Bacterial DNA on the skin surface overrepresents the viable skin microbiome
- Department of Molecular Biology, Princeton University, United States
- Lewis-Sigler Institute for Integrative Genomics, Princeton University, United States
- Department of Pathology, Immunology and Microbiology, Vanderbilt University Medical Center, United States
- Vanderbilt Institute for Infection, Immunology and Inflammation,, United States
- Department of Dermatology, University of Pennsylvania, United States
Figures
Figure 1 with 1 supplement
Bacterial fluorescence in situ hybridization (FISH) staining of human tissue.
(A–C) Scale bar = 20 µm. The bottom-left corner of each diagram shows a schematic of the hair follicle in white and the anatomic location of each image frame in yellow. DAPI staining is shown in …
Figure 1—figure supplement 1
Fluorescence in situ hybridization (FISH) on stationary-phase skin microbiome bacterial species.
All bacterial species were grown in appropriate conditions to stationary phase. The pan-bacterial FISH probe EUB338 was used. Hybridization is indicated in red.
Figure 2 with 2 supplements
Propidium monoazide-droplet digital PCR (PMA-ddPCR) and viability scores for human skin and non-skin microbiomes.
(A) Schematic of the PMA-ddPCR workflow. (B) Sampling scheme showing each skin site that was sampled. Colors indicate site type (sebaceous in blue, moist in green, dry in red). (C) PMA-ddPCR on skin …
Figure 2—figure supplement 1
Propidium monoazide-droplet digital PCR (PMA-ddPCR) and sampling controls.
(A) PMA-ddPCR validation controls on known ratios of heat-killed and exponentially growing E. coli cultures. PMA-ddPCR performed on a population of exponentially growing cells resulted in a …
Figure 2—figure supplement 2
Copies per 20 µL droplet digital PCR (ddPCR) reaction without (A) and with (B) the use of propidium monoazide (PMA).
Data in (A) and (B) were used to calculate the viability score shown in Figure 2. Error bars represent standard deviation.
Figure 3 with 1 supplement
Relative abundance and change in richness and diversity of traditional sequencing compared to PMA-seq.
(A) Relative abundance of all sequenced bacterial taxa at the family level. Paired bars represent data from traditional sequencing (left) and PMA-seq (right). Samples are ordered by increasing …
Figure 3—figure supplement 1
Full list of identified taxa with corresponding colors.
(A) Full list of identified bacterial groups shown in Figure 3.
Figure 4 with 1 supplement
Propidium monoazide (PMA) index and change in relative abundance between traditional and PMA-seq.
(A) The PMA index for each bacterial taxon that was present in at least four samples is shown here as an average between samples of the same sample site shown in Figure 3. Color indicates PMA index …
Figure 4—figure supplement 1
Viability score for three skin sites using lysostaphin and Staphylococcus-specific PCR primers.
(A) The three skin sites with the most abundant bacterial DNA are shown. Half of each sample was treated with lysostaphin prior to DNA isolation to assess how the viability score would change. …
Figure 5
Bacterial fluorescence in situ hybridization (FISH) staining of mouse tissue (A–D) and comparison of mouse viability scores and human viability scores.
(A). Tissues from a K14-H2B-GFP mouse stained with EUB338 show abundant bacterial signal in hair follicles but not on the skin surface. (B) Tissues from SKH1-Hrhr Elite nude mice also show bacterial …
Figure 6 with 3 supplements
Skin microbiome perturbation and recovery.
(A) Bacterial relative abundance in each individual over the 48 hr following perturbation. 0 hr represents baseline, pre-perturbation community. Whether a sample was treated with propidium monoazide …
Figure 6—figure supplement 1
List of bacteria identified in perturbation recovery.
The bacterial groups listed here correspond to the entire sequencing dataset shown in Figure 6. Bacteria are listed in order of relative abundance.
Figure 6—figure supplement 2
Skin microbiome perturbation and recovery.
(A) Bray–Curtis dissimilarity of each individual over the 48 hr following perturbation. Red data points are comparing propidium monoazide (PMA)-treated samples to the PMA-untreated baseline sample. …
Figure 6—figure supplement 3
Contamination removal performed on 600 nt sequencing data.
The Shannon diversity changes between traditional sequencing (Htrad) and PMA-seq (HPMA) are demonstrated by plotting the change in diversity (∆H) against Htrad (A–C). The richness changes between …
Additional files
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Supplementary file 1
Nucleotide sequences used in this study.
- https://cdn.elifesciences.org/articles/87192/elife-87192-supp1-v1.docx
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Supplementary file 2
- https://cdn.elifesciences.org/articles/87192/elife-87192-supp2-v1.docx
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Supplementary file 3
Sequence counts in perturbation recovery.
- https://cdn.elifesciences.org/articles/87192/elife-87192-supp3-v1.docx
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MDAR checklist
- https://cdn.elifesciences.org/articles/87192/elife-87192-mdarchecklist1-v1.docx
