Host and antibiotic jointly select for greater virulence in Staphylococcus aureus

Michelle Su
Kim L Hoang
McKenna Penley
Michelle H Davis
Jennifer D Gresham
Levi T Morran
Timothy D Read author has email address

Emory University School of Medicine, Division of Infectious Diseases, Atlanta, United States
Department of Biology, Emory University, Atlanta, United States
Science Department, Penn State Scranton, Dunmore, United States

https://doi.org/10.7554/eLife.107936.1

Open access
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Figures and data

Experimental evolution design.
A) Growth curves and B) total growth of ancestral MRSA and MSSA populations in vitro with and without sub-MIC oxacillin. Different letters indicate significant differences. C) MRSA and MSSA were passaged 12 times with or without hosts, in the presence or absence of a sub-MIC of the antibiotic oxacillin. Each treatment consisted of six independently evolving replicate populations. Experimental evolution treatment abbreviations are indicated in the purple and yellow boxes. Error bars indicate standard errors.

Experimental evolution design.
A) Growth curves and B) total growth of ancestral MRSA and MSSA populations in vitro with and without sub-MIC oxacillin. Different letters indicate significant differences. C) MRSA and MSSA were passaged 12 times with or without hosts, in the presence or absence of a sub-MIC of the antibiotic oxacillin. Each treatment consisted of six independently evolving replicate populations. Experimental evolution treatment abbreviations are indicated in the purple and yellow boxes. Error bars indicate standard errors.

Evolution of virulence is facilitated by exposure to both host and sub-MIC antibiotic.
A) Virulence in terms of C. elegans mortality. Dashed lines indicate respective ancestral virulence. Different letters indicate significant differences. B) Virulence in terms of the ability to hemolyze sheep’s blood, assayed at the population level. The y-axis indicates the number of evolved populations for each category. C). Host mortality from A) grouped by hemolysis status in B). D) The proportion of colonies sampled from each evolved population that are able to hemolyze sheep’s blood. Error bars indicate standard errors. *P < 0.05

Host and sub-MIC antibiotic selection facilitated pathogen growth in antibiotics.
Pathogen in vitro growth A) without oxacillin and B) in sub-MIC oxacillin. Different letters indicate significant differences. C). Oxacillin MIC of evolved populations. The y-axis indicates the number of evolved populations for each category. Error bars indicate standard errors. **P < 0.01

Regulatory genes likely played an important role in pathogen adaptation.
A) Mutations swept to fixation, excluding intergenic and synonymous mutations, grouped by general function. The size of each point indicates how many populations had acquired at least one mutation in the gene. Colored shapes next to genes indicate whether these genes are regulatory or have been implicated in virulence or antibiotic resistance in the literature (see Table S1). B) In vitro growth of populations with SCCmec and ACME deletions with or without sub-MIC oxacillin. C) Change from ancestral pathogen-induced mortality vs. mutations. D) Hemolysis status vs. mutations. E) Oxacillin MIC vs. mutations. F) Biofilm production of evolved populations. Different letters indicate significant differences. The column widths in D and E corresponds to the number of mutations. Error bars indicate standard errors. All evolved populations were sequenced except for one population from the -host-ox treatment. *P < 0.05, **P < 0.01

Regulatory genes likely played an important role in pathogen adaptation.
A) Mutations swept to fixation, excluding intergenic and synonymous mutations, grouped by general function. The size of each point indicates how many populations had acquired at least one mutation in the gene. Colored shapes next to genes indicate whether these genes are regulatory or have been implicated in virulence or antibiotic resistance in the literature (see Table S1). B) In vitro growth of populations with SCCmec and ACME deletions with or without sub-MIC oxacillin. C) Change from ancestral pathogen-induced mortality vs. mutations. D) Hemolysis status vs. mutations. E) Oxacillin MIC vs. mutations. F) Biofilm production of evolved populations. Different letters indicate significant differences. The column widths in D and E corresponds to the number of mutations. Error bars indicate standard errors. All evolved populations were sequenced except for one population from the -host-ox treatment. *P < 0.05, **P < 0.01

Mutations that arose during experiment are enriched in isolates associated with systemic infection in humans.
A) Number of genomes in our public S. aureus genome dataset (see Methods) grouped by isolation source. General host-association refers to descriptions not specific enough to assign to other categories. We excluded samples that were ambiguously specified or missing information. B) to H) Number of genomes in the database containing mutations in the respective gene. I) Number of genomes in the dataset containing the mutations arisen in agr. This gene is separate from the others to facilitate ease of visualization due to the magnitude of the y-axis. Asterisks indicate significant difference in the proportion of blood/systemic-associated genomes compared to the expected distribution in the dataset. *P < 0.05, **P < 0.01, ***P < 0.001

MRSA and MSSA underwent distinct evolutionary trajectories.
Principal component analysis of traits evolved from A) MRSA and B) MSSA ancestors. Phylogenetic tree constructed from frequencies of mutations in populations evolving from C) MRSA and D) MSSA ancestors. Scale bar indicates Euclidean distance.

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