Toward an understanding of changes in diversity associated with fecal microbiome transplantation based on 16S rRNA gene deep sequencing - PubMed (original) (raw)

Toward an understanding of changes in diversity associated with fecal microbiome transplantation based on 16S rRNA gene deep sequencing

Dea Shahinas et al. mBio. 2012.

Abstract

Fecal microbiome transplantation by low-volume enema is an effective, safe, and inexpensive alternative to antibiotic therapy for patients with chronic relapsing Clostridium difficile infection (CDI). We explored the microbial diversity of pre- and posttransplant stool specimens from CDI patients (n = 6) using deep sequencing of the 16S rRNA gene. While interindividual variability in microbiota change occurs with fecal transplantation and vancomycin exposure, in this pilot study we note that clinical cure of CDI is associated with an increase in diversity and richness. Genus- and species-level analysis may reveal a cocktail of microorganisms or products thereof that will ultimately be used as a probiotic to treat CDI. IMPORTANCE Antibiotic-associated diarrhea (AAD) due to Clostridium difficile is a widespread phenomenon in hospitals today. Despite the use of antibiotics, up to 30% of patients are unable to clear the infection and suffer recurrent bouts of diarrheal disease. As a result, clinicians have resorted to fecal microbiome transplantation (FT). Donor stool for this type of therapy is typically obtained from a spouse or close relative and thoroughly tested for various pathogenic microorganisms prior to infusion. Anecdotal reports suggest a very high success rate of FT in patients who fail antibiotic treatment (>90%). We used deep-sequencing technology to explore the human microbial diversity in patients with Clostridium difficile infection (CDI) disease after FT. Genus- and species-level analysis revealed a cocktail of microorganisms in the Bacteroidetes and Firmicutes phyla that may ultimately be used as a probiotic to treat CDI.

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Figures

FIG 1

FIG 1

Rank abundance curve comparison of refOTUs for all donors and recipients pre- and post-FT. Abundance has been shown in log scale on the y axis, and the rank order of the refOTUs is shown on the x axis. The dashed blue line corresponds (99% detection) to an abundance of 5. Summary statistics are tabulated in Table S1 in the supplemental material. D#-S, donor of successful transplant; D#-F, donor of failed transplant; R# Pre, recipient pre-FT; R# Post-S, recipient post-successful FT.

FIG 2

FIG 2

Rarefaction curve comparison of refOTUs for all donors and recipients pre- and post-FT. Phylogenetic diversity is plotted against the number of V5-V6 sequence reads for each donor and recipient pre- and post-FT. D-S, donor of successful transplant; D-F, donor of failed transplant; R Pre, recipient pre-FT; R Post-S, recipient post-successful FT; R Post-F, recipient post-failed FT. Each 30th point was plotted with the confidence interval at 95%.

FIG 3

FIG 3

Diversity statistics observed for all donors and recipients pre- and post-FT. Observed taxon richness, Shannon diversity, and Simpson diversity for each sample are shown. The lines in the plots represent the medians for each group. The asterisks denote the failed-FT samples. R Pre samples are characterized by fewer taxa and lower diversity than those of post-FT and donor samples. R Pre, recipient pre-FT samples; R Post, recipient post-FT samples; D, donor samples.

FIG 4

FIG 4

(A) PCoA. PCoA represents multivariate analysis, which explores community similarity by measuring phylogenetic branch length (UniFrac) of 16S rRNA tag sequences. The dots represent the projected shadows of the samples on the respective two-dimensional planes (xy, blue; yz, red; xz, green). The x axis, y axis, and z axis represent 27.1%, 24.0%, and 20.7% of the intersample variability, respectively. R Pre, recipient pre-FT; R Post-F, recipient post-failed FT; D-S, donor of successful transplant; R Post-S, recipient post-successful FT; D-F, donor of failed transplant. (B) PCoA axes in relation to observed variability. The percentage of variability observed is depicted in relation to the first five axes. The dashed line at 5.26% represents the amount of variability expected for each axis based on random events.

FIG 5

FIG 5

Phylum-level differences for each FT set. Donor (D) and post-successful (Post-S)-FT microbial populations are distinctive from pre-FT (R Pre) and post-failed (Post-F)-FT samples by higher levels of Bacteroidetes and Firmicutes and lower levels of Proteobacteria. At the phylum level, there is no difference observed between the failed-FT donor samples and the successful-FT donor samples.

FIG 6

FIG 6

(A) Comparison of refOTU structures across the stool microbial communities. The Venn diagram displays the overlapping refOTUs for pooled samples belonging to each sample type. Each of the communities is characterized by more unique than overlapping refOTUs. The highest overlap is seen between R Pre and R Post-F samples as well as between R Post-S and D-S samples. R Pre, recipient pre-FT samples; R Post, recipient post-FT samples; D-S, donor samples. (B) Relative contributions of significant genera in pooled specimens before and after FT. Genera shown as a percentage of the total genera for each sample type are depicted. Only genera which change significantly (P < 0.05, EDGE analysis) following FT are depicted. R Pre, recipient pre-FT samples; R Post-S, recipient post-FT samples; D-S, donor samples. (C) Relative percentage of species in pooled specimens before and after FT. Species which increased significantly (P < 0.05, EDGE analysis) following FT are shown as absolute counts (number of V5-V6 sequence reads) for each sample type. R Pre, recipient pre-FT samples; R Post, recipient post-FT samples; D, donor samples.

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