A longitudinal study of humoral immune responses induced by a 3-dose inactivated COVID-19 vaccine in an observational, prospective cohort.

BACKGROUND: To determine the dynamic SARS-CoV-2 specific antibody levels induced by 3 doses of an inactivated COVID-19 vaccine, CoronaVac. An observational, prospective cohort study was performed with 93 healthy healthcare workers from a tertiary hospital in Nanjing, China. Serum SARS-CoV-2 specific IgM, IgG, and neutralizing antibodies (NAb) were measured at different time points among participants who received 3 doses of inactivated COVID-19 vaccine. RESULTS: 91.3% (85/93) and 100% (72/72) participants showed positive both for SARS-CoV-2 specific IgG and NAb after 2-dose CoronaVac and after 3-dose CoronaVac, respectively. Anti-SARS-CoV-2 IgG responses reached 91.21 (55.66-152.06) AU/mL, and surrogate NAb was 47.60 (25.96-100.81) IU/mL on day 14 after the second dose. Anti-SARS-CoV-2 IgG responses reached 218.29 (167.53-292.16) AU/mL and surrogate NAb was 445.54 (171.54-810.90) IU/mL on day 14 after the third dose. Additionally, SARS-CoV-2 specific surrogate neutralizing antibody titers were highly correlated with serum neutralization activities against Ancestral, Omicron, and Delta strains. Moreover, significantly higher SARS-CoV-2 IgG responses, but not NAb responses, were found in individuals with breakthrough infection when compared to that of 3-dose CoronaVac recipients. CONCLUSIONS: CoronaVac elicited robust SARS-CoV-2 specific humoral responses. Surrogate NAb assay might substitute for pseudovirus neutralization assay. Monitoring SARS-CoV-2 antibody responses induced by vaccination would provide important guidance for the optimization of COVID-19 vaccines.

Friend or Foe? Implication of the autophagy-lysosome pathway in SARS-CoV-2 infection and COVID-19.

There is increasing amount of evidence indicating the close interplays between the replication cycle of SARS-CoV-2 and the autophagy-lysosome pathway in the host cells. While autophagy machinery is known to either assist or inhibit the viral replication process, the reciprocal effects of the SARS-CoV-2 on the autophagy-lysosome pathway have also been increasingly appreciated. More importantly, despite the disappointing results from the clinical trials of chloroquine and hydroxychloroquine in treatment of COVID-19, there is still ongoing effort in discovering new therapeutics targeting the autophagy-lysosome pathway. In this review, we provide an update-to-date summary of the interplays between the autophagy-lysosome pathway in the host cells and the pathogen SARS-CoV-2 at the molecular level, to highlight the prognostic value of autophagy markers in COVID-19 patients and to discuss the potential of developing novel therapeutic strategies for COVID-19 by targeting the autophagy-lysosome pathway. Thus, understanding the nature of such interactions between SARS-CoV-2 and the autophagy-lysosome pathway in the host cells is expected to provide novel strategies in battling against this global pandemic.

Autophagy in health and disease: From molecular mechanisms to therapeutic target.

Macroautophagy/autophagy is an evolutionally conserved catabolic process in which cytosolic contents, such as aggregated proteins, dysfunctional organelle, or invading pathogens, are sequestered by the double-membrane structure termed autophagosome and delivered to lysosome for degradation. Over the past two decades, autophagy has been extensively studied, from the molecular mechanisms, biological functions, implications in various human diseases, to development of autophagy-related therapeutics. This review will focus on the latest development of autophagy research, covering molecular mechanisms in control of autophagosome biogenesis and autophagosome-lysosome fusion, and the upstream regulatory pathways including the AMPK and MTORC1 pathways. We will also provide a systematic discussion on the implication of autophagy in various human diseases, including cancer, neurodegenerative disorders (Alzheimer disease, Parkinson disease, Huntington's disease, and Amyotrophic lateral sclerosis), metabolic diseases (obesity and diabetes), viral infection especially SARS-Cov-2 and COVID-19, cardiovascular diseases (cardiac ischemia/reperfusion and cardiomyopathy), and aging. Finally, we will also summarize the development of pharmacological agents that have therapeutic potential for clinical applications via targeting the autophagy pathway. It is believed that decades of hard work on autophagy research is eventually to bring real and tangible benefits for improvement of human health and control of human diseases.

The Third dose of CoronVac vaccination induces broad and potent adaptive immune responses that recognize SARS-CoV-2 Delta and Omicron variants.

The waning humoral immunity and emerging contagious severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants resulted in the necessity of the booster vaccination of coronavirus disease 2019 (COVID-19). The inactivated vaccine, CoronaVac, is the most widely supplied COVID-19 vaccine globally. Whether the CoronaVac booster elicited adaptive responses that cross-recognize SARS-CoV-2 variants of concern (VoCs) among 77 healthy subjects receiving the third dose of CoronaVac were explored. After the boost, remarkable elevated spike-specific IgG and IgA responses, as well as boosted neutralization activities, were observed, despite 3.0-fold and 5.9-fold reduced neutralization activities against Delta and Omicron strains compared to that of the ancestral strain. Furthermore, the booster dose induced potent B cells and memory B cells that cross-bound receptor-binding domain (RBD) proteins derived from VoCs, while Delta and Omicron RBD-specific memory B cell recognitions were reduced by 2.7-fold and 4.2-fold compared to that of ancestral strain, respectively. Consistently, spike-specific circulating follicular helper T cells (cTfh) significantly increased and remained stable after the boost, with a predominant expansion towards cTfh17 subpopulations. Moreover, SARS-CoV-2-specific CD4+ and CD8+ T cells peaked and sustained after the booster. Notably, CD4+ and CD8+ T cell recognition of VoC spike was largely preserved compared to the ancestral strain. Individuals without generating Delta or Omicron neutralization activities had comparable levels of CD4+ and CD8+ T cells responses as those with detectable neutralizing activities. Our study demonstrated that the CoronaVac booster induced broad and potent adaptive immune responses that could be effective in controlling SARS-CoV-2 Delta and Omicron variants.

Self-reported symptom study of COVID-19 chemosensory dysfunction in Malaysia.

Alterations in the three chemosensory modalities-smell, taste, and chemesthesis-have been implicated in Coronavirus Disease 2019 (COVID-19), yet emerging data suggest a wide geographic and ethnic variation in the prevalence of these symptoms. Studies on chemosensory disorders in COVID-19 have predominantly focused on Caucasian populations whereas Asians remain understudied. We conducted a nationwide, multicentre cross-sectional study using an online questionnaire on a cohort of RT-PCR-confirmed adult COVID-19 patients in Malaysia between 6 June and 30 November 2020. The aim of our study was to investigate their presenting symptoms and assess their chemosensory function using self-ratings of perceived smell, taste, chemesthesis, and nasal blockage. In this cohort of 498 patients, 41.4% reported smell and/or taste loss when diagnosed with COVID-19, which was the commonest symptom. Blocked nose, loss of appetite, and gastrointestinal disturbances were independent predictors of smell and/or taste loss on multivariate analysis. Self-ratings of chemosensory function revealed a reduction in smell, taste, and chemesthesis across the entire cohort of patients that was more profound among those reporting smell and/or taste loss as their presenting symptom. Perceived nasal obstruction accounted for only a small proportion of changes in smell and taste, but not for chemesthesis, supporting viral disruption of sensorineural mechanisms as the dominant aetiology of chemosensory dysfunction. Our study suggests that chemosensory dysfunction in COVID-19 is more widespread than previously reported among Asians and may be related to the infectivity of viral strains.Study Registration: NMRR-20-934-54803 and NCT04390165.

Dynamic SARS-CoV-2-specific B-cell and T-cell responses following immunization with an inactivated COVID-19 vaccine.

OBJECTIVE: The dynamic adaptive immune responses elicited by the inactivated virus vaccine CoronaVac remain elusive. METHODS: In a prospective cohort of 100 healthcare professionals naïve for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) who received two doses of CoronaVac, we analysed SARS-CoV-2-specific humoral and cellular responses at four different timepoints, including before vaccination (T1), 2 weeks after the first dose (T2), 2 weeks after the booster dose (T3), and 8-10 weeks after the booster dose (T4). SARS-CoV-2-specific antibodies, serum neutralizing activities, peripheral B cells, CD4+ and CD8+ T cells and their memory subsets were simultaneously measured in this cohort. RESULTS: SARS-CoV-2 spike-specific IgG responses reached a peak (geometric mean titre (GMT) 54827, 30969-97065) after two doses and rapidly declined (GMT 502, 212-1190) at T4, whereas suboptimal IgA responses were detected (GMT 5, 2-9). Spike-specific circulating B cells (0.60%, 0.46-0.73% of total B cells) and memory B cells (1.18%, 0.92-1.44% of total memory B cells) were effectively induced at T3 and sustained over time (0.33%, 0.23-0.43%; 0.87%, 0.05-1.67%, respectively). SARS-CoV-2-specific circulating CD4+ T cells (0.57%, 0.47-0.66%) and CD8+ T cells (1.29%, 1.04-1.54%) were detected at T3. At T4, 0.78% (0.43-1.20%) of memory CD4+ T cells and 0.68% (0.29-1.30%) of memory CD8+ T cells were identified as SARS-CoV-2-specific, while 0.62% (0.51-0.75%) of CD4+ T cells and 0.47% (0.38-0.58%) of CD8+ T cells were SARS-CoV-2-specific terminally differentiated effector memory cells. Furthermore, age and interval between doses affected the magnitude of CoronaVac-induced immune responses. SARS-CoV-2 memory CD4+ T cells were strongly associated with both receptor binding domain (RBD)-specific memory B cells (r 0.87, p <0.0001) and SARS-CoV-2-specific memory CD8+ T cells (r 0.48, p <0.0001). CONCLUSIONS: CoronaVac induced robust circulating and memory B cell and T cell responses. Our study offers new insight into the underlying immunobiology of inactivated virus vaccines in humans and may have implications for vaccine strategies in the future.

Hydroxychloroquine/Chloroquine as Therapeutics for COVID-19: Truth under the Mystery.

The outbreak of coronavirus disease-19 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly evolved into a global pandemic. One major challenge in the battle against this deadly disease is to find effective therapy. Due to the availability and proven clinical record of hydroxychloroquine (HCQ) and chloroquine (CQ) in various human diseases, there have been enormous efforts in repurposing these two drugs as therapeutics for COVID-19. To date, substantial amount of work at cellular, animal models and clinical trials have been performed to verify their therapeutic potential against COVID-19. However, neither lab-based studies nor clinical trials have provided consistent and convincing evidence to support the therapeutic value of HCQ/CQ in the treatment of COVID-19. In this mini review we provide a systematic summary on this important topic and aim to reveal some truth covered by the mystery regarding the therapeutic value of HCQ/CQ in COVID-19.

Implications of cardiac markers in risk-stratification and management for COVID-19 patients.

BACKGROUND: COVID-19 has resulted in high mortality worldwide. Information regarding cardiac markers for precise risk-stratification is limited. We aim to discover sensitive and reliable early-warning biomarkers for optimizing management and improving the prognosis of COVID-19 patients. METHODS: A total of 2954 consecutive COVID-19 patients who were receiving treatment from the Wuhan Huoshenshan Hospital in China from February 4 to April 10 were included in this retrospective cohort. Serum levels of cardiac markers were collected after admission. Coronary artery disease diagnosis and survival status were recorded. Single-cell RNA-sequencing and bulk RNA-sequencing from different cohorts of non-COVID-19 were performed to analyze SARS-CoV-2 receptor expression. RESULTS: Among 2954 COVID-19 patients in the analysis, the median age was 60 years (50-68 years), 1461 (49.5%) were female, and 1515 (51.3%) were severe/critical. Compared to mild/moderate (1439, 48.7%) patients, severe/critical patients showed significantly higher levels of cardiac markers within the first week after admission. In severe/critical COVID-19 patients, those with abnormal serum levels of BNP (42 [24.6%] vs 7 [1.1%]), hs-TNI (38 [48.1%] vs 6 [1.0%]), α- HBDH (55 [10.4%] vs 2 [0.2%]), CK-MB (45 [36.3%] vs 12 [0.9%]), and LDH (56 [12.5%] vs 1 [0.1%]) had a significantly higher mortality rate compared to patients with normal levels. The same trend was observed in the ICU admission rate. Severe/critical COVID-19 patients with pre-existing coronary artery disease (165/1,155 [10.9%]) had more cases of BNP (52 [46.5%] vs 119 [16.5%]), hs-TNI (24 [26.7%] vs 9.6 [%], α- HBDH (86 [55.5%] vs 443 [34.4%]), CK-MB (27 [17.4%] vs 97 [7.5%]), and LDH (65 [41.9%] vs 382 [29.7%]), when compared with those without coronary artery disease. There was enhanced SARS-CoV-2 receptor expression in coronary artery disease compared with healthy controls. From regression analysis, patients with five elevated cardiac markers were at a higher risk of death (hazards ratio 3.4 [95% CI 2.4-4.8]). CONCLUSIONS: COVID-19 patients with pre-existing coronary artery disease represented a higher abnormal percentage of cardiac markers, accompanied by high mortality and ICU admission rate. BNP together with hs-TNI, α- HBDH, CK-MB and LDH act as a prognostic biomarker in COVID-19 patients with or without pre-existing coronary artery disease.

A comprehensive, longitudinal analysis of humoral responses specific to four recombinant antigens of SARS-CoV-2 in severe and non-severe COVID-19 patients.

There is an urgent need for effective treatment and preventive vaccine to contain this devastating global pandemic, which requires a comprehensive understanding of humoral responses specific to SARS-CoV-2 during the disease progression and convalescent phase of COVID-19 patients. We continuously monitored the serum IgM and IgG responses specific to four SARS-CoV-2 related antigens, including the nucleoprotein (NP), receptor binding domain (RBD), S1 protein, and ectodomain (ECD) of the spike protein among non-severe and severe COVID-19 patients for seven weeks since disease onset. Most patients generated humoral responses against NP and spike protein-related antigens but with their distinct kinetics profiles. Combined detection of NP and ECD antigens as detecting antigen synergistically improved the sensitivity of the serological assay, compared to that of using NP or RBD as detection antigen. 80.7% of convalescent sera from COVID-19 patients revealed that the varying extents of neutralization activities against SARS-CoV-2. S1-specific and ECD-specific IgA responses were strongly correlated with the neutralization activities in non-severe patients, but not in severe patients. Moreover, the neutralizing activities of the convalescent sera were shown to significantly decline during the period between 21 days to 28 days after hospital discharge, accompanied by a substantial drop in RBD-specific IgA response. Our data provide evidence that are crucial for serological testing, antibody-based intervention, and vaccine design of COVID-19.

Longitudinal anti-SARS-CoV-2 antibody profile and neutralization activity of a COVID-19 patient.

We followed-up a mild COVID-19 patient for 91 days and serially monitored his serum antibodies to four SARS-CoV-2 related antigens (NP, RBD, S1 and ECD) and neutralization activities. Our data revealed a profile of serial antibody responses during the progress and a quick decline of neutralization activities after discharge.

High SARS-CoV-2 antibody prevalence among healthcare workers exposed to COVID-19 patients.

The seroprevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was examined among 105 healthcare workers (HCWs) exposed to four patients who were laboratory confirmed with coronavirus disease 2019 (COVID-19), the disease caused by SARS-CoV-2 infection. These HCWs were immediately under quarantine for 14 days as soon as they were identified as close contacts. The nasopharyngeal swab samples were collected on the first and 14th day of the quarantine, while the serum samples were obtained on the 14th day of the quarantine. With the assay of enzyme immunoassay (EIA) and microneutralization assay, 17.14% (18/105) of HCWs were seropositive, while their swab samples were found to be SARS-CoV-2 RNA negative. Risk analysis revealed that wearing face mask could reduce the infection risk (odds ratio [OR], 0.127, 95% confidence interval [CI] 0.017, 0.968), while when exposed to COVID-19 patients, doctors might have higher risk of seroconversion (OR, 346.837, 95% CI 8.924, 13479.434), compared with HCWs exposed to colleagues as well as nurses and general service assistants who exposed to patients. Our study revealed that the serological testing is useful for the identification of asymptomatic or subclinical infection of SARS-CoV-2 among close contacts with COVID-19 patients.

Different longitudinal patterns of nucleic acid and serology testing results based on disease severity of COVID-19 patients.

Effective strategy to mitigate the ongoing pandemic of 2019 novel coronavirus (COVID-19) require a comprehensive understanding of humoral responses against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the emerging virus causing COVID-19. The dynamic profile of viral replication and shedding along with viral antigen specific antibody responses among COVID-19 patients started to be reported but there is no consensus on their patterns. Here, we conducted a serial investigation on 21 individuals infected with SARS-CoV-2 in two medical centres from Jiangsu Province, including 11 non-severe COVID-19 patients, and 5 severe COVID-19 patients and 5 asymptomatic carriers based on nucleic acid test and clinical symptoms. The longitudinal swab samples and sera were collected from these people for viral RNA testing and antibody responses, respectively. Our data revealed different pattern of seroconversion among these groups. All 11 non-severe COVID-19 patients and 5 severe COVID-19 patients were seroconverted during hospitalization or follow-up period, suggesting that serological testing is a complementary assay to nucleic acid test for those symptomatic COVID-19 patients. Of note, immediate antibody responses were identified among severe cases, compared to non-severe cases. On the other hand, only one were seroconverted for asymptomatic carriers. The SARS-CoV-2 specific antibody responses were well-maintained during the observation period. Such information is of immediate relevance and would assist COVID-19 clinical diagnosis, prognosis and vaccine design.

Targeting the Endocytic Pathway and Autophagy Process as a Novel Therapeutic Strategy in COVID-19.

Coronaviruses (CoVs) are a group of enveloped, single-stranded positive genomic RNA viruses and some of them are known to cause severe respiratory diseases in human, including Severe Acute Respiratory Syndrome (SARS), Middle East Respiratory Syndrome (MERS) and the ongoing coronavirus disease-19 (COVID-19). One key element in viral infection is the process of viral entry into the host cells. In the last two decades, there is increasing understanding on the importance of the endocytic pathway and the autophagy process in viral entry and replication. As a result, the endocytic pathway including endosome and lysosome has become important targets for development of therapeutic strategies in combating diseases caused by CoVs. In this mini-review, we will focus on the importance of the endocytic pathway as well as the autophagy process in viral infection of several pathogenic CoVs inclusive of SARS-CoV, MERS-CoV and the new CoV named as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and discuss the development of therapeutic agents by targeting these processes. Such knowledge will provide important clues for control of the ongoing epidemic of SARS-CoV-2 infection and treatment of COVID-19.