Type

Journal Article

Authors

Paul W O’ Toole
R Paul Ross
Willem M. de Vos
Hauka Smidt
Julian R. Marchesi
Janne Nikkila
Qiong Wang
Orla O'Sullivan
Marcus J. Claesson

Subjects

Microbiology

Topics
rna ribosomal 16s aged intestines microbiology bacteria classification community bacteria phylogeny dna bacterial oligonucleotide array sequence analysis humans intestines structures phylogeny bacteria genes bacterial genetics microbiology microbiota molecular profiling 16s rrna phylogenetic microarray pyrosequencing classification human intestinal microbiota gene expression profiling microarray analysis

Comparative Analysis of Pyrosequencing and a Phylogenetic Microarray for Exploring Microbial Community Structures in the Human Distal Intestine (2009)

Abstract Background: Variations in the composition of the human intestinal microbiota are linked to diverse health conditions. Highthroughputmolecular technologies have recently elucidated microbial community structure at much higher resolution thanwas previously possible. Here we compare two such methods, pyrosequencing and a phylogenetic array, and evaluateclassifications based on two variable 16S rRNA gene regions.Methods and Findings: Over 1.75 million amplicon sequences were generated from the V4 and V6 regions of 16S rRNAgenes in bacterial DNA extracted from four fecal samples of elderly individuals. The phylotype richness, for individualsamples, was 1,400–1,800 for V4 reads and 12,500 for V6 reads, and 5,200 unique phylotypes when combining V4 readsfrom all samples. The RDP-classifier was more efficient for the V4 than for the far less conserved and shorter V6 region, butdifferences in community structure also affected efficiency. Even when analyzing only 20% of the reads, the majority of themicrobial diversity was captured in two samples tested. DNA from the four samples was hybridized against the HumanIntestinal Tract (HIT) Chip, a phylogenetic microarray for community profiling. Comparison of clustering of genus countsfrom pyrosequencing and HITChip data revealed highly similar profiles. Furthermore, correlations of sequence abundanceand hybridization signal intensities were very high for lower-order ranks, but lower at family-level, which was probably dueto ambiguous taxonomic groupings.Conclusions: The RDP-classifier consistently assigned most V4 sequences from human intestinal samples down to genuslevelwith good accuracy and speed. This is the deepest sequencing of single gastrointestinal samples reported to date, butmicrobial richness levels have still not leveled out. A majority of these diversities can also be captured with five times lowersampling-depth. HITChip hybridizations and resulting community profiles correlate well with pyrosequencing-basedcompositions, especially for lower-order ranks, indicating high robustness of both approaches. However, incompatiblegrouping schemes make exact comparison difficult.
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Full list of authors on original publication

Paul W O’ Toole, R Paul Ross, Willem M. de Vos, Hauka Smidt, Julian R. Marchesi, Janne Nikkila, Qiong Wang, Orla O'Sullivan, Marcus J. Claesson

Experts in our system

1
R Paul Ross
Teagasc
Total Publications: 499
 
2
Willem M. de Vos
University College Cork
Total Publications: 12
 
3
Julian R Marchesi
University College Cork
Total Publications: 45
 
4
Orla O'Sullivan
Teagasc
Total Publications: 103
 
5
Marcus J. Claesson
University College Cork
Total Publications: 53