Parallel evolution of Pseudomonas aeruginosa phage resistance and virulence loss in response to phage treatment in vivo and in vitro
- College of Life and Environmental Sciences, Environment and Sustainability Institute, University of Exeter, United Kingdom
- Department of Biology, University of York, United Kingdom
- Laboratory for Molecular and Cellular Technology, Queen Astrid Military Hospital, Belgium
Figures
Figure 1
Time-shift assays of bacterial evolution and phage during phage therapy.
The proportion of phage resistant bacterial isolates from (a) ancestral (pre-phage therapy, “Anc.”), and days 2 (T1) and 4 (T2) of in vivo (clinical) phage therapy (n=8) tested against ancestral …
Figure 2
Survival curves of Galleria mellonella inoculated with Pseudomonas aeruginosa isolates isolated (a) in vivo and (b, c) in vitro.
(a) Virulence of bacterial isolates isolated from before (black) and after (gray) phage therapy. (b) Virulence of bacteria in control populations where no phage is present (yellow) is much greater …
Figure 3 with 1 supplement
Phenotypic changes underpinning the resistance—virulence trade-off of bacterial populations.
Populations were evolved (a, c) in vivo and (b, d) in vitro. (a, b) Changes in bacterial growth were estimated for populations isolated (a) before and after phage treatment, and (b) between control …
Figure 3—figure supplement 1
Growth curves of bacterial clones as measured by change in log-transformed ocular density (OD600) over time (hr).
Measures from the first 2.5 hr were excluded (blue line) as a rise in OD was observed independent of bacterial logistic growth which may skew the rolling regression process. Growth curves were …
Figure 4
Significant genetic changes identified in vitro and in vivo.
The frequency of genes that differed significantly between treatments in vitro is shown. Genetic changes that were already present in the ancestral population (first column) are shown in blue, while …
Figure 5 with 2 supplements
Genomic changes for populations evolved in vitro.
(a) The divergence of each treatment group as indicated by Euclidean distances of each gene from the ancestral isolates. Genetic changes were characterized as a change in frequency of an existing …
Figure 5—figure supplement 1
Genomic changes for populations evolved in vitro, including genetic variants occurring in genes of known and unknown function.
(a) The divergence of each treatment group as indicated by Euclidean distances of each gene from the ancestral isolates. Genetic changes were characterized as a change in frequency of an existing …
Figure 5—figure supplement 2
Associations between genetic similarity and phenotypic traits for in vivo clones.
Individual points represent individual clones isolated at day 2 (n=8) and day 4 (n=2) of phage treatment. Genes that diverged in frequency in vitro from the ancestor are indicated in gray while in …
Figure 6
Selection process of colonies used for time-series assays measuring resistance-infectivity (co)evolution and phenotypic assays (end-point resistance, virulence, growth rate, and biofilm) for isolates evolved in vitro.
For time-series assays, 16 bacterial colonies were randomly selected from each treatment replicate (phage added once or repeatedly, n=6 each) and time point (T1–T3), alongside a phage extract that …
Additional files
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Supplementary file 1
Pairwise comparisons of resistance of in vivo clones from day two against phage from contemporary (day 2) and two future time points (days 4 and 7).
P-values adjusted using the Tukey method of comparing a family of three estimates. “SE” = standard error.
- https://cdn.elifesciences.org/articles/73679/elife-73679-supp1-v2.docx
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Supplementary file 2
Susceptible colonies picked from in vitro phage treatments for phenotypic assays and sequencing.
- https://cdn.elifesciences.org/articles/73679/elife-73679-supp2-v2.docx
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Supplementary file 3
The effect of outlier removal on the significance of model terms (treatment and phage resistance).
Outlier removed states in which treatment each outlier was identified and subsequently removed. Significant p-values in bold.
- https://cdn.elifesciences.org/articles/73679/elife-73679-supp3-v2.docx
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Supplementary file 4
The effect of outlier removal on Tukey HSD contrasts comparing biofilm productivity between phage resistance groups.
Significant p-values highlighted in bold.
- https://cdn.elifesciences.org/articles/73679/elife-73679-supp4-v2.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/73679/elife-73679-transrepform1-v2.docx
