir are initiated before infection and viral kinetics are evaluated during 7 to 15 days after infection. Favipiravir leads to a statistically significant reduction in plasma Zika viral load compared to untreated animals. However, favipiravir has no effects on SARS-CoV-2 viral kinetics, and 4 treated animals have to be euthanized due to rapid clinical deterioration, suggesting a potential role of favipiravir in disease worsening in SARS-CoV-2 infected animals. To summarize, favipiravir has an antiviral activity against Zika virus but not against SARS-CoV-2 infection in the cynomolgus macaque model. Our results support the clinical evaluation of favipiravir against Zika virus but they advocate against its use against SARS-CoV-2 infection.

Chikungunya virus (CHIKV) is a reemerging mosquito-borne alphavirus responsible for numerous outbreaks. Chikungunya can cause debilitating acute and chronic disease. Thus, the development of a safe and effective CHIKV vaccine is an urgent global health priority. This study evaluated the effectiveness of the live-attenuated CHIKV vaccine VLA1553 against WT CHIKV infection by using passive transfer of sera from vaccinated volunteers to nonhuman primates (NHP) subsequently exposed to WT CHIKV and established a serological surrogate of protection. We demonstrated that human VLA1553 sera transferred to NHPs conferred complete protection from CHIKV viremia and fever after challenge with homologous WT CHIKV. In addition, serum transfer protected animals from other CHIKV-associated clinical symptoms and from CHIKV persistence in tissue. Based on this passive transfer study, a 50% micro–plaque reduction neutralization test titer of ≥ 150 was determined as a surrogate of protection, which was supported by analysis of samples from a seroepidemiological study. In conclusion, considering the unfeasibility of an efficacy trial due to the unpredictability and explosive, rapidly moving nature of chikungunya outbreaks, the definition of a surrogate of protection for VLA1553 is an important step toward vaccine licensure to reduce the medical burden caused by chikungunya.

The COVID-19 pandemic continues to have devastating consequences on health and economy, even after the approval of safe and effective vaccines. Waning immunity, the emergence of variants of concern, breakthrough infections, and lack of global vaccine access and acceptance perpetuate the epidemic. Here, we demonstrate that a single injection of an adenoassociated virus (AAV)-based COVID-19 vaccine elicits at least 17-month-long neutralizing antibody responses in non-human primates at levels that were previously shown to protect from viral challenge. To improve the scalability of this durable vaccine candidate, we further optimized the vector design for greater potency at a reduced dose in mice and non-human primates. Finally, we show that the platform can be rapidly adapted to other variants of concern to robustly maintain immunogenicity and protect from challenge. In summary, we demonstrate this class of AAV can provide durable immunogenicity, provide protection at dose that is low and scalable, and be adapted readily to novel emerging vaccine antigens thus may provide a potent tool in the ongoing fight against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).

Summary: Non-human primates (NHPs) are particularly relevant as preclinical models for SARS-CoV-2 infection and nuclear imaging may represent a valuable tool for monitoring infection in this species. We investigated the benefit of computed X-ray tomography (CT) and [18F]-FDG positron emission tomography (PET) to monitor the early phase of the disease in a large cohort (n = 76) of SARS-CoV-2 infected macaques.Following infection, animals showed mild COVID-19 symptoms including typical lung lesions. CT scores at the acute phase reflect the heterogeneity of lung burden following infection. Moreover, [18F]-FDG PET revealed that FDG uptake was significantly higher in the lungs, nasal cavities, lung-draining lymph nodes, and spleen of NHPs by 5 days postinfection compared to pre-infection levels, indicating early local inflammation. The comparison of CT and PET data from previous COVID-19 treatments or vaccines we tested in NHP, to this large cohort of untreated animals demonstrated the value of in vivo imaging in preclinical trials.

Older age is an important risk factor for severe COVID-19 disease. Understanding the biological mechanisms that link aging to the pathogenesis of COVID-19 is essential for developing of therapeutic strategies. We hypothesized that cell senescence, a basic aging process that plays a pivotal role in lung diseases, is involved in the pathogenesis of COVID-19 including the development of long-lasting lung alterations. To evaluate the impact of SARS-CoV-2 infection on cell senescence, we (1) analyzed publicly available datasets of scRNA-seq performed in BALF cells from patients with moderate or severe/critical COVID-19; (2) investigated lung samples from cynomolgus macaques infected with 10 6 pfu of a SARS-CoV-2 clinical isolate. Two macaques were sacrificed at 4 days post-infection (dpi.) and two others at 30 dpi. In BALF obtained within 10 days after symptom onset, the expression of several senescence markers, i.e., CDKN2A, CDKN1A (encoding p21), uPAR, CXCL8, IGFBP3, and GDF15 was significantly increased in epithelial cells in BALF from patients with severe COVID-19, suggesting that lung-cell senescence induction was contemporary of viral detection. Next, we investigated macaques at 4 and 30 dpi, corresponding respectively to the viral load peak and to the absence of detectable viral RNA in BALF (1). Immunohistochemical analysis revealed numerous SARS-CoV-2 antigen-stained cells, also co-stained for senescence markers p16- and p21. The lungs at 30 dpi no longer contained the consolidated parenchymal areas seen at 4 dpi but showed extensive lung parenchyma remodelling, with thickening of the alveoli and pulmonary vessel walls and abundant extracellular matrix deposits as assessed by collagen staining. These lesions were accompanied with massive accumulation of p16- and p21-positive cells, mostly pneumocytes II and ECs. Of note, p16 staining of most ECs was seen in pulmonary vessels, notably those occluded by thrombosis and showing intraluminal vWF staining. Cells stained for p16 were alsostained for the DNA damage markers γ-H2AX protein and p53-binding protein [1] . Our data constitute the first evidence of temporal and topographic relations between senescent-cell accumulation and pulmonary lesions induced by SARS-CoV-2 infection.