Structural Requirements and Plasticity of Receptor-Binding Domain in Human Coronavirus Spike

The most recent human coronaviruses including severe acute respiratory syndrome coronavirus-2 causing severe respiratory tract infection and high pathogenicity bring significant global public health concerns. Infections are initiated by recognizing host cell receptors by coronavirus spike protein S1 subunit, and then S2 mediates membrane fusion. However, human coronavirus spikes undergo frequent mutation, which may result in diverse pathogenesis and infectivity. In this review, we summarize some of these recent structural and mutational characteristics of RBD of human coronavirus spike protein and their interaction with specific human cell receptors and analyze the structural requirements and plasticity of RBD. Stability of spike protein, affinity toward receptor, virus fitness, and infectivity are the factors controlling the viral tropisms. Thus, understanding the molecular details of RBDs and their mutations is critical in deciphering virus evolution. Structural information of spike and receptors of human coronaviruses not only reveals the molecular mechanism of host–microbe interaction and pathogenesis but also helps develop effective drug to control these infectious pathogens and cope with the future emerging coronavirus outbreaks.

The AST/ALT Ratio (De Ritis Ratio) Represents an Unfavorable Prognosis in Patients in Early-Stage SFTS: An Observational Cohort Study.

BACKGROUND: Severe fever with thrombocytopenia syndrome (SFTS), a widely prevalent infectious disease caused by severe fever with thrombocytopenia syndrome virus (SFTSV) that carries with it a high mortality rate, has emerged to be a public health concern. This study aimed to investigate the epidemiological and clinical characteristics of patients infected with SFTSV, seeking novel prognostic risk factors for SFTS. METHODS: In this retrospective and cross-sectional study, confirmed SFTS patients from the First Affiliated Hospital of Anhui Medical University were enrolled from September 1, 2019, to December 12, 2020. Cases were analyzed for epidemiological, demographic, clinical, and laboratory data. Logistic regression models were used to assess the association between predictors and outcome variables. A generalized additive mixed model (GAMM) was conducted to analyze the trending shift of aspartate aminotransferase/alanine transaminase-ratio (AST/ALT-ratio) and platelet (PLT) in SFTS patients treated with ribavirin. p values ≤ 0.05 were considered statistically significant. RESULTS: Clinical and laboratory results of 107 hospitalized patients with SFTSV infection were retrospectively described. The mean age at onset of disease was 60.38 ± 11.29 years old and the ratio between male and female was 1:1.2. Fever and thrombocytopenia are hallmark features of SFTS. Furthermore, multiple cases also experienced neurological complications, gastrointestinal/skeletal muscle symptoms together with other non-specific clinical manifestations; laboratory dataset outcomes reported dysregulated levels for routine blood biomarkers, coagulation function, and biochemistry. Overall, 107 patients were segregated into two groups according to patient condition at the clinical endpoint (survivors/non-survivors). SFTS survivors had a higher level of PLT- counts, total protein (TP), and estimated glomerular filtration rate (eGFR), while levels of activated partial thromboplastin time (APTT), thrombin time (TT), D-dimer (D-D), fibrinogen degradation products (FDP), ALT, AST, AST/ALT-ratio, creatinine (Cr), creatine phosphokinase (CK) and procalcitonin (PCT) was higher in non-survivors. Results from univariate Cox regression revealed that elevated levels of FDP, TT, AST/ALT-ratio, PCT, as well as decreased eGFR level and presence of central nervous system symptoms (CNS), were significant predictors for SFTS prognostic, results from multivariate logistic regression analysis in three adjusted models showed AST/ALT-ratio and PCT were independent risk factors for the prognosis of SFTS patients. Kaplan-Meier survival analysis showed that SFTS patients with AST/ALT-ratio >2.683 were associated with a shorter futime (means survival time), therefore indicating an unfavorable prognosis. Treatment with ribavirin could increase PLT count while decreasing AST/ALT-ratio within SFTS patients. CONCLUSION: SFTS is an emerging infectious disease, possibly leading to multiple-organ injury; AST/ALT-ratio was an independent risk factor for the prognosis of SFTS patients. Further investigation should be performed in order to gain more knowledge on this disease and guide clinical management.

Study on the Effect and Application Value of Heat-Inactivated Serum on the Detection of Thyroid Function, Tumor Markers, and Cytokines During the SARS-CoV-2 Pandemic.

Purpose: The current explored the impact of heat inactivation of blood samples on the results of a particular clinical test and its potential application value during the SARS-CoV-2 pandemic. We have aimed at providing a reference for clinical testing methods during the pandemic. Methods: Blood samples were selected from our department's routine clinical examination between January 2021 and June 2021. The levels of these samples for quantitative detection of these indicators in each group (n = 90 cases/group) covered normal reference ranges and medically determined levels. For qualitative testing of the indicators, the specimens were additionally classified as negative, weakly positive, and positive (n = 20 cases/group). The specimens were then inactivated, and the differences in relevant indicators before and after inactivation were evaluated. Results: A statistically significant difference was evident between the levels of TSH, T3, FT4, FT3, AFP, NSE, CYFRA211, IRI, IL-1ß, IL-6, IL-8, IL-10, IL-2R, and TNF-α in the non-inactivated group 1 and the inactivated group 1 (P < 0.05). Among them, there was a strong correlation between TSH, T3, FT4, FT3, CYFRA211, IRI, IL-1ß, IL-6, IL-8, and IL-2R levels in the two groups (P < 0.05), however, there was no correlation between AFP (P = 0.256) and NSE (P = 0.352) levels between the two groups (P > 0.05). The detected values of low-level AFP (<4 ng/mL), IL-10, and TNF-α after inactivation were all lower than the detection limit. There was not any statistically significant difference in the levels of tumor markers, such as CEA, CA125, CA724, CA199, CA153, and the quantitative levels of T4, Vit. D, HCG, CPS, and five items of hepatitis B virus (P > 0.05). The positive rate of anti-nuclear antibodies after inactivation was not statistically different from the ones observed before inactivation (P > 0.05). Upon correction by the regression equation, the observed levels of TSH, T3, FT4, FT3, CYFRA211, IRI, IL-1ß, IL-6, IL-8, and IL-2R were not significantly different from those before inactivation (P > 0.05). Conclusion: The heat inactivation of blood samples had different various effects on different test indicators, and some indicators could be corrected by employing regression equations. This detection method could potentially be employed during the SARS-CoV-2 pandemic, thereby effectively preventing iatrogenic infections.

Swab and Sputum SARS-CoV-2 RNA-Negative, CT-Positive, Symptomatic Contacts of COVID-19 Cases: A Hypothesis-Generating Prospective Population-Based Cohort Study of Eight Clusters.

Background: While some contacts of COVID-19 cases become symptomatic and radiographically abnormal, their SARS-CoV-2 RNA tests remain negative throughout the disease course. This prospective population-based cohort study aimed to explore their characteristics and significances. Methods: From January 22, 2020, when the first COVID-19 case was identified in Hefei, China, until July 3, a total of 14,839 people in Feidong, Hefei, with a population of ~1,081,000 underwent SARS-CoV-2 RNA testing, where 36 cases (0.2%) with confirmed COVID-19 infection (Group 1) and 27 close contacts (0.2%) testing negative for SARS-CoV-2 RNA but having both positive COVID-19 exposure histories and CT findings (Group 2) from eight clusters were prospectively identified. Another 62 non-COVID-19 pneumonia cases without any exposure history (Group 3) were enrolled, and characteristics of the three groups were described and compared. We further described a cluster with an unusual transmission pattern. Results: Fever was more common in Group 2 than Groups 1 and 3. Frequency of diarrhea in Group 1 was higher than in Groups 2 and 3. Median leucocyte, neutrophil, monocyte, and eosinophil counts were all lower in Groups 1 and 2 than in Group 3. Median D-dimer level was lower in Group 1 than in Groups 2 and 3. Total protein and albumin levels were higher in Groups 1 and 2 than in Group 3. C-reactive protein level was lower and erythrocyte sedimentation rate slower in Groups 1 and 2 than in Group 3. Combination antibacterial therapy and levofloxacin were more often used in Group 3 than in Groups 1 and 2. Lopinavir/ritonavir was more often administered in Groups 1 and 2 than in Group 3. Group 1 received more often corticosteroids than Groups 2 and 3. Group 2 received less often oxygen therapy than Groups 1 and 3. Median duration from illness onset to discharge was longer in Group 1 (27 d) than Groups 2 and 3 (both 17 d). Among contacts of a confirmed COVID-19 patient, only one had a positive virus RNA test but remained asymptomatic and had negative CT findings, and three had negative virus RNA tests but had symptoms and positive CT findings, one of whom transmitted COVID-19 to another asymptomatic laboratory-confirmed patient who had no other exposures. Conclusions: Among close contacts of confirmed COVID-19 cases, some present with positive symptoms and CT findings but test negative for SARS-CoV-2 RNA using common respiratory (throat swab and sputum) specimens; they have features more similar to confirmed COVID-19 cases than non-COVID-19 pneumonia cases and might have transmitted SARS-CoV-2 to others. Such cases might add to the complexity and difficulty of COVID-19 control. Our hypothesis-generating study might suggest that SARS-CoV-2 RNA testing by rRT-PCR assays of common respiratory (throat swab and sputum) specimens alone, the widely accepted "golden standard" for diagnosing COVID-19, might be sometimes insufficient, and that further studies with some further procedures (e.g., testing via bronchoalveolar lavage or specific antibodies) would be warranted for Group 2-like patients, namely, the SARS-CoV-2 RNA-negative (tested using common respiratory specimens), radiographically positive, symptomatic contacts of COVID-19 cases, to further reveal their nature.

Development of a SARS-CoV-2 rapid antibody detection kit and study on dynamic changes in antibodies in infected patients.

INTRODUCTION: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread rapidly to 185 regions and countries around the world with more than 2.8 million confirmed infections and 203,044 deaths. Respiratory diseases caused by SARS-CoV-2 are serious threats to human health. OBJECTIVES: To develop a rapid detection kit for new coronavirus antibodies and use it to study the dynamic changes in antibodies in clinically confirmed SARS-CoV-2-infected patients. METHODS: The SARS-CoV-2 IgM/IgG antibody test kit (colloidal gold method) was developed. Serum SARS-CoV-2 IgM and IgG antibodies were tested in SARS-CoV-2- and non-SARS-CoV-2-infected persons, respectively. RESULTS AND CONCLUSION: The sensitivities of the SARS-CoV-2 IgM/IgG antibody test kit (colloidal gold method) were 50%, 70%, 92.5% and 97.5% after 1-3 days, 4-6 days, 7-9 days and >9 days of admission, respectively, and the specificities of the IgM, IgG and IgM + IgG antibodies were all 100%. Using the SARS-CoV-2 IgM/IgG antibody test kit (colloidal gold method), the positive rates of SARS-CoV-2 IgM and IgG antibodies increased from 50% to 92.5% after 1-3 days, 4-6 days and 7-9 days of admission, which showed an increasing trend. The titers of the SARS-CoV-2 IgM and IgG antibodies in the positive specimens increased with the length of admission.

Changes of blood routine examination and lymphocyte subsets in patients with COVID-19

Objective: To investigate the changes of blood routine examination and lymphocyte subsets in patients with coronavirus disease 2019 (COVID-19).

Serum SARS-COV-2 Nucleocapsid Protein: A Sensitivity and Specificity Early Diagnostic Marker for SARS-COV-2 Infection.

Objective: To explore the diagnostic value of serum severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein assay in the early stages of SARS-COV-2 infection. Methods: Serum N protein level in SARS-COV-2 infected patients and non-SARS-COV-2 infected population was measured by enzyme-linked immunosorbent assay (ELISA) double antibody sandwich assay. Colloidal gold immunochromatography assay was used to detect serum N protein antibodies in the above populations. Results: Fifty cases of SARS-CoV-2 nucleic acid-positive and SARS-CoV-2 antibody-negative patients had a serum N protein positivity rate of 76%. Thirty-seven patients who were positive for serum SARS-CoV-2 antibody after infection had a serum SARS-CoV-2 N protein positivity rate of 2.7%. Serum N protein test results of 633 non-SARS-COV-2 infected patients, including pregnant women, patients with other respiratory infections, and individuals with increased rheumatoid factor were all negative, with serum N protein concentration <10.00 pg/mL at 100% specificity. Using SPSS 19.0 to calculate the receiver operating characteristic curve, the area under the curve was determined to be 0.9756 (95% confidence interval 0.9485-1.000, p < 0.0001), and sensitivity and specificity were 92% (95% confidence interval 81.16-96.85%) and 96.84% (95% confidence interval 95.17-97.15%), respectively. The best CUT-OFF value was 1.850 pg/mL. Conclusion: The measurement of serum SARS-COV-2 N protein has a high diagnostic value for infected patients before the antibody appears and shortens the window period of serological diagnosis. It is recommended that the manufacturer establish two different CUT-OFF values according to the purpose of the application. One CUT-OFF value is used for the diagnosis of clinical SARS-COV-2 infection, and the other is used to screen out as many suspected cases as possible.

A Low Viral Dose in COVID-19 Patient: A Case Report.

SARS-CoV-2 outbreak has attracted global attention. Verifying the presence of viral RNA is the gold standard for the diagnosis of COVID-19. However, RT-qPCR diagnosis often fails to catch infected patients, because of inconsistent swab sample collection. Here we report a case that showed 5 consecutive negative and 1 low-viral- dose RT-qPCR results during illness spanning over 20 days. Clinical symptoms suggest SARS-CoV-2 infection with typical ground glass like a lung in computed tomography. SARS-CoV-2 infection was serologically confirmed by the presence of anti-SARS-CoV-2 specific antibodies in patients' serum. Finally, a high level of protective IgG was produced after the patient recovered. Surprisingly, as a barber and a housewife staying at home for the first 2 weeks after the onset of illness, none of the close contacts were infected, showing a case of low viral load and low infectivity in this patient.

Characteristics of laboratory indexes in COVID-19 patients with non-severe symptoms in Hefei City, China: diagnostic value in organ injuries.

This study compared the laboratory indexes in 40 non-severe COVID-19 patients with those in 57 healthy controls. In the peripheral blood system of non-severe symptom COVID-19 patients, lymphocytes, eosinophils, basophils, total procollagen type 1 amino-terminal propeptide, osteocalcin N-terminal, thyroid-stimulating hormone, growth hormone, and insulin-like growth factor-binding protein 3 significantly decreased, and total protein, albumin, alanine transaminase, alkaline phosphatase, γ-glutamyl transferase, activated partial thromboplastin time, prothrombin time, fibrinogen, D-dimer, fibrinogen degradation products, human epididymal protein 4, serum ferritin, and C-reactive protein were elevated. SARS-CoV-2 infection can affect hematopoiesis, hemostasis, coagulation, fibrinolysis, bone metabolism, thyroid, parathyroid glands, the liver, and the reproductive system.

Lopinavir/ritonavir and interferon combination therapy may help shorten the duration of viral shedding in patients with COVID-19: A retrospective study in two designated hospitals in Anhui, China.

Prolonged viral shedding may pose a threat to the control of coronavirus disease-2019 (COVID-19), and data on the duration of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) shedding are still limited, with the associated factors being unknown. All adult patients with laboratory-confirmed COVID-19 were included in this retrospective cross-sectional study in two designated hospitals during 21 January 2020 to 16 March 2020 in Anhui, China. In all patients, data on the duration of SARS-CoV-2 RNA shedding were analyzed by reviewing all RNA detection results during hospitalization. In addition, demographic, clinical, treatment, laboratory, and outcome data were also collected from electronic medical records. Factors associated with prolonged viral shedding were analyzed with the Cox proportional hazards model. Among 181 patients, the mean age was 44.3 ± 13.2 years, and 55.2% were male. The median duration of viral shedding from illness onset was 18.0 days (interquartile range [IQR], 15.0-24.0). Prolonged viral shedding was associated with longer hospital stays (P < .001) and higher medical costs (P < .001). The severity of COVID-19 had nothing to do with prolonged shedding. Moreover, the median time from onset to antiviral treatment initiation was 5.0 days (IQR, 3.0-7.0). Delayed antiviral treatment (hazard ratio [HR], 0.976; 95% confidence interval [CI], 0.962-0.990]) and lopinavir/ritonavir + interferon-α (IFN-α) combination therapy as the initial antiviral treatment (HR 1.649; 95% CI, 1.162-2.339) were independent factors associated with prolonged SARS-CoV-2 RNA shedding. SARS-CoV-2 showed prolonged viral shedding, causing increased hospital stays and medical costs. Early initiation of lopinavir/ritonavir + IFN-α combination therapy may help shorten the duration of SARS-CoV-2 shedding.

Rapid asymptomatic transmission of COVID-19 during the incubation period demonstrating strong infectivity in a cluster of youngsters aged 16-23 years outside Wuhan and characteristics of young patients with COVID-19: A prospective contact-tracing study.

BACKGROUND: The outbreak of coronavirus-disease-2019 (COVID-19) has rapidly spread to many places outside Wuhan. Previous studies on COVID-19 mostly included older hospitalized-adults. Little information on infectivity among and characteristics of youngsters with COVID-19 is available. METHODS: A cluster of 22 close-contacts of a 22-year-old male (Patient-Index) including youngsters with laboratory-confirmed COVID-19 and hospitalized close-contacts testing negative for severe-acute-respiratory-syndrome-coronavirus-2 (SARS-CoV-2) in Anhui Province, China was prospectively-traced. RESULTS: Since January 23, 2020, we enrolled a cluster of eight youngsters with COVID-19 (median age [range], 22 [16-23] years; six males) originating from Patient-Index returning from Wuhan to Hefei on January 19. Patient-Index visited his 16-year-old female cousin in the evening on his return, and met 15 previous classmates in a get-together on January 21. He reported being totally asymptomatic and were described by all his contacts as healthy on January 19-21. His very first symptoms were itchy eyes and fever developed at noon and in the afternoon on January 22, respectively. Seven youngsters (his cousin and six classmates) became infected with COVID-19 after a-few-hour-contact with Patient-Index. None of the patients and contacts had visited Wuhan (except Patient-Index), or had any exposure to wet-markets, wild-animals, or medical-institutes within three months. For affected youngsters, the median incubation-period was 2 days (range, 1-4). The median serial-interval was 1 day (range, 0-4). Half or more of the eight COVID-19-infected youngsters had fever, cough, sputum production, nasal congestion, and fatigue on admission. All patients had mild conditions. Six patients developed pneumonia (all mild; one bilateral) on admission. As of February 20, four patients were discharged. CONCLUSIONS: SARS-CoV-2-infection presented strong infectivity during the incubation-period with rapid transmission in this cluster of youngsters outside Wuhan. COVID-19 developed in these youngsters had fast onset and various nonspecific atypical manifestations, and were much milder than in older patients as previously reported.

Diagnostic utility of clinical laboratory data determinations for patients with the severe COVID-19.

The role of clinical laboratory data in the differential diagnosis of the severe forms of COVID-19 has not been definitely established. The aim of this study was to look for the warning index in severe COVID-19 patients. We investigated 43 adult patients with COVID-19. The patients were classified into mild group (28 patients) and severe group (15 patients). A comparison of the hematological parameters between the mild and severe groups showed significant differences in interleukin-6 (IL-6), d-dimer (d-D), glucose, thrombin time, fibrinogen, and C-reactive protein (P < .05). The optimal threshold and area under the receiver operator characteristic curve (ROC) of IL-6 were 24.3 and 0.795 µg/L, respectively, while those of d-D were 0.28 and 0.750 µg/L, respectively. The area under the ROC curve of IL-6 combined with d-D was 0.840. The specificity of predicting the severity of COVID-19 during IL-6 and d-D tandem testing was up to 93.3%, while the sensitivity of IL-6 and d-D by parallel test in the severe COVID-19 was 96.4%. IL-6 and d-D were closely related to the occurrence of severe COVID-19 in the adult patients, and their combined detection had the highest specificity and sensitivity for early prediction of the severity of COVID-19 patients, which has important clinical value.