CRISPR-Cas12a-assisted elimination of the non-specific signal from non-specific amplification in the Exponential Amplification Reaction.

Non-specific amplification is a major problem in nucleic acid amplification resulting in false-positive results, especially for exponential amplification reactions (EXPAR). Although efforts were made to suppress the influence of non-specific amplification, such as chemical blocking of the template's 3'-ends and sequence-independent weakening of template-template interactions, it is still a common problem in many conventional EXPAR reactions. In this study, we propose a novel strategy to eliminate the non-specific signal from non-specific amplification by integrating the CRISPR-Cas12a system into two-templates EXPAR. An EXPAR-Cas12a strategy named EXPCas was developed, where the Cas12a system acted as a filter to filter out non-specific amplificons in EXPAR, suppressing and eliminating the influence of non-specific amplification. As a result, the signal-to-background ratio was improved from 1.3 to 15.4 using this method. With microRNA-21 (miRNA-21) as a target, the detection can be finished in 40 min with a LOD of 103 fM and no non-specific amplification was observed.

Modular characterization of SARS-CoV-2 nucleocapsid protein domain functions in nucleocapsid-like assembly.

SARS-CoV-2 and its variants, with the Omicron subvariant XBB currently prevailing the global infections, continue to pose threats on public health worldwide. This non-segmented positive-stranded RNA virus encodes the multi-functional nucleocapsid protein (N) that plays key roles in viral infection, replication, genome packaging and budding. N protein consists of two structural domains, NTD and CTD, and three intrinsically disordered regions (IDRs) including the NIDR, the serine/arginine rich motif (SRIDR), and the CIDR. Previous studies revealed functions of N protein in RNA binding, oligomerization, and liquid-liquid phase separation (LLPS), however, characterizations of individual domains and their dissected contributions to N protein functions remain incomplete. In particular, little is known about N protein assembly that may play essential roles in viral replication and genome packing. Here, we present a modular approach to dissect functional roles of individual domains in SARS-CoV-2 N protein that reveals inhibitory or augmented modulations of protein assembly and LLPS in the presence of viral RNAs. Intriguingly, full-length N protein (NFL) assembles into ring-like architecture whereas the truncated SRIDR-CTD-CIDR (N182-419) promotes filamentous assembly. Moreover, LLPS droplets of NFL and N182-419 are significantly enlarged in the presence of viral RNAs, and we observed filamentous structures in the N182-419 droplets using correlative light and electron microscopy (CLEM), suggesting that the formation of LLPS droplets may promote higher-order assembly of N protein for transcription, replication and packaging. Together this study expands our understanding of the multiple functions of N protein in SARS-CoV-2.

Functional metal-organic framework as high-performance adsorbent for selective enrichment of pharmaceutical contaminants in aqueous samples

Facile and sensitive analysis methods for pharmaceutical contaminants in aqueous environment are of vital importance for water safety, especially when large amounts of anti-viral drugs are being used, discharged and accumulated. In this work, we used functional metal-organic framework (MOF) as high-performance adsorbent for selective enrichment of such pharmaceutical contaminants in aqueous samples. The MOF was synthesized via a new synthesis method previously developed by our group and immobilized on paper membrane to be used in solid-phase extraction (SPE) device. Different metal ions were anchored by MOF to screen out the adsorbent with the best affinity. The targets were a potential anti-COVID-19 drug favipiravir, and its structural and functional analogues (ingredients or intermediates, other anti-viral drugs). To deeply understand the adsorption mechanisms, quantum calculation and computational fluid dynamics (CFD) simulation were both applied. The experimental and in-silico results together demonstrated that the as-prepared MOF adsorbent possessed high affinity and fast dynamics. The established SPE-based liquid chromatography (LC) method worked well in the range of 10-1000 ng/mL, with only 3 mg of adsorbent per device and 5 mL sample needed, and no mass spectrometer (MS) included, which was very efficient compared to commercial adsorbents. The results met the current detection needs in the application scenario, and inspirable for later design of well-behaved adsorbents.

Expression of COVID-19 virus N gene in insect cells

In order to obtain COVID-19 nucleocapsid protein with similar function and activity to natural protein and apply it to practical detection. Firstly, according to Bac-to-bac insect expression system and synthetic COVID-19 nucleocapsid protein(N protein)sequence, BamH I and Xba I on pFastBacTMHTB vector were added to upstream and downstream primers respectively. The N gene was amplified by PCR technology, and T-Vector pMD19(simple)vector and pFastBacTMHTB vector were connected successively and recombinant plasmids pMD19-T(simple)-COV19-N and pFastBacTMHTB-COV19-N, and finally construct recombinant bacmid DH10 Bac-pFastBacTMHTB-COV19-N in DH10 Bac cells was expressed in insect cell Sf9. The recombinant protein was obtained and analyzed by SDS-PAGE and WB. The recombinant plasmid pMD19-T(simple)-COV19-N was identified by PCR and double enzyme digestion. The recombinant bacmid DH10 Bac-pFastBacTMHTB-COV19-N was constructed in DH10 Bac cells was identified by PCR and the expected two bands were 2 430,3 690 bp, respectively, which proved that the recombinant bacmid was successfully obtained. The recombinant bacmid was transfected into Sf9 insect cells. At the same time, the recombinant GFP protein control group was established. After 120 h of transfection, the recombinant N protein and recombinant GFP protein were collected and samples were prepared;SDS-PAGE and WB analysis were carried out respectively. HRP-His labeled antibody was used to verify that the transfection was successful, and both recombinant N protein and recombinant GFP protein were successfully expressed in Sf9 cells. The experimental results were consistent with the expectation, and the size of recombinant N protein band was about 46 ku. The eukaryotic expression vector of respiratory coronavirus N gene was successfully constructed and successfully expressed in insect cells, which provides an experimental basis for the establishment of ELISA detection methods and other related research.

Psychological disorders and related factors among Chinese adolescent earthquake survivors during the COVID-19 pandemic

We studied the mental health of adolescent earthquake survivors during the COVID-19 pandemic by measuring their psychological disorders with the Zung Self-Rating Depression Scale (SDS) and their risk factors with the Zung Self-Rating Anxiety Scale (SAS). We conducted an exploratory factor analysis and logistic regression modeling and the results revealed that there were high levels of depression and anxiety among the 385 participants. Significant indicators of depression as measured by the SDS were family monthly income, years of education of the adolescents, and marital status of parents. The SAS measured family monthly income as a significant factor predicting anxiety. This study will provide references for future psychological support and early interventions for disadvantaged groups during the ongoing COVID-19 pandemic.

Investigating the Role of Perceived Information Overload on COVID-19 Fear: A Moderation Role of Fake News Related to COVID-19

During crises and uncertain situations such as the coronavirus disease 2019 (COVID-19) pandemic, social media plays a key function because it allows people to seek and share news, as well as personal views and ideas with each other in real time globally. Past research has highlighted the implications of social media during disease outbreaks;nevertheless, this study refers to the possible negative effects of social media usage by individuals in the developing country during the COVID-19 epidemic lockdown. Specifically, this study investigates the COVID-19 fear using the survey data collected from a developing country. In total, 880 entries were used to analyze the COVID-19 fear using the AMOS software. Findings indicated that information-seeking and sharing behavior of individuals on social media has a significant impact on perceived COVID-19 information overload. Perceived COVID-19 information overload has a positive impact on COVID-19 fear. In addition, fake news related to COVID-19 strengthens the relationship between perceived COVID-19 information overload and COVID-19 fear. The implication and limitations of the study are also discussed in the final section of the study.

E-waste dismantling-related occupational and routine exposure to melamine and its derivatives: Estimating exposure via dust ingestion and hand-to-mouth contact.

Melamine (MEL) and its derivatives are increasingly applied as nitrogenous flame retardants in consumer products. Nevertheless, limited information is available on their environmental occurrence and subsequent human exposure via multiple exposure pathways. In this study, we analysed MEL and its derivatives in dust (indication of the dust ingestion route) and hand wipe samples (indication of the hand-to-mouth route) collected in various microenvironments. The levels of ∑MELs in both dust (median: 24,100 ng/g) and participant hand samples (803 ng/m2) collected in e-waste dismantling workshops were significantly higher than those in samples collected in homes (15,600 ng/g and 196 ng/m2, respectively), dormitories (13,100 ng/g and 227 ng/m2, respectively) and hotel rooms (11,800 ng/g and 154 ng/m2, respectively). Generally, MEL dominated in dust samples collected in e-waste dismantling workshops, whereas cyanuric acid dominated in hand wipe samples. This may occur partly because the latter is an ingredient in disinfection products, which are more frequently employed in daily lives during the COVID-19 pandemic. Exposure assessment suggests that dust ingestion is an important exposure pathway among dismantling workers and the general population, whereas hand-to-mouth contact could not be overlooked in certain populations, such as children and dismantling workers not wear gloves at work.

Functional metal-organic framework as high-performance adsorbent for selective enrichment of pharmaceutical contaminants in aqueous samples

Facile and sensitive analysis methods for pharmaceutical contaminants in aqueous environment are of vital importance for water safety, especially when large amounts of anti-viral drugs are being used, discharged and accumulated. In this work, we used functional metal-organic framework (MOF) as high-performance adsorbent for selective enrichment of such pharmaceutical contaminants in aqueous samples. The MOF was synthesized via a new synthesis method previously developed by our group and immobilized on paper membrane to be used in solid-phase extraction (SPE) device. Different metal ions were anchored by MOF to screen out the adsorbent with the best affinity. The targets were a potential anti-COVID-19 drug favipiravir, and its structural and functional analogues (ingredients or intermediates, other anti-viral drugs). To deeply understand the adsorption mechanisms, quantum calculation and computational fluid dynamics (CFD) simulation were both applied. The experimental and in-silico results together demonstrated that the as-prepared MOF adsorbent possessed high affinity and fast dynamics. The established SPE-based liquid chromatography (LC) method worked well in the range of 10-1000 ng/mL, with only 3 mg of adsorbent per device and 5 mL sample needed, and no mass spectrometer (MS) included, which was very efficient compared to commercial adsorbents. The results met the current detection needs in the application scenario, and inspirable for later design of well-behaved adsorbents.

Detections of major respiratory pathogens in pneumonia patients in early phase of COVID-19 epidemic in Tongzhou, Beijing

Objective: To understand the infection status of major respiratory pathogens in pneumonia patients in the early phase of coronavirus disease 2019 (COVID-19) epidemic (January-March, 2020) in Tongzhou district of Beijing.

SARS-CoV-2 impairs the disassembly of stress granules and promotes ALS-associated amyloid aggregation.

The nucleocapsid (N) protein of SARS-CoV-2 has been reported to have a high ability of liquid-liquid phase separation, which enables its incorporation into stress granules (SGs) of host cells. However, whether SG invasion by N protein occurs in the scenario of SARS-CoV-2 infection is unknow, neither do we know its consequence. Here, we used SARS-CoV-2 to infect mammalian cells and observed the incorporation of N protein into SGs, which resulted in markedly impaired self-disassembly but stimulated cell cellular clearance of SGs. NMR experiments further showed that N protein binds to the SG-related amyloid proteins via non-specific transient interactions, which not only expedites the phase transition of these proteins to aberrant amyloid aggregation in vitro, but also promotes the aggregation of FUS with ALS-associated P525L mutation in cells. In addition, we found that ACE2 is not necessary for the infection of SARS-CoV-2 to mammalian cells. Our work indicates that SARS-CoV-2 infection can impair the disassembly of host SGs and promote the aggregation of SG-related amyloid proteins, which may lead to an increased risk of neurodegeneration.

Probability-Based Estimates of Severe Acute Respiratory Syndrome Coronavirus 2 Seroprevalence and Detection Fraction, Utah, USA.

We aimed to generate an unbiased estimate of the incidence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in 4 urban counties in Utah, USA. We used a multistage sampling design to randomly select community-representative participants >12 years of age. During May 4-June 30, 2020, we collected serum samples and survey responses from 8,108 persons belonging to 5,125 households. We used a qualitative chemiluminescent microparticle immunoassay to detect SARS-CoV-2 IgG in serum samples. We estimated the overall seroprevalence to be 0.8%. The estimated seroprevalence-to-case count ratio was 2.5, corresponding to a detection fraction of 40%. Only 0.2% of participants from whom we collected nasopharyngeal swab samples had SARS-CoV-2-positive reverse transcription PCR results. SARS-CoV-2 antibody prevalence during the study was low, and prevalence of PCR-positive cases was even lower. The comparatively high SARS-CoV-2 detection rate (40%) demonstrates the effectiveness of Utah's testing strategy and public health response.

AXL is a candidate receptor for SARS-CoV-2 that promotes infection of pulmonary and bronchial epithelial cells.

The current coronavirus disease 2019 (COVID-19) pandemic presents a global public health challenge. The viral pathogen responsible, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), binds to the host receptor ACE2 through its spike (S) glycoprotein, which mediates membrane fusion and viral entry. Although the role of ACE2 as a receptor for SARS-CoV-2 is clear, studies have shown that ACE2 expression is extremely low in various human tissues, especially in the respiratory tract. Thus, other host receptors and/or co-receptors that promote the entry of SARS-CoV-2 into cells of the respiratory system may exist. In this study, we found that the tyrosine-protein kinase receptor UFO (AXL) specifically interacts with the N-terminal domain of SARS-CoV-2 S. Using both a SARS-CoV-2 virus pseudotype and authentic SARS-CoV-2, we found that overexpression of AXL in HEK293T cells promotes SARS-CoV-2 entry as efficiently as overexpression of ACE2, while knocking out AXL significantly reduces SARS-CoV-2 infection in H1299 pulmonary cells and in human primary lung epithelial cells. Soluble human recombinant AXL blocks SARS-CoV-2 infection in cells expressing high levels of AXL. The AXL expression level is well correlated with SARS-CoV-2 S level in bronchoalveolar lavage fluid cells from COVID-19 patients. Taken together, our findings suggest that AXL is a novel candidate receptor for SARS-CoV-2 which may play an important role in promoting viral infection of the human respiratory system and indicate that it is a potential target for future clinical intervention strategies.

Therapeutic experience of an 89-year-old high-risk patient with incarcerated cholecystolithiasis: A case report and literature review.

BACKGROUND: The global pandemic of coronavirus disease 2019 pneumonia poses a particular challenge to the emergency surgical treatment of elderly patients with high-risk acute abdominal diseases. Elderly patients are a high-risk group for surgical treatment. If the incarceration of gallstones cannot be relieved, emergency surgery is unavoidable. CASE SUMMARY: We report an 89-year-old male patient with acute gangrenous cholecystitis and septic shock induced by incarcerated cholecystolithiasis. He had several coexisting, high-risk underlying diseases, had a history of radical gastrectomy for gastric cancer, and was taking aspirin before the operation. Nevertheless, he underwent emergency laparoscopic cholecystectomy, with maintenance of postoperative heart and lung function, successfully recovered, and was discharged on day 8 after the operation. CONCLUSION: Emergency surgery for elderly patients with acute abdominal disease is safe and feasible during the coronavirus disease 2019 pandemic, the key is to abide strictly by the hospital's epidemic prevention regulations, fully implement the epidemic prevention procedure for emergency surgery, fully prepare before the operation, accurately perform the operation, and carefully manage the patient postoperatively.

Expert consensus on protocol of rehabilitation for COVID-19 patients using framework and approaches of WHO International Family Classifications.

Coronavirus disease 2019 (COVID-19) has widely spread all over the world and the numbers of patients and deaths are increasing. According to the epidemiology, virology, and clinical practice, there are varying degrees of changes in patients, involving the human body structure and function and the activity and participation. Based on the World Health Organization (WHO) International Classification of Functioning, Disability and Health (ICF) and its biopsychosocial model of functioning, we use the WHO Family of International Classifications (WHO-FICs) framework to form an expert consensus on the COVID-19 rehabilitation program, focusing on the diagnosis and evaluation of disease and functioning, and service delivery of rehabilitation, and to establish a standard rehabilitation framework, terminology system, and evaluation and intervention systems based the WHO-FICs.