Detection of influenza A and B in respiratory secretions with the polymerase chain reaction.

Donofrio, James C.; Coonrod, J D; Davidson, James; Betts, Robert F.

doi:10.1101/gr.1.4.263

Cited by 51 publications

(39 citation statements)

References 8 publications

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“…Despite the highly conserved nature of the M-gene sequence among influenza A viruses (18), the region encompassing the 5 amino acid markers of adamantane resistance in M2 is variable among the different genetic lineages and subtypes (14). While the assay was primarily designed for the detection of markers of adamantane resistance, the variability in the target sequences also provides information on the M-gene lineage [i.e., if the M gene is of seasonal human A(H1N1), A(H3N2), swine, or avian A(H5N1) origin (14)].…”

Section: Resultsmentioning

confidence: 99%

Detection of Molecular Markers of Drug Resistance in 2009 Pandemic Influenza A (H1N1) Viruses by Pyrosequencing

Deyde

Sheu

Trujillo

et al. 2010

Antimicrob Agents Chemother

113

121

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The M2 blockers amantadine and rimantadine and the neuraminidase (NA) inhibitors (NAIs) oseltamivir and zanamivir are approved by the FDA for use for the control of influenza A virus infections. The 2009 pandemic influenza A (H1N1) viruses (H1N1pdm) are reassortants that acquired M and NA gene segments from a Eurasian adamantane-resistant swine influenza virus. NAI resistance in the H1N1pdm viruses has been rare, and its occurrence is mainly limited to oseltamivir-exposed patients. The pyrosequencing assay has been proven to be a useful tool in surveillance for drug resistance in seasonal influenza A viruses. We provide a protocol which allows the detection of adamantane resistance markers as well as the I43T change, which is unique to the H1N1pdm M2 protein. The protocol also allows the detection of changes at residues V116, I117, E119, Q136, K150, D151, D199, I223, H275, and N295 in the NA, known to alter NAI drug susceptibility. We report on the detection of the first cases of the oseltamivir resistance-conferring mutation H275Y and the I223V change in viruses from the United States using the approach described in this study. Moreover, the assay permits the quick identification of the major NA group (V106/N248, I106/D248, or I106/N248) to which a pandemic virus belongs. Pyrosequencing is well suited for the detection of drug resistance markers and signature mutations in the M and NA gene segments of the pandemic H1N1 influenza viruses.

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Section: Resultsmentioning

confidence: 99%

Detection of Molecular Markers of Drug Resistance in 2009 Pandemic Influenza A (H1N1) Viruses by Pyrosequencing

Deyde

Sheu

Trujillo

et al. 2010

Antimicrob Agents Chemother

113

121

View full text Add to dashboard Cite

show abstract

“…While this assay has been found to be most useful as an initial screening test to confirm a diagnosis of EI during an outbreak, its limited sensitivity does not make it an ideal method for the diagnosis of EIV infection on an individual animal basis (34). There have been several reports of the use of RT-PCR assays for the detection of influenza virus in clinical specimens (13)(14)(15)30); however, such assays were not widely used for the routine diagnosis of this disease. This changed, however, following the introduction of EI into Australia in 2007, when an rRT-PCR developed to detect the avian influenza virus matrix gene was used as the molecular diagnostic method of choice for EI (15,39).…”

mentioning

confidence: 99%

Development and Evaluation of One-Step TaqMan Real-Time Reverse Transcription-PCR Assays Targeting Nucleoprotein, Matrix, and Hemagglutinin Genes of Equine Influenza Virus

Lu¹,

Chambers²,

Boliar³

et al. 2009

J Clin Microbiol

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The objective of this study was to develop and evaluate new TaqMan real-time reverse transcription-PCR (rRT-PCR) assays by the use of the minor groove binding probe to detect a wide range of equine influenza virus (EIV) strains comprising both subtypes of the virus (H3N8 and H7N7). A total of eight rRT-PCR assays were developed, targeting the nucleoprotein (NP), matrix (M), and hemagglutinin (HA) genes of the two EIV subtypes. None of the eight assays cross-reacted with any of the other known equine respiratory viruses. Three rRT-PCR assays (EqFlu NP, M, and HA3) which can detect strains of the H3N8 subtype were evaluated using nasal swabs received for routine diagnosis and swabs collected from experimentally inoculated horses. All three rRT-PCR assays have greater specificity and sensitivity than virus isolation by egg inoculation (93%, 89%, and 87% sensitivity for EqFlu NP, EqFlu M, and EqFlu HA3 assays, respectively). These assays had analytical sensitivities of >10 EIV RNA molecules. Comparison of the sensitivities of rRT-PCR assays targeting the NP and M genes of both subtypes with egg inoculation and the Directigen Flu A test clearly shows that molecular assays provide the highest sensitivity. The EqFlu HA7 assay targeting the H7 HA gene is highly specific for the H7N7 subtype of EIV. It should enable highly reliable surveillance for the H7N7 subtype, which is thought to be extinct or possibly still circulating at a very low level in nature. The assays that we developed provide a fast and reliable means of EIV diagnosis and subtype identification of EIV subtypes.

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“…They have shown that RT-PCRmethods are effective methods for rapid and sensitive diagnosing for influenza infections (4,24,25). For rapid detection of influenza virus in clinical specimens, the commercially available kit of influenza A virus (Directigen Flu Aá", Beckon Dickinson, Maryland, USA) has been used widely in Japan from 1999/2000 influenza season.…”

Section: Discussionmentioning

confidence: 99%

Two Cases of Severe Bronchopneumonia due to Influenza A (H3N2) Virus. Detection of Influenza Virus Gene Using Reverse Transcription Polymerase Chain Reaction.

Doi

Takao²,

Kaneko³

et al. 2001

Intern. Med.

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Wereport two cases of severe bronchopneumoniadue to influenza A (H3N2) virus. The severity of the disease necessitated initiation of empiric therapy based on the present illness and clinical data on admission. Both patients were improved by artificial ventilation with positive end-expiratory pressures and administration of broad spectrum antibiotics and corticosteroids before confirming the diagnosis of viral bronchopneumonia using viral culture and serological tests. Within 24 hours, influenza A (H3N2) virus was identified by amplification of the pathogen genes by reverse transcription polymerase chain reaction (RT-PCR) using the stored bronchoalveolar lavage (BAL) fluids of both cases. This suggests that a combination of detection methods of pathogens using RT-PCRand BALfluid will facilitate determination of rational treatment aimed at influenza A virus. (Internal Medicine 40: 61-67, 2001)

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Detection of influenza A and B in respiratory secretions with the polymerase chain reaction.

Cited by 51 publications

References 8 publications

Detection of Molecular Markers of Drug Resistance in 2009 Pandemic Influenza A (H1N1) Viruses by Pyrosequencing

Detection of Molecular Markers of Drug Resistance in 2009 Pandemic Influenza A (H1N1) Viruses by Pyrosequencing

Development and Evaluation of One-Step TaqMan Real-Time Reverse Transcription-PCR Assays Targeting Nucleoprotein, Matrix, and Hemagglutinin Genes of Equine Influenza Virus

Two Cases of Severe Bronchopneumonia due to Influenza A (H3N2) Virus. Detection of Influenza Virus Gene Using Reverse Transcription Polymerase Chain Reaction.

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