Bacterial–fungal interactions in the neonatal gut influence asthma outcomes later in life
- Department of Microbiology and Immunology, University of British Columbia, Canada
- Michael Smith Laboratories, University of British Columbia, Canada
- Children’s Hospital Research Institute of Manitoba and Department of Pediatrics and Child Health, University of Manitoba, Canada
- Department of Pediatrics, University of Alberta, Canada
- School of Public Health, University of Alberta, Canada
- The Hospital for Sick Children, Canada
- Department of Medicine, McMaster University, Canada
- Department of Pediatrics, University of Toronto, Canada
- Department of Biomedical Engineering, University of British Columbia, Canada
- Department of Medical Genetics University of British Columbia, Canada
- Department of Pediatrics, University of British Columbia, Canada
- Department of Biochemistry and Molecular Biology, University of British Columbia, Canada
Figures
Figure 1
Canadian infants at high risk of asthma demonstrate increased levels of fecal fungi.
Quantitative (q) PCR quantification (standard curve method) of DNA in feces collected at 3 months of age from infants in the CHILD Cohort Study (control subjects, n = 115; Atopy+Wheeze at age 5 years…
-
Figure 1—source data 1
qPCR quantification of Pichia kudriavzevii DNA in CHILD Cohort participants.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig1-data1-v3.zip
-
Figure 1—source data 2
qPCR quantification of fungal DNA in CHILD Cohort participants.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig1-data2-v3.zip
Figure 2 with 1 supplement
Mice neonatally exposed to Pichia kudriavzevii demonstrate increased inflammatory responses during allergic airway disease later in life.
(a) Experimental procedure for neonatal exposure to P. kudriavzevii (Pichia; numbers indicate days of life) and (b) house dust mite (HDM) model of allergic airway disease (AAD). Dashed lines …
-
Figure 2—source data 1
Neonatal exposure lung cell counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-data1-v3.xlsx
-
Figure 2—source data 2
Neonatal exposure serum IgE.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-data2-v3.xlsx
-
Figure 2—source data 3
Neonatal exposure lung histology scoring.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-data3-v3.xlsx
-
Figure 2—source data 4
Neonatal exposure lung histology.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-data4-v3.xlsx
-
Figure 2—source data 5
Neonatal exposure lung pathology.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-data5-v3.xlsx
Figure 2—figure supplement 1
Adolescent exposure to Pichia kudriavzevii does not alter inflammatory responses during allergic airway disease later in life.
(a) Experimental procedure for adolescent exposure to P. kudriavzevii (Pichia; numbers indicate days of life) and house dust mite (HDM) model of allergic airway disease (AAD). (b–d) Control (n = 4) …
-
Figure 2—figure supplement 1—source data 1
Adolescent exposure lung cell counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-figsupp1-data1-v3.xlsx
-
Figure 2—figure supplement 1—source data 2
Adolescent exposure histology scoring.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig2-figsupp1-data2-v3.xlsx
Figure 3 with 4 supplements
Mice transiently colonized with Pichia kudriavzevii in neonatal life mount an immune response to this yeast and exhibit persistent alterations to their gut bacterial populations.
(a–c) Mice neonatally exposed to PBS (control) or P. kudriavzevii (Pichia) via suckling during the first 2 weeks of life were sacrificed at day 16 of life to assess for colonization by P. …
-
Figure 3—source data 1
Neonatal exposure colony counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-data1-v3.xlsx
-
Figure 3—source data 2
Neonatal exposure Pichia-specific IgG.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-data2-v3.xlsx
-
Figure 3—source data 3
Neonatal exposure colony counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-data3-v3.xlsx
-
Figure 3—source data 4
Neonatal exposure colony counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-data4-v3.xlsx
Figure 3—figure supplement 1
Mice neonatally exposed to Pichia kudriavzevii are transiently colonized and mount an immune response in the gut.
(a) Colony counts of Pi.kudriavzevii (Pichia) in fecal samples collected on day 14 (n = 7), day 16 (n = 10), day 18 (n = 15), and day 21 (n = 15) from pups neonatally exposed to P. kudriavzevii via …
-
Figure 3—figure supplement 1—source data 1
Pichia colonization time course.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-figsupp1-data1-v3.xlsx
Figure 3—figure supplement 2
Gut bacterial populations from mice neonatally exposed to Pichia kudriavzevii in PBS or PBS alone are faithfully transplanted into germ-free mice.
(a–f) Bacterial microbiota of germ-free mice given a fecal transplant with samples collected at day 28 from mice treated as described in Figure 2a. (a) Experimental design (numbers indicate days of …
Figure 3—figure supplement 3
Relative abundance of the bacterial genera identified in fecal samples based on 16S rDNA amplicon sequencing from (a) 4-week-old mice (day 28) described in Figure 2a and (b) ex-germ-free mice given a fecal transplant as described in Figure 3—figure supplement 1a 2 weeks after the fecal transplant.
Figure 3—figure supplement 4
Changes to gut bacteria resulting from neonatal colonization with Pichia kudriavzevii are not responsible for increased lung inflammation observed during allergic airway disease later in life.
(a–k) Severity of allergic airway disease (AAD) induced following the procedure in Figure 2b with house dust mite does not differ in germ-free mice given a fecal transplant with samples collected at …
-
Figure 3—figure supplement 4—source data 1
Germ-free mice lung cell counts.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-figsupp4-data1-v3.xlsx
-
Figure 3—figure supplement 4—source data 2
Germ-free mice serum IgE.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig3-figsupp4-data2-v3.xlsx
Figure 4 with 1 supplement
Short-chain fatty acids inhibit the growth of Pichia kudriavzevii.
(a–d) Growth over time (top; mean of three biological replicates) and optical density (OD) at 600 nm at 15 hr (bottom) of P. kudriavzevii grown in yeast peptone dextrose (YPD) broth supplemented …
-
Figure 4—source data 1
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data1-v3.xlsx
-
Figure 4—source data 2
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data2-v3.xlsx
-
Figure 4—source data 3
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data3-v3.xlsx
-
Figure 4—source data 4
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data4-v3.xlsx
-
Figure 4—source data 5
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data5-v3.xlsx
-
Figure 4—source data 6
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data6-v3.xlsx
-
Figure 4—source data 7
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data7-v3.xlsx
-
Figure 4—source data 8
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data8-v3.xlsx
-
Figure 4—source data 9
SCFA growth curve.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-data9-v3.xlsx
Figure 4—figure supplement 1
Mice supplemented with short-chain fatty acids (SCFAs) exhibit reduced colonization by Pichia kudriavzevii.
(a and b) Experimental design (a) and colony counts (b) in fecal samples collected from animals given an oral gavage of P. kudriavzevii 48 hr previously and maintained on regular (control; n = 15) …
-
Figure 4—figure supplement 1—source data 1
SCFA supplementation colonization.
- https://cdn.elifesciences.org/articles/67740/elife-67740-fig4-figsupp1-data1-v3.xlsx
Figure 5
Schematic summary of the hypothesized sequence of events by which early life gut fungal dysbiosis associated with increased susceptibility to asthma in childhood occurs.
Author response image 1
Author response image 2
Author response image 3
Author response image 4
Author response image 5
Author response image 6
Author response image 7
Author response image 8
Author response image 9
Goblet cell numbers quantified in primary (A), secondary (B), and tertiary (C) airways of mice in Figure 1.
(D) Shows the average goblet cell index for all airways in each animal. Figure illustrates combined results of three experiments.
Author response image 10
Author response image 11
(A-C) Growth over time (top) and optical density (OD) at 600nm at 18 hours (bottom) of Pichia kudriavzevii grown in Yeast Peptone Dextrose (YPD) broth supplemented with the sodium salts of the short chain fatty acids (SCFA) acetate (A), butyrate (B), or propionate (C) at the indicated concentrations.
(D-G) Scanning electron microscopy of P. kudriavzevii grown in YPD (D) or the sodium salts of acetate (E), butyrate (F), or propionate (G). (A-C) Data represent results from three independent …
Author response image 12
Number of eosinophils (left) and percentage of IL-17+ (middle) and ICOS+ (right) T cells in the lungs of mice given allergic airway disease via house dust mite extract sensitization and challenge in adulthood.
Gating strategies are indicated on the y-axis. Animals were neonatally exposed to the yeast Pichia kudriavzevii (Pichia) or PBC (control) and born to dams supplemented with either both acetate and …
Tables
Table 1
Anti-mouse flow cytometry antibodies used in this study.
| Cell marker | Fluorophore | Source | Clone |
|---|---|---|---|
| CD3† | eFluor450 | eBioscience | 17A2 |
| CD4 | PerCP-Cy5.5 | TONBO biosciences | RM4-5 |
| CD4 | FITC | TONBO biosciences | RM4-5 |
| FOXP3 | PE | eBioscience | FJK-16s |
| RORγt | APC | eBioscience | B2D |
| ICOS | FITC | eBioscience | 7E.17G9 |
| GATA3 | PE-Cy7 | eBioscience | TWAJ |
| CD11b† | eFluor450 | eBioscience | M1/70 |
| CD11b | APC | eBioscience | M1/70 |
| CD11c | eFluor450 | eBioscience | N418 |
| F4/80 | FITC | eBioscience | BM8 |
| CD45 | PerCPCy5.5 | eBiosceicne | 30-F11 |
| SinglecF | PE | BD Biosciences | E50-2440 |
| GR-1 | PE-Cy7 | eBioscience | RB6-8C5 |
| IL-13 | APC-eFluor780 | eBioscience | eBio13A |
| IL-5 | PE | eBioscience | TRFK5 |
| IFN-γ | AlexaFluor700 | eBioscience | XMG1.2 |
| IFN-γ | PE | TONBO biosciences | XMG1.2 |
| IL-4 | APC | eBioscience | 11B11 |
| IL-17A | PE-Cy7 | eBioscience | eBio17B7 |
| B220† | eFluor450 | eBioscience | RA3-6B2 |
| NK1.1† | eFluor450 | eBioscience | PK136 |
| FcεRI† | eFluor450 | eBioscience | MAR-1 |
-
*Cy = cyanine.
†Used to define lineage + cells.
Additional files
-
Source data 1
R code for RNA-seq data.
- https://cdn.elifesciences.org/articles/67740/elife-67740-data1-v3.zip
-
Transparent reporting form
- https://cdn.elifesciences.org/articles/67740/elife-67740-transrepform-v3.docx
