Rapid Detection of Predominant SARS-CoV-2 Variants Using Multiplex High-Resolution Melting Analysis.

Coronavirus disease 2019, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), poses a considerable threat to global public health. This study developed and evaluated a rapid, low-cost, expandable, and sequencing-free high-resolution melting (HRM) assay for the direct detection of SARS-CoV-2 variants. A panel of 64 common bacterial and viral pathogens that can cause respiratory tract infections was employed to evaluate our method's specificity. Serial dilutions of viral isolates determined the sensitivity of the method. Finally, the assay's clinical performance was assessed using 324 clinical samples with potential SARS-CoV-2 infection. Multiplex HRM analysis accurately identified SARS-CoV-2 (as confirmed with parallel reverse transcription-quantitative PCR [qRT-PCR] tests), differentiating between mutations at each marker site within approximately 2 h. For each target, the limit of detection (LOD) was lower than 10 copies/reaction (the LOD of N, G142D, R158G, Y505H, V213G, G446S, S413R, F486V, and S704L was 7.38, 9.72, 9.96, 9.96, 9.50, 7.80, 9.33, 8.25, and 8.25 copies/reaction, respectively). No cross-reactivity occurred with organisms of the specificity testing panel. In terms of variant detection, our results had a 97.9% (47/48) rate of agreement with standard Sanger sequencing. The multiplex HRM assay therefore offers a rapid and simple procedure for detecting SARS-CoV-2 variants. IMPORTANCE In the face of the current severe situation of increasing SARS-CoV-2 variants, we developed an upgraded multiplex HRM method for the predominant SARS-CoV-2 variants based on our original research. This method not only could identify the variants but also could be utilized in subsequent detection of novel variants since the assay has great performance in terms of flexibility. In summary, the upgraded multiplex HRM assay is a rapid, reliable, and economical detection method, which could better screen prevalent virus strains, monitor the epidemic situation, and help to develop measures for the prevention and control of SARS-CoV-2.

Influences of Problem-Based Online Learning on the Learning Outcomes of Learners

Influenced by the COVID-19 pandemic, online learning has become a major learning mode for most university students in China. For high-quality online teaching, teachers must optimize the teaching mode and strengthen the enthusiasm of students in classroom learning. In this study, a comparative experiment was carried out based on the course Cross-border E-commerce Customer Services for sophomore E-commerce majors at Huanghuai University in Zhumadian City, Henan Province, China. In the experiment, the teaching effect of the problem-based learning (PBL) mode in higher education was verified and gender differences in learning outcomes of learners who accepted PBL were analyzed. Results showed a significant difference at the 0.01 level between pre- and post-test results of the experimental group (t=−11.367, p=0.000), proving the teaching effect of PBL. The academic performances of the experimental class are significantly better compared with those of the control class, indicating that PBL is more beneficial to improve learning outcomes of students than traditional teaching methods. Gender has significant influences on post-test results at the 0.01 level (p=0.002<0.01). The median of final exam scores of male students is obviously higher than that of female students. Research conclusions can provide important references to test the effectiveness of PBL in learning outcomes of a specific subject, construct a PBL mode that can promote deep learning, and facilitate development of university teaching from knowledge teaching to core quality training of students.

Variation in synonymous evolutionary rates in the SARS-CoV-2 genome.

Introduction: Coronavirus disease 2019 is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Influential variants and mutants of this virus continue to emerge, and more effective virus-related information is urgently required for identifying and predicting new mutants. According to earlier reports, synonymous substitutions were considered phenotypically silent; thus, such mutations were frequently ignored in studies of viral mutations because they did not directly cause amino acid changes. However, recent studies have shown that synonymous substitutions are not completely silent, and their patterns and potential functional correlations should thus be delineated for better control of the pandemic. Methods: In this study, we estimated the synonymous evolutionary rate (SER) across the SARS-CoV-2 genome and used it to infer the relationship between the viral RNA and host protein. We also assessed the patterns of characteristic mutations found in different viral lineages. Results: We found that the SER varies across the genome and that the variation is primarily influenced by codon-related factors. Moreover, the conserved motifs identified based on the SER were found to be related to host RNA transport and regulation. Importantly, the majority of the existing fixed-characteristic mutations for five important virus lineages (Alpha, Beta, Gamma, Delta, and Omicron) were significantly enriched in partially constrained regions. Discussion: Taken together, our results provide unique information on the evolutionary and functional dynamics of SARS-CoV-2 based on synonymous mutations and offer potentially useful information for better control of the SARS-CoV-2 pandemic.

The impact of COVID-19 on kidney transplant care.

The SARS-CoV-2 virus precipitated the coronavirus 2019 (COVID-19) pandemic, which placed considerable strain on healthcare systems and necessitated immediate and rapid alterations in the delivery of healthcare. In the transplant population, COVID-19 directly impacts an inherently vulnerable population in the setting of immunosuppression and co-morbidities, but also further complicates the clinical evaluation and management of kidney transplant candidates and recipients in a strained healthcare environment being challenged by the pandemic. Many transplant centers around the world saw mortality rate spikes in organ recipients related to COVID-19, and changes in care delivery abound. This review evaluates the care of the kidney transplant patient through all phases of the process including pre-operative evaluations, perioperative care, post-transplantation considerations, and how the global pandemic has changed the way we care for our patients.

Rational Design and Synthesis of D-galactosyl Lysophospholipids as Selective Substrates and non-ATP-competitive Inhibitors of Phosphatidylinositol Phosphate Kinases.

Phosphatidylinositol phosphate kinases (PIPKs) produce lipid signaling molecules and have been attracting increasing attention as drug targets for cancer, neurodegenerative diseases, and viral infection. Given the potential cross-inhibition of kinases and other ATP-utilizing enzymes by ATP-competitive inhibitors, targeting the unique lipid substrate binding site represents a superior strategy for PIPK inhibition. Here, by taking advantage of the nearly identical stereochemistry between myo-inositol and D-galactose, we designed and synthesized a panel of D-galactosyl lysophospholipids, one of which was found to be a selective substrate of phosphatidylinositol 4-phosphate 5-kinase. Derivatization of this compound led to the discovery of a human PIKfyve inhibitor with an apparent IC50 of 6.2 µM, which significantly potentiated the inhibitory effect of Apilimod, an ATP-competitive PIKfyve inhibitor under clinical trials against SARS-CoV-2 infection and amyotrophic lateral sclerosis. Our results provide the proof of concept that D-galactose-based phosphoinositide mimetics can be developed into artificial substrates and new inhibitors of PIPKs.

The impact of COVID-19 on kidney transplant care

The SARS-CoV-2 virus precipitated the coronavirus 2019 (COVID-19) pandemic, which placed considerable strain on healthcare systems and necessitated immediate and rapid alterations in the delivery of healthcare. In the transplant population, COVID-19 directly impacts an inherently vulnerable population in the setting of immunosuppression and co-morbidities, but also further complicates the clinical evaluation and management of kidney transplant candidates and recipients in a strained healthcare environment being challenged by the pandemic. Many transplant centers around the world saw mortality rate spikes in organ recipients related to COVID-19, and changes in care delivery abound. This review evaluates the care of the kidney transplant patient through all phases of the process including pre-operative evaluations, perioperative care, post-transplantation considerations, and how the global pandemic has changed the way we care for our patients.

Broadly neutralizing aptamers to SARS-CoV-2: A diverse panel of modified DNA antiviral agents.

Since its discovery, COVID-19 has rapidly spread across the globe and has had a massive toll on human health, with infection mortality rates as high as 10%, and a crippling impact on the world economy. Despite numerous advances, there remains an urgent need for accurate and rapid point-of-care diagnostic tests and better therapeutic treatment options. To contribute chemically distinct, non-protein-based affinity reagents, we report here the identification of modified DNA-based aptamers that selectively bind to the S1, S2, or receptor-binding domain of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein. Several aptamers inhibit the binding of the spike protein to its cell-surface receptor angiotensin-converting enzyme 2 (ACE2) and neutralize authentic SARS-CoV-2 virus in vitro, including all variants of concern. With a high degree of nuclease resistance imparted by the base modifications, these reagents represent a new class of molecules with potential for further development as diagnostics or therapeutics.

Molecular evolution of human coronavirus-NL63, -229E, -HKU1 and -OC43 in hospitalized children in China.

Human coronaviruses (HCoVs) HCoV-NL63, HCoV-229E, HCoV-HKU1 and HCoV-OC43 have been circulated in the human population worldwide, and they are associated with a broad range of respiratory diseases with varying severity. However, there are neither effective therapeutic drugs nor licensed vaccines available for the treatment and prevention of infections by the four HCoVs. In this study, we collected nasopharyngeal aspirates of children hospitalized for respiratory tract infection in China during 2014-2018 and conducted next-generation sequencing. Sequences of four HCoVs were then selected for an in-depth analysis. Genome sequences of 2 HCoV-NL63, 8 HCoV-229E, 2 HCoV-HKU1, and 6 HCoV-OC43 were obtained. Based on the full-length S gene, a strong temporal signal was found in HCoV-229E and the molecular evolutionary rate was 6 × 10-4 substitutions/site/year. Based on the maximum-likelihood (ML) phylogenetic tree of complete S gene, we designated H78 as a new sub-genotype C2 of HCoV-HKU1, and the obtained P43 sequence was grouped into the reported novel genotype K of HCoV-OC43 circulating in Guangzhou, China. Based on the complete genome, potential recombination events were found to occur as two phenomena, namely intraspecies and interspecies. Moreover, we observed two amino acid substitutions in the S1 subunit of obtained HCoV-NL63 (G534V) and HCoV-HKU1 (H512R), while residues 534 and 512 are important for the binding of angiotensin-converting enzyme 2 and neutralizing antibodies, respectively. Our findings might provide a clue for the molecular evolution of the four HCoVs and help in the early diagnosis, treatment and prevention of broad-spectrum HCoV infection.

A network view of human immune system and virus-human interaction.

The immune system is highly networked and complex, which is continuously changing as encountering old and new pathogens. However, reductionism-based researches do not give a systematic understanding of the molecular mechanism of the immune response and viral pathogenesis. Here, we present HUMPPI-2022, a high-quality human protein-protein interaction (PPI) network, containing > 11,000 protein-coding genes with > 78,000 interactions. The network topology and functional characteristics analyses of the immune-related genes (IRGs) reveal that IRGs are mostly located in the center of the network and link genes of diverse biological processes, which may reflect the gene pleiotropy phenomenon. Moreover, the virus-human interactions reveal that pan-viral targets are mostly hubs, located in the center of the network and enriched in fundamental biological processes, but not for coronavirus. Finally, gene age effect was analyzed from the view of the host network for IRGs and virally-targeted genes (VTGs) during evolution, with IRGs gradually became hubs and integrated into host network through bridging functionally differentiated modules. Briefly, HUMPPI-2022 serves as a valuable resource for gaining a better understanding of the composition and evolution of human immune system, as well as the pathogenesis of viruses.

Clinical value of admission blood glucose level on prognosis of COVID-19 patients

Objective: To explore the clinical value of admission blood glucose level on prognosis of COVID- 19 patients. Methods A total of 420 novel coronavirus pneumonia (COVID-19) patients admitted to Tongji Hospital of Tongji MedicalCollege from January 18, 2020 to February 26, 2020 were selected as the subjects of study. The data of diabetes or not, admissionblood glucose level(GLU), clinical severity grade were collected through the electronic medical record system, and the outcome, which defined as in-hospital motality, was also monitored. The patients were divided into diabetes group and non-diabetes groupin terms of the complication of diabetes, and then, firstly, stratified these two groups into survival subgroup and non-survivalsubgroup in according to the event of in-hospital motality, GLU between these two subgroups were compared. Secondly, according to the clinical severity grade, these two groups were stratified into moderate subgroup, severe subgroup and criticalsubgroup, and GLU among these subgroups were also compared. Thirdly, according to the admission blood glucose level, stratified these two groups into GLU 3.9~7.8 mmol /L subgroup, GLU 7.8~10.0 mmol/L subgroup and GLU>10.0 mmol/Lsubgroup, the in-hospital motality rates among these subgroups were compared. Finally, the multivariate logistic regression wasused to explore whether increased GLU were independent risk factor for in-hospital motality in diabetes group and non-diabetesgroup when adjusted for sex, age and underlying disease. Results In non-diabetes group, compared with Survival subgroup, GLUwas significantly elevated in non-Survival subgroup[6.96(5.95, 8.23)mmol/L vs 5.96 (5.32, 6.92) mmol/L], the difference wasstatistically significant(U=6047.0, P < 0.001), but in diabetes group, there was no significant difference between non-survivalsubgroup and Survival subgroup [12.42(8.41, 18.17) mmol/L vs 9.88 (7.79, 14.02) mmol/L], the difference was statisticallysignificant(U=1 200.5, P=0.059).In Non-diabetes group, GLU elevated remarkably along with the clinical severity gradeincreased, moderate subgroup, severe subgroup, critical subgroup GLU were 5.87(5.24, 6.69) mmol/L, 6.94(5.95, 7.90) mmol/L,9.73 (6.22, 11.64) mmol/L, the difference were statistically significant, respectively(U=723.0~4978.0, all P < 0.01). However indiabetes group, there was no significant difference on GLU when the clinical severity grade increased, moderate subgroup, severesubgroup, critical subgroup GLU were 9.88(7.81, 11.93)mmol/L, 12.42(8.43, 16.94)mmol/L, 11.43(7.89, 18.76)mmol/L, the difference were statistically significant, respectively (U=262.0~946.5, all P>0.05).In non-diabetes group, GLU> 10.0 mmol/L subgroup had the hightest in-hospital motality rate (72.0%) among all three subgroups, the differences were statisticallysignificant(X2=24.607, 9.625, all P < 0.01), when compared between GLU 3.9~7.8 mmol/L subgroup (in-hospital motality rate24.8%) and GLU 7.8~10.0 mmol/L subgroup (in-hospital motality rate 30.0%), there was no significant difference on in-hospitalmotality rate (X2=0.383, P > 0.05). However, in diabetes group, along with GLU increased, it had no significant difference on inhospitalmotality rate, GLU 3.9~7.8 mmol/L subgroup, GLU 7.8~10.0 mmol/L subgroup, GLU> 10.0 mmol/L subgroup, the inhospitalmotality rate were 34.8%, 41.4%, 49.2%, respectively(X2=0.236~1.380, all P> 0.05). Multivariate logistic regressionshowed, in non-diabets group, GLU>10.0 mmol/L was the independent risk factor when adjusted for sex, age and underlyingdisease, odds ratio was 7.969, and 95% confidence interval was 3.022~21.013, but in diabets group.It seemed that GLU>10.0 mmol/L was not the independent risk factor. Conclusion Admission blood glucose is a good predictor for disease severity andoutcome in non-diabetes patients with COVID-19. When admission hyperglycemia occurs, it tends to predict a poor prognosis.

Research on older people's health information search behavior based on risk perception in social networks-A case study in China during COVID-19.

Objective: COVID-19 has caused great loss of human life and livelihoods. The dissemination of health information in online social networks increased during the pandemic's quarantine. Older people are the most vulnerable group in sudden public health emergencies, and they have the disadvantage of infection rates and online search for health information. This study explores the relationship between the health risk perception and health information search behavior of older people in social networks, to help them make better use of the positive role of social networks in public health emergencies. Method: Based on the Risk Information Search and Processing model, and in the specific context of COVID-19, this study redefines health risk perception as a second-order construct of four first-order factors (perceived probability, perceived severity, perceived controllability, and perceived familiarity), and constructs a research model of the health risk perception and health information search behavior of older people. An online survey of people over 55 years old was conducted through convenience sampling in China from February 2020 to March 2020. Results: A total of 646 older adults completed the survey. The structural equation model showed that health risk perception is a second-order factor (H1), that health risk perception has significant positive effects on health information search behavior (H2: ß = 0.470, T = 11.577, P < 0.001), and that health risk perception has significant positive effects on affective response (H3: ß = 0.536, T = 17.356, P < 0.001). In addition, affective response has a significant positive mediating effect on information sufficiency (H4: ß = 0.435, T = 12.231, P < 0.001), and information sufficiency has a significant positive mediating effect on health information search behavior (H5: ß = 0.136, T = 3.081, P = 0.002). Conclusion: The study results indicate that the health risk perception of older people during the COVID-19 outbreak not only directly affected their health information search behavior, but also had an indirect impact on their health information search behavior by affecting affective response and information sufficiency.

SARS-CoV-2 antibody persistence after five and twelve months: A cohort study from South-Eastern Norway.

OBJECTIVES: To assess total antibody levels against Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS CoV-2) spike protein up to 12 months after Coronavirus Disease (COVID-19) infection in non-vaccinated individuals and the possible predictors of antibody persistence. METHODS: This is the first part of a prospective multi-centre cohort study. PARTICIPANTS: The study included SARS-CoV-2 real-time polymerase chain reaction (RT-PCR) positive and negative participants in South-Eastern Norway from February to December 2020. Possible predictors of SARS-CoV-2 total antibody persistence was assessed. The SARS-CoV-2 total antibody levels against spike protein were measured three to five months after PCR in 391 PCR-positive and 703 PCR-negative participants; 212 PCR-positive participants were included in follow-up measurements at 10 to 12 months. The participants completed a questionnaire including information about symptoms, comorbidities, allergies, body mass index (BMI), and hospitalisation. PRIMARY OUTCOME: The SARS-CoV-2 total antibody levels against spike protein three to five and 10 to 12 months after PCR positive tests. RESULTS: SARS-CoV-2 total antibodies against spike protein were present in 366 (94%) non-vaccinated PCR-positive participants after three to five months, compared with nine (1%) PCR-negative participants. After 10 to 12 months, antibodies were present in 204 (96%) non-vaccinated PCR-positive participants. Of the PCR-positive participants, 369 (94%) were not hospitalised. The mean age of the PCR-positive participants was 48 years (SD 15, range 20-85) and 50% of them were male. BMI ≥ 25 kg/m2 was positively associated with decreased antibody levels (OR 2.34, 95% CI 1.06 to 5.42). Participants with higher age and self-reported initial fever with chills or sweating were less likely to have decreased antibody levels (age: OR 0.97, 95% CI 0.94 to 0.99; fever: OR 0.33, 95% CI 0.13 to 0.75). CONCLUSION: Our results indicate that the level of SARS-CoV-2 total antibodies against spike protein persists for the vast majority of non-vaccinated PCR-positive persons at least 10 to 12 months after mild COVID-19.

Behavioral Intention and Its Predictors toward COVID-19 Booster Vaccination among Chinese Parents: Applying Two Behavioral Theories.

The booster vaccination of COVID-19 is being implemented in most parts of the world. This study used behavioral psychology to investigate the predictors of parents' intentions regarding the COVID-19 booster vaccination for their children. This is a cross-sectional study with a self-designed questionnaire based on two behavioral theories-protective motivation theory (PMT) and theory of planned behavior (TPB). A stratified multi-stage sampling procedure was conducted in Nanjing, China, and multivariable regression analyses were applied to examine the parents' intentions. The intention rate was 87.3%. The response efficacy (ORa = 2.238, 95% CI: 1.360-3.682) and response cost (ORa = 0.484, 95% CI: 0.319-0.732) in the PMT, were significant psychological predictors of parents' intentions, and so were the attitude (ORa = 2.619, 95% CI: 1.480-4.636) and behavioral control (ORa = 3.743, 95% CI: 2.165-6.471) in the TPB. The findings of crucial independent predictors in the PMT and TPB constructs inform the evidence-based formulation and implementation of strategies for booster vaccination in children.

Behavioral Intention and Its Predictors toward COVID-19 Booster Vaccination among Chinese Parents: Applying Two Behavioral Theories

The booster vaccination of COVID-19 is being implemented in most parts of the world. This study used behavioral psychology to investigate the predictors of parents' intentions regarding the COVID-19 booster vaccination for their children. This is a cross-sectional study with a self-designed questionnaire based on two behavioral theories-protective motivation theory (PMT) and theory of planned behavior (TPB). A stratified multi-stage sampling procedure was conducted in Nanjing, China, and multivariable regression analyses were applied to examine the parents' intentions. The intention rate was 87.3%. The response efficacy (ORa = 2.238, 95% CI: 1.360–3.682) and response cost (ORa = 0.484, 95% CI: 0.319–0.732) in the PMT, were significant psychological predictors of parents' intentions, and so were the attitude (ORa = 2.619, 95% CI: 1.480–4.636) and behavioral control (ORa = 3.743, 95% CI: 2.165–6.471) in the TPB. The findings of crucial independent predictors in the PMT and TPB constructs inform the evidence-based formulation and implementation of strategies for booster vaccination in children.

Associations Between Online Learning, Smartphone Addiction Problems, and Psychological Symptoms in Chinese College Students After the COVID-19 Pandemic.

Background: Smartphone-based online education gained popularity during and after the COVID-19 pandemic. Although recent studies have highlighted the association between problematic smartphone use (PSU) and mental health symptoms, the potential role of online learning in this relationship remains unclear. This study aimed to analyze the relationships between higher education modes, PSU, and related psychological symptoms in university students. Methods: A total of 1,629 Chinese university students from five provinces completed a web-based questionnaire survey between March 2020 and October 2021. Demographic characteristics and learning conditions were recorded. All participants completed the Smartphone Addiction Scale-Short Version, Patient Health Questionnaire, Generalized Anxiety Disorder Scale, and Athens Insomnia Scale. Multiple regressions models and stratified analyses were used to examine the association between online education mode, PSU, and symptoms of depression, anxiety, and insomnia. Results: The prevalence of PSU was 58.5%. Students who relied primarily on online learning had a higher prevalence of depressive symptoms (29.95% vs. 22.24%), anxiety symptoms (25.13% vs. 18.91%), and insomnia symptoms (75.89% vs. 70.27%) than those who relied on traditional face-to-face learning (Ps < 0.05). After adjusting for covariates, subjects with PSU were more likely to report depressive symptoms (AdjOR = 3.14, 95% CI = 2.26-4.37), anxiety symptoms (AdjOR = 3.73, 95% CI = 2.13-4.59), and insomnia symptoms (AdjOR = 2.96, 95% CI = 2.23-3.92) than those without PSU. Furthermore, the associations of PSU with depressive symptoms (OR = 4.66 vs. 2.33, P for interaction = 0.015) and anxiety symptoms (OR = 6.05 vs. 2.94, P for interaction = 0.021) were more pronounced in the online learning group. Conclusion: Our study provides preliminary evidence that Chinese university students have serious smartphone addiction problems, which are associated with depressive, anxiety, and insomnia symptoms. Online learning is found to exacerbate PSU and mental health problems. Our findings provide valuable information for targeted psychological interventions in the post-COVID-19 era.

Research on antibody changes and nucleic acid clearance in COVID-19 patients treated with convalescent plasma

Purpose: To investigate changes in the production of IgM and IgG antibodies and the negative transformation of viral nucleic acids in COVID-19 patients after convalescent plasma therapy, and also to discuss the clinical therapeutic effect, so as to provide a basis for the treatment of COVID-19 using specific antibodies. Methods: The convalescent plasma of recovered patients from COVID-19 was used to treat other patients, and the levels of antibodies IgM and IgG and the nucleic acid genes ORF1ab and N in the patients were tested regularly for statistical comparison and analysis. Results: In general, the Ct value and concentration of IgM and IgG antibodies in the plasma infusion group were significantly higher (1-3 times higher) than those in the non-plasma infusion group, respectively, but these differences were not significant (P>0.05). However, the content of antibodies in severe patients in the plasma transfusion group was significantly higher than those in the non-plasma transfusion group at discharge, the results being statistically significant (P<0.05). Conclusions: The application of convalescent plasma significantly increases the antibody content in severe and critical inpatients, effectively enhances immune function, accelerates the clearance of virus and the nucleic acid negative conversion rate, and significantly promotes early improvement in COVID-19 patients.

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