Comparative analysis of pyrosequencing and a phylogenetic microarray for exploring microbial community structures in the human distal intestine.
Comparative analysis of pyrosequencing and a phylogenetic microarray for exploring microbial community structures in the human distal intestine.
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DOI：
10.1371/journal.pone.0006669
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发表时间：
2009-08-20
期刊：
影响因子：
3.7
通讯作者：
O'Toole PW
中科院分区：
文献类型：
--
作者：
Claesson MJ;O'Sullivan O;Wang Q;Nikkilä J;Marchesi JR;Smidt H;de Vos WM;Ross RP;O'Toole PW
人类肠道微生物的组成的变化与潜水员的健康状况有关。基于两个变量16S rRNA基因区域的评估分类。
超过175万个放大器序列是从16S rRNA基因的V4和V6区域中产生的细菌DNA中从四个粪便样品中提取的细菌DNA，用于单个样本的系统型，V4读数为1,400-1,800。与所有样品中的V4读取时，RDP分类器的v4读取相比较短的V6区域，但社区结构的差异也影响了效率，即使仅分析了20％的读数，大多数微生物的多样性也被捕获在两个样品中。 ）芯片，一种用于社区分析的微阵列。对于低阶等级而言，序列丰度和杂交信号强度的相关性非常高，但在家庭层面上较低，这可能是由于分类分组的模棱两可。
RDP分类器始终以良好的精度和速度分配到人类肠道样品到属级别的大多数V4序列这些多样性也可以通过降低五倍的采样深度来捕获。组成，尤其是对于低阶等级，表明两种方法的鲁棒性高。
Variations in the composition of the human intestinal microbiota are linked to diverse health conditions. High-throughput molecular technologies have recently elucidated microbial community structure at much higher resolution than was previously possible. Here we compare two such methods, pyrosequencing and a phylogenetic array, and evaluate classifications based on two variable 16S rRNA gene regions. Over 1.75 million amplicon sequences were generated from the V4 and V6 regions of 16S rRNA genes in bacterial DNA extracted from four fecal samples of elderly individuals. The phylotype richness, for individual samples, was 1,400–1,800 for V4 reads and 12,500 for V6 reads, and 5,200 unique phylotypes when combining V4 reads from all samples. The RDP-classifier was more efficient for the V4 than for the far less conserved and shorter V6 region, but differences in community structure also affected efficiency. Even when analyzing only 20% of the reads, the majority of the microbial diversity was captured in two samples tested. DNA from the four samples was hybridized against the Human Intestinal Tract (HIT) Chip, a phylogenetic microarray for community profiling. Comparison of clustering of genus counts from pyrosequencing and HITChip data revealed highly similar profiles. Furthermore, correlations of sequence abundance and hybridization signal intensities were very high for lower-order ranks, but lower at family-level, which was probably due to ambiguous taxonomic groupings. The RDP-classifier consistently assigned most V4 sequences from human intestinal samples down to genus-level with good accuracy and speed. This is the deepest sequencing of single gastrointestinal samples reported to date, but microbial richness levels have still not leveled out. A majority of these diversities can also be captured with five times lower sampling-depth. HITChip hybridizations and resulting community profiles correlate well with pyrosequencing-based compositions, especially for lower-order ranks, indicating high robustness of both approaches. However, incompatible grouping schemes make exact comparison difficult.
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影响因子：
12.3
作者：
Huse SM;Huber JA;Morrison HG;Sogin ML;Welch DM
通讯作者：
Welch DM
影响因子：
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作者：
Guschin, DY;Mobarry, BK;Mirzabekov, AD
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Mirzabekov, AD
影响因子：
56.9
作者：
Gill, Steven R.;Pop, Mihai;Nelson, Karen E.
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Nelson, Karen E.
影响因子：
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作者：
Andersson, Anders F.;Lindberg, Mathilda;Jakobsson, Hedvig;Backhed, Fredrik;Nyren, Pal;Engstrand, Lars
通讯作者：
Engstrand, Lars
影响因子：
14.9
作者：
Cole JR;Wang Q;Cardenas E;Fish J;Chai B;Farris RJ;Kulam-Syed-Mohideen AS;McGarrell DM;Marsh T;Garrity GM;Tiedje JM
通讯作者：
Tiedje JM