Correlation coefficient-directed label-free characterization of native proteins by surface-enhanced Raman spectroscopy.

Investigation of proteins in their native state is the core of proteomics towards better understanding of their structures and functions. Surface-enhanced Raman spectroscopy (SERS) has shown its unique advantages in protein characterization with fingerprint information and high sensitivity, which makes it a promising tool for proteomics. It is still challenging to obtain SERS spectra of proteins in the native state and evaluate the native degree. Here, we constructed 3D physiological hotspots for a label-free dynamic SERS characterization of a native protein with iodide-modified 140 nm Au nanoparticles. We further introduced the correlation coefficient to quantitatively evaluate the variation of the native degree, whose quantitative nature allows us to explicitly investigate the Hofmeister effect on the protein structure. We realized the classification of a protein of SARS-CoV-2 variants in 15 min, which has not been achieved before. This study offers an effective tool for tracking the dynamic structure of proteins and biomedical research.

Changes in the mental health status of the general Chinese population during the COVID-19 pandemic: A longitudinal study.

The study is based on a longitudinal evaluation of the public, during the initial COVID-19 outbreak in China and 8 months after. It aimed to explore the changes in the mental health of the public at the beginning of the pandemic and during the regular epidemic prevention and control. An online survey questionnaire was used to collect data during the initial COVID-19 outbreak (February 10, 2020-February 18, 2020; T1) and 8 months after the outbreak (October 21, 2020-December 29, 2020; T2). Psychological distress was assessed using the Patient Health Questionnaire-9 (PHQ-9), Self-rating Anxiety Scale (SAS), and Post-traumatic Stress Disorder Checklist (PCL-5). A chi-square test was used to compare the changes in the depression and anxiety scores at T1 and T2, and the correlation between symptoms was analyzed through Spearman's rank correlation. In T1, 1,200 people were recruited, while 168 people responded in T2. Depression (48.2-31.0%; p=0.001) and anxiety (17.9-9.5%; p = 0.026) symptoms decreased over time; two participants developed post-traumatic stress disorder (PTSD) in T2. The scores of the PHQ-9 scale and the SAS scale were both positively correlated with the score of the PCL-5 scale and negatively correlated with sleep time. During the COVID-19 pandemic, part of the general population's anxiety and depression significantly reduced with time, and they rarely developed PTSD. PTSD occurrence was related to severe depression and anxiety.

Rapid Adaptation of Established High-Throughput Molecular Testing Infrastructure for Monkeypox Virus Detection.

Beginning in May 2022, a rising number of monkeypox cases were reported in non-monkeypox-endemic countries in the Northern Hemisphere. We adapted 2 published quantitative PCRs for use as a dual-target monkeypox virus test on widely used automated high-throughput PCR systems. We determined analytic performance by serial dilutions of monkeypox virus reference material, which we quantified by digital PCR. We found the lower limit of detection for the combined assays was 4.795 (95% CI 3.6-8.6) copies/mL. We compared clinical performance against a commercial manual orthopoxvirus research use only PCR kit by using clinical remnant swab samples. Our assay showed 100% positive (n = 11) and 100% negative (n = 56) agreement. Timely and scalable PCR tests are crucial for limiting further spread of monkeypox. The assay we provide streamlines high-throughput molecular testing for monkeypox virus on existing broadly established platforms used for SARS-CoV-2 diagnostic testing.

An Analysis of the Measurement of Symbiosis Intensity in Scenic Spots and the Influence Mechanism

Scenic symbiosis, which is of great research significance, is an important way to achieve high-quality coordinated development of regional tourism. Based on the symbiosis theory, this paper defines the symbiosis intensity of scenic spots and establishes a calculation scheme. On the one hand, it measures the spatiotemporal evolution characteristics of symbiosis intensity of 281 scenic spots in Hunan Province from September 2018 to December 2021. On the other hand, it uses a multiple linear regression model to quantitatively explore its influence mechanism. The results indicate that: (1) From September 2018 to December 2021, the tourism demand in Hunan Province presented fluctuations due to the impact of the COVID-19 pandemic, which can be divided into three stages: 'slight increase before the COVID-19 pandemic';, 'rapid decline during the COVID-19 pandemic';, and 'slow recovery after the COVID-19 pandemic';;(2) The symbiosis intensity of scenic spots is mainly at a low, low-middle level;with Changsha City and the Western Hunan Area as dual cores, it reflects obvious 'core-edge';characteristics;(3) Quality index, market attention level, connection breadth, and distance index jointly affect the symbiosis intensity of scenic spots. Among them, the quality index, market attention level, and connection breadth all have positive impacts on the symbiosis intensity, whereas the distance index has a negative impact on it. Finally, this paper discusses how to improve the symbiosis intensity of scenic spots and promote the mutual benefit and symbiosis development to provide a theoretical basis and practical reference for the promotion of the sustainable development of regional tourism.

Clinical Evaluation of a Fully-Automated High-Throughput Multiplex Screening-Assay to Detect and Differentiate the SARS-CoV-2 B.1.1.529 (Omicron) and B.1.617.2 (Delta) Lineage Variants.

BACKGROUND: The recently emerged SARS-CoV-2 B.1.1.529 lineage and its sublineages (Omicron variant) pose a new challenge to healthcare systems worldwide due to its ability to efficiently spread in immunized populations and its resistance to currently available monoclonal antibody therapies. RT-PCR-based variant tests can be used to screen large sample-sets rapidly and accurately for relevant variants of concern (VOC). The aim of this study was to establish and validate a multiplex assay on the cobas 6800/8800 systems to allow discrimination between the two currently circulating VOCs, Omicron and Delta, in clinical samples. METHODS: Primers and probes were evaluated for multiplex compatibility. Analytic performance was assessed using cell culture supernatant of an Omicron variant isolate and a clinical Delta variant sample, normalized to WHO-Standard. Clinical performance of the multiplex assay was benchmarked against NGS results. RESULTS: In silico testing of all oligos showed no interactions with a high risk of primer-dimer formation or amplification of human DNA/RNA. Over 99.9% of all currently available Omicron variant sequences are a perfect match for at least one of the three Omicron targets included in the multiplex. Analytic sensitivity was determined as 19.0 IU/mL (CI95%: 12.9-132.2 IU/mL) for the A67V + del-HV69-70 target, 193.9 IU/mL (CI95%: 144.7-334.7 IU/mL) for the E484A target, 35.5 IU/mL (CI95%: 23.3-158.0 IU/mL) for the N679K + P681H target and 105.0 IU/mL (CI95%: 80.7-129.3 IU/mL) for the P681R target. All sequence variances were correctly detected in the clinical sample set (225/225 Targets). CONCLUSION: RT-PCR-based variant screening compared to whole genome sequencing is both rapid and reliable in detecting relevant sequence variations in SARS-CoV-2 positive samples to exclude or verify relevant VOCs. This allows short-term decision-making, e.g., for patient treatment or public health measures.

Flavonoids in Ampelopsis grossedentata as covalent inhibitors of SARS-CoV-2 3CLpro: Inhibition potentials, covalent binding sites and inhibitory mechanisms.

Coronavirus 3C-like protease (3CLpro) is a crucial target for treating coronavirus diseases including COVID-19. Our preliminary screening showed that Ampelopsis grossedentata extract (AGE) displayed potent SARS-CoV-2-3CLpro inhibitory activity, but the key constituents with SARS-CoV-2-3CLpro inhibitory effect and their mechanisms were unrevealed. Herein, a practical strategy via integrating bioactivity-guided fractionation and purification, mass spectrometry-based peptide profiling and time-dependent biochemical assay, was applied to identify the crucial constituents in AGE and to uncover their inhibitory mechanisms. The results demonstrated that the flavonoid-rich fractions (10-17.5 min) displayed strong SARS-CoV-2-3CLpro inhibitory activities, while the constituents in these fractions were isolated and their SARS-CoV-2-3CLpro inhibitory activities were investigated. Among all isolated flavonoids, dihydromyricetin, isodihydromyricetin and myricetin strongly inhibited SARS-CoV-2 3CLpro in a time-dependent manner. Further investigations demonstrated that myricetin could covalently bind on SARS-CoV-2 3CLpro at Cys300 and Cys44, while dihydromyricetin and isodihydromyricetin covalently bound at Cys300. Covalent docking coupling with molecular dynamics simulations showed the detailed interactions between the orthoquinone form of myricetin and two covalent binding sites (surrounding Cys300 and Cys44) of SARS-CoV-2 3CLpro. Collectively, the flavonoids in AGE strongly and time-dependently inhibit SARS-CoV-2 3CLpro, while the newly identified SARS-CoV-2 3CLpro inhibitors in AGE offer promising lead compounds for developing novel antiviral agents.

Rapid Automated Screening for SARS-CoV-2 B.1.617 Lineage Variants (Delta/Kappa) through a Versatile Toolset of qPCR-Based SNP Detection.

BACKGROUND: The recent emergence of distinct and highly successful SARS-CoV-2 lineages has substantial implications for individual patients and public health measures. While next-generation-sequencing is routinely performed for surveillance purposes, RT-qPCR can be used to rapidly rule-in or rule-out relevant variants, e.g., in outbreak scenarios. The objective of this study was to create an adaptable and comprehensive toolset for multiplexed Spike-gene SNP detection, which was applied to screen for SARS-CoV-2 B.1.617 lineage variants. METHODS: We created a broad set of single nucleotide polymorphism (SNP)-assays including del-Y144/145, E484K, E484Q, P681H, P681R, L452R, and V1176F based on a highly specific multi-LNA (locked nucleic acid)-probe design to maximize mismatch discrimination. As proof-of-concept, a multiplex-test was compiled and validated (SCOV2-617VOC-UCT) including SNP-detection for L452R, P681R, E484K, and E484Q to provide rapid screening capabilities for the novel B.1.617 lineages. RESULTS: For the multiplex-test (SCOV2-617VOC-UCT), the analytic lower limit of detection was determined as 182 IU/mL for L452R, 144 IU/mL for P681R, and 79 IU/mL for E484Q. A total of 233 clinical samples were tested with the assay, including various on-target and off-target sequences. All SNPs (179/179 positive) were correctly identified as determined by SARS-CoV-2 whole genome sequencing. CONCLUSION: The recurrence of SNP locations and flexibility of methodology presented in this study allows for rapid adaptation to current and future variants. Furthermore, the ability to multiplex various SNP-assays into screening panels improves speed and efficiency for variant testing. We show 100% concordance with whole genome sequencing for a B.1.617.2 screening assay on the cobas6800 high-throughput system.

Sex Differences in Psychological Status and Fatigue of Frontline Staff After the COVID-19 Outbreak in China: A Cross-Sectional Study.

Background: The coronavirus disease 2019 (COVID-19) is spreading globally, and it is significant to pay attention to the mental health of frontline staff in this pandemic. This study is aimed to explore the sex difference among the frontline staff in demographics, characteristics of mental state, and the potential relationship between them. Method: A total of 2,614 Chinese frontline staff were recruited. The Self-Rating Anxiety Scale (SAS) and the Patient Health Questionnaire-9 (PHQ-9) were used for assessing the mental status of frontline staff, and the Fatigue Self-Assessment Scale (FSAS) was used for detecting fatigue. Result: The prevalence rate of anxiety for female frontline staff is higher than that of male (P = 0.003), and the prevalence rate of depression is similar between them (P = 0.091). After comparing the risk factors of unhealthy mental state between different sexes, it is found that family income below 100,000 [depression: odds ratio (OR) 1.37; 95% CI, 1.08-1.73; anxiety: OR 1.99; 95% CI, 1.44-2.75], unsupported of family (depression: OR 10.94; 95% CI, 1.39-85.79; anxiety: OR 11.92; 95% CI, 3.80-37.36), and marriage (depression: OR 1.67; 95% CI, 1.15-2.43) are risk factors for male, and total fatigue (depression: OR 2.96; 95% CI, 1.46-6.02) is risk factor for female. Conclusions: This study found that depression and anxiety are widespread among the frontline staff of COVID-19, and anxiety showed a higher prevalence rate among female frontline staff. From the sex difference in risk factors, the focus of psychological interventions may differ between genders. Men with low family income, unsupported by family or marriage, and women with a high score of total fatigue required particular attention to their psychological status.

Risk factors for the mortality of hemodialysis patients with COVID-19: A multicenter study from the overall hemodialysis population in Wuhan.

INTRODUCTION: Maintenance hemodialysis (MHD) patients are highly threatened in the novel coronavirus disease 2019 (COVID-19) pandemic, but evidence of risk factors for mortality in this population is still lacking. METHODS: We followed outcomes of the overall MHD population of Wuhan, including 7154 MHD patients from 65 hemodialysis centers, from January 1 to May 4, 2020. Among them, 130 were diagnosed with COVID-19. The demographic and clinical data of them were collected and compared between survivors and nonsurvivors. RESULTS: Compared to the corresponding period of last year, the all-cause mortality rate of the Wuhan MHD population significantly rose in February, and dropped down in March 2020. Of the 130 COVID-19 cases, 51 (39.2%) were deceased. Advanced age, decreased oxygen saturation, low diastolic blood pressure (DBP) on admission, and complications including acute cardiac injury (HR 5.03 [95% CI 2.21-11.14], p < 0.001), cerebrovascular event (HR 2.80 [95% CI 1.14-6.86], p = 0.025) and acute respiratory distress syndrome (HR 3.50 [95% CI 1.63-7.51], p = 0.001) were identified as independent risk factors for the death of COVID-19. The median virus shedding period of survivors was 25 days, longer than the general population. CONCLUSIONS: Maintenance hemodialysis patients are a highly vulnerable population at increased risk of mortality and prolonged virus shedding period in the ongoing COVID-19 pandemic. Advanced age, decreased oxygen saturation, low DBP on admission, and complications like acute cardiac injury are parameters independently associated with poor prognosis.

Discovery of naturally occurring inhibitors against SARS-CoV-2 3CLpro from Ginkgo biloba leaves via large-scale screening.

3-Chymotrypsin-like protease (3CLpro) is a virally encoded main proteinase that is pivotal for the viral replication across a broad spectrum of coronaviruses. This study aims to discover the naturally occurring SARS-CoV-2 3CLpro inhibitors from herbal constituents, as well as to investigate the inhibitory mechanism of the newly identified efficacious SARS-CoV-2 3CLpro inhibitors. Following screening of the inhibitory potentials of eighty herbal products against SARS-CoV-2 3CLpro, Ginkgo biloba leaves extract (GBLE) was found with the most potent SARS-CoV-2 3CLpro inhibition activity (IC50 = 6.68 µg/mL). Inhibition assays demonstrated that the ginkgolic acids (GAs) and the bioflavones isolated from GBLE displayed relatively strong SARS-CoV-2 3CLpro inhibition activities (IC50 < 10 µM). Among all tested constituents, GA C15:0, GA C17:1 and sciadopitysin displayed potent 3CLpro inhibition activities, with IC50 values of less than 2 µM. Further inhibition kinetic studies and docking simulations clearly demonstrated that two GAs and sciadopitysin strongly inhibit SARS-CoV-2 3CLprovia a reversible and mixed inhibition manner. Collectively, this study found that both GBLE and the major constituents in this herbal product exhibit strong SARS-CoV-2 3CLpro inhibition activities, which offer several promising leading compounds for developing novel anti-COVID-19 medications via targeting on 3CLpro.

Mutations in spike protein and allele variations in ACE2 impact targeted therapy strategies against SARS-CoV-2.

Coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread rapidly worldwide with high rates of transmission and substantial mortality. To date, however, no effective treatments or enough vaccines for COVID-19 are available. The roles of angiotensin converting enzyme 2 (ACE2) and spike protein in the treatment of COVID-19 are major areas of research. In this study, we explored the potential of ACE2 and spike protein as targets for the development of antiviral agents against SARS-CoV-2. We analyzed clinical data, genetic data, and receptor binding capability. Clinical data revealed that COVID-19 patients with comorbidities related to an abnormal renin-angiotensin system exhibited more early symptoms and poorer prognoses. However, the relationship between ACE2 expression and COVID-19 progression is still not clear. Furthermore, if ACE2 is not a good targetable protein, it would not be applicable across a wide range of populations. The spike-S1 receptor-binding domain that interacts with ACE2 showed various amino acid mutations based on sequence analysis. We identified two spike-S1 point mutations (V354F and V470A) by receptor-ligand docking and binding enzyme-linked immunosorbent assays. These variants enhanced the binding of the spike protein to ACE2 receptors and were potentially associated with increased infectivity. Importantly, the number of patients infected with the V354F and V470A mutants has increased with the development of the SARS-CoV-2 pandemic. These results suggest that ACE2 and spike-S1 are likely not ideal targets for the design of peptide drugs to treat COVID-19 in different populations.

Mental health of front-line staff in prevention of coronavirus disease 2019. / 2019å† çŠ¶ç— æ¯’ç— é˜²æŽ§ä¸€çº¿äººå‘˜çš„å¿ƒç†çŠ¶å†µ.

OBJECTIVES: The coronavirus disease 2019 (COVID-19) pandemic is a global public health crisis, which elicits psychological problems in different population. This study is to investigate the impact of COVID-19 on mental health in the front-line staff. METHODS: Patient Health Questionnare-9 (PHQ-9), Self-Rating Anxiety Scale (SAS), and Fatigue Self-assessment Scale (FSAS) were used to assess the depression, anxiety, and fatigue in front-line staff. RESULTS: The detection rates of depression, anxiety, and fatigue were 49.1%, 21.8%, and 76.0% among the front-line staff. The rates of depression, anxiety, and fatigue in community workers were higher than those in medical workers and other occupational staff (P<0.01). The PHQ-9 of front-line staffs was negatively correlated with age, family income, family members' support, satisfaction of service objects, and sleep quality (all P<0.01), while positively correlated with education level, fatigue, fear of pneumonia, and the duration of daily attention to the COVID-19 (all P<0.01). SAS was negatively correlated with age, family income, family support, satisfaction of objects service, and sleep quality (all P<0.01), while positively correlated with gender, fatigue, fear of pneumonia, and duration of daily attention to the COVID-19 (all P<0.01). CONCLUSIONS: The front-line workers should manage work and rest time reasonably to adjust their negative mood and fatigue. The government and the society should pay more attention to the psychological state of the front-line staff, particularly for the staff working in the community or villages and towns in preventing the COVID-19 pandemic. Thus, front-line staff can be obtained mental intervention or be taken a rest from the high-intensive work.

Serologic Detection of SARS-CoV-2 Infections in Hemodialysis Centers: A Multicenter Retrospective Study in Wuhan, China.

RATIONALE & OBJECTIVE: Patients receiving maintenance hemodialysis (MHD) are highly vulnerable to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The current study was designed to evaluate the prevalence of SARS-CoV-2 infection based on both nucleic acid testing (NAT) and antibody testing in Chinese patients receiving MHD. STUDY DESIGN: Cross-sectional study. SETTING & PARTICIPANTS: From December 1, 2019, to March 31, 2020, a total of 1,027 MHD patients in 5 large hemodialysis centers in Wuhan, China, were enrolled. Patients were screened for SARS-CoV-2 infection by symptoms and initial computed tomography (CT) of the chest. If patients developed symptoms after the initial screening was negative, repeat CT was performed. Patients suspected of being infected with SARS-CoV-2 were tested with 2 consecutive throat swabs for viral RNA. In mid-March 2020, antibody testing for SARS-CoV-2 was obtained for all MHD patients. EXPOSURE: NAT and antibody testing results for SARS-CoV-2. OUTCOMES: Morbidity, clinical features, and laboratory and radiologic findings. ANALYTICAL APPROACH: Differences between groups were examined using t test or Mann-Whitney U test, comparing those not infected with those infected and comparing those with infection detected using NAT with those with infection detected by positive serology test results. RESULTS: Among 1,027 patients receiving MHD, 99 were identified as having SARS-CoV-2 infection, for a prevalence of 9.6%. Among the 99 cases, 52 (53%) were initially diagnosed with SARS-CoV-2 infection by positive NAT; 47 (47%) were identified later by positive immunoglobulin G (IgG) or IgM antibodies against SARS-CoV-2. There was a spectrum of antibody profiles in these 47 patients: IgM antibodies in 5 (11%), IgG antibodies in 35 (74%), and both IgM and IgG antibodies in 7 (15%). Of the 99 cases, 51% were asymptomatic during the epidemic; 61% had ground-glass or patchy opacities on CT of the chest compared with 11.6% among uninfected patients (P<0.001). Patients with hypertensive kidney disease were more often found to have SARS-CoV-2 infection and were more likely to be symptomatic than patients with another primary cause of kidney failure. LIMITATIONS: Possible false-positive and false-negative results for both NAT and antibody testing; possible lack of generalizability to other dialysis populations. CONCLUSIONS: Half the SARS-CoV-2 infections in patients receiving MHD were subclinical and were not identified by universal CT of the chest and selective NAT. Serologic testing may help evaluate the overall prevalence and understand the diversity of clinical courses among patients receiving MHD who are infected with SARS-CoV-2.

Sleep of preschoolers during the coronavirus disease 2019 (COVID-19) outbreak.

Coronavirus disease 2019 (COVID-19) has resulted in a significantly large number of psychological consequences, including sleep health. The present study evaluated sleep patterns, sleep disturbances, and associated factors in Chinese preschoolers confined at home during the COVID-19 outbreak. Caregivers of 1619 preschoolers (aged 4-6 years) recruited from 11 preschools in Zunyi, Guizhou province completed the Children's Sleep Habit Questionnaire (CSHQ) between 17th and 19th February 2020. Data were compared to a sociodemographically similar sample of preschoolers (included in the 11 preschools) in 2018. Compared to the 2018 sample, the confined preschoolers demonstrated changes in sleep patterns characterized by later bedtimes and wake times, longer nocturnal and shorter nap sleep durations, comparable 24-hr sleep duration, and fewer caregiver-reported sleep disturbances. Moreover, behavioural practices (sleeping arrangement, reduced electronic device use, regular diet) and parenting practices (harmonious family atmosphere and increased parent-child communication) were associated with less sleep disturbances in the confined sample. The present study provides the first description of the impact of prolonged home confinement during the COVID-19 outbreak on sleep patterns and sleep disturbances in preschoolers, as well as highlighting the importance of the link between sleep health and family factors. Given that disrupted and insufficient sleep has been linked to immune system dysfunction, our findings also have potential implications for resilience to infection in young children during the COVID-19 pandemic. Future studies should further explore deficient sleep as a risk factor for coronavirus infection.

Clinical Characteristics of and Medical Interventions for COVID-19 in Hemodialysis Patients in Wuhan, China.

BACKGROUND: Reports indicate that those most vulnerable to developing severe coronavirus disease 2019 (COVID-19) are older adults and those with underlying illnesses, such as diabetes mellitus, hypertension, or cardiovascular disease, which are common comorbidities among patients undergoing maintenance hemodialysis. However, there is limited information about the clinical characteristics of hemodialysis patients with COVID-19 or about interventions to control COVID-19 in hemodialysis centers. METHODS: We collected data retrospectively through an online registration system that includes all patients receiving maintenance hemodialysis at 65 centers in Wuhan, China. We reviewed epidemiologic and clinical data of patients with laboratory-confirmed COVID-19 between January 1, 2020 and March 10, 2020. RESULTS: Of 7154 patients undergoing hemodialysis, 154 had laboratory-confirmed COVID-19. The mean age of the 131 patients in our analysis was 63.2 years; 57.3% were men. Many had underlying comorbidities, with cardiovascular disease (including hypertension) being the most common (68.7%). Only 51.9% of patients manifested fever; 21.4% of infected patients were asymptomatic. The most common finding on chest computed tomography (CT) was ground-grass or patchy opacity (82.1%). After initiating comprehensive interventions-including entrance screening of body temperature and symptoms, universal chest CT and blood tests, and other measures-new patients presenting with COVID-19 peaked at 10 per day on January 30, decreasing to 4 per day on February 11. No new cases occurred between February 26 and March 10, 2020. CONCLUSIONS: We found that patients receiving maintenance hemodialysis were susceptible to COVID-19 and that hemodialysis centers were high-risk settings during the epidemic. Increasing prevention efforts, instituting universal screening, and isolating patients with COVID-19 and directing them to designated hemodialysis centers were effective in preventing the spread of COVID-19 in hemodialysis centers.