Lung-Targeted Transgene Expression of Nanocomplexed Ad5 Enhances Immune Response in the Presence of Preexisting Immunity.

Recombinant adenovirus serotype 5 (Ad5) vector has been widely applied in vaccine development targeting infectious diseases, such as Ebola virus disease and coronavirus disease 2019 (COVID-19). However, the high prevalence of preexisting anti-vector immunity compromises the immunogenicity of Ad5-based vaccines. Thus, there is a substantial unmet need to minimize preexisting immunity while improving the insert-induced immunity of Ad5 vectors. Herein, we address this need by utilizing biocompatible nanoparticles to modulate Ad5-host interactions. We show that positively charged human serum albumin nanoparticles ((+)HSAnp), which are capable of forming a complex with Ad5, significantly increase the transgene expression of Ad5 in both coxsackievirus-adenovirus receptor-positive and -negative cells. Furthermore, in charge- and dose-dependent manners, Ad5/(+)HSAnp complexes achieve robust (up to 227-fold higher) and long-term (up to 60 days) transgene expression in the lungs of mice following intranasal instillation. Importantly, in the presence of preexisting anti-Ad5 immunity, complexed Ad5-based Ebola and COVID-19 vaccines significantly enhance antigen-specific humoral response and mucosal immunity. These findings suggest that viral aggregation and charge modification could be leveraged to engineer enhanced viral vectors for vaccines and gene therapies.

Highly pathogenic coronavirus N protein aggravates inflammation by MASP-2-mediated lectin complement pathway overactivation.

Excessive inflammatory responses contribute to the pathogenesis and lethality of highly pathogenic human coronaviruses, but the underlying mechanism remains unclear. In this study, the N proteins of highly pathogenic human coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV), middle east respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), were found to bind MASP-2, a key serine protease in the lectin pathway of complement activation, resulting in excessive complement activation by potentiating MBL-dependent MASP-2 activation, and the deposition of MASP-2, C4b, activated C3 and C5b-9. Aggravated inflammatory lung injury was observed in mice infected with adenovirus expressing the N protein. Complement hyperactivation was also observed in SARS-CoV-2-infected patients. Either blocking the N protein:MASP-2 interaction, MASP-2 depletion or suppressing complement activation can significantly alleviate N protein-induced complement hyperactivation and lung injury in vitro and in vivo. Altogether, these data suggested that complement suppression may represent a novel therapeutic approach for pneumonia induced by these highly pathogenic coronaviruses.

Detection of SARS-CoV-2 in Simultaneously Collected Tear and Throat Swab Samples from the Patients with 2019- new SARS-CoV-2 Infection Disease: A Single Center Cross-sectional Study.

Objecive: This study aimed to evaluate whether Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) can be detected in the tears in the eyes of Coronavirus Disease 2019 (COVID-2019) infected patients and compare the detection consistency of two simultaneously collected samples, from the tears and throat swabs.Methods: A total of 35 COVID-2019 patients were included in this cross-sectional case series study. Throat samples from all enrolled patients were collected with sampling swab, and simultaneously, tear samples were collected with sampling swab from 9 patients (No.1-9) and with Schirmer's strip from the remaining patients (No.10-35) (bilateral eyes for all patients). Sample collecting and testing were performed in three separate time points: first from patients No.1-9, second from patients No.10-29, and third from patients No. 30-35. Reverse-transcriptase-polymerase-chain-reaction (RT-PCR) assay was performed.Results: Among enrolled patients, 29 (No.1-29) had mild or moderate clinical symptoms and 6 (No.30-35) had severe symptoms. The mean time interval from the sample collection day to diagnosis confirmation day was 9.71 ± 6.50 days (ranged from 3 to 29 days). None of the patients had conjunctivitis. Nineteen out of 35 (54.3%) throat samples presented positive Rt-PCR results. Three (no.13,21,31) out of 35 (8.6%) tear samples presented positive RT-PCR results. Two (no.21, 31) of these three patients were throat swab positive and one (No. 13) was negative. The consistency analysis indicated that tears and throat samples showed poor consistency (Kappa = -0.009, P = .9).The cycle threshold value (Ct-value) of tear samples collected by sampling swab was significantly higher than that by Schirmer's strip (t = 2.288, P = .03).Conclusion: In spite of the low SARS-CoV-2 positive detection rate of tear samples from COVID-2019 patients, we cannot fully rule out the transmission by ocular surface. Whether tear testing can be used as an aid in judging of SARS-CoV-2 infection need further investigation.

Erratum: Author Correction: Immune cell profiling of COVID-19 patients in the recovery stage by single-cell sequencing.

[This corrects the article DOI: 10.1038/s41421-020-0168-9.].

Immune cell profiling of COVID-19 patients in the recovery stage by single-cell sequencing.

COVID-19, caused by SARS-CoV-2, has recently affected over 1,200,000 people and killed more than 60,000. The key immune cell subsets change and their states during the course of COVID-19 remain unclear. We sought to comprehensively characterize the transcriptional changes in peripheral blood mononuclear cells during the recovery stage of COVID-19 by single-cell RNA sequencing technique. It was found that T cells decreased remarkably, whereas monocytes increased in patients in the early recovery stage (ERS) of COVID-19. There was an increased ratio of classical CD14++ monocytes with high inflammatory gene expression as well as a greater abundance of CD14++IL1ß+ monocytes in the ERS. CD4+ T cells and CD8+ T cells decreased significantly and expressed high levels of inflammatory genes in the ERS. Among the B cells, the plasma cells increased remarkably, whereas the naïve B cells decreased. Several novel B cell-receptor (BCR) changes were identified, such as IGHV3-23 and IGHV3-7, and isotypes (IGHV3-15, IGHV3-30, and IGKV3-11) previously used for virus vaccine development were confirmed. The strongest pairing frequencies, IGHV3-23-IGHJ4, indicated a monoclonal state associated with SARS-CoV-2 specificity, which had not been reported yet. Furthermore, integrated analysis predicted that IL-1ß and M-CSF may be novel candidate target genes for inflammatory storm and that TNFSF13, IL-18, IL-2, and IL-4 may be beneficial for the recovery of COVID-19 patients. Our study provides the first evidence of an inflammatory immune signature in the ERS, suggesting COVID-19 patients are still vulnerable after hospital discharge. Identification of novel BCR signaling may lead to the development of vaccines and antibodies for the treatment of COVID-19.