Data from: Cross-feeding constrains the evolution of antibiotic resistance in Escherichia coli
Abstract
Dataset DOI: 10.5061/dryad.wm37pvn2b
Description of the data and file structure
Reference Information
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Provenance for this README
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* File name: README_file.txt
File/Folder Details
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Details for: Data.pdf
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* Description: a series of tables containing population densities, population minimum antibiotic inhibitory concentration of kanamycin, amino acid sercetion concentration, relative gene expression levels and area under the growth curve of bacterial population in the study.
* Format(s): .pdf
* Size(s): 1.14 MB
Details for: Data.pdf__Figure1
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*Explanation:
This is the design of the evolutionary experiment, and it does not include the experimental data.
Details for: Data.pdf__Figure2
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*Methods:
Monoculture and coculture populations were transferred into fresh M9 medium every 72 h with kanamycin or without knanmycin, all the populations were subjected to five serial passages over 15 days.
Minimum antibiotic inhibitory concentration (MIC) of kanamycin were measured at both the initial and the final transfer stages.
* Variables:
* Group: Mono-culture of ΔL, mono-culture of ΔA, and a co-culture of ΔL and ΔA
* Kanamycin concentration used in each transfer: transfer 1 were 0.25μg/mL and 0.5μg/mL, transfer 2 were 0.5μg/mL and 1μg/mL, transfer 3 were 1μg/mL and 2μg/mL, transfer 4 were 2μg/mL and 4μg/mL, transfer 5 were 4μg/mL and 8μg/mL.
* ΔMIC was calculated by subtracting the MIC values of the initial transfer populations from those of their respective descendants (the final transfer population).
Details for: Data.pdf__Figure3
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*methods:
ΔL and ΔA strains were separate out from the monocultures and cocultures which evolved under kanamycin.
ΔL and ΔA strains were cocultured with or without supplementing with amino acids.
MIC values were determined in different cocultures.
* Variables:
*ΔL+ΔA-CO: the ΔL and ΔA strain was all separate out from the cocultures.
*ΔL+ΔA-MO: the ΔL and ΔA strain was all separate out from the monocultures.
+: the coculture pupolation was supplemented with amino acids.
Details for: Data.pdf__Figure4
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*methods:
ΔL and ΔA strains were separate out from the monocultures and cocultures which evolved under kanamycin, then monocultured in M9 medium with supplementing amino acids for 24 h and 800 rpm at 37 °C in a multifunctional microplate reader to generate growth curves.
The growth rate and doubling time were calculated from the growth curves using r software.
Real-time quantitative PCR was performed using a QuantiNovaTM SYBR® Green PCR kit to quantify the expression of key genes in the arginine and lysine metabolism pathway.
The arginine production levels of the ΔL strain evolved under antibiotic stress were measured using high performance liquid chromatography.
* Variables:
*ΔL-MO: ΔL strain was separate out from the monocultures.
*ΔA-MO: ΔA strain was separate out from the monocultures.
*ΔL-CO: ΔL strain was separate out from the cocultures.
*ΔA-CO: ΔA strain was separate out from the cocultures.
Details for: Data.pdf__Figure5
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*methods:
ΔL and ΔA strains were separate out from the monocultures and cocultures which evolved in the absence of kanamycin.
ΔL and ΔA strains were cocultured with or without supplementing with amino acids.
MIC values were determined in different cocultures.
* Variables:
*ΔL+ΔA-CO: the ΔL and ΔA strain was all separate out from the cocultures.
*ΔL+ΔA-MO: the ΔL and ΔA strain was all separate out from the monocultures.
+: the coculture pupolation was supplemented with amino acids.
Details for: Data.pdf__Figure6
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*methods:
Area under the growth curve of ΔL and ΔA strains in both mono-cultures and co-cultures with and without supplemental amino acids were calculated.
The ΔL and ΔA strains that coevolved in the absence of the antibiotic were pre-cultured overnight, washed three times with 0.85% NaCl to thoroughly remove any residual amino acids from the pre-culture medium, and resuspended in 0.85% NaCl to the same optical density (OD600).
Then the cultures were monocultured and cocultured for 36 h in M9 medium supplemented with and without lysine or arginine.
The corresponding abundance of each strain (CFU) was estimated by plating these cultures onto selective agar culture medium over the course of 36 h.
* Variables:
*ΔL-MO: ΔL strain was separate out from the monocultures.
*ΔA-MO: ΔA strain was separate out from the monocultures.
*ΔL-CO: ΔL strain was separate out from the cocultures.
*ΔA-CO: ΔA strain was separate out from the cocultures.
+: the pupolation was supplemented with amino acids.
*AUC: area under the growth curve, which was caculated by GraphPad Prism 8.
Details for: Data.pdf__FigureS1
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*methods:
The ancestral ΔL and ΔA strain were monocultured in M9 medium which contained 0.08 g/L, 0.09 g/L, 0.1 g/L, 0.11 g/L, 0.12 g/L, 0.13 g/L, 0.14 g/L, 0.15 g/L, 0.20 g/L, 0.25 g/L and 0.30 g/L lysine or arginine.
All populations were cultured for 36 h and 800 rpm at 37 °C in a multifunctional microplate reader, the OD600 value was measured at hourly intervals.
The concentration that reached the maximum optical density (Max OD600) was considered as the most appropriate supplement concentration of exogenous amino acid.
* Variables:
*Max OD600: the maximum optical density.
Details for: Data.pdf__FigureS2
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*methods:
The ancestral ΔL strain、ancestral ΔA strain and wild type strain were monocultured in M9 medium without supplementing amino acids.
All populations were cultured for 24 h and 800 rpm at 37 °C in a multifunctional microplate reader, the OD600 value was measured at hourly intervals.
* Variables:
*ΔL-ANC: ancestral ΔL strain.
*ΔA-ANC: ancestral ΔA strain.
*Wild-type: Escherichia coli K-12 MG1655
Details for: Data.pdf__FigureS3
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*methods:
Molecular validation of ΔL and ΔA strain.
Gel was run at 150 V for 15min, the wild type gene band of lysA and argH were at 1762 bp and 1873 bp, ΔL and ΔA strains gene band of were at 500bp.
* Variables:
*ΔL-ANC: ancestral ΔL strain.
*ΔA-ANC: ancestral ΔA strain.
*Wild-type: Escherichia coli K-12 MG1655
*ΔL-K5-C: ΔL strain was separate out from the cocultures under kanamycin at the final transfer.
*ΔL-K5-M: ΔL strain was separate out from the monocultures under kanamycin at the final transfer.
*ΔA-K5-C: ΔA strain was separate out from the cocultures under kanamycin at the final transfer.
*ΔA-K5-M: ΔA strain was separate out from the monocultures under kanamycin at the final transfer.
*ΔL-T5-C: ΔL strain was separate out from the cocultures without kanamycin at the final transfer.
*ΔL-T5-M: ΔL strain was separate out from the monocultures without kanamycin at the final transfer.
*ΔA-T5-C: ΔA strain was separate out from the cocultures without kanamycin at the final transfer.
*ΔA-T5-M: ΔA strain was separate out from the monocultures without kanamycin at the final transfer.
Details for: Data.pdf__FigureS4
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*methods:
The ancestral ΔL and ΔA strain were monocultured or cocultuerd in M9 medium.
All populations were cultured for 64 h and 800 rpm at 37 °C in a multifunctional microplate reader, the OD600 value was measured at hourly intervals.
Doubling time were derived from specific growth rates used the package Growthcurver in R-version 4.3.1, relative fitness was expressed as the ratio of the Malthusian parameter (m) in which the initial culture OD600 was annotated as N0 and the culture OD600 at time i was annotated as Ni.
mi=[ln(Ni/N0)]/i
* Variables:
*ΔL-ANC: monocultures of ancestral ΔL strain.
*ΔA-ANC: monocultures of ancestral ΔA strain.
*ΔL+ΔA-ANC: cocultures of ancestral ΔA and ΔLstrain.
+: the pupolation was supplemented with amino acids.
Details for: Data.pdf__FigureS5
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*methods:
Ancestral ΔL and ΔA strain were monocultured and cocultured under 12 different kanamycin concentrations.
After 36 h incubation, the OD600 were measured using a microplate reader, and the MIC was determined by GraphPad Prism 8 software.
* Variables:
*ΔL-ANC: monocultures of ancestral ΔL strain.
*ΔA-ANC: monocultures of ancestral ΔA strain.
*ΔL+ΔA-ANC: cocultures of ancestral ΔA and ΔLstrain.
+: the pupolation was supplemented with amino acids.
Details for: Data.pdf__FigureS6
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*methods:
Ancestral ΔL and ΔA strain, ΔL and ΔA strain which evolved under kanamycin at final transfer, were cocultured for 36h, the OD600 value was measured at hourly intervals.
Growth rate were caculated using the package Growthcurver in R-version 4.3.1.
* Variables:
*ΔL+ΔA-ANC: cocultures of ancestral ΔA and ΔLstrain.
*ΔL+ΔA-CO: cocultures of ΔA and ΔLstrain, which coevolved under kanamycin at the final transfer.
*ΔL+ΔA-MO: cocultures of ΔA and ΔLstrain, which monoevolved under kanamycin at the final transfer.
*Max OD600: maximum optical density
Details for: Data.pdf__FigureS7
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*methods:
ΔL and ΔA strain which derived from kanamycin at the initial and final transfer were cocultured for 24h, the OD600 value was measured at hourly intervals.
* Variables:
*ΔL+ΔA-CO: cocultures of ΔA and ΔLstrain, which coevolved under kanamycin.
*ΔL+ΔA-MO: cocultures of ΔA and ΔLstrain, which monoevolved under kanamycin.
+: the pupolation was supplemented with amino acids.
*Kanamycin: the strains were evolved under kanamycin.
Details for: Data.pdf__FigureS8
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*methods:
ΔL and ΔA strain which derived from kanamycin at the final transfer were monocultured or cocultured for 24h, the OD600 value was measured at hourly intervals.
* Variables:
*ΔL+ΔA-CO: cocultures of ΔA and ΔLstrain, which coevolved under kanamycin.
*ΔL+ΔA-MO: cocultures of ΔA and ΔLstrain, which monoevolved under kanamycin.
*Kanamycin: the strains were evolved under kanamycin.
Details for: Data.pdf__FigureS9
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*methods:
RNA extraction of ΔL that evolved under antibiotic was performed using the RNA iso plus extraction solution.
The relative expression levels of the target genes were normalized using a reference gene (16S rRNA).
Gene expression was measured using 2-ΔΔCT method.
* Variables:
*ΔL-CO: ΔL strain was separate out from the cocultures which evolved under kanamycin.
*ΔA-CO: ΔA strain was separate out from the cocultures which evolved under kanamycin.
Details for: Data.pdf__FigureS10
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*methods:
RNA extraction of ΔL and ΔLA that evolved without antibiotic was performed using the RNA iso plus extraction solution.
The relative expression levels of the target genes were normalized using a reference gene (16S rRNA).
Gene expression was measured using 2-ΔΔCT method.
* Variables:
*ΔL-CO: ΔL strain was separate out from the cocultures which evolved without kanamycin.
*ΔA-CO: ΔA strain was separate out from the cocultures which evolved without kanamycin.
Files and variables
File: Data.pdf
Description: a series of tables containing population densities, population minimum antibiotic inhibitory concentration of kanamycin, amino acid serection concentration, relative gene expression levels and area under the growth curve of bacterial population in the study.