Association of Perceived Stress With Depression Among Vaccinated Healthcare Workers During the SARS-CoV-2 Variant Outbreak: The Mediating Role of Compassion Fatigue.

OBJECTIVE: Depression was common during coronavirus disease-2019 (COVID-19) pandemic, while the association of perceived stress with depression among vaccinated healthcare workers has not been investigated. This study aimed to address this issue. METHODS: We included a total of 898 fully vaccinated healthcare workers during the outbreak of severe acute respiratory syndrome coronavirus 2 Delta variant in Nanjing, 2021. Depression was ascertained by Patient Health Questionnaire-9, with a cut-off score of ≥5 indicative of mild-to-severe depression. Perceived stress, resilience, and compassion fatigue were assessed by Perceived Stress Scale-10, Resilience Scale-25, and Professional Quality of Life Scale version-5, respectively. Logistic regression analyses were used to estimate the odds ratio (OR) and 95% confidence interval (CI), along with subgroup and mediation analyses. RESULTS: The prevalence of mild-to-severe depression was 41.1% in vaccinated healthcare workers. The odd of mild-to-severe depression was increased with higher perceived stress. Compared with vaccinated healthcare workers with the lowest tertile of perceived stress, those with the highest tertile had increased odds of mild-to-severe depression by 120% (OR 2.20, 95% CI 1.46 to 3.31) after multivariable-adjustment. However, perceived stress was not associated with mild-to-severe depression in vaccinated healthcare workers with strong resilience, but was in those with weak resilience (pinteraction=0.004). Further analysis showed that compassion fatigue mediated the relationship between perceived stress and mild-to-severe depression, with a mediating effect of 49.7%. CONCLUSION: Perceived stress was related to an increased odd of mild-to-severe depression in vaccinated healthcare workers during COVID-19 pandemic, and this relationship might be explained by compassion fatigue.

Neuropilin-1 Facilitates Pseudorabies Virus Replication and Viral Glycoprotein B Promotes Its Degradation in a Furin-Dependent Manner.

Pseudorabies virus (PRV), which is extremely infectious and can infect numerous mammals, has a risk of spillover into humans. Virus-host interactions determine viral entry and spreading. Here, we showed that neuropilin-1 (NRP1) significantly potentiates PRV infection. Mechanistically, NRP1 promoted PRV attachment and entry, and enhanced cell-to-cell fusion mediated by viral glycoprotein B (gB), gD, gH, and gL. Furthermore, through in vitro coimmunoprecipitation (Co-IP) and bimolecular fluorescence complementation (BiFC) assays, NRP1 was found to physically interact with gB, gD, and gH, and these interactions were C-end Rule (CendR) motif independent, in contrast to currently known viruses. Remarkably, we illustrated that the viral protein gB promotes NRP1 degradation via a lysosome-dependent pathway. We further demonstrate that gB promotes NRP1 degradation in a furin-cleavage-dependent manner. Interestingly, in this study, we generated gB furin cleavage site (FCS)-knockout PRV (Δfurin PRV) and evaluated its pathogenesis; in vivo, we found that Δfurin PRV virulence was significantly attenuated in mice. Together, our findings demonstrated that NRP1 is an important host factor for PRV and that NRP1 may be a potential target for antiviral intervention. IMPORTANCE Recent studies have shown accelerated PRV cross-species spillover and that PRV poses a potential threat to humans. PRV infection in humans always manifests as a high fever, tonic-clonic seizures, and encephalitis. Therefore, understanding the interaction between PRV and host factors may contribute to the development of new antiviral strategies against PRV. NRP1 has been demonstrated to be a receptor for several viruses that harbor CendR, including SARS-CoV-2. However, the relationships between NRP1 and PRV are poorly understood. Here, we found that NRP1 significantly potentiated PRV infection by promoting PRV attachment and enhanced cell-to-cell fusion. For the first time, we demonstrated that gB promotes NRP1 degradation via a lysosome-dependent pathway. Last, in vivo, Δfurin PRV virulence was significantly attenuated in mice. Therefore, NRP1 is an important host factor for PRV, and NRP1 may be a potential target for antiviral drug development.

Nurse managers' burnout and organizational support: The serial mediating role of leadership and resilience.

AIMS: To investigate the relationship between organizational support and nurse managers' burnout and the serial multiple mediating effects of leadership and resilience. BACKGROUND: Nurse managers are at a high risk of burnout, especially during the COVID-19 pandemic. However, no research has been done to examine the associations between nurse managers' organizational support, leadership, resilience and burnout. METHODS: This cross-sectional study recruited 458 nurse managers from 13 tertiary public hospitals in Jiangsu, China. They completed the Survey of Perceived Organizational Support, the Clinical Leadership Survey, the Resilience Scale and the Maslach Burnout Inventor-Human Service Survey. The serial mediating effect of individual leadership and resilience was estimated using the structural equation modelling method via Mplus 7.0. RESULTS: There were direct and indirect effects of organizational support on burnout, controlling for work variables. Leadership and resilience serially mediate the association between organizational support and burnout (ß = -.051, 95% confidence interval: -0.093 to -0.020). CONCLUSIONS: Among nurse managers, organizational support may be sequentially associated with improved leadership first and then resilience, which in turn is related to decreased burnout. IMPLICATIONS FOR NURSING MANAGEMENT: We recommend that hospital administrators incorporate leadership, resilience and burnout assessment in the routine psychological screening of nurse managers and creatively apply the organizational interventions to decrease nurse managers' burnout.

SARS-CoV-2 hijacks macropinocytosis to facilitate its entry and promote viral spike-mediated cell-to-cell fusion.

Revealing the mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry and cell-to-cell spread might provide insights for understanding the underlying mechanisms of viral pathogenesis, tropism, and virulence. The signaling pathways involved in SARS-CoV-2 entry and viral spike-mediated cell-to-cell fusion remain elusive. In the current study, we found that macropinocytosis inhibitors significantly suppressed SARS-CoV-2 infection at both the entry and viral spike-mediated cell-to-cell fusion steps. We demonstrated that SARS-CoV-2 entry required the small GTPase Rac1 and its effector kinase p21-activated kinase 1 by dominant-negative and RNAi assays in human embryonic kidney 293T-angiotensin-converting enzyme 2 cells and that the serine protease transmembrane serine protease 2 reversed the decrease in SARS-CoV-2 entry caused by the macropinocytosis inhibitors. Moreover, in the cell-to-cell fusion assay, we confirmed that macropinocytosis inhibitors significantly decreased viral spike-mediated cell-to-cell fusion. Overall, we provided evidence that SARS-CoV-2 utilizes a macropinocytosis pathway to enter target cells and to efficiently promote viral spike-mediated cell-to-cell fusion.

Integration of protein context improves protein-based COVID-19 patient stratification.

BACKGROUND: Classification of disease severity is crucial for the management of COVID-19. Several studies have shown that individual proteins can be used to classify the severity of COVID-19. Here, we aimed to investigate whether integrating four types of protein context data, namely, protein complexes, stoichiometric ratios, pathways and network degrees will improve the severity classification of COVID-19. METHODS: We performed machine learning based on three previously published datasets. The first was a SWATH (sequential window acquisition of all theoretical fragment ion spectra) MS (mass spectrometry) based proteomic dataset. The second was a TMTpro 16plex labeled shotgun proteomics dataset. The third was a SWATH dataset of an independent patient cohort. RESULTS: Besides twelve proteins, machine learning also prioritized two complexes, one stoichiometric ratio, five pathways, and five network degrees, resulting a 25-feature panel. As a result, a model based on the 25 features led to effective classification of severe cases with an AUC of 0.965, outperforming the models with proteins only. Complement component C9, transthyretin (TTR) and TTR-RBP (transthyretin-retinol binding protein) complex, the stoichiometric ratio of SAA2 (serum amyloid A proteins 2)/YLPM1 (YLP Motif Containing 1), and the network degree of SIRT7 (Sirtuin 7) and A2M (alpha-2-macroglobulin) were highlighted as potential markers by this classifier. This classifier was further validated with a TMT-based proteomic data set from the same cohort (test dataset 1) and an independent SWATH-based proteomic data set from Germany (test dataset 2), reaching an AUC of 0.900 and 0.908, respectively. Machine learning models integrating protein context information achieved higher AUCs than models with only one feature type. CONCLUSION: Our results show that the integration of protein context including protein complexes, stoichiometric ratios, pathways, network degrees, and proteins improves phenotype prediction.

Enhanced inflammation and suppressed adaptive immunity in COVID-19 with prolonged RNA shedding.

Little is known regarding why a subset of COVID-19 patients exhibited prolonged positivity of SARS-CoV-2 infection. Here, we found that patients with long viral RNA course (LC) exhibited prolonged high-level IgG antibodies and higher regulatory T (Treg) cell counts compared to those with short viral RNA course (SC) in terms of viral load. Longitudinal proteomics and metabolomics analyses of the patient sera uncovered that prolonged viral RNA shedding was associated with inhibition of the liver X receptor/retinoid X receptor (LXR/RXR) pathway, substantial suppression of diverse metabolites, activation of the complement system, suppressed cell migration, and enhanced viral replication. Furthermore, a ten-molecule learning model was established which could potentially predict viral RNA shedding period. In summary, this study uncovered enhanced inflammation and suppressed adaptive immunity in COVID-19 patients with prolonged viral RNA shedding, and proposed a multi-omic classifier for viral RNA shedding prediction.

Protein sensors combining both on-and-off model for antibody homogeneous assay.

Protein sensors based on allosteric enzymes responding to target binding with rapid changes in enzymatic activity are potential tools for homogeneous assays. However, a high signal-to-noise ratio (S/N) is difficult to achieve in their construction. A high S/N is critical to discriminate signals from the background, a phenomenon that might largely vary among serum samples from different individuals. Herein, based on the modularized luciferase NanoLuc, we designed a novel biosensor called NanoSwitch. This sensor allows direct detection of antibodies in 1 µl serum in 45 min without washing steps. In the detection of Flag and HA antibodies, NanoSwitches respond to antibodies with S/N ratios of 33-fold and 42-fold, respectively. Further, we constructed a NanoSwitch for detecting SARS-CoV-2-specific antibodies, which showed over 200-fold S/N in serum samples. High S/N was achieved by a new working model, combining the turn-off of the sensor with human serum albumin and turn-on with a specific antibody. Also, we constructed NanoSwitches for detecting antibodies against the core protein of hepatitis C virus (HCV) and gp41 of the human immunodeficiency virus (HIV). Interestingly, these sensors demonstrated a high S/N and good performance in the assays of clinical samples; this was partly attributed to the combination of off-and-on models. In summary, we provide a novel type of protein sensor and a working model that potentially guides new sensor design with better performance.

Gut microbiota, inflammation, and molecular signatures of host response to infection.

Gut microbial dysbiosis has been linked to many noncommunicable diseases. However, little is known about specific gut microbiota composition and its correlated metabolites associated with molecular signatures underlying host response to infection. Here, we describe the construction of a proteomic risk score based on 20 blood proteomic biomarkers, which have recently been identified as molecular signatures predicting the progression of the COVID-19. We demonstrate that in our cohort of 990 healthy individuals without infection, this proteomic risk score is positively associated with proinflammatory cytokines mainly among older, but not younger, individuals. We further discover that a core set of gut microbiota can accurately predict the above proteomic biomarkers among 301 individuals using a machine learning model and that these gut microbiota features are highly correlated with proinflammatory cytokines in another independent set of 366 individuals. Fecal metabolomics analysis suggests potential amino acid-related pathways linking gut microbiota to host metabolism and inflammation. Overall, our multi-omics analyses suggest that gut microbiota composition and function are closely related to inflammation and molecular signatures of host response to infection among healthy individuals. These results may provide novel insights into the cross-talk between gut microbiota and host immune system.

Rapid visual detection of porcine reproductive and respiratory syndrome virus via recombinase polymerase amplification combined with a lateral flow dipstick.

Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically devastating infectious diseases in the global swine industry. A rapid and sensitive on-site detection method for PRRS virus (PRRSV) is critically important for diagnosing PRRS. In this study, we established a method that combines reverse transcription recombinase polymerase amplification (RT-RPA) with a lateral flow dipstick (LFD) for detecting North American PRRSV (PRRSV-2). The primers and probe were designed based on the conserved region of all complete PRRSV-2 genomic sequences available in China (n = 512) from 1996 to 2020. The detection limit of the assay was 5.6 × 10-1 median tissue culture infection dose (TCID50) per reaction within 30 min at 42 °C, which was more sensitive than that of reverse transcription polymerase chain reaction (RT-PCR) (5.6 TCID50 per reaction). The assay was highly specific for the epidemic lineages of PRRSV-2 in China and did not cross-react with pseudorabies virus, porcine circovirus 2, classical swine fever virus, or porcine epidemic diarrhea virus. The assay performance was evaluated by testing 179 samples and comparing the results with those of quantitative RT-PCR (RT-qPCR). The results showed that the detection coincidence rate of RT-RPA and RT-qPCR was 100% when the cycle threshold values of RT-qPCR were < 32. The assay provides a new alternative for simple and reliable detection of PRRSV-2 and has great potential for application in the field.

Antibody responses to SARS-CoV-2 in patients with COVID-19.

We report acute antibody responses to SARS-CoV-2 in 285 patients with COVID-19. Within 19 days after symptom onset, 100% of patients tested positive for antiviral immunoglobulin-G (IgG). Seroconversion for IgG and IgM occurred simultaneously or sequentially. Both IgG and IgM titers plateaued within 6 days after seroconversion. Serological testing may be helpful for the diagnosis of suspected patients with negative RT-PCR results and for the identification of asymptomatic infections.

Potential Use of Serum Proteomics for Monitoring COVID-19 Progression to Complement RT-PCR Detection.

RT-PCR is the primary method to diagnose COVID-19 and is also used to monitor the disease course. This approach, however, suffers from false negatives due to RNA instability and poses a high risk to medical practitioners. Here, we investigated the potential of using serum proteomics to predict viral nucleic acid positivity during COVID-19. We analyzed the proteome of 275 inactivated serum samples from 54 out of 144 COVID-19 patients and shortlisted 42 regulated proteins in the severe group and 12 in the non-severe group. Using these regulated proteins and several key clinical indexes, including days after symptoms onset, platelet counts, and magnesium, we developed two machine learning models to predict nucleic acid positivity, with an AUC of 0.94 in severe cases and 0.89 in non-severe cases, respectively. Our data suggest the potential of using a serum protein-based machine learning model to monitor COVID-19 progression, thus complementing swab RT-PCR tests. More efforts are required to promote this approach into clinical practice since mass spectrometry-based protein measurement is not currently widely accessible in clinic.

Immune memory in convalescent patients with asymptomatic or mild COVID-19.

It is important to evaluate the durability of the protective immune response elicited by primary infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we systematically evaluated the SARS-CoV-2-specific memory B cell and T cell responses in healthy controls and individuals recovered from asymptomatic or symptomatic infection approximately 6 months prior. Comparatively low frequencies of memory B cells specific for the receptor-binding domain (RBD) of spike glycoprotein (S) persisted in the peripheral blood of individuals who recovered from infection (median 0.62%, interquartile range 0.48-0.69). The SARS-CoV-2 RBD-specific memory B cell response was detected in 2 of 13 individuals who recovered from asymptomatic infection and 10 of 20 individuals who recovered from symptomatic infection. T cell responses induced by S, membrane (M), and nucleocapsid (N) peptide libraries from SARS-CoV-2 were observed in individuals recovered from coronavirus disease 2019 (COVID-19), and cross-reactive T cell responses to SARS-CoV-2 were also detected in healthy controls.

Clofazimine broadly inhibits coronaviruses including SARS-CoV-2.

The COVID-19 pandemic is the third outbreak this century of a zoonotic disease caused by a coronavirus, following the emergence of severe acute respiratory syndrome (SARS) in 20031 and Middle East respiratory syndrome (MERS) in 20122. Treatment options for coronaviruses are limited. Here we show that clofazimine-an anti-leprosy drug with a favourable safety profile3-possesses inhibitory activity against several coronaviruses, and can antagonize the replication of SARS-CoV-2 and MERS-CoV in a range of in vitro systems. We found that this molecule, which has been approved by the US Food and Drug Administration, inhibits cell fusion mediated by the viral spike glycoprotein, as well as activity of the viral helicase. Prophylactic or therapeutic administration of clofazimine in a hamster model of SARS-CoV-2 pathogenesis led to reduced viral loads in the lung and viral shedding in faeces, and also alleviated the inflammation associated with viral infection. Combinations of clofazimine and remdesivir exhibited antiviral synergy in vitro and in vivo, and restricted viral shedding from the upper respiratory tract. Clofazimine, which is orally bioavailable and comparatively cheap to manufacture, is an attractive clinical candidate for the treatment of outpatients and-when combined with remdesivir-in therapy for hospitalized patients with COVID-19, particularly in contexts in which costs are an important factor or specialized medical facilities are limited. Our data provide evidence that clofazimine may have a role in the control of the current pandemic of COVID-19 and-possibly more importantly-in dealing with coronavirus diseases that may emerge in the future.

A retrospective analysis of clinical efficacy of ribavirin in adults hospitalized with severe COVID-19.

INTRODUCTION: Coronavirus disease-2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) swept rapidly throughout the world. So far, no therapeutics have yet proven to be effective. Ribavirin was recommended for the treatment of COVID-19 in China because of its in vitro activity. However, evidence supporting its clinical use with good efficacy is still lacking. METHODS: A total of 208 confirmed severe COVID-19 patients who were hospitalized in Wuhan Union West Campus between 1 February 2020 and 10 March 2020 were enrolled in the retrospective study. Patients were divided into two groups based on the use of ribavirin. The primary endpoint was the time to clinical improvement. The secondary endpoints included mortality, survival time, time to throat swab SARS-CoV-2 nucleic acid negative conversion, and the length of hospital stay. RESULTS: 68 patients were treated with ribavirin while 140 not. There were no significant between-group differences in demographic characteristics, baseline laboratory test results, treatment, and distribution of ordinal scale scores at enrollment, except for coexisting diseases especially cancer (ribavirin group vs no ribavirin group, P = 0.01). Treatment with ribavirin was not associated with a difference in the time to clinical improvement (P = 0.48, HR = 0.88, 95% CI = 0.63-1.25). There were also no significant differences between-group in SARS-CoV-2 nucleic acid negative conversion, mortality, survival time, and the length of hospital stay. CONCLUSIONS: In hospitalized adult patients with severe COVID-19, no significant benefit was observed with ribavirin treatment.

Multi-organ proteomic landscape of COVID-19 autopsies.

The molecular pathology of multi-organ injuries in COVID-19 patients remains unclear, preventing effective therapeutics development. Here, we report a proteomic analysis of 144 autopsy samples from seven organs in 19 COVID-19 patients. We quantified 11,394 proteins in these samples, in which 5,336 were perturbed in the COVID-19 patients compared to controls. Our data showed that cathepsin L1, rather than ACE2, was significantly upregulated in the lung from the COVID-19 patients. Systemic hyperinflammation and dysregulation of glucose and fatty acid metabolism were detected in multiple organs. We also observed dysregulation of key factors involved in hypoxia, angiogenesis, blood coagulation, and fibrosis in multiple organs from the COVID-19 patients. Evidence for testicular injuries includes reduced Leydig cells, suppressed cholesterol biosynthesis, and sperm mobility. In summary, this study depicts a multi-organ proteomic landscape of COVID-19 autopsies that furthers our understanding of the biological basis of COVID-19 pathology.

A Peptide-Based Magnetic Chemiluminescence Enzyme Immunoassay for Serological Diagnosis of Coronavirus Disease 2019.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel ß-coronavirus, causes severe pneumonia and has spread throughout the globe rapidly. The disease associated with SARS-CoV-2 infection is named coronavirus disease 2019 (COVID-19). To date, real-time reverse-transcription polymerase chain reaction (RT-PCR) is the only test able to confirm this infection. However, the accuracy of RT-PCR depends on several factors; variations in these factors might significantly lower the sensitivity of detection. METHODS: In this study, we developed a peptide-based luminescent immunoassay that detected immunoglobulin (Ig)G and IgM. The assay cutoff value was determined by evaluating the sera from healthy and infected patients for pathogens other than SARS-CoV-2. RESULTS: To evaluate assay performance, we detected IgG and IgM in the sera from confirmed patients. The positive rate of IgG and IgM was 71.4% and 57.2%, respectively. CONCLUSIONS: Therefore, combining our immunoassay with real-time RT-PCR might enhance the diagnostic accuracy of COVID-19.

Proteomic and Metabolomic Characterization of COVID-19 Patient Sera.

Early detection and effective treatment of severe COVID-19 patients remain major challenges. Here, we performed proteomic and metabolomic profiling of sera from 46 COVID-19 and 53 control individuals. We then trained a machine learning model using proteomic and metabolomic measurements from a training cohort of 18 non-severe and 13 severe patients. The model was validated using 10 independent patients, 7 of which were correctly classified. Targeted proteomics and metabolomics assays were employed to further validate this molecular classifier in a second test cohort of 19 COVID-19 patients, leading to 16 correct assignments. We identified molecular changes in the sera of COVID-19 patients compared to other groups implicating dysregulation of macrophage, platelet degranulation, complement system pathways, and massive metabolic suppression. This study revealed characteristic protein and metabolite changes in the sera of severe COVID-19 patients, which might be used in selection of potential blood biomarkers for severity evaluation.

The clinical and immunological features of pediatric COVID-19 patients in China.

In December 2019, the corona virus disease 2019 (COVID-19) caused by novel coronavirus (SARS-CoV-2) emerged in Wuhan, China and rapidly spread worldwide. Few information on clinical features and immunological profile of COVID-19 in paediatrics. The clinical features and treatment outcomes of twelve paediatric patients confirmed as COVID-19 were analyzed. The immunological features of children patients was investigated and compared with twenty adult patients. The median age was 14.5-years (range from 0.64 to 17), and six of the patients were male. The average incubation period was 8 days. Clinically, cough (9/12, 75%) and fever (7/12, 58.3%) were the most common symptoms. Four patients (33.3%) had diarrhea during the disease. As to the immune profile, children had higher amount of total T cell, CD8+ T cell and B cell but lower CRP levels than adults (P < 0.05). Ground-glass opacity (GGO) and local patchy shadowing were the typical radiological findings on chest CT scan. All patients received antiviral and symptomatic treatment and the symptom relieved in 3-4 days after admitted to hospital. The paediatric patients showed mild symptom but with longer incubation period. Children infected with SARS-CoV-2 had different immune profile with higher T cell amount and low inflammatory factors level, which might ascribed to the mild clinical symptom. We advise that nucleic acid test or examination of serum IgM/IgG antibodies against SARS-CoV-2 should be taken for children with exposure history regardless of clinical symptom.