scholarly journals Nucleic Acid-Based Sensing Techniques for Diagnostics and Surveillance of Influenza

Biosensors ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 47 ◽  
Author(s):  
Samantha J. Courtney ◽  
Zachary R. Stromberg ◽  
Jessica Z. Kubicek-Sutherland
Keyword(s):  
Influenza Virus ◽  
Nucleic Acid ◽  
Diagnostic Test ◽  
Influenza A ◽  
Direct Detection ◽  
Point Of Care ◽  
Influenza Viruses ◽  
Nucleic Acid Amplification ◽  
High Genetic Diversity ◽  
Mild Disease

Influenza virus poses a threat to global health by causing seasonal outbreaks as well as three pandemics in the 20th century. In humans, disease is primarily caused by influenza A and B viruses, while influenza C virus causes mild disease mostly in children. Influenza D is an emerging virus found in cattle and pigs. To mitigate the morbidity and mortality associated with influenza, rapid and accurate diagnostic tests need to be deployed. However, the high genetic diversity displayed by influenza viruses presents a challenge to the development of a robust diagnostic test. Nucleic acid-based tests are more accurate than rapid antigen tests for influenza and are therefore better candidates to be used in both diagnostic and surveillance applications. Here, we review various nucleic acid-based techniques that have been applied towards the detection of influenza viruses in order to evaluate their utility as both diagnostic and surveillance tools. We discuss both traditional as well as novel methods to detect influenza viruses by covering techniques that require nucleic acid amplification or direct detection of viral RNA as well as comparing advantages and limitations for each method. There has been substantial progress in the development of nucleic acid-based sensing techniques for the detection of influenza virus. However, there is still an urgent need for a rapid and reliable influenza diagnostic test that can be used at point-of-care in order to enhance responsiveness to both seasonal and pandemic influenza outbreaks.

scholarly journals A comparative study of isothermal nucleic acid amplification methods for SARS-CoV-2 detection at point of care

2020 ◽  
Author(s):  
Diem Hong Tran ◽  
Hoang Quoc Cuong ◽  
Hau Thi Tran ◽  
Uyen Phuong Le ◽  
Hoang Dang Khoa Do ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Limit Of Detection ◽  
Direct Detection ◽  
Point Of Care ◽  
Nucleic Acid Amplification ◽  
Gold Standard Method ◽  
Synthetic Dna ◽  
Isothermal Nucleic Acid Amplification ◽  
Freeze Dried ◽  
Test Kit

ABSTRACTThe COVID-19, caused by the novel coronavirus SARS-CoV-2, has broken out of control all over the globe and put the majority of the world under lockdown. There have been no specific antiviral medications for SARS-CoV-2 while vaccines are still under development. Thus, rapid diagnosis and necessary public health measures are currently key parts to contain the pandemic. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is the gold standard method for SARS-CoV-2 detection. However, this method is not suitable for point-of-care (POC) diagnosis because of the timeconsuming procedure, the requirements of biosafety conditions and expensive equipment. In this study, the colorimetric isothermal nucleic acid amplification tests (iNAATs) for SARS-CoV-2 based on loop-mediated isothermal amplification (LAMP), cross-priming amplification (CPA), and polymerase spiral reaction (PSR) were developed and compared. The three methods exhibited similar performance with the limit of detection (LOD) as low as just 1 copy per reaction when evaluated on the synthetic DNA fragments. The results can be read with naked eyes within 30 minutes without crossreactivity to closely related coronaviruses. When tested with SARS-CoV-2 extracted genomic-RNA, LAMP outperformed both CPA and PSR assays. Moreover, the direct detection of SARS-CoV-2 in simulated patient samples (oropharyngeal and nasopharyngeal swabs) by colorimetric iNAATs was also successful. Further preparation of the lyophilized reagents for LAMP reactions revealed that the freeze-dried, ready-to-use kit maintained the sensitivity and LOD value of the liquid assays. These results strongly indicate that the colorimetric lyophilized LAMP test kit developed herein is highly suitable for detecting SARS-CoV-2 at POC.

scholarly journals Comparison of the ID Now Influenza A & B 2, Cobas Influenza A/B, and Xpert Xpress Flu Point-of-Care Nucleic Acid Amplification Tests for Influenza A/B Virus Detection in Children

2020 ◽  
Vol 58 (3) ◽  
Author(s):  
Neena Kanwar ◽  
Jeffrey Michael ◽  
Kathryn Doran ◽  
Emily Montgomery ◽  
Rangaraj Selvarangan
Keyword(s):  
Nucleic Acid ◽  
Influenza A ◽  
Point Of Care ◽  
Virus Detection ◽  
Reference Method ◽  
Nucleic Acid Amplification ◽  
Molecular Assays ◽  
B Virus ◽  
Flu Virus ◽  
Xpert Assay

ABSTRACT Early diagnosis of influenza (Flu) is critical for patient management and infection control. The ID Now influenza A & B 2 (ID Now) assay (Abbott Laboratories), Cobas influenza A/B nucleic acid test (LIAT; Roche Molecular Systems, Inc.), and Xpert Xpress Flu (Xpert; Cepheid) are rapid, point-of-care molecular assays for Flu virus detection. The study aim was to compare the performances of these three commercially available Clinical Laboratory Improvement Amendments (CLIA)-waived Flu virus assays. We prospectively enrolled 201 children <18 years old from January to April 2018 and collected nasopharyngeal swab specimens in viral medium. Aliquots were frozen for testing on different diagnostic platforms, as per the manufacturers’ instructions. CDC Flu A/B PCR was used as a reference method to evaluate the performances of these three platforms. Among the 201 specimens tested, the CDC Flu A/B PCR assay detected Flu A/B virus in 107 samples (Flu A virus, 73 samples; Flu B virus, 36 samples; dual Flu A/B virus positive, 2 samples), while the ID Now virus detected 102 samples (Flu A virus, 69 samples; Flu B virus, 37 samples; dual Flu A/B virus positive, 4 samples; invalid rate, 1/201 [0.5%]), the LIAT detected 112 samples (Flu A virus, 74 samples; Flu B virus, 38 samples; invalid rate, 11/201 [5.5%]), and the Xpert assay detected 112 samples (Flu A virus, 76 samples; Flu B virus, 36 samples; invalid rate, 6/201 [3.0%]). The overall sensitivities for the ID Now assay, LIAT, and Xpert assay for Flu A virus detection (93.2%, 100%, and 100%, respectively) and Flu B virus detection (97.2%, 94.4%, and 91.7%, respectively) were comparable. The specificity for Flu A and B virus detection by all methods was >97%. These molecular assays had higher sensitivity than did a historical standard-of-care test from the BD Veritor antigen test (Flu A virus, 79.5%; Flu B virus, 66.7%).

scholarly journals Specific Detection of Influenza A and B Viruses by CRISPR-Cas12a-Based Assay

Biosensors ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 88
Author(s):  
Bum Ju Park ◽  
Man Seong Park ◽  
Jae Myun Lee ◽  
Yoon Jae Song
Keyword(s):  
Diagnostic Test ◽  
Reverse Transcription ◽  
Influenza A ◽  
High Sensitivity ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Influenza Viruses ◽  
Nucleic Acid Amplification ◽  
Lateral Flow Strip ◽  
Influenza Pandemics

A rapid and accurate on-site diagnostic test for pathogens including influenza viruses is critical for preventing the spread of infectious diseases. Two types of influenza virus, A and B cause seasonal flu epidemics, whereas type A can cause influenza pandemics. To specifically detect influenza A (IAV) and B (IBV) viruses, we developed a clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated (Cas) system-based assay. By coupling reverse transcription recombinase polymerase amplification (RT-RPA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP), a CRISPR-Cas12a DNA endonuclease-targeted CRISPR trans-reporter (DETECTR) detected IAV and IBV titers as low as 1 × 100 plaque forming units (PFUs) per reaction without exhibiting cross-reactivity. Only 75 to 85 min were required to detect IAV and IBV, depending on isothermal nucleic acid amplification methods, and results were verified using a lateral flow strip assay that does not require additional analytic equipment. Taken together, our findings establish RT-RPA and RT-LAMP-coupled DETECTR-based diagnostic tests for rapid, specific and high-sensitivity detection of IAV and IBV using fluorescence and lateral flow assays. The diagnostic test developed in this study can be used to distinguish IAV and IBV infections, a capability that is necessary for monitoring and preventing the spread of influenza epidemics and pandemics.

scholarly journals Clinical evaluation of fully automated molecular diagnostic system “Simprova” for influenza virus, respiratory syncytial virus, and human metapneumovirus

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ikuyo Takayama ◽  
Shohei Semba ◽  
Kota Yokono ◽  
Shinji Saito ◽  
Mina Nakauchi ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Respiratory Syncytial Virus ◽  
Nucleic Acid ◽  
Point Of Care ◽  
Antigen Detection ◽  
Human Metapneumovirus ◽  
Nucleic Acid Amplification ◽  
Clinical Specimens ◽  
Nucleic Acid Amplification Tests ◽  
Syncytial Virus

Abstract Influenza virus, respiratory syncytial virus, and human metapneumovirus commonly cause acute upper and lower respiratory tract infections, especially in children and the elderly. Although rapid antigen detection tests for detecting these infections have been introduced recently, these are less sensitive than nucleic acid amplification tests. More recently, highly sensitive point-of-care testings (POCTs) have been developed based on nucleic acid amplification tests, which are easy to use in clinical settings. In this study, loop-mediated isothermal amplification (LAMP)-based POCT “Simprova” to detect influenza A and B viruses, respiratory syncytial virus, and human metapneumovirus was developed. Simprova system is fully automated and does not require skilled personnel. In addition, positive results can be achieved faster than with PCR. In this study, the accuracy of the POCT was retrospectively analyzed using 241 frozen stocked specimens. Additionally, the usability of the Simprova at clinical sites was assessed in a prospective clinical study using 380 clinical specimens and compared to those of real-time PCR and rapid antigen detection test. The novel LAMP-based POCT demonstrated high sensitivity and specificity in characterizing clinical specimens from patients with influenza-like illnesses. The Simprova is a powerful tool for early diagnosis of respiratory viral infections in point-of-care settings.

scholarly journals Point-Counterpoint: Can Newly Developed, Rapid Immunochromatographic Antigen Detection Tests Be Reliably Used for the Laboratory Diagnosis of Influenza Virus Infections?: TABLE 1

2014 ◽  
Vol 53 (6) ◽  
pp. 1790-1796 ◽  
Author(s):  
James J. Dunn ◽  
Christine C. Ginocchio
Keyword(s):  
Influenza Virus ◽  
Influenza A ◽  
Influenza Infection ◽  
Rapid Development ◽  
Hospital Setting ◽  
Antigen Detection ◽  
Influenza Viruses ◽  
Nucleic Acid Amplification ◽  
Infectious Disease Diagnostics

Five years ago, the Point-Counterpoint series was launched. The initial article asked about the role of rapid immunochromatographic antigen testing in the diagnosis of influenza A virus 2009 H1N1 infection (D. F. Welch and C. C. Ginocchio, J Clin Microbiol 48:22–25, 2010,http://dx.doi.org/10.1128/JCM.02268-09). Since that article, not only have major changes been made in immunochromatographic antigen detection (IAD) testing for the influenza viruses, but there has also been rapid development of commercially available nucleic acid amplification tests (NAATs) for influenza virus detection. Further, a novel variant of influenza A, H7N9, has emerged in Asia, and H5N1 is also reemergent. In that initial article, the editor of this series, Peter Gilligan, identified two issues that required further consideration. One was how well IAD tests worked in clinical settings, especially in times of antigen drift and shift. The other was the role of future iterations of influenza NAATs and whether this testing would be available in a community hospital setting. James Dunn, who is Director of Medical Microbiology and Virology at Texas Children's Hospital, has extensive experience using IAD tests for diagnosing influenza. He will discuss the application and value of these tests in influenza diagnosis. Christine Ginocchio, who recently retired as the Senior Medical Director, Division of Infectious Disease Diagnostics, North Shore-LIJ Health System, and now is Vice President for Global Microbiology Affairs at bioMérieux, Durham, NC, wrote the initial counterpoint in this series, where she advocated the use of NAATs for influenza diagnosis. She will update us on the commercially available NAAT systems and explain what their role should be in the diagnosis of influenza infection.

scholarly journals Understanding Nucleic Acid Amplification Techniques in the Detection of Influenza viruses in Developing Countries

2019 ◽  
Vol 6 (1) ◽  
Author(s):  
Abdulazeez A. Anjorin ◽  
Olumuyiwa B. Salu ◽  
Robert K. Obi ◽  
Bamidele O. Oke ◽  
Akeeb O. Oyefolu ◽  
...  
Keyword(s):  
Developing Countries ◽  
Influenza Virus ◽  
Nucleic Acid ◽  
Influenza A ◽  
Virus Detection ◽  
Nucleic Acid Amplification ◽  
Chain Reaction ◽  
Nucleic Acid Amplification Techniques ◽  
Influenza Virus Detection ◽  
Polymerase Chain

Introduction: Early detection of emerging influenza virus variant is a key factor in the WHO influenza Global strategies for prevention and control. Rapid, accurate, inexpensive and portable detection systems are needed for influenza virus diagnosis and surveillance. Such a detection system should easily identify all the subtypes of influenza virus. Degenerate primers and probes designed from evolutionally conserved regions for known influenza A viruses present the best way to identify unknown subtypes of influenza A virus by polymerase chain reaction PCR and array techniques. The isothermal reactions, Nucleic Acid Sequencing Based Amplification (NASBA) and Loop-mediated isothermal Amplification (LAMP) possess great potential for influenza A virus detection especially in developing countries. However, multiplex real-time (rT) or quantitative (q) polymerase chain reaction (qPCR) remains a rapid, accurate and timesaving technique used for influenza virus detection. Aim: This manuscript explained the principles of nucleic acid amplification techniques commonly used in developing countries. Methods: Literature search was done in NCBI PUBMED, PUBMED Central and Google Scholar using words and phrases including “Influenzamolecular diagnosis, NAAT”, Molecular techniques/ methods, PCR, qPCR, NASBA, LAMP, and DNA microarray. Results: The underlining principles and basic processes involved in the application of nucleic acid amplification techniques for the detection and epidemiological surveillance of influenza virus were identified and grouped under PCR (RT-PCR and qRT-PCR) and Non-PCR (LCR, pyrosequencing, NASBA, LAMP and DNA microarray) amplifications. Conclusion: It is hoped that by understanding the techniques and basic principles of Nucleic acid amplifications, less expensive, and more convenient protocols for influenza virus detection and surveillance can be developed Keywords: Influenza, NAAT, Molecular, PCR, qPCR, Viral diagnosis.

Evaluation of a rapid isothermal nucleic acid amplification kit, Alere™ i Influenza A&B, for the detection of avian influenza viruses

2019 ◽  
Vol 265 ◽  
pp. 121-125
Author(s):  
Enkhbold Bazarragchaa ◽  
Masatoshi Okamatsu ◽  
Ankhanbaatar Ulaankhuu ◽  
Augustin Tshibwabwa Twabela ◽  
Keita Matsuno ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Avian Influenza ◽  
Influenza A ◽  
Influenza Viruses ◽  
Nucleic Acid Amplification ◽  
Avian Influenza Viruses ◽  
Isothermal Nucleic Acid Amplification

scholarly journals 689. Nasal versus Nasopharyngeal Sample Collection for Diagnostic Nucleic Acid Amplification Testing for Influenza A, Influenza B, and Respiratory Syncytial Viruses

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S398-S398
Author(s):  
Grace A Schaack ◽  
Brittney Jung-Hynes ◽  
Derrick Chen
Keyword(s):  
Nucleic Acid ◽  
Influenza A ◽  
False Negative ◽  
Influenza Viruses ◽  
Viral Rna ◽  
Nucleic Acid Amplification ◽  
Influenza B ◽  
Sample Collection ◽  
Negative Results ◽  
False Negative Results

Abstract Background Nucleic acid amplification testing (NAAT) for influenza A virus (IAV), influenza B virus (IBV), and respiratory syncytial virus (RSV) is a standard component of diagnosis of infections with these pathogens. At our institution, current standard of practice is to collect nasopharyngeal (NP) samples for such NAAT. In an effort to provide clinicians and patients a simpler, more comfortable sample collection option, we evaluated the use of nasal samples for NAAT, compared to NP samples. Methods Both nasal and NP specimens were collected from each of 58 patients seen in our emergency department (January – March 2020). NP samples were collected using minitip FLOQswabs; nasal samples were collected using regular or minitip FLOQswabs. Nasal and NP samples were processed using the same protocol and tested for influenza viruses and RSV using the Cepheid GeneXpert (Xpress Flu/RSV) platform. Results In total, 20 NP samples tested negative for virus and 38 tested positive (16 IAV-positive, 14 IBV-positive, 8 RSV-positive). There were 3 cases (5% of total cases) in which qualitative (positive/negative) results from the corresponding nasal samples were not in agreement with results derived from NP samples. These were considered false-negative results; one such discrepancy was resolved upon re-testing the same samples. Overall positive percent agreement between nasal- and NP-derived results was 92% (35/38), and negative percent agreement was 100% (20/20). Among samples testing positive for virus by both NP and nasal sampling methods, we found that the average cycle threshold (Ct) value for IAV detection was 5.1 cycles (n = 16, SEM = 0.83) higher for nasal samples than for NP samples. The average Ct for IBV detection was 3.3 cycles (n = 12, SEM = 1.87) higher for nasal than for NP samples. The average Ct for RSV detection was 1.9 cycles (n = 7, SEM = 1.57) higher for nasal than for NP samples. Conclusion These results suggest that recovery of viral RNA from nasal samples is lower than that from nasopharyngeal samples, particularly for influenza viruses. This decreased detection of viral RNA from nasal samples may explain the false-negative results seen in our discrepant cases. These data suggest that a decrease in recovery of viral RNA by nasal sampling may translate to decreased diagnostic accuracy. Disclosures All Authors: No reported disclosures

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