Microbial diversity in the sputum of a cystic fibrosis patient studied with 16S rDNA pyrosequencing

Microbial diversity in the sputum of a cystic fibrosis patient studied with 16S rDNA pyrosequencing Recent studies using 16S rRNA gene amplification followed by clonal Sanger sequencing in cystic fibrosis demonstrated that cultured microorganisms are only part of the infecting flora. The purpose of this paper was to compare pyrosequencing and clonal Sanger sequencing on sputum. The sputum of a patient with cystic fibrosis was analysed by culture, Sanger clone sequencing and pyrosequencing after 16S rRNA gene amplification. A total of 4,499 sequencing reads were obtained, which could be attributed to six consensus sequences, but the length of reads leads to fastidious data analysis. Compared to clonal Sanger sequencing and to cultivation results, pyrosequencing recovers greater species richness and gives a more reliable estimate of the relative abundance of bacterial species. The 16S pyrosequencing approach expands our knowledge of the microbial diversity of cystic fibrosis sputum. The current lack of phylogenetic resolution at the species level for the GS 20 sequencing reads will be overcome with the next generation of pyrosequencing apparatus. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png European Journal of Clinical Microbiology Infectious Diseases Springer Journals

Microbial diversity in the sputum of a cystic fibrosis patient studied with 16S rDNA pyrosequencing

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Publisher
Springer Journals
Copyright
Copyright © 2009 by Springer-Verlag
Subject
Biomedicine; Medical Microbiology ; Internal Medicine
ISSN
0934-9723
eISSN
1435-4373
DOI
10.1007/s10096-009-0749-x
Publisher site
See Article on Publisher Site

Abstract

Recent studies using 16S rRNA gene amplification followed by clonal Sanger sequencing in cystic fibrosis demonstrated that cultured microorganisms are only part of the infecting flora. The purpose of this paper was to compare pyrosequencing and clonal Sanger sequencing on sputum. The sputum of a patient with cystic fibrosis was analysed by culture, Sanger clone sequencing and pyrosequencing after 16S rRNA gene amplification. A total of 4,499 sequencing reads were obtained, which could be attributed to six consensus sequences, but the length of reads leads to fastidious data analysis. Compared to clonal Sanger sequencing and to cultivation results, pyrosequencing recovers greater species richness and gives a more reliable estimate of the relative abundance of bacterial species. The 16S pyrosequencing approach expands our knowledge of the microbial diversity of cystic fibrosis sputum. The current lack of phylogenetic resolution at the species level for the GS 20 sequencing reads will be overcome with the next generation of pyrosequencing apparatus.

Journal

European Journal of Clinical Microbiology Infectious DiseasesSpringer Journals

Published: May 16, 2009

References

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