scholarly journals CRISPR/Cas9-Based Lateral Flow and Fluorescence Diagnostics

2021 ◽  
Vol 8 (2) ◽  
pp. 23
Author(s):  
Mark J. Osborn ◽  
Akshay Bhardwaj ◽  
Samuel P. Bingea ◽  
Friederike Knipping ◽  
Colby J. Feser ◽  
...  
Keyword(s):  
Influenza A ◽  
Point Of Care ◽  
Lateral Flow ◽  
Great Promise ◽  
Rna Sequences ◽  
Dna And Rna ◽  
Cas9 Nuclease ◽  
Base Specificity ◽  
Short Palindromic Repeat ◽  
Syncytial Virus

Clustered regularly interspaced short palindromic repeat (CRISPR/Cas) proteins can be designed to bind specified DNA and RNA sequences and hold great promise for the accurate detection of nucleic acids for diagnostics. We integrated commercially available reagents into a CRISPR/Cas9-based lateral flow assay that can detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequences with single-base specificity. This approach requires minimal equipment and represents a simplified platform for field-based deployment. We also developed a rapid, multiplex fluorescence CRISPR/Cas9 nuclease cleavage assay capable of detecting and differentiating SARS-CoV-2, influenza A and B, and respiratory syncytial virus in a single reaction. Our findings provide proof-of-principle for CRISPR/Cas9 point-of-care diagnosis as well as a scalable fluorescent platform for identifying respiratory viral pathogens with overlapping symptomology.

scholarly journals Influenza and RSV incidence during COVID-19 pandemic—an observational study from in-hospital point-of-care testing

2021 ◽  
Author(s):  
Paul Stamm ◽  
Ingo Sagoschen ◽  
Kerstin Weise ◽  
Bodo Plachter ◽  
Thomas Münzel ◽  
...  
Keyword(s):  
Emergency Room ◽  
Influenza A ◽  
Point Of Care ◽  
Respiratory Viruses ◽  
Pediatric Emergency ◽  
Influenza B ◽  
Complete Suppression ◽  
Emergency Room Patients ◽  
Syncytial Virus ◽  
Point Of Care Tests

AbstractThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has forced the implementation of unprecedented public health measures strategies which might also have a significant impact on the spreading of other viral pathogens such as influenza and Respiratory Syncytial Virus (RSV) . The present study compares the incidences of the most relevant respiratory viruses before and during the SARS-CoV-2 pandemic in emergency room patients. We analyzed the results of in total 14,946 polymerase chain reaction point-of-care tests (POCT-PCR) for Influenza A, Influenza B, RSV and SARS-CoV-2 in an adult and a pediatric emergency room between December 1, 2018 and March 31, 2021. Despite a fivefold increase in the number of tests performed, the positivity rate for Influenza A dropped from 19.32% (165 positives of 854 tests in 2018/19), 14.57% (149 positives of 1023 in 2019–20) to 0% (0 positives of 4915 tests) in 2020/21. In analogy, the positivity rate for Influenza B and RSV dropped from 0.35 to 1.47%, respectively, 10.65–21.08% to 0% for both in 2020/21. The positivity rate for SARS-CoV2 reached 9.74% (110 of 1129 tests performed) during the so-called second wave in December 2020. Compared to the two previous years, seasonal influenza and RSV incidence was eliminated during the COVID-19 pandemic. Corona-related measures and human behavior patterns could lead to a significant decline or even complete suppression of other respiratory viruses such as influenza and RSV.

scholarly journals Detection of Influenza A and B Viruses and Respiratory Syncytial Virus by Use of Clinical Laboratory Improvement Amendments of 1988 (CLIA)-Waived Point-of-Care Assays: a Paradigm Shift to Molecular Tests

2018 ◽  
Vol 56 (7) ◽  
Author(s):  
Marwan M. Azar ◽  
Marie L. Landry
Keyword(s):  
Respiratory Syncytial Virus ◽  
Antiviral Therapy ◽  
Influenza A ◽  
Clinical Laboratory ◽  
Point Of Care ◽  
The United States ◽  
Nucleic Acid Amplification ◽  
Molecular Tests ◽  
Syncytial Virus ◽  
Clinical Laboratory Improvement Amendments

ABSTRACT An accurate laboratory diagnosis of influenza, respiratory syncytial virus (RSV), and other respiratory viruses can help to guide patient management, antiviral therapy, infection prevention strategies, and epidemiologic monitoring. Influenza has been the primary driver of rapid laboratory testing due to its morbidity and mortality across all ages, the availability of antiviral therapy, which must be given early to have an effect, and the constant threat of new pandemic strains. Over the past 30 years, there has been an evolution in viral diagnostic testing, from viral culture to rapid antigen detection, and more recently, to highly sensitive nucleic acid amplification tests (NAAT), as well as a trend to testing at the point of care (POC). Simple rapid antigen immunoassays have long been the mainstay for POC testing for influenza A and B viruses and respiratory syncytial virus (RSV) but have been faulted for low sensitivity. In 2015, the first POC NAAT for the detection of influenza was approved by the Food and Drug Administration (FDA), ushering in a new era. In 2017, the FDA reclassified rapid influenza diagnostic tests (RIDTs) from class I to class II devices with new minimum performance standards and a requirement for annual reactivity testing. Consequently, many previously available RIDTs can no longer be purchased in the United States. In this review, recent developments in Clinical Laboratory Improvement Amendments of 1988 (CLIA)-waived testing for respiratory virus infections will be presented, with the focus on currently available FDA-cleared rapid antigen and molecular tests primarily for influenza A and B viruses and RSV.

scholarly journals Impact of a multiplex PCR point-of-care test for influenza A/B and respiratory syncytial virus on an acute pediatric hospital ward

2018 ◽  
Vol 91 (4) ◽  
pp. 331-335 ◽  
Author(s):  
Andres I. Vecino-Ortiz ◽  
Simon D. Goldenberg ◽  
Sam T. Douthwaite ◽  
Chih-Yuan Cheng ◽  
Rebecca E. Glover ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Multiplex Pcr ◽  
Influenza A ◽  
Point Of Care ◽  
Hospital Ward ◽  
Pediatric Hospital ◽  
Point Of Care Test ◽  
Syncytial Virus

scholarly journals Performance of a Novel Point-of-Care Molecular Assay for Detection of Influenza A and B Viruses and Respiratory Syncytial Virus (Enigma MiniLab) in Children with Acute Respiratory Infection

2015 ◽  
Vol 54 (1) ◽  
pp. 212-215 ◽  
Author(s):  
Sam T. Douthwaite ◽  
Charlotte Walker ◽  
Elisabeth J. Adams ◽  
Catherine Mak ◽  
Andres Vecino Ortiz ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Influenza A ◽  
Respiratory Virus ◽  
Point Of Care ◽  
Influenza B Virus ◽  
Influenza B ◽  
Molecular Assay ◽  
B Virus ◽  
Syncytial Virus ◽  
Virus Influenza

The performance of the Enigma MiniLab assay for influenza A and B viruses and respiratory syncytial virus (RSV) was compared to a centralized laboratory respiratory virus panel. The positive and negative percent agreement for influenza A virus, influenza B virus, and RSV were 79.2% (95% confidence interval [95% CI], 57.8 to 92.9%) and 99.4% (95% CI, 98.4 to 99.9), 100% (95% CI, 47.8 to 100%) and 100% (95% CI, 99.3 to 100%), 98.5% (95% CI, 94.6 to 99.8%) and 94.5% (95% CI, 91.9 to 96.4%), respectively.

scholarly journals Diagnostic and economic evaluation of a point-of-care test for respiratory syncytial virus

2020 ◽  
Vol 6 (3) ◽  
pp. 00018-2020
Author(s):  
A. Joy Allen ◽  
Andrea Gonzalez-Ciscar ◽  
Clare Lendrem ◽  
Jana Suklan ◽  
Karen Allen ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Diagnostic Accuracy ◽  
Influenza A ◽  
Diagnostic Yield ◽  
Point Of Care ◽  
Cost Savings ◽  
Sample Type ◽  
Standard Laboratory ◽  
Point Of Care Test ◽  
Syncytial Virus

Respiratory syncytial virus is a common cause of bronchiolitis. Historically, point-of-care tests have involved antigen detection technology with limited sensitivity. The aim of this study was to prospectively evaluate the diagnostic accuracy and model the economic impact of the Roche cobas® Liat® point-of-care influenza A/B and respiratory syncytial virus test.The “DEC-RSV” study was a multi-centre, prospective, observational study in children under 2 years presenting with viral respiratory symptoms. A nasopharyngeal aspirate sample was tested using the point-of-care test and standard laboratory-based procedures. The primary outcome was accuracy of respiratory syncytial virus detection. The cost implications of adopting a point-of-care test were modelled using study data.A total of 186 participants were recruited, with both tests performed on 177 samples. The point-of-care test was invalid for 16 samples (diagnostic yield 91%) leaving 161 available for primary analysis. After resolving discrepancies, the cobas® Liat® respiratory syncytial virus test had 100.00% (95% CI 96.07%–100.00%) sensitivity and 98.53% (95% CI 92.08%–99.96%) specificity. Median time to result was 0.6 h (interquartile range (IQR) 0.5–1) for point-of-care testing and 28.9 h (IQR 26.3–48.1) for standard laboratory testing. Estimated non-diagnostic cost savings for 1000 patients, based on isolation decision-making on point-of-care test result, were £57 010, which would increase to £94 847 when cohort nursing is used.In young children the cobas® Liat® point-of-care respiratory syncytial virus test has high diagnostic accuracy using nasopharyngeal aspirates (currently an off-licence sample type). Time to result is clinically important and was favourable compared to laboratory-based testing. The potential exists for cost savings when adopting the point-of-care test.

scholarly journals Molecular point-of-care testing for influenza A/B and respiratory syncytial virus: comparison of workflow parameters for the ID Now and cobas Liat systems

2019 ◽  
Author(s):  
Stephen Young ◽  
Jamie Phillips ◽  
Christen Griego-Fullbright ◽  
Aaron Wagner ◽  
Patricia Jim ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Influenza A ◽  
Point Of Care ◽  
Turnaround Time ◽  
Walk Away ◽  
Molecular Tests ◽  
Syncytial Virus ◽  
Improve Patient Management ◽  
Parallel Workflow ◽  
Improve Patient

ABSTRACTAimsPoint-of-care (POC) tests for influenza and respiratory syncytial virus (RSV) offer the potential to improve patient management and antimicrobial stewardship. Studies have focused on performance; however, no workflow assessments have been published comparing POC molecular tests. This study compared the Liat and ID Now systems workflow, to assist end-users in selecting an influenza and/or RSV POC test.MethodsStaffing, walk-away, and turnaround time (TAT) of the Liat and ID Now systems were determined using 40 nasopharyngeal samples, positive for influenza or RSV. The ID Now system requires separate tests for influenza and RSV, so parallel (two instruments) and sequential (one instrument) workflows were evaluated.ResultsThe ID Now ranged 4.1–6.2 minutes for staffing, 1.9–10.9 minutes for walk-away and 6.4–15.8 minutes for TAT per result. The Liat ranged 1.1–1.8 minutes for staffing, 20.0–20.5 minutes for walk-away and 21.3–22.0 minutes for TAT. Mean walk-away time comprised 38.0% (influenza positive) and 68.1% (influenza negative) of TAT for ID Now and 93.7% (influenza/RSV) for Liat. The ID Now parallel workflow resulted in medians of 5.9 minutes for staffing, 9.7 minutes for walk-away, and 15.6 minutes for TAT. Assuming prevalence of 20% influenza and 20% RSV, the ID Now sequential workflow resulted in medians of 9.4 minutes for staffing, 17.4 minutes for walk-away, and 27.1 minutes for TAT.ConclusionsThe ID Now and Liat systems offer different workflow characteristics. Key considerations for implementation include value of both influenza and RSV results, clinical setting, staffing capacity, and instrument(s) placement.

Helicase dissociation and annealing of RNA-DNA hybrids by Escherichia coli Cas3 protein

2011 ◽  
Vol 439 (1) ◽  
pp. 85-95 ◽  
Author(s):  
Jamieson A. L. Howard ◽  
Stephane Delmas ◽  
Ivana Ivančić-Baće ◽  
Edward L. Bolt
Keyword(s):  
Escherichia Coli ◽  
Processing System ◽  
Loop Formation ◽  
Rna Sequences ◽  
Crispr Locus ◽  
E Coli ◽  
Dna And Rna ◽  
Acid Processing ◽  
Short Palindromic Repeat ◽  
K 12

CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) is a nucleic acid processing system in bacteria and archaea that interacts with mobile genetic elements. CRISPR DNA and RNA sequences are processed by Cas proteins: in Escherichia coli K-12, one CRISPR locus links to eight cas genes (cas1, 2, 3 and casABCDE), whose protein products promote protection against phage. In the present paper, we report that purified E. coli Cas3 catalyses ATP-independent annealing of RNA with DNA forming R-loops, hybrids of RNA base-paired into duplex DNA. ATP abolishes Cas3 R-loop formation and instead powers Cas3 helicase unwinding of the invading RNA strand of a model R-loop substrate. R-loop formation by Cas3 requires magnesium as a co-factor and is inactivated by mutagenesis of a conserved amino acid motif. Cells expressing the mutant Cas3 protein are more sensitive to plaque formation by the phage λvir. A complex of CasABCDE (‘Cascade’) also promotes R-loop formation and we discuss possible overlapping roles of Cas3 and Cascade in E. coli, and the apparently antagonistic roles of Cas3 catalysing RNA–DNA annealing and ATP-dependent helicase unwinding.

scholarly journals Prognosis of hospitalized children under 2 years of age with co-detection of influenza A and respiratory syncytial virus at the healthcare facility

2021 ◽  
Vol 21 (2) ◽  
pp. 531-537
Author(s):  
Elisa Teixeira Mendes ◽  
Hadassa L. Paranhos ◽  
Isabela C. M. Santos ◽  
Lais Bomediano de Souza ◽  
José Luis Braga de Aquino ◽  
...  
Keyword(s):  
Influenza A ◽  
Point Of Care ◽  
Rapid Test ◽  
Healthcare Facility ◽  
Orotracheal Intubation ◽  
Younger Age ◽  
Co Detection ◽  
Syncytial Virus ◽  
The Impact ◽  
Mean Time

Abstract Objectives: the aim of this study is to evaluate the impact of co-detection of Flu A and RSV using rapid immunochromatographic tests at the point of care, in pediatric patients under 2 years of age in a general hospital. Methods: a retrospective cohort study was conducted to analyze clinical outcomes in hospitalized infants with viral respiratory disease with positive results of rapid immunochromatographic test for RSV and/or Flu-A, from 2013 to 2018. A logistic regression model was adjusted to analyze predictors of orotracheal intubation during hospitalization. Results: we analyzed 220 cases: RSV (192), Flu-A (9), co-detection (19). Lethality rate was 1.8% (2 cases), and 88% (194) were under 1 year of age. Mean time of hospitalizations was higher in patients with co-detection. Variables significantly associated with orotracheal intubation were: younger age in months, comorbidities, RSV and Flu-A co-detection, and bacterial pneumonia during hospitalization. Conclusions: RSV and Flu-Aco-detection was associated with the least favorable clinical prognoses in this study. Rapid test diagnosis may provide important information at the point of care, because molecular panels are not widely accessible in general hospitals. Rapid diagnosis allows timely evaluation and treatment.

scholarly journals Molecular point-of-care testing for influenza A/B and respiratory syncytial virus: comparison of workflow parameters for the ID Now and cobas Liat systems

2019 ◽  
Vol 73 (6) ◽  
pp. 328-334 ◽  
Author(s):  
Stephen Young ◽  
Jamie Phillips ◽  
Christen Griego-Fullbright ◽  
Aaron Wagner ◽  
Patricia Jim ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Influenza A ◽  
Point Of Care ◽  
Turnaround Time ◽  
Walk Away ◽  
Molecular Tests ◽  
Syncytial Virus ◽  
Improve Patient Management ◽  
Parallel Workflow ◽  
Improve Patient

AimsPoint-of-care (POC) tests for influenza and respiratory syncytial virus (RSV) offer the potential to improve patient management and antimicrobial stewardship. Studies have focused on performance; however, no workflow assessments have been published comparing POC molecular tests. This study compared the Liat and ID Now systems workflow, to assist end-users in selecting an influenza and/or RSV POC test.MethodsStaffing, walk-away and turnaround time (TAT) of the Liat and ID Now systems were determined using 40 nasopharyngeal samples, positive for influenza or RSV. The ID Now system requires separate tests for influenza and RSV, so parallel (two instruments) and sequential (one instrument) workflows were evaluated.ResultsThe ID Now ranged 4.1–6.2 min for staffing, 1.9–10.9 min for walk-away and 6.4–15.8 min for TAT per result. The Liat ranged 1.1–1.8 min for staffing, 20.0–20.5 min for walk-away and 21.3–22.0 min for TAT. Mean walk-away time comprised 38.0% (influenza positive) and 68.1% (influenza negative) of TAT for ID Now and 93.7% (influenza/RSV) for Liat. The ID Now parallel workflow resulted in medians of 5.9 min for staffing, 9.7 min for walk-away and 15.6 min for TAT. Assuming prevalence of 20% influenza and 20% RSV, the ID Now sequential workflow resulted in medians of 9.4 min for staffing, 17.4 min for walk-away, and 27.1 min for TAT.ConclusionsThe ID Now and Liat systems offer different workflow characteristics. Key considerations for implementation include value of both influenza and RSV results, clinical setting, staffing capacity, and instrument(s) placement.

scholarly journals Accurate PCR Detection of Influenza A/B and Respiratory Syncytial Viruses by Use of Cepheid Xpert Flu+RSV Xpress Assay in Point-of-Care Settings: Comparison to Prodesse ProFlu+

2017 ◽  
Vol 56 (2) ◽  
Author(s):  
Daniel M. Cohen ◽  
Jennifer Kline ◽  
Larissa S. May ◽  
Glenn Eric Harnett ◽  
Jane Gibson ◽  
...  
Keyword(s):  
Influenza A ◽  
Clinical Laboratory ◽  
Point Of Care ◽  
Reverse Transcription Pcr ◽  
Qualitative Detection ◽  
High Concordance ◽  
Respiratory Syncytial Viruses ◽  
Syncytial Virus ◽  
Clinical Laboratory Improvement Amendments

ABSTRACT The Xpert Flu+RSV Xpress Assay is a fast, automated in vitro diagnostic test for qualitative detection and differentiation of influenza A and B viruses and respiratory syncytial virus (RSV) performed on the Cepheid GeneXpert Xpress System. The objective of this study was to establish performance characteristics of the Xpert Flu+RSV Xpress Assay compared to those of the Prodesse ProFlu+ real-time reverse transcription-PCR (RT-PCR) assay (ProFlu+) for the detection of influenza A and B viruses as well as RSV in a Clinical Laboratory Improvement Amendments (CLIA)-waived (CW) setting. Overall, the assay, using fresh and frozen nasopharyngeal (NP) swabs, demonstrated high concordance with results of the ProFlu+ assay in the combined CW and non-CW settings with positive percent agreements (PPA) (100%, 100%, and 97.1%) and negative percent agreements (NPA) (95.2%, 99.5%, and 99.6%) for influenza A and B viruses and RSV, respectively. In conclusion, this multicenter study using the Cepheid Xpert Flu+RSV Xpress Assay demonstrated high sensitivities and specificities for influenza A and B viruses and RSV in ∼60 min for use at the point-of-care in the CW setting.

Sign in / Sign up

Export Citation Format

Share Document