Reevaluation of von Willebrand disease diagnosis in a Croatian paediatric cohort combining bleeding scores,phenotypic laboratory assays and next generation sequencing: a pilot study

IntroductionThis study reevaluated von Willebrand disease (vWD) diagnosis in a Croatian paediatric cohort by combining bleeding scores (BS), phenotypic laboratory testing, and next-generation sequencing (NGS).Materials and methodsA total of 25 children (11 males and 14 females, median age 10 years, from 2 to 17) previously diagnosed with vWD were included. BS were calculated using an online bleeding assessment tool. Phenotypic laboratory analyses included platelet count, platelet function analyser closure times, prothrombin time, activated partial thromboplastin time, von Willebrand factor antigen (vWF:Ag), vWF gain-of-function mutant glycoprotein Ib binding activity (vWF:GPIbM), vWF collagen binding activity (vWF:CBA), factor VIII activity (FVIII:C) and multimeric analysis. Next-generation sequencing covered regions of both vWF and FVIII genes and was performed on MiSeq (Illumina, San Diego, USA).ResultsDisease-associated variants identified in 15 patients comprised 11 distinct heterozygous vWF gene variants in 13 patients and one novel FVIII gene variant (p.Glu2085Lys) in two male siblings. Four vWF variants were novel (p.Gln499Pro, p.Asp1277Tyr, p.Asp1277His, p.Lys1491Glu). Three patients without distinctive variants had vWF:GPIbM between 30 and 50%. Patients with identified vWF gene variants had statistically significant lower values of vWF:GPIbM (P = 0.002), vWF:Ag (P = 0.007), vWF:CBA (P < 0.001) and FVIII:C (P = 0.002), compared to those without. Correlations between BS and phenotypic laboratory test results were not statistically significant for either of the tests.ConclusionThe applied diagnostic approach confirmed the diagnosis of vWD in 13 patients and mild haemophilia A in two. Limited utility of BS in the paediatric population was evidenced.     

Current status of the lateral flow immunoassay for the detection of SARS-CoV-2 in nasopharyngeal swabs

Early detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and diagnosis of coronavirus disease 2019 (COVID-19) are priorities during the pandemic. Symptomatic and suspected asymptomatic individuals should be tested for COVID-19 to confirm infection and to be excluded from social interactions. As molecular testing capacity is overloaded during the pandemic, rapid antigen tests, such as lateral flow immunoassays (LFIAs), can be a useful tool as they allow greater test availability and obtain results in a very short time. This short review aims to present the analytical properties of LFIAs in the detection of SARS-CoV-2 in nasopharyngeal swabs. Lateral flow immunoassay is a method that combines thin-layer chromatography and indirect immunochemical sandwich method and allows the detection of a specific SARS-CoV-2 antigen in nasopharyngeal swabs. Swab specimens should be adequately collected and tested as soon as possible. Users should pay attention to quality control and possible interferences. Antigen tests for SARS-CoV-2 show high sensitivity and specificity in cases with high viral loads, and should be used up to five days after the onset of the first symptoms of COVID-19. False positive results may be obtained when screening large populations with a low prevalence of COVID-19 infection, while false negative results may happen due to improper specimen collection or insufficient amount of antigen in the specimen. So as to achieve reliable results, a diagnostic accuracy study of a specific rapid antigen test should be performed.     

Comparison of diagnostic accuracy for eight SARS-CoV-2 serological assays

IntroductionSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) serological tests have been suggested as an additional diagnostic tool in highly suspected cases with a negative molecular test and determination of seroprevalence in population. We compared the diagnostic performance of eight commercial serological assays for IgA, IgM, and IgG antibodies to the SARS-CoV-2 virus.Materials and methodsThe comparison study was performed on a total of 76 serum samples: 30 SARS-CoV-2 polymerase chain reaction (PCR)-negative and 46 SARS-CoV-2 PCR-positive patients with asymptomatic to severe disease and symptoms duration from 3-30 days. The study included: three rapid lateral flow immunochromatographic assays (LFIC), two enzyme-linked immunosorbent assays (ELISA), and three chemiluminescence immunoassays (CLIA).ResultsAgreement between IgM assays were minimal to moderate (kappa 0.26 to 0.63) and for IgG moderate to excellent (kappa 0.72 to 0.92). Sensitivities improved with > 10 days of symptoms and were: 30% to 89% for IgM; 89% to 100% for IgG; 96% for IgA; 100% for IgA/IgM combination; 96% for total antibodies. Overall specificities were: 90% to 100% for IgM; 85% to 100% for IgG; 90% for IgA; 70% for IgA/IgM combination; 100% for total antibodies. Diagnostic accuracy for IgG ELISA and CIA assays were excellent (AUC ≥ 0.90), without significant difference. IgA showed significantly better diagnostic accuracy than IgM (P < 0.001).ConclusionThere is high variability between IgM assays independently of the assay format, while IgG assays showed moderate to perfect agreement. The appropriate time for testing is crucial for the proper immunity investigation.     

Identification of Cyanobacteria-Based Natural Inhibitors Against SARS-CoV-2 Druggable Target ACE2 Using Molecular Docking Study,ADME and Toxicity Analysis

In 2019–2020, the novel “severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)” had emerged as the biggest challenge for humanity, causing “coronavirus disease 19 (COVID-19)”. Scientists around the world have been putting continuous efforts to unfold potential inhibitors of SARS-CoV-2. We have performed computational studies that help us to identify cyanobacterial photoprotective compounds as potential inhibitors against SARS-CoV-2 druggable target human angiotensin-converting enzyme (ACE2), which plays a vital role in the attachment and entry of the virus into the cell. Blocking the receptor-binding domain of ACE2 can prevent the access of the virus into the compartment. A molecular docking study was performed between photoprotective compounds mycosporine-like amino acids, scytonemins and ACE2 protein using AutoDock tools. Among sixteen molecularly docked metabolites, seven compounds were selected with binding energy < 6.8 kcal/mol. Afterwards, drug-likeness and toxicity of the top candidate were predicted using Swiss ADME and Pro Tox-II online servers. All top hits show desirable drug-likeness properties, but toxicity pattern analysis discloses the toxic effect of scytonemin and its derivatives, resulting in the elimination from the screening pipeline. Further molecular interaction study of the rest two ligands, mycosporine–glycine–valine and shinorine with ACE2 was performed using PyMol, Biovia Discovery studio and LigPlot+. Lastly biological activity of both the ligands was predicted by using the PASS online server. Combining the docking score and other studied properties, we believe that mycosporine–glycine–valine and shinorine have potential to be potent inhibitors of ACE2 and can be explored further to use against COVID-19.     

Separation of antihemophilic factor VII from human plasma by column chromatography

TMAE-Fractogel 650 (M) (Trimethylaminoethyl) is an ion exchange medium can be used to capture factor VIII (F VIII) directly from plasma. Previous reports have focused on the use of DEAE-Fractogel 650 (M) (Dimethylaminoethyl) ion exchange medium to capture F VIII from cryoprecipitate and plasma. Our main objectives were (I) to standardise the purification of FVIII from human plasma by column chromatographic technique. (II) to study the recovery of FVIII activity in purified fraction at 18–20°C process condition. (III) to study the effect of virucidal step on recovery of FVIII activity and (IV) to study the effect of lyophilisation on FVIII activity. In this report, Citrate Phosphate Dextrose (CPD) plasma was batch stirred with dry DEAE-Sephadex A50, filtered, diluted, loaded on to a column packed with TMAE-Fractogel and chromatographed. Most of the unwanted proteins flowed through the gel unadsorbed. Bound F VIII was eluted by increasing the ionic strength of the buffer. This purification step gave an overall 80% recovery from the plasma with a specific activity of 0.97 IU FVIII/mg protein. The purified F VIII fraction was made virus safe by employing the virucidal technique developed by New York Blood Centre (NYBC). There was 48.43% loss of FVIII activity in Virus inactivation treatment and the loss of FVIII activity in lyophilisation was 8.45% which is acceptable. This method of purification gave a higher yield of FVIII than cryoprecipitation, and is a promising alternative method to cryoprecipitation of F VIII.     

Replication,pathogenicity, and transmission of SARS-CoV-2 in minks

Minks are raised in many countries and have transmitted severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to humans. However, the biologic properties of SARS-CoV-2 in minks are largely unknown. Here, we investigated and found that SARS-CoV-2 replicates efficiently in both the upper and lower respiratory tracts, and transmits efficiently in minks via respiratory droplets; pulmonary lesions caused by SARS-CoV-2 in minks are similar to those seen in humans with COVID-19. We further found that a spike protein-based subunit vaccine largely prevented SARS-CoV-2 replication and lung damage caused by SARS-CoV-2 infection in minks. Our study indicates that minks are a useful animal model for evaluating the efficacy of drugs or vaccines against COVID-19 and that vaccination is a potential strategy to prevent minks from transmitting SARS-CoV-2.     

An Integrated Approach of the Potential Underlying Molecular Mechanistic Paradigms of SARS-CoV-2-Mediated Coagulopathy

Coronavirus disease 2019 (Covid-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a pandemic disease which has affected more than 6.2 million people globally, with numbers mounting considerably daily. However, till date, no specific treatment modalities are available for Covid-19 and also not much information is known about this disease. Recent studies have revealed that SARS-CoV-2 infection is associated with the generation of thrombosis and coagulopathy. Fundamentally, it has been believed that a diverse array of signalling pathways might be responsible for the activation of coagulation cascade during SARS-CoV-2 infection. Henceforth, a detailed understanding of these probable underlying molecular mechanistic pathways causing thrombosis in Covid-19 disease deserves an urgent exploration. Therefore, in this review, the hypothetical crosstalk between distinct signalling pathways including apoptosis, inflammation, hypoxia and angiogenesis attributable for the commencement of thrombotic events during SARS-CoV-2 infection has been addressed which might further unravel promising therapeutic targets in Covid-19 disease.     

A Case of Sanfillippo’s Disease Correlating Clinical and Biochemical Findings

The mucopolysaccharidoses (MPSs) are a heterogeneous group of rare inherited disorders caused by deficiency of lysosomal enzyme necessary to break down mucopolysaccharides or glycosaminoglycans (GAGs). We had combined clinical findings and ophthalmological features. Biochemical test for urine glycosaminoglycans was done for confirmation of diagnosis in the patient. The case of Sanfillippo’s disease was characterized by delayed development, hyperactivity with aggressive behaviour. Coarse facial feature, hirsutism and sleep disorder. Urine GAG tests for MPS was positive in the case. Based on clinical findings and biochemical tests for MPS, this case was diagnosed as a type III mucopolysaccharidoses. Urinary GAG’s electrophoresis is an important screening test for MPS suspected cases.     

Double lock of a potent human therapeutic monoclonal antibody against SARS-CoV-2

Receptor recognition and subsequent membrane fusion are essential for the establishment of successful infection by SARS-CoV-2. Halting these steps can cure COVID-19. Here we have identified and characterized a potent human monoclonal antibody, HB27, that blocks SARS-CoV-2 attachment to its cellular receptor at sub-nM concentrations. Remarkably, HB27 can also prevent SARS-CoV-2 membrane fusion. Consequently, a single dose of HB27 conferred effective protection against SARS-CoV-2 in two established mouse models. Rhesus macaques showed no obvious adverse events when administrated with 10 times the effective dose of HB27. Cryo-EM studies on complex of SARS-CoV-2 trimeric S with HB27 Fab reveal that three Fab fragments work synergistically to occlude SARS-CoV-2 from binding to the ACE2 receptor. Binding of the antibody also restrains any further conformational changes of the receptor binding domain, possibly interfering with progression from the prefusion to the postfusion stage. These results suggest that HB27 is a promising candidate for immuno-therapies against COVID-19.     

Polydimethyl siloxane wet etching for three dimensional fabrication of microneedle array and high-aspect-ratio micropillars

Among various transdermal drug delivery (TDD) approaches, utilizing the microneedles (MNs) not only can penetrate the skin but also deliver the drug with reduced tissue damage, reduced pain, and no bleeding. However, the MNs with larger height are required to overcome the skin barrier for effective TDD. Unlike 2D patterning, etching polydimethyl siloxane (PDMS) micropillars for fabrication of 3D microstructures is presented. The PDMS micropillars were first constructed by casting PDMS on the computer numerical control-machined cylindrical microwells, which then went through etching process to obtain the MNs for subsequent fabrication of polymer MNs or high aspect ratio micropillars.     

Determining the SARS-CoV-2 serological immunoassay test performance indices based on the test results frequency distribution

IntroductionCoronavirus disease 2019 (COVID-19) is known to induce robust antibody response in most of the affected individuals. The objective of the study was to determine if we can harvest the test sensitivity and specificity of a commercial serologic immunoassay merely based on the frequency distribution of the SARS-CoV-2 immunoglobulin (Ig) G concentrations measured in a population-based seroprevalence study.Materials and methodsThe current study was conducted on a subset of a previously published dataset from the canton of Geneva. Data were taken from two non-consecutive weeks (774 samples from May 4-9, and 658 from June 1-6, 2020). Assuming that the frequency distribution of the measured SARS-CoV-2 IgG is binormal (an educated guess), using a non-linear regression, we decomposed the distribution into its two Gaussian components. Based on the obtained regression coefficients, we calculated the prevalence of SARS-CoV-2 infection, the sensitivity and specificity, and the most appropriate cut-off value for the test. The obtained results were compared with those obtained from a validity study and a seroprevalence population-based study.ResultsThe model could predict more than 90% of the variance observed in the SARS-CoV-2 IgG distribution. The results derived from our model were in good agreement with the results obtained from the seroprevalence and validity studies. Altogether 138 of 1432 people had SARS-CoV-2 IgG ≥ 0.90, the cut-off value which maximized the Youden’s index. This translates into a true prevalence of 7.0% (95% confidence interval 5.4% to 8.6%), which is in keeping with the estimated prevalence of 7.7% derived from our model. Our model can provide the true prevalence.ConclusionsHaving an educated guess about the distribution of test results, the test performance indices can be derived with acceptable accuracy merely based on the test results frequency distribution without the need for conducting a validity study and comparing the test results against a gold-standard test.     

The missing slope: paradoxical shortening of activated partial thromboplastin time in a patient on unfractionated heparin therapy

This case report describes false shortening of activated partial thromboplastin time (aPTT) due to erroneous optical reading of the clotting point in the presence of unfractionated heparin (UFH), and a biphasic waveform. Activated partial thromboplastin time performed on a coagulometer with photo-optical detection yielded an ambiguous clotting curve characterized by an early and steady decrease in light transmittance throughout the whole measuring range, with the clotting point read at 65 seconds. Further investigations included measurement of aPTT by means of a mechanical clot detection method as well as determination of another heparin-sensitive coagulation assay, that is thrombin time (TT), both being unmeasurably prolonged (> 150 seconds). Communication with clinicians revealed that the patient was on continuous UFH therapy and had an underlying sepsis, with highly elevated C-reactive protein (289 mg/L). The aPTT measurements requested at three timepoints later during the same day revealed gradual aPTT shortening and unveiled a peculiar biphasic waveform pattern. In this case, unmeasurably prolonged aPTT due to UFH therapy was masked by a biphasic aPTT curve pattern making only the first slope of the biphasic waveform visible within the measuring range. The early decrease in plasma light transmittance mimicked optical changes related to clot formation, thus causing erroneous optical reading and yielding a falsely shortened aPTT. This case emphasizes that such a pattern should be carefully inspected, especially when a combination of a critically ill condition and UFH therapy is present, in order to prevent erroneous reporting of aPTT and potential adverse effects on patient care.     

Notable and Emerging Variants of SARS-CoV-2 Virus: A Quick Glance

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the etiological agent of coronavirus disease-2019 (COVID-19), is a highly contagious pathogenic coronavirus to emerge and spread in human populations. Although substantial exertions have been laid to avert spread of COVID-19 by therapeutic and preventive countermeasures, but emergence of SARS-CoV-2 variants as a result of mutations make the infection more ominous. New viral confers a higher nasopharyngeal viral load, increased viral transmissibility, higher infectiousness, immune escape, increased resistance to monoclonal/polyclonal antibodies from convalescence sera/vaccine, and an enhanced virulence. Thus, it is pertinent to monitor evolving mutations and genetic diversity of SARS-CoV-2 as it is decisive for understanding the viral variants. In this review we provide an overview of colloquial nomenclature and the genetic characteristics of different SARS-CoV-2 variants in the context of mutational changes of the circulating strains, transmissibility potential, virulence and infectivity.     

A CRISPR/Cas12a-empowered surface plasmon resonance platform for rapid and specific diagnosis of the Omicron variant of SARS-CoV-2

The outbreak of the COVID-19 pandemic was partially due to the challenge of identifying asymptomatic and presymptomatic carriers of the virus, and thus highlights a strong motivation for diagnostics with high sensitivity that can be rapidly deployed. On the other hand, several concerning SARS-CoV-2 variants, including Omicron, are required to be identified as soon as the samples are identified as ‘positive’. Unfortunately, a traditional PCR test does not allow their specific identification. Herein, for the first time, we have developed MOPCS (Methodologies of Photonic CRISPR Sensing), which combines an optical sensing technology-surface plasmon resonance (SPR) with the ‘gene scissors’ clustered regularly interspaced short palindromic repeat (CRISPR) technique to achieve both high sensitivity and specificity when it comes to measurement of viral variants. MOPCS is a low-cost, CRISPR/Cas12a-system-empowered SPR gene-detecting platform that can analyze viral RNA, without the need for amplification, within 38 min from sample input to results output, and achieve a limit of detection of 15 fM. MOPCS achieves a highly sensitive analysis of SARS-CoV-2, and mutations appear in variants B.1.617.2 (Delta), B.1.1.529 (Omicron) and BA.1 (a subtype of Omicron). This platform was also used to analyze some recently collected patient samples from a local outbreak in China, identified by the Centers for Disease Control and Prevention. This innovative CRISPR-empowered SPR platform will further contribute to the fast, sensitive and accurate detection of target nucleic acid sequences with single-base mutations.     

COVID-19 and Gut Microbiota: A Potential Connection

Currently, world is facing a global outbreak causing a pandemic threat known as COVID-19. This infectious disease is triggered by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Gut microbiota harbours multi species community with a strong impact on host immune homeostasis. However, our knowledge about this gut microbiota and its symbiotic relationship with immune activation in association with SARS-CoV-2 is limited. Unbalanced bacterial flora with too many opportunistic infections can shift immune system towards a cascade of inflammatory responses leading to multi organ damage. This review will highlight immune-regulation via various mechanisms in SARS-CoV-2 infection. Diet has an unbelievable influence on gut microbiome that allows a new state of homeostasis to be reached through timing, frequency and duration of intake. This review article focuses on gut, lung microbiota and immunomodulation with specific attention on immune activation by gut microbiota.     

Multiple Myeloma: A Case of Atypical Presentation on Protein Electrophoresis

Multiple myeloma is a group of B-cell disorders resulting in the secretion of a specific and unique monoclonal immunoglobulin (M-protein). Protein electrophoresis is advised whenever multiple myeloma is suspected. The monoclonal protein migrates as a single entity in the electric field and is detected by the non-specific protein stain as a more intensely stained band superimposed on the usual protein pattern. The M-protein usually migrates in the gamma or beta region of the normal protein pattern; very rarely it may appear in the α2 or even in α1 region. Here we have given an atypical case presentation where the patient with multiple myeloma presented with two M-spike one each in α2 and β-globulin region on agarose gel protein electrophoresis with hypoglobulinemia but with reversed A:G ratio.     

A proof of concept for neutralizing antibody-guided vaccine design against SARS-CoV-2

Mutations and transient conformational movements of the receptor binding domain (RBD) that make neutralizing epitopes momentarily unavailable present immune escape routes for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To mitigate viral escape, we developed a cocktail of neutralizing antibodies (NAbs) targeting epitopes located on different domains of spike (S) protein. Screening of a library of monoclonal antibodies generated from peripheral blood mononuclear cells of COVID-19 convalescent patients yielded potent NAbs, targeting the N-terminal domain (NTD) and RBD domain of S, effective at nM concentrations. Remarkably, a combination of RBD-targeting NAbs and NTD-binding NAbs, FC05, enhanced the neutralization potency in cell-based assays and an animal model. Results of competitive surface plasmon resonance assays and cryo-electron microscopy (cryo-EM) structures of antigen-binding fragments bound to S unveil determinants of immunogenicity. Combinations of immunogens, identified in the NTD and RBD of S, when immunized in rabbits and macaques, elicited potent protective immune responses against SARS-CoV-2. More importantly, two immunizations of this combination of NTD and RBD immunogens provided complete protection in macaques against a SARS-CoV-2 challenge, without observable antibody-dependent enhancement of infection. These results provide a proof of concept for neutralization-based immunogen design targeting SARS-CoV-2 NTD and RBD.     

Utility of P-SEP,sTREM-1 and suPAR as Novel Sepsis Biomarkers in SARS-CoV-2 Infection

The coronavirus disease 2019 is a highly contagious viral infection caused by SARS-CoV-2 virus, member of coronaviridae family. It causes life threatening complications due to complexity and rapid onset course of the disease. Early identification of high-risk patients who require close monitoring and aggressive treatment remains challengeable till date. Novel biomarkers which help to identify high risk patients at the early stage is high priority. Objective of this review to find utility of P-SEP, sTREM-1 and suPAR for diagnosis, risk stratification and prognosis of SARS-CoV-2 infected cases. Soluble receptors like, P-SEP, sTREM-1 and suPAR have been involved in immune regulation in SARS-CoV-2 infection and elevate more in severe cases. A comprehensive research of databases like PubMed, EMBASE, CNKI and Web of Science was performed for relevant studies. A total of nine out of fifteen research literature in initial screening were included for this review. Interestingly all studies have reported high levels of P-SEP, sTREM-1 and suPAR in SARS-CoV-2 infected cases and the biomarkers positively correlated with severity of infection. This implies that P-SEP, sTREM-1 and suPAR can be implemented as surrogate marker in blood profile for early diagnosis, risk stratification and prognosis in SARS-CoV-2 for better management in Indian population at the current situation.     

Contribution of the laboratory to a diagnosis process by sequential reflective testing: Paraprotein interference on a direct bilirubin assay

Errors in laboratory medicine occur in the preanalytical, analytical, and postanalytical phases. The errors are mostly detected in the preanalytical period. However, analytical errors are still an important source of error, despite their frequency is reduced significantly in years thanks to developments in laboratories. In this case, an analytical error was noticed during the verification of a patient’s results. The direct bilirubin of a 66-year-old male patient admitted to the emergency department was higher than the total bilirubin. The patient’s symptoms were fatigue and dyspnoea. Albumin and haemoglobin (Hb) concentrations of the patient were significantly low. After considering the patient’s demographics and laboratory results, the laboratory specialist suspected a paraproteinemia interference. Total protein was performed as a reflective test. The albumin/globulin ratio was reversed. Thereafter, serum protein electrophoresis (SPEP) and immunofixation electrophoresis (IFE) were performed as another reflective tests, respectively. SPEP and IFE results were in favour of monoclonal gammopathy. The patient was directed to a haematologist, underwent a bone marrow biopsy, and the result was reported as Waldenstrom’s macroglobulinemia with plasma cell differentiation expressing IgM-Kappa. The patient went on a chemotherapy protocol, and his condition has been improved in subsequent months. Detection of analytical errors is of great importance, like in our case, and may be used as a tool to identify patients who have not yet been diagnosed. The laboratory specialist must dominate the entire process of each test in the laboratory, be aware of the limitations of tests, and turn these disadvantages into advantages when necessary.     

Laboratory medicine in pandemic of COVID-19

After the outbreak in China in the year 2019, severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) quickly spread around the world causing a protracted pandemic. Approximately one-third of infections appear to be asymptomatic. Symptomatic disease is characterized primarily by symptoms of respiratory tract infection of varying severity. But Coronavirus Disease 2019 (COVID-19) is much more than an acute respiratory disease because SARS-CoV-2 affects many organs inducing a vast number of symptoms such as cardiovascular, neurological, gastrointestinal, dermatological, with numerous complications. Short and long-term effects of infection, severe ones, and especially mild forms of the disease which affect a huge number of patients need to be further investigated. Laboratory medicine has a crucial role in early diagnosis of the disease, recognition of the patients who need hospital care, and close monitoring of hospitalized patients to timely identify associated clinical complications as well as follow-up of patients with long-term COVID-19.