Rapid, sensitive detection of intact SARS-CoV-2 using DNA nets and a smartphone-linked fluorimeter

We report a single-step, room-temperature, 5-10 minute SARS-CoV-2 saliva self-monitoring method that overcomes the limitations of existing approaches through the use of fluorophore-releasing Designer DNA Nanostructures (DDNs) that bind with the multivalent pattern of spike proteins on the exterior intact virions and an inexpensive smartphone-linked, pocket-size fluorimeter, called a "V-Pod” for its resemblance to an Apple AirPod™ headphone case. We characterize the V-Pod fluorimeter performance and the DDN-based assay to demonstrate a clinically relevant detection limit of 104 virus particles/mL for pseudo-typed WT SARS-CoV-2 and 105 virus particles/mL for real pathogenic variants, including Delta, Omicron, and D614g. © 2023 SPIE.

Point-of-Care Testing for SARS-CoV-2: A Prospective Study in a Primary Health Centre.

BACKGROUND: In 2020, health systems across the world responded to the COVID-19 pandemic by making rapid changes to reduce the risk of exposure in patients and healthcare professionals. The use of point-of-care tests (POCT) has been a central strategy in dealing with the COVID-19 pandemic. The aims of this study were to evaluate the impact of POCT strategy (1) on maintaining elective surgeries by removing the risk of delayed pre-appointment testing and turn-around times and (2) on time dedicated for end-to-end appointment and management, and (3) to assess the feasibility of using the ID NOW® among healthcare professionals and patients in a primary care setting, requiring pre-surgical appointment and minor ENT surgery in the Townsend House Medical Centre (THMC), Devon, United Kingdom. METHODS: A logistic regression was performed to identify factors associated with the risk of canceled or delayed surgeries and medical appointments. Second, a multivariate linear regression analysis was conducted to calculate changes in the time dedicated to administrative tasks. A questionnaire was developed to assess the acceptance of POCT in patients and staff. RESULTS: 274 patients were included in this study; 174 (63.5%) in Group 1 (Usual Care) and 100 (36.5%) in Group 2 (Point of Care). Multivariate logistic regression showed that the percentage of postponed or canceled appointments was similar between the two groups (adjusted OR = 0.65, [95%CI: 0.22-1.88]; p = 0.42). Similar results were observed for the percentage of postponed or canceled scheduled surgeries (adjusted OR = 0.47, [95%CI: 0.15-1.47]; p = 0.19). The time dedicated to administrative tasks was significantly lowered by 24.7 min in G2 compared to G1 (p < 0.001). 79 patients in G2 (79.0%) completed the survey, and the majority agreed or strongly agreed that it improved care management (79.7%), decreased administrative time (65.8%), reduced the risk of canceled appointments (74.7%) and the traveling time to do COVID-19 test (91.1%). Having point-of-care testing in the clinic in the future seemed more than welcome by 96.6% of patients; 93.6% declared to be less stressed by having the test at the clinic than waiting for the results of the test realized elsewhere. The five healthcare professionals of the primary care center completed the survey, and all agreed that the POCT positively influences the workflow and can be successfully implemented into routine primary care. CONCLUSIONS: Our study shows that NAAT-based point-of-care SARS-CoV-2 testing significantly improved flow management in a primary care setting. POC testing was a feasible and well-accepted strategy by patients and providers.

Community pharmacist-provided test and treat programs for acute infectious conditions

Community pharmacists' roles have expanded in recent years to include offering test and treat programs where they perform testing on Clinical Laboratory Improvement Amendment (CLIA)-waived point-of-care testing (POCT) devices to diagnose specific acute infectious conditions, such as influenza and group A streptococcus (GAS) pharyngitis, and then potentially prescribe and dispense appropriate antimicrobials. Availability of these services in pharmacies has several benefits, including increased access to care, decreased overutilization of other health care services, and decreased antimicrobial resistance. States have different requirements for collaborative practice agreements and reimbursement for these clinical services in community pharmacies. Several studies have looked at outcomes related to community pharmacies implementing test and treat programs for influenza and/or GAS. Other studies looked at outcomes related to implementing testing for SARS-CoV-2 and referring for treatment. Most studies described successful implementation and barriers to integration of these programs into pharmacy workflow. Some studies showed that patients want these services to be offered in community pharmacies and are willing to pay for the services. Data show that these services are cost effective compared to physician provider-based treatment. Newer CLIA-waived POCT technology may increase implementation of these services, but studies are needed to evaluate their utility in community pharmacies. Pharmacy schools should implement widespread training on these devices, and research should continue in this area to test the use of newer technology (i.e., multiplexed devices) and their economic impact.Copyright © 2023 Pharmacotherapy Publications, Inc.

Evaluation of Urine L-FABP Point of Care Kit in the Philippines as Predictive Marker of Clinical Severity of COVID-19 (EPOCH COVID study)

Background: The search for simple clinical and laboratory markers to help predict the clinical severity of patients presenting with COVID-19 has prompted this study to look at the predictive value of urine L-FABP (Liver Type-Fatty Acid Binding Protein) point-of-care test kit at the initial presentation of COVID-19 patients to the hospital. Method(s): The validation study prospectively included 109 consecutive patients with mild to moderate COVID-19, mean age of 52.2 years (range 19-84) presenting at the Emergency Rooms of 4 participating Metro-Manila hospitals from February to April 2021, with available data for analysis for 103 patients. Urine L-FABP POC (Point-of-Care) test and other clinical parameters and the level of severity of COVID-19 were determined at Day 0, Day 4 and Day 7. Computations for Sensitivity, Specificity, Positive and Negative Predictive values and Likelihood ratios were performed Results: Twenty-three patients tested positive for urine L-FABP, out of the 103 patients analyzed, while 80 tested negative. Of the 23 patients who tested positive for urine L-FABP, 6 has progressed in severity, while 17 did not progressed. Of the 80 patients who tested negative for urine L-FABP, 13 progressed, while 67 did not progressed in severity. Giving a Sensitivity of 31.58%, Specificity of 79.76%, Positive predictive value of 26.09%, Negative predictive value of 83.75%. Combining urine L-FABP and initial clinical parameters like SIRS (Systemic Inflammatory Response Syndrome) criteria to predict progression of severity yielded a higher Specificity of 91.67 % and Negative Predictive value of 84.62%. Conclusion(s): The study shows the utility of initial urine L-FABP POC test as a negative screening test in triaging adult patients presenting to the ER with mild to moderate COVID-19. Patients at the ER with a negative urine L-FABP test, will most likely not progressed to severe COVID-19. Combining clinical parameters like SIRS Criteria with the urine L-FABP result can increase the negative predictive value.Copyright © The PHILIPPINE JOURNAL OF INTERNAL MEDICINE is a peer reviewed journal and a copyrighted publication of the Philippine College of Physicians.

An update on lateral flow immunoassay for the rapid detection of SARS-CoV-2 antibodies.

Over the last three years, after the outbreak of the COVID-19 pandemic, an unprecedented number of novel diagnostic tests have been developed. Assays to evaluate the immune response to SARS-CoV-2 have been widely considered as part of the control strategy. The lateral flow immunoassay (LFIA), to detect both IgM and IgG against SARS-CoV-2, has been widely studied as a point-of-care (POC) test. Compared to laboratory tests, LFIAs are faster, cheaper and user-friendly, thus available also in areas with low economic resources. Soon after the onset of the pandemic, numerous kits for rapid antibody detection were put on the market with an emergency use authorization. However, since then, scientists have tried to better define the accuracy of these tests and their usefulness in different contexts. In fact, while during the first phase of the pandemic LFIAs for antibody detection were auxiliary to molecular tests for the diagnosis of COVID-19, successively these tests became a tool of seroprevalence surveillance to address infection control policies. When in 2021 a massive vaccination campaign was implemented worldwide, the interest in LFIA reemerged due to the need to establish the extent and the longevity of immunization in the vaccinated population and to establish priorities to guide health policies in low-income countries with limited access to vaccines. Here, we summarize the accuracy, the advantages and limits of LFIAs as POC tests for antibody detection, highlighting the efforts that have been made to improve this technology over the last few years.

Detection and Diagnosis Technologies for SARS-CoV-2

Detection and diagnosis platforms play key roles in early warning, outbreak control and exit strategy for any pandemic, and they are especially pertinent for the Coronavirus disease 2019 (COVID-19) pandemic. The challenges posed by the speed and extent of severe acute respiratory syndrome Coronavirus-2 (SARS-CoV-2) spread around the globe also offered unprecedented opportunities for the development and deployment of novel strategies and products - not only vaccines and therapeutics, but also diagnostics. This chapter provides a brief summary of the vast array of molecular, serological, cell-based and other diagnostic tools for the specific detection of SARS-CoV-2 infections and immune responses. The focus is on the principles and applications of each platform, while detailed protocols can be found in the cited references. © 2023 by World Scientific Publishing Co. Pte. Ltd.

CRISPR techniques and potential for the detection and discrimination of SARS-CoV-2 variants of concern.

The continuing evolution of the SARS-CoV-2 virus has led to the emergence of many variants, including variants of concern (VOCs). CRISPR-Cas systems have been used to develop techniques for the detection of variants. These techniques have focused on the detection of variant-specific mutations in the spike protein gene of SARS-CoV-2. These sequences mostly carry single-nucleotide mutations and are difficult to differentiate using a single CRISPR-based assay. Here we discuss the specificity of the Cas9, Cas12, and Cas13 systems, important considerations of mutation sites, design of guide RNA, and recent progress in CRISPR-based assays for SARS-CoV-2 variants. Strategies for discriminating single-nucleotide mutations include optimizing the position of mismatches, modifying nucleotides in the guide RNA, and using two guide RNAs to recognize the specific mutation sequence and a conservative sequence. Further research is needed to confront challenges in the detection and differentiation of variants and sublineages of SARS-CoV-2 in clinical diagnostic and point-of-care applications.

Rapid detection of intact SARS-CoV-2 using designer DNA Nets and a pocket-size smartphone-linked fluorimeter.

Rapid, sensitive, and inexpensive point-of-care diagnosis is vital to controlling highly infectious diseases, including COVID-19. Here, we report the design and characterization of a compact fluorimeter called a "Virus Pod" (V-Pod) that enables sensitive self-testing of SARS-CoV-2 viral load in saliva. The rechargeable battery-operated device reads the fluorescence generated by Designer DNA Nanostructures (DDN) when they specifically interact with intact SARS-CoV-2 virions. DDNs are net-shaped self-assembling nucleic acid constructs that provide an array of highly specific aptamer-fluorescent quencher duplexes located at precise positions that match the pattern of spike proteins. The room-temperature assay is performed by mixing the test sample with DNA Net sensor in a conventional PCR tube and placing the tube into the V-Pod. Fluorescent signals are generated when multivalent aptamer-spike binding releases fluorescent quenchers, resulting in rapid (5-min) generation of dose-dependent output. The V-Pod instrument performs laser excitation, fluorescence intensity quantitation, and secure transmission of data to an App via Bluetooth™. We show that the V-Pod and DNA Net assay achieves clinically relevant detection limits of 3.92 × 103 viral-genome-copies/mL for pseudo-typed wild-type SARS-CoV-2 and 1.84 × 104, 9.69 × 104, 6.99 × 104 viral-genome-copies/mL for pathogenic Delta, Omicron, and D614G variants, representing sensitivity similar to laboratory-based PCR. The pocket-sized instrument (∼$294), inexpensive reagent-cost/test ($1.26), single-step, rapid sample-to-answer, and quantitative output represent a capability that is compatible with the needs of frequent self-testing in a consumer-friendly format that can link with medical service systems such as healthcare providers, contact tracing, and infectious disease reporting.

Ultrafast Plasmonic Nucleic Acid Amplification and Real-Time Quantification for Decentralized Molecular Diagnostics.

Point-of-care real-time reverse-transcription polymerase chain reaction (RT-PCR) facilitates the widespread use of rapid, accurate, and cost-effective near-patient testing that is available to the public. Here, we report ultrafast plasmonic nucleic acid amplification and real-time quantification for decentralized molecular diagnostics. The plasmonic real-time RT-PCR system features an ultrafast plasmonic thermocycler (PTC), a disposable plastic-on-metal (PoM) cartridge, and an ultrathin microlens array fluorescence (MAF) microscope. The PTC provides ultrafast photothermal cycling under white-light-emitting diode illumination and precise temperature monitoring with an integrated resistance temperature detector. The PoM thin film cartridge allows rapid heat transfer as well as complete light blocking from the photothermal excitation source, resulting in real-time and highly efficient PCR quantification. Besides, the MAF microscope exhibits close-up and high-contrast fluorescence microscopic imaging. All of the systems were fully packaged in a palm size for point-of-care testing. The real-time RT-PCR system demonstrates the rapid diagnosis of coronavirus disease-19 RNA virus within 10 min and yields 95.6% of amplification efficiency, 96.6% of classification accuracy for preoperational test, and 91% of total percent agreement for clinical diagnostic test. The ultrafast and compact PCR system can decentralize point-of-care molecular diagnostic testing in primary care and developing countries.

Performance of point-of care molecular and antigen-based tests for SARS-CoV-2: a living systematic review and meta-analysis.

BACKGROUND: Molecular and antigen point-of-care tests (POCTs) have augmented our ability to rapidly identify and manage SARS-CoV-2 infection. However, their clinical performance varies among individual studies. OBJECTIVES: The evaluation of the performance of molecular and antigen-based POCTs in confirmed, suspected, or probable COVID-19 cases compared with that of laboratory-based RT-PCR in real-life settings. DATA SOURCES: MEDLINE/PubMed, Scopus, Embase, Web of Science, Cochrane Library, Cochrane COVID-19 study register, and COVID-19 Living Evidence Database from the University of Bern. STUDY ELIGIBILITY CRITERIA: Peer-reviewed or preprint observational studies or randomized controlled trials that evaluated any type of commercially available antigen and/or molecular POCTs for SARS-CoV-2, including multiplex PCR panels, approved by the United States Food and Drug Administration, with Emergency Use Authorization, and/or marked with Conformitè Europëenne from European Commission/European Union. PARTICIPANTS: Close contacts and/or patients with symptomatic and/or asymptomatic confirmed, suspected, or probable COVID-19 infection of any age. TEST/S: Molecular and/or antigen-based SARS-CoV-2 POCTs. REFERENCE STANDARD: Laboratory-based SARS-CoV-2 RT-PCR. ASSESSMENT OF RISK OF BIAS: Eligible studies were subjected to quality-control and risk-of-bias assessment using the Quality Assessment of Diagnostic Accuracy Studies 2 tool. METHODS OF DATA SYNTHESIS: Summary sensitivities and specificities with their 95% CIs were estimated using a bivariate model. Subgroup analysis was performed when at least three studies informed the outcome. RESULTS: A total of 123 eligible publications (97 and 26 studies assessing antigen-based and molecular POCTs, respectively) were retrieved from 4674 initial records. The pooled sensitivity and specificity for 13 molecular-based POCTs were 92.8% (95% CI, 88.9-95.4%) and 97.6% (95% CI, 96.6-98.3%), respectively. The sensitivity of antigen-based POCTs pooled from 138 individual evaluations was considerably lower than that of molecular POCTs; the pooled sensitivity and specificity rates were 70.6% (95% CI, 67.2-73.8%) and 98.9% (95% CI, 98.5-99.2%), respectively. DISCUSSION: Further studies are needed to evaluate the performance of molecular and antigen-based POCTs in underrepresented patient subgroups and different respiratory samples.

Recent Advances in Colorimetric Sensors Based on Gold Nanoparticles for Pathogen Detection.

Infectious pathogens cause severe threats to public health due to their frightening infectivity and lethal capacity. Rapid and accurate detection of pathogens is of great significance for preventing their infection. Gold nanoparticles have drawn considerable attention in colorimetric biosensing during the past decades due to their unique physicochemical properties. Colorimetric diagnosis platforms based on functionalized AuNPs are emerging as a promising pathogen-analysis technique with the merits of high sensitivity, low-cost, and easy operation. This review summarizes the recent development in this field. We first introduce the significance of detecting pathogens and the characteristics of gold nanoparticles. Four types of colorimetric strategies, including the application of indirect target-mediated aggregation, chromogenic substrate-mediated catalytic activity, point-of-care testing (POCT) devices, and machine learning-assisted colorimetric sensor arrays, are systematically introduced. In particular, three biomolecule-functionalized AuNP-based colorimetric sensors are described in detail. Finally, we conclude by presenting our subjective views on the present challenges and some appropriate suggestions for future research directions of colorimetric sensors.

Evaluation of STANDARDTM M10 SARS-CoV-2 assay as a diagnostic tool for SARS-CoV-2 in nasopharyngeal or oropharyngeal swab samples.

The SARS-CoV-2 pandemic led to an urgent need for rapid diagnostic testing in order to inform timely patients' management. This study aimed to assess the performance of the STANDARD™ M10 SARS-CoV-2 assay as a diagnostic tool for COVID-19. A total of 400 nasopharyngeal or oropharyngeal swabs were tested against a reference real-time RT-PCR, including 200 positive samples spanning the full range of observed Ct values. The sensitivity of the STANDARD™ M10 SARS-CoV-2 assay was 98.00% (95% CI 94.96% to 99.45%, 196/200), while the specificity was also estimated at 97.50% (95% CI 94.26% to 99.18%, 195/200). The assay proved highly efficient for the detection of SARS-CoV-2, even in samples with low viral load (Ct>25), presenting lower Ct values compared to the reference method. We concluded that the STANDARD™ M10 SARS-CoV-2 assay has a similar performance compared to the reference method and other molecular point-of-care assays and can be a valuable tool for rapid and accurate diagnosis.

A Cross-Sectional Study of SARS-CoV-2 Antibodies and Risk Factors for Seropositivity in Staff in Day Care Facilities and Preschools in Denmark.

The aim of this study was to provide information about immunity against COVID-19 along with risk factors and behavior among employees in day care facilities and preschools (DCS) in Denmark. In collaboration with the Danish Union of Pedagogues, during February and March 2021, 47,810 members were offered a point-of-care rapid SARS-CoV-2 antibody test (POCT) at work and were invited to fill in an electronic questionnaire covering COVID-19 exposure. Seroprevalence data from Danish blood donors (total Ig enzyme-linked immunosorbent assay [ELISA]) were used as a proxy for the Danish population. A total of 21,018 (45%) DCS employees completed the questionnaire and reported their POCT result {median age, 44.3 years (interquartile range [IQR], [32.7 to 53.6]); females, 84.1%}, of which 20,267 (96.4%) were unvaccinated and included in analysis. A total of 1,857 (9.2%) participants tested seropositive, significantly higher than a seroprevalence at 7.6% (risk ratio [RR], 1.2; 95% confidence interval [CI], 1.14 to 1.27) among 40,541 healthy blood donors (median age, 42 years [IQR, 28 to 53]; males, 51.3%). Exposure at work (RR, 2.9; 95% CI, 2.3 to 3.6) was less of a risk factor than exposure within the household (RR, 12.7; 95% CI, 10.2 to 15.8). Less than 25% of participants reported wearing face protection at work. Most of the participants expressed some degree of fear of contracting COVID-19 both at work and outside work. SARS-CoV-2 seroprevalence was slightly higher in DCS staff than in blood donors, but possible exposure at home was associated with a higher risk than at work. DCS staff expressed fear of contracting COVID-19, though there was limited use of face protection at work. IMPORTANCE Identifying at-risk groups and evaluating preventive interventions in at-risk groups is imperative for the ongoing pandemic as well as for the control of future epidemics. Although DCS staff have a much higher risk of being infected within their own household than at their workplace, most are fearful of being infected with COVID-19 or bringing COVID-19 to work. This represents an interesting dilemma and an important issue which should be addressed by public health authorities for risk communication and pandemic planning. This study design can be used in a strategy for ongoing surveillance of COVID-19 immunity or other infections in the population. The findings of this study can be used to assess the need for future preventive interventions in DCS, such as the use of personal protective equipment.

Performance of Rapid, Antigen-Detecting Point-of-Care Test for Suspected SARS-CoV-2 Patients Presented to the Emergency Department

Patients who present to the emergency department (ED) with SARS-CoV-2 are highly contagious. Early diagnosis is necessary to reduce the transmission of this disease. We aimed to determine the performance of a rapid, antigen-detecting point-of-care test in symptomatic patients with suspected SARS-CoV-2 infection, using rt-PCR as the reference standard. A single-centered, cross-sectional retrospective study from data registry was conducted in the ED. Patients who were 18 years or older and had symptoms that were compatible with COVID-19 infection were eligible for inclusion. All patients were tested with rapid, antigen-detecting point-of-care tests and rt-PCR in the same visit. The diagnostic performances of the test were demonstrated with sensitivity, specificity, and predictive value, including positive predictive value (PPV) and negative predictive value (NPV). In total, 703 eligible patients were included in the analysis. Of these, 120 patients infected with SARS-CoV-2 were confirmed by rt-PCR. Total prevalence was 17.07%. The rapid antigen test demonstrated high sensitivity (87.5%), very high specificity (99.8%), PPV (99.1%), and NPV (97.5%). The test demonstrated high performance in patients with high viral loads (sensitivity of 94.2-100% in Ct values ≤ 25). However, the test showed a decline performance in patients with higher Ct values (sensitivity of 40 – 85.7% in Ct values > 25). In conclusion, the rapid, antigen-detecting point-of-care test showed high sensitivity and very high specificity in symptomatic ED patients, as compared to rt-PCR. This test can be used as an initial screening tool to detect and guide clinical decisions for suspected COVID-19 patients who present to the ED. © 2022, Thammasat University. All rights reserved.

A novel paper-based lysis strip for SARS-CoV-2 RNA detection at low resource settings.

Infectious respiratory diseases such as COVID-19 are serious and global concerns from the past to the present. To isolate the spread of infectious diseases even in the absence of a health system, a simple, inexpensive, reliable, sensitive, and selective molecular diagnosis platform for Point of Care Test (POCT) is required. Especially, the nucleic acid extraction step is difficult to perform out of laboratory. Here, we propose a paper-based lysis (PBL) strip for nucleic acid extraction, especially in low-resource settings (LRS). PBL strips are suitable for isolating RNA from viruses with biological interference and inhibitors. We optimized the buffer compositions and membranes of the strip. A simple preparation method using a PBL strip could obtain an eluent for downstream inspection within 20 min. Overall, 104 copies/swaps were detected for 20 min for amplification in combination with Reverse Transcription Loop-Mediated Amplification (RT-LAMP).

Integrating CRISPR and isothermal amplification reactions in single-tubes for ultrasensitive detection of nucleic acids: the SARS-CoV-2 RNA example

Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein (CRISPR-Cas) systems have advanced rapidly for the detection of nucleic acids and molecular diagnoses. The sensitivity of techniques directly using CRISPR-Cas systems for target recognition and signal generation is limited by the kinetics of trans-cleavage. Thus, CRISPR-Cas systems have been coupled with isothermal amplification techniques. One strategy for integrating CRISPR-Cas and amplification reactions into a single-tube is to place reagents in separate locations within the tube, maintaining optimum conditions for each reaction. A more challenging strategy is to mix all reagents and allow nucleic acid amplification and CRISPR-based detection to proceed in a homogeneous solution. This desirable approach requires substantial understanding of the compatibility of enzymatic reactions, systematic optimization, and appropriate adjustments of the integrated reactions to ensure high sensitivity. Ultrasensitive techniques have been developed for the detection of SARS-CoV-2 in single-tubes. In this review, we highlight the principle, research needs, and challenges of ultrasensitive single-tube RNA detection using CRISPR technology. We stress the importance of understanding the kinetics of trans-cleavage activity of CRISPR-Cas systems. © 2022 Scientia Sinica Chimica. All rights reserved.

Investigation of differences in coagulation characteristics between hospitalized patients with SARS-CoV-2 Alpha, Delta, and Omicron variant infection using rotational thromboelastometry (ROTEM): A single-center, retrospective, observational study.

BACKGROUND: The severe acute respiratory syndrome coronavirus 2 Omicron variant has a low rate of serious illness, is highly contagious, and has spread rapidly since January 2022. The number of severe cases and deaths remains problematic. Here, we aimed to elucidate the coagulation pathology of Omicron-infected patients using rotational thromboelastometry. METHODS: Patients with coronavirus disease 2019, hospitalized and treated from January 2021 to April 2022, were included. The Alpha-Delta and Omicron groups were defined during admission. Blood tests, clinical course, and rotational thromboelastometry measurements were compared using a propensity score-matched cohort. RESULTS: Both groups had 21 patients each. Lactate dehydrogenase (Alpha-Delta group [interquartile range] vs. Omicron group [interquartile range]; 449 [368-518] U/L vs. 241 [196-398] U/L, p = 0.01) and ferritin (1428 [1145-3061] ng/dl vs. 481 [188-881] ng/dl, p = 0.0002) levels were significantly lower in the Omicron group. In rotational thromboelastometry, the thrombus hardness indexes FIBTEM A5 (29 [23-34] mm vs. 23 [18-28] mm, p = 0.034) and maximum clot firmness (34 [27-40] mm vs. 26 [21-33] mm, p = 0.021) were significantly lower in the Omicron group, whereas the fibrinolysis index FIBTEM LI60 (98 [92-100] % vs. 100 [100-100] %, p = 0.0082) was higher. CONCLUSION: Severe coagulation abnormalities may be less likely in Omicron-infected patients than in those infected with the previous Alpha and Delta variants.

Plasmonic nanostructure-enhanced Raman scattering for detection of SARS-CoV-2 nucleocapsid protein and spike protein variants.

Epidemiological control and public health monitoring during the outbreaks of infectious viral diseases rely on the ability to detect viral pathogens. Here we demonstrate a rapid, sensitive, and selective nanotechnology-enhanced severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection based on the surface-enhanced Raman scattering (SERS) responses from the plasma-engineered, variant-specific antibody-functionalized silver microplasma-engineered nanoassemblies (AgMEN) interacting with the SARS-CoV-2 spike (S) and nucleocapsid (N) proteins. The three-dimensional (3D) porous AgMEN with plasmonic-active nanostructures provide a high sensitivity to virus detection via the remarkable SERS signal collection. Moreover, the variant-specific antibody-functionalization on the SERS-active AgMEN enabled the high selectivity of the SARS-CoV-2 S variants, including wild-type, Alpha, Delta, and Omicron, under the simulated human saliva conditions. The exceptional ultrahigh sensitivity of our SERS biosensor was demonstrated via SARS-CoV-2 S and N proteins at the detection limit of 1 fg mL-1 and 0.1 pg mL-1, respectively. Our work demonstrates a versatile SERS-based detection platform can be applied for the ultrasensitive detection of virus variants, infectious diseases, and cancer biomarkers.

Clinical Feasibility of a Saliva-Based Antigen Qualitative Test for Severe Acute Respiratory Syndrome Coronavirus 2.

BACKGROUND: Nasopharyngeal swabs (NPS) are generally used as specimen samples for antigen qualitative tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The principle of the reaction to the antigen protein is the same when saliva is used, and saliva samples were reported to be as accurate as NPS for real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) testing to identify SARS-CoV-2. Unlike NPS collection, self-collected saliva does not expose healthcare workers to the risk of infection. In this study, we evaluated the feasibility of using saliva samples for a SARS-CoV-2 antigen qualitative test (TA2107SA) under development. METHODS: Saliva samples were collected from patients with confirmed or suspected COVID-19 infection and analyzed. The sensitivity, specificity, and concordance index of the antigen qualitative test were calculated using an RT-qPCR test as reference. RESULTS: Saliva samples were collected from 105 patients. The mean interval from onset to specimen collection was 5.7 days. The mean cycle threshold (Ct) value of RT-qPCR was 31.3. The sensitivity, specificity, and concordance index were 70.7%, 100%, and 0.85, respectively. In 33 patients with Ct values <30, the results of both the RT-qPCR and antigen tests were positive. The sensitivity of the saliva-based TA2107SA SARS-CoV-2 antigen qualitative test was slightly lower than that of the conventional antigen qualitative test using NPS samples from the same patient. CONCLUSION: Saliva-based antigen qualitative tests for SARS-CoV-2 are an alternative option during a pandemic.

Ultrafast PCR Detection of COVID-19 by Using a Microfluidic Chip-Based System.

With the evolution of the pandemic caused by the Coronavirus disease of 2019 (COVID-19), reverse transcriptase-polymerase chain reactions (RT-PCR) have invariably been a golden standard in clinical diagnosis. Nevertheless, the traditional polymerase chain reaction (PCR) is not feasible for field application due to its drawbacks, such as time-consuming and laboratory-based dependence. To overcome these challenges, a microchip-based ultrafast PCR system called SWM-02 was proposed to make PCR assay in a rapid, portable, and low-cost strategy. This novel platform can perform 6-sample detection per run using multiple fluorescent channels and complete an ultrafast COVID-19 RT-PCR test within 40 min. Here, we evaluated the performance of the microdevice using the gradient-diluted COVID-19 reference samples and commercial PCR kit and determined its limit-of-detection (LoD) as 500 copies/mL, whose variation coefficients for the nucleocapsid (N) gene and open reading frame 1 ab region (ORF1ab) gene are 1.427% and 0.7872%, respectively. The system also revealed an excellent linear correlation between cycle threshold (Ct) values and dilution factors (R2 > 0.99). Additionally, we successfully detected the target RNAs and internal gene in the clinical samples by fast PCR, which shows strong consistency with conventional PCR protocol. Hence, with compact dimension, user-friendly design, and fast processing time, SWM-02 has the capability of offering timely and sensitive on-site molecular diagnosis for prevention and control of pathogen transmission.