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. 2009 Jul;11(7):1736-51.
doi: 10.1111/j.1462-2920.2009.01900.x. Epub 2009 Mar 11.

"V体育2025版" Development and application of the human intestinal tract chip, a phylogenetic microarray: analysis of universally conserved phylotypes in the abundant microbiota of young and elderly adults

Mirjana Rajilić-Stojanović  1 , Hans G H J Heilig, Douwe Molenaar, Kajsa Kajander, Anu Surakka, Hauke Smidt, Willem M de Vos
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Free PMC article

Development and application of the human intestinal tract chip, a phylogenetic microarray: analysis of universally conserved phylotypes in the abundant microbiota of young and elderly adults

Mirjana Rajilić-Stojanović et al. Environ Microbiol. 2009 Jul.
Free PMC article

"V体育ios版" Abstract

In this paper we present the in silico assessment of the diversity of variable regions of the small subunit ribosomal RNA (SSU rRNA) gene based on an ecosystem-specific curated database, describe a probe design procedure based on two hypervariable regions with minimal redundancy and test the potential of such probe design strategy for the design of a flexible microarray platform. This resulted in the development and application of a phylogenetic microarray for studying the human gastrointestinal microbiota--referred as the human intestinal tract chip (HITChip). Over 4800 dedicated tiling oligonucleotide probes were designed based on two hypervariable regions of the SSU rRNA gene of 1140 unique microbial phylotypes (< 98% identity) following analysis of over 16,000 human intestinal SSU rRNA sequences. These HITChip probes were hybridized to a diverse set of human intestinal samples and SSU rRNA clones to validate its fingerprinting and quantification potential. Excellent reproducibility (median Pearson's correlation of 0. 99) was obtained following hybridization with T7 polymerase transcripts generated in vitro from SSU rRNA gene amplicons. A linear dose-response was observed with artificial mixtures of 40 different representative amplicons with relative abundances as low as 0. 1% of total microbiota. Analysis of three consecutively collected faecal samples from ten individuals (five young and five elderly adults) revealed temporal dynamics and confirmed that the adult intestinal microbiota is an individual-specific and relatively stable ecosystem. Further analysis of the stable part allowed for the identification of a universal microbiota core at the approximate genus level (90% sequence similarity). This core consists of members of Actinobacteria, Bacteroidetes and Firmicutes. Used as a phylogenetic fingerprinting tool with the possibility for relative quantification, the HITChip has the potential to bridge the gaps in our knowledge in the quantitative and qualitative description of the human gastrointestinal microbiota composition VSports手机版. .

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Figures (VSports app下载)

Fig. 2
Fig. 2
Correlation between relative abundance of ten phylogenetic groups assessed as average of the group-specific HITChip hybridization signals (empty columns) and FISH quantification (filled columns) based on the analysis of faecal microbiota of five subjects. Error bars indicate standard deviations.
Fig. 1
Fig. 1
Correlation of results obtained by the Bifidobacterium-specific FISH quantification and sum of the Bifidobacterium-specific HITChip hybridization signals for 60 faecal samples. Faecal samples obtained from 20 individuals in three time points were analysed, of which four samples that did not reach detection limit with the FISH are not plotted on the graph.
Fig. 3
Fig. 3
Clustering of the HITChip phylogenetic fingerprints of the human gastrointestinal microbiota of three faecal samples of ten subjects collected during period of at least two months. Samples are encoded by YA1–YA5 for younger adults, and by EA1–EA5 for elderly adults, while 1, 2 or 3 subsequent to slash sign indicate sample collection sequence. The highest phylogenetic level of specificity of probes (level 1) is depicted on the right panel of the figure.
Fig. 4
Fig. 4
Similarity of the intestinal microbiota profiles for ten individuals calculated for 1 (grey bars) and two month (open bars) span. Results for total microbiota and six level 1 groups of the microbiota (phyla Actinobacteria, Bacteroidetes and Proteobacteria, and three groups within the Firmicutes phylum) are presented as box whisker plot. The box extends from the first quartile until the third quartile, with a line at the median, while whiskers are indicating minimal and maximal observation in the data set.
Fig. 5
Fig. 5
Venn diagram showing the distribution of the HITChip probes that showed significant hybridization signal when analysing five younger adults (A), five elderly adults (B) and between the core of younger and elderly adults (C). Only the core probes (those that were corresponding in all three analysed samples per subject) were taken into consideration for this analysis. For subject codes see Fig. 3. The image was generated using AutoFocus software (Aduna B.V., the Netherlands).
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