POCT Detection of 14 Respiratory Viruses Using Multiplex RT-PCR

Because respiratory viruses play an important role in the causation and pathogenesis of acute otitis media (AOM), determining which virus has infected a child is important with respect to vaccines and antiviral drugs. In some instances, this information might be used to prevent the occurrence of AOM. We used a rapid, economical, and sensitive diagnostic system involving a multiplex nested reverse transcription–polymerase chain reaction (RT-PCR) assay to detect various respiratory viruses in clinical specimens of middle ear fluid (MEF) from children with AOM in our hospital. Multiplex RT-PCR was completed on 40 MEF samples from 28 infants and children less than 6 years old with AOM. Viral RNA was detected in 17 MEF samples (43%). Respiratory syncytial virus type A was present in 12 samples, adenovirus in 3, rhinovirus in 2, and influenza A (H3N2) in 1. The multiplex RT-PCR assay is recommended to clinical laboratories that are considering adoption of a molecular technique for viral diagnosis.

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Self-Sampling of Oropharyngeal Swabs Among Healthcare Workers for Molecular Detection of Respiratory Viruses: A Valuable Approach for Epidemiological Studies and Surveillance Programs

Frontiers in Public Health ◽

10.3389/fpubh.2020.511669 ◽

2020 ◽

Vol 8 ◽

Author(s):

Cristina Galli ◽

Laura Pellegrinelli ◽

Gabriele Del Castillo ◽

Giovanni Forni ◽

Cecilia Eugenia Gandolfi ◽

...

Keyword(s):

Real Time ◽

Healthcare Workers ◽

Molecular Detection ◽

Respiratory Viruses ◽

Epidemiological Studies ◽

Influenza Viruses ◽

Rt Pcr ◽

Surveillance Programs ◽

Group 2 ◽

Group 1

This study aimed at assessing the validity of self-collected (self-sampled) oropharyngeal (OP) swabs among healthcare workers compared to those collected by trained sentinel general practitioners (GP-sampled) from individuals with influenza-like illness (ILI), to be implemented in epidemiological studies and/or surveillance programs of viral pathogens involved in community respiratory infections. In our study, OP swabs were collected from adults (>18 years) with ILI during the 2018–2019 influenza season. Two groups of samples were considered: group 1−131 self-sampled OP swabs collected by healthcare workers after being trained on the sampling procedure; group 2−131 GP-sampled OP swabs collected from outpatients by sentinel GPs operating within the Italian Influenza Surveillance Network. To assess swabbing quality, following RNA extraction, each sample was tested for the presence of the human ribonuclease P gene (RNP) by in-house real-time reverse transcriptase–polymerase chain reaction (RT-PCR). Samples with a cycle threshold (Ct) <35 were considered adequate for further virological analysis. Influenza viruses (IVs), respiratory syncytial virus (RSV), and rhinovirus (RV) genomes were detected by in-house real-time RT-PCR. All samples were positive to RNP detection with Ct <35. The mean Ct value was similar in the two groups (group 1 vs. group 2: 25.93 ± 2.22 vs. 25.46 ± 2.40; p = 0.10). IVs, RSV, and RV positivity rates were 26.7 vs. 52.7% (p < 0.01), 7.6 vs. 9.9% (p = 0.52), and 21.4 vs. 19.9% (p = 0.76), respectively. Self-sampled OP swabs resulted as valid as GP-sampled OP swabs for molecular detection of respiratory viruses. Self-swabbing can thus be a worthwhile strategy for sample collection to implement molecular surveillance of respiratory pathogens and carry out epidemiological studies, easily reaching a larger population size.

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Metagenomic Sequencing To Detect Respiratory Viruses in Persons under Investigation for COVID-19

Journal of Clinical Microbiology ◽

10.1128/jcm.02142-20 ◽

2020 ◽

Vol 59 (1) ◽

pp. e02142-20

Author(s):

Ahmed Babiker ◽

Heath L. Bradley ◽

Victoria D. Stittleburg ◽

Jessica M. Ingersoll ◽

Autum Key ◽

...

Keyword(s):

Influenza A ◽

Viral Infections ◽

Respiratory Viruses ◽

Human Metapneumovirus ◽

Molecular Testing ◽

Metagenomic Sequencing ◽

Rt Pcr ◽

Human Coronavirus ◽

Testing Algorithms ◽

Syncytial Virus

ABSTRACTBroad testing for respiratory viruses among persons under investigation (PUIs) for SARS-CoV-2 has been performed inconsistently, limiting our understanding of alternative viral infections and coinfections in these patients. RNA metagenomic next-generation sequencing (mNGS) offers an agnostic tool for the detection of both SARS-CoV-2 and other RNA respiratory viruses in PUIs. Here, we used RNA mNGS to assess the frequencies of alternative viral infections in SARS-CoV-2 RT-PCR-negative PUIs (n = 30) and viral coinfections in SARS-CoV-2 RT-PCR-positive PUIs (n = 45). mNGS identified all viruses detected by routine clinical testing (influenza A [n = 3], human metapneumovirus [n = 2], and human coronavirus OC43 [n = 2], and human coronavirus HKU1 [n = 1]). mNGS also identified both coinfections (1, 2.2%) and alternative viral infections (4, 13.3%) that were not detected by routine clinical workup (respiratory syncytial virus [n = 3], human metapneumovirus [n = 1], and human coronavirus NL63 [n = 1]). Among SARS-CoV-2 RT-PCR-positive PUIs, lower cycle threshold (CT) values correlated with greater SARS-CoV-2 read recovery by mNGS (R2, 0.65; P < 0.001). Our results suggest that current broad-spectrum molecular testing algorithms identify most respiratory viral infections among SARS-CoV-2 PUIs, when available and implemented consistently.

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A novel multiplex real-time RT-PCR assay with FRET hybridization probes for the detection and quantitation of 13 respiratory viruses

Journal of Virological Methods ◽

10.1016/j.jviromet.2010.02.005 ◽

2010 ◽

Vol 165 (2) ◽

pp. 254-260 ◽

Cited By ~ 44

Author(s):

R. Lassaunière ◽

T. Kresfelder ◽

M. Venter

Keyword(s):

Real Time ◽

Respiratory Viruses ◽

Rt Pcr ◽

Pcr Assay ◽

Hybridization Probes

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Comparison of Luminex xTAG® RVP fast assay and real time RT-PCR for the detection of respiratory viruses in adults with community-acquired pneumonia

Journal of Medical Virology ◽

10.1002/jmv.24463 ◽

2016 ◽

Vol 88 (7) ◽

pp. 1173-1179 ◽

Cited By ~ 4

Author(s):

Vivian Luchsinger ◽

Yara Prades ◽

Mauricio Ruiz ◽

Rolando Pizarro ◽

Patricio Rossi ◽

...

Keyword(s):

Real Time ◽

Respiratory Viruses ◽

Community Acquired Pneumonia ◽

Rt Pcr

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Surveillance of Respiratory Viruses in Long Term Care Facilities

Online Journal of Public Health Informatics ◽

10.5210/ojphi.v11i1.9893 ◽

2019 ◽

Vol 11 (1) ◽

Author(s):

Mary Checivich ◽

Shari Barlow ◽

Peter Shult ◽

Erik Residorf ◽

Jonathan L. Temte

Keyword(s):

Influenza A ◽

Nasal Swab ◽

Long Term Care ◽

Respiratory Infections ◽

Respiratory Viruses ◽

Influenza B ◽

Rt Pcr ◽

Term Care ◽

Care Facilities

ObjectiveTo assess the feasibility of conducting respiratory virus surveillance for residents of long term care facilities (LTCF) using simple nasal swab specimens and to describe the virology of acute respiratory infections (ARI) in LCTFs.IntroductionAlthough residents of LTCFs have high morbidity and mortality associated with ARIs, there is very limited information on the virology of ARI in LTCFs.[1,2] Moreover, most virological testing of LCTF residents is reactive and is triggered by a resident meeting selected surveillance criteria. We report on incidental findings from a prospective trial of introducing rapid influenza diagnostic testing (RIDT) in ten Wisconsin LTCFs over a two-year period with an approach of testing any resident with ARI.MethodsAny resident with new onset of respiratory symptoms consistent with ARI had a nasal swab specimen collected for RIDT by nursing staff. Following processing for RIDT (Quidel Sofia Influenza A+B FIA), the residual swab was placed into viral transport medium and forwarded to the Wisconsin State Laboratory of Hygiene and tested for influenza using RT-PCR (IVD CDC Human Influenza Virus Real-Time RT-PCR Diagnostic Panel), and for 17 viruses (Luminex NxTAG Respiratory Pathogen Panel [RPP]). The numbers of viruses in each of 7 categories [influenza A (FluA ), influenza B (FluB), coronaviruses (COR), human metapneumovirus (hMPV), parainfluenza (PARA), respiratory syncytial virus (RSV) and rhinovirus/enterovirus (R/E)], across the two years were compared using chi-square.ResultsTotals of 164 and 190 specimens were submitted during 2016-2017 and 2017-2018, respectively. RPP identified viruses in 56.2% of specimens, with no difference in capture rate between years (55.5% vs. 56.8%). Influenza A (21.5%), influenza B (16.5%), RSV (19.0%) and hMPV (16.5%) accounted for 73.5% of all detections, while coronaviruses (15.5%), rhino/enteroviruses (8.5%) and parainfluenza (2.5%) were less common. Specific distribution of viruses varied significantly across the two years (Table: X2=48.1, df=6; p<0.001).ConclusionsSurveillance in LTCFs using nasal swabs collected for RIDT is highly feasible and yields virus identification rates similar to those obtained in clinical surveillance of ARI with collection of nasopharyngeal specimens by clinicians and those obtained in a school-based surveillance project of ARI with collection of combined nasal and oropharyngeal specimens collected by trained research assistants. Significant differences in virus composition occurred across the two study years. RSV varied little between years while hMPV demonstrated wide variation. Simple approaches to surveillance may provide a more comprehensive assessment of respiratory viruses in LTCF settings.References(1) Uršič T, Gorišek Miksić N, Lusa L, Strle F, Petrovec M. Viral respiratory infections in a nursing home: a six-month prospective study. BMC Infect Dis. 2016; 16: 637. Published online 2016 Nov 4. doi: 10.1186/s12879-016-1962-8(2) Masse S, Capai L, Falchi A. Epidemiology of Respiratory Pathogens among Elderly Nursing Home Residents with Acute Respiratory Infections in Corsica, France, 2013–2017. Biomed Res Int. 2017; 2017: 1423718. Published online 2017 Dec 17. doi: 10.1155/2017/1423718

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Co-infections of MERS-CoV with other respiratory viruses in Saudi Arabia

African Journal of Clinical and Experimental Microbiology ◽

10.4314/ajcem.v21i4.4 ◽

2020 ◽

Vol 21 (4) ◽

pp. 284-289

Author(s):

K. Al-Quthami ◽

W.S. Al-Waneen ◽

B.O. Al Johnyi

Keyword(s):

Saudi Arabia ◽

Middle East ◽

Fast Track ◽

Respiratory Viruses ◽

Middle East Respiratory Syndrome ◽

Respiratory Pathogen ◽

Fisher Exact Test ◽

Rt Pcr ◽

Pcr Multiplex ◽

Arabie Saoudite

Background: The Middle East Respiratory Syndrome (MERS) is a viral respiratory disease caused by a member of the coronaviruses called Middle East Respiratory Syndrome Coronavirus (MERS-CoV). The co-infections of MERS-CoV with other respiratory viruses have been documented in rare cases in the scientific literature. This study was carried out to determine whether confection of MERS-CoV occurs with other respiratory viruses in Saudi Arabia.Methods: Nasopharyngeal swabs samples of 57 MERS-CoV positive outpatients were collected using flocked swabs. Nucleic acid was extracted from each sample using commercial NucliSens easyMAG system. Amplification was performed by multiplex RT-PCR using Fast Track Diagnostics Respiratory Pathogen 33. Data were analyzed with SPSS software version 19 and comparison of variables was done with Fisher Exact test, with p value <0.05 considered significant.Results: Six of the total 57 MERS-COV patients (35 males, 22 females) were positive for co-infection of MERS CoV with other respiratory viruses, giving a prevalence rate of 10.5%, with 14.5% (5/35) in males and 4.5% (1/22) in females (OR=3.500, 95% CI=0.3806-32.188, p=0.3889). The prevalence of co-infections was significantly higher among non-Saudis (23.8%, 5/21) than Saudis (2.8%, 1/36) (OR=0.09143, 95% CI=0.009855-0.8485, p=0.0217), and among the age group 18-34 years (25%, 3/12) than other age groups (X2=3.649, p=0.1613). Human rhinovirus (HRV) was found in 2 of the 6 (33.3%) patients with co-infection while the other viruses were found in each of the remaining 4 patients.Conclusion: Our study confirms that MERS-CoV co-infects with other respiratory viruses in Saudi Arabia. Keywords: MERS-CoV; URTI; Co-infection; Coronavirus French title: Co-infections de MERS-CoV avec d'autres virus respiratoires en Arabie saoudite Contexte: Le syndrome respiratoire du Moyen-Orient (MERS) est une maladie respiratoire virale causée par un membre des coronavirus appelé coronavirus du syndrome respiratoire du Moyen-Orient (MERS-CoV). Les co-infections de MERS-CoV avec d'autres virus respiratoires ont été documentées dans de rares cas dans la littérature scientifique. Cette étude a été réalisée pour déterminer si la confection du MERS-CoV se produit avec d'autres virus respiratoires en Arabie saoudite. Méthodes: Des écouvillons nasopharyngés de 57 patients ambulatoires positifs au MERS-CoV ont été prélevés à l'aide d'écouvillons floqués. L'acide nucléique a été extrait de chaque échantillon en utilisant le système NucliSens easyMAG commercial. L'amplification a été réalisée par RT-PCR multiplex en utilisant Fast Track Diagnostics Respiratory Pathogen 33. Les données ont été analysées avec le logiciel SPSS version 19 et la comparaison des variables a été effectuée avec le test Fisher Exact, avec une valeur p<0,05 considérée comme significative. Résultats: Six des 57 patients MERS-COV (35 hommes, 22 femmes) étaient positifs pour la co-infection de MERS CoV avec d'autres virus respiratoires, donnant un taux de prévalence de 10,5%, avec 14,5% (5/35) chez les hommes et 4,5% (1/22) chez les femelles (OR 3.500, 95% CI 0.3806-32.188, p=0.3889). La prévalence des co-infections était plus élevée chez les non-saoudiens (23.8%, 5/21) que chez les saoudiens (2.8%, 1/36) (OR=0.09143, 95% CI=0.009855-0.8485, p=0.0217) et parmi le groupe d'âge de 18 à 34 ans (25%, 3/12) que dans les autres groupes d'âge (X2=3.649, p=0.1613). Le rhinovirus humain (VRC) a été trouvé chez 2 des 6 (33,3%) patients co-infectés tandis que les autres virus ont été trouvés chez chacun des 4 patients restants. Conclusion: Notre étude confirme que le MERS-CoV co-infecte avec d'autres virus respiratoires en Arabie Saoudite. Mots-clés: MERS-CoV; URTI; Co-infection; Coronavirus

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