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Sillanpää S, Kramna L, Oikarinen S, et al. Next-Generation Sequencing Combined with Specific PCR Assays To Determine the Bacterial 16S rRNA Gene Profiles of Middle Ear Fluid Collected from Children with Acute Otitis Media. mSphere. 2017;2(2)doi: 10.1128/mSphere.00006-17.
Sillanpää, S., Kramna, L., Oikarinen, S., Sipilä, M., Rautiainen, M., Aittoniemi, J., Laranne, J., Hyöty, H., & Cinek, O. (2017). Next-Generation Sequencing Combined with Specific PCR Assays To Determine the Bacterial 16S rRNA Gene Profiles of Middle Ear Fluid Collected from Children with Acute Otitis Media. mSphere, 2(2), . https://doi.org/10.1128/mSphere.00006-17
Sillanpää, Saara, et al. "Next-Generation Sequencing Combined with Specific PCR Assays To Determine the Bacterial 16S rRNA Gene Profiles of Middle Ear Fluid Collected from Children with Acute Otitis Media." mSphere vol. 2,2 (2017). doi: https://doi.org/10.1128/mSphere.00006-17
Sillanpää S, Kramna L, Oikarinen S, Sipilä M, Rautiainen M, Aittoniemi J, Laranne J, Hyöty H, Cinek O. Next-Generation Sequencing Combined with Specific PCR Assays To Determine the Bacterial 16S rRNA Gene Profiles of Middle Ear Fluid Collected from Children with Acute Otitis Media. mSphere. 2017 Mar 22;2(2). doi: 10.1128/mSphere.00006-17. eCollection 2017. PMID: 28357413; PMCID: PMC5362748.
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mSphere. 2017 Mar 22;2(2). doi: 10.1128/mSphere.00006-17. eCollection 2017.

Next-Generation Sequencing Combined with Specific PCR Assays To Determine the Bacterial 16S rRNA Gene Profiles of Middle Ear Fluid Collected from Children with Acute Otitis Media.

mSphere

Saara Sillanpää, Lenka Kramna, Sami Oikarinen, Markku Sipilä, Markus Rautiainen, Janne Aittoniemi, Jussi Laranne, Heikki Hyöty, Ondrej Cinek

Affiliations

  1. Department of Otorhinolaryngology and Head and Neck Surgery, Tampere University Hospital and School of Medicine, University of Tampere, Tampere, Finland.
  2. Department of Pediatrics, 2nd Faculty of Medicine, Charles University in Prague and University Hospital Motol, Prague, Czech Republic.
  3. Department of Virology, School of Medicine, University of Tampere, Tampere, Finland.
  4. Fimlab Laboratories, Tampere, Finland.
  5. Department of Virology, School of Medicine, University of Tampere, Tampere, Finland; Fimlab Laboratories, Tampere, Finland.

PMID: 28357413 PMCID: PMC5362748 DOI: 10.1128/mSphere.00006-17

Abstract

The aim of the study was to analyze the bacteriome of acute otitis media with a novel modification of next-generation sequencing techniques. Outpatient children with acute otitis media were enrolled in the study, and middle ear fluids were collected during 90 episodes from 79 subjects aged 5 to 42 months (median age, 19 months). The bacteriome profiles of middle ear fluid samples were determined by a nested-PCR amplification of the 16S rRNA gene (V4 region), followed by mass sequencing. The profiling results were compared to the results of specific PCR assays targeting selected prevalent pathogens. Bacteriome profiling using nested amplification of low-volume samples was aided by a bioinformatic subtraction of signal contaminants from the recombinant polymerase, achieving a sensitivity slightly lower than that of specific PCR detection.

Keywords: 16S profiling; acute otitis media; bacteriome profiling; mass sequencing; next-generation sequencing

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