Rapid Differentiation of Seasonal and Pandemic H1N1 Influenza through Proteotyping of Viral Neuraminidase with Mass Spectrometry

Schwahn, Alexander B.; Wong, Jason; Downard, Kevin M.

doi:10.1021/ac100594j

Cited by 32 publications

(42 citation statements)

References 24 publications

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“…The most prominent bands were detected for nucleoprotein and matrix M1 protein in accord with their higher copy numbers per virion, as evident in our previous studies [18], [19]. The matrix protein constitutes some 40% of the total viral protein while the hemagglutinin and neuraminidase surface antigens constitute only 25% and 5% of all viral protein respectively.…”

Section: Resultssupporting

confidence: 80%

“…It complements related studies that have employed proteomics methods and mass spectrometry to characterise the antigenicity of the influenza virus [15]–[18]. More recently, it has been shown that the proteotyping approach can distinguish seasonal from pandemic type A H1N1 influenza strains [19] and also assign the lineage of human strains of type A H1N1 and type B influenza virus [20].…”

Section: Introductionmentioning

confidence: 80%

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Proteotyping to Establish Gene Origin within Reassortant Influenza Viruses

Schwahn

Downard

2011

PLoS ONE

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The application of a rapid and direct proteotyping approach with which to identify the gene origin of viral antigens in a reassortant influenza strain is demonstrated. The reassortant strain, constructed for a vaccine against type A 2009 H1N1 pandemic influenza, contains genes derived from a wild-type pandemic strain (A/California/7/2009) and an egg adapted high-growth strain (denoted NYMC X-157) derived from an earlier A/Puerto Rico/8/34 strain. The proteotyping approach employs modern proteomics methods and high resolution mass spectrometry to correctly establish that the genes of the surface antigens, hemagglutinin and neuraminidase, are derived from the A/California/7/2009 strain while those for nucleoprotein and matrix protein M1 antigens are derived from the NYMC X-157 strain. This is achieved for both gel-separated antigens and those from a whole vaccine digest. Furthermore, signature peptides detected in the mass spectra of the digested antigens enable the engineered reassortant strain to be identified as a type A virus of the H1N1 subtype in accord with earlier studies. The results demonstrate that proteotyping approach provides a more direct and rapid approach over RT-PCR with which to characterize reassortant strains of the influenza virus at the molecular protein level. Given that these strains pose the greatest risk to human and animal health and have been responsible for all human pandemics of the 20th and 21st centuries, there is a vital need for the origins and evolutionary history of these strains to be rapidly established.

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Section: Resultssupporting

confidence: 80%

Section: Introductionmentioning

confidence: 80%

Proteotyping to Establish Gene Origin within Reassortant Influenza Viruses

Schwahn

Downard

2011

PLoS ONE

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“…Solutions of viral peptides (1 l) were diluted with a solution (4 l) of matrix (5 mg/ml ␣-cyano-4-hydroxycinnaminic acid in 50% acetonitrile with 0.1% trifluoroacetic acid), and 1 l was spotted onto a matrix-assisted laser desorption ionization-Fourier-transform ion cyclotron resonance (MALDI-FT-ICR) mass spectrometry sample plate (MTP AnchorChip 400/384 TF; Bruker Daltonics, Billerica, MA). MALDI-FT-ICR mass spectra were recorded with a 7T Bruker APEX-Qe instrument (Bruker Daltonics, Billerica, MA) in the positive-ion mode, as described previously (8)(9)(10)(11). The mass spectra shown are representative of mass spectra obtained from three replicate experiments.…”

Section: Growth Of Clinical Virus Specimensmentioning

confidence: 99%

“…The approach has been shown to be able to distinguish seasonal and pandemic influenza strains (10), to determine strain lineages (11), and to identify reassortant viruses that pose the greatest pandemic risk (12). The practicality of this proteotyping approach for the surveillance of circulating strains of the virus, in terms of primary clinical specimens grown in cell culture, is demonstrated here for the first time.…”

mentioning

confidence: 97%

Incorporation of a Proteotyping Approach Using Mass Spectrometry for Surveillance of Influenza Virus in Cell-Cultured Strains

Fernandes

Downard

2014

J Clin Microbiol

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The reemergence of deadly pandemic influenza virus strains has necessitated the development of improved methods for rapid detection and subtyping of influenza viruses that will enable more strains to be characterized at the molecular level. Representative circulating strains of human influenza viruses from primary clinical specimens were grown in cell culture, purified through polyethylene glycol precipitation, proteolytically digested with an endoproteinase, and analyzed and identified by high-resolution mass spectrometry using unique signature peptides that are characteristic of type A H1N1 and H3N2 and type B influenza viruses. This proteotyping approach enabled circulating strains of type A influenza virus to be typed and subtyped, cocirculating seasonal and pandemic H1N1 viruses to be differentiated, and the lineage of type B viruses to be determined through single-ion detection by high-resolution mass spectrometry. Results were obtained using virus titers comparable to those used in reverse transcription (RT)-PCR assays with clinical specimens grown in cell cultures. The methodology represents a more rapid and direct approach than RT-PCR and can be integrated into existing procedures currently used for the surveillance of emerging pandemic and seasonal influenza viruses.

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“…The high-resolution MS of whole antigen, virus or vaccine digests allows a rapid subtyping of influenza strains using the neuraminidase as marker. Even, the pandemic (H1N1) 2009 influenza strain has an unique signature peptides different from seasonal type A (H1N1) influenza strains [151].…”

Section: Virus Strain Typingmentioning

confidence: 99%