Integron activity accelerates the evolution of antibiotic resistance
- University of Oxford, Department of Zoology, United Kingdom
- Universidad Complutense de Madrid, Departamento de Sanidad Animal and VISAVET, Spain
Figures
Figure 1 with 1 supplement
Overview of integron system.
(a) Diagram of the integron mechanism: the integron consists of an integrase gene, intI, followed by an array of promoterless gene cassettes (represented here by arrows). Cassettes are expressed …
Figure 1—figure supplement 1
Transcriptional and translational origin of the aadB expression gradient.
(A) Transcript levels of the aadB cassette in the different arrays. Transcription levels are normalized relatively to the best transcribed array (WTA6). Error bars represent the standard error of …
Figure 2
Integrase activity can increase bacteria evolvability against antibiotics.
(a) Schematic representation of the experimental evolution protocol. (b) Top: Representation of the WTA3 integron. Bottom: Survival curves of the PA01:WTA3 and PA01:ΔintI1A3 populations during …
Figure 3 with 3 supplements
Extensive cassette re-arrangements are linked with integrase activity.
(a,b) Distribution of cassette re-arrangements at ×4 MIC (a) and ×1024 MIC (b) time points in the WTA3 populations. Homogeneous populations represent populations where only one type of array could …
Figure 3—figure supplement 1
Plasmid mutations and re-arrangements at (a) ×4 MIC and (b) ×1024 MIC.
Representation of the plasmid mutations and re-arrangements in PA01:WTA3 and PA01: ΔintI1A3 populations, mapped to the plasmid reference sequence. Each circle represents a separate population. …
Figure 3—figure supplement 2
Re-arrangements detection by PCR in ×1024 MIC WTA3 populations.
(A) Top: Primer binding sites and amplicons used in the screen for integrase size and cassette re-arrangements. Bottom: Full gels of the PCR screen for cassettes re-arrangements of the ×1024 MIC …
Figure 3—figure supplement 3
Re-arrangements in the plasmid backbone of the WT populations at ×1024 MIC.
Left: Re-arrangements in the plasmid backbone of the WT populations at ×1024 MIC. Each junction site is indicated by a letter. Right: junction sequences for each re-arrangement. The junction site is …
Figure 4 with 2 supplements
Chromosome evolution.
(a) Summary of the chromosomal mutations at ×4 MIC (left) and at ×1024 MIC (right) mapped to the PAO1 reference sequence. Each circle represents a summary of each genotype. The type (indel, …
Figure 4—figure supplement 1
Summary statistics of mutations in the ×1024 MIC populations.
Box plots representing the average cumulative mutation frequencies for each population at the ×1024 MIC time point for (A) all mutations (B) per mutation type (C) per mutation effect (SNP only). The …
Figure 4—figure supplement 2
Summary statistics of mutations in the ×4 MIC populations.
Box plots representing the average cumulative mutation frequencies for each population at the ×4 MIC time point for (A) all mutations (B) per mutation type (C) per mutation effect (SNP only). The …
Figure 5
pAMBL1 re-arrangements.
(a) Representation of the pAMBL1 plasmid. The integron is highlighted in color. (b) Survival curve of the 30 PA01:pAMBL1 populations under ramping treatment. The black arrow indicates the time point …
Figure 6
Representation of the proposed reactions leading to the various arrays observed in this study.
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain, strain background (Pseudomonas aeruginosa) | PA01 | Lab strain | NC_002516 | |
| Strain, strain background (Escherichia coli) | MG-1 | Poirel et al., 1999 | AF205943 | E. coli clinical isolate containing a qacI–aadB–aacA1/orfG–blaVEB1–aadB–arr2–cmIA5–blaOXA–10/aadA1 integron array |
| Strain, strain background (Escherichia coli) | EIEC-4 | Gassama et al., 2004 | E. coli clinical isolate containing a dfrA5 integron cassette | |
| Recombinant DNA reagent | R388 | Avila and de la Cruz, 1988 | NC_028464.1 | |
| Recombinant DNA reagent | WTA1 | This study | Custom integron array drfA5–blaVEB1–aadB on R388 plasmid backbone | |
| Recombinant DNA reagent | WTA2 | This study | Custom integron array blaVEB1– aadB–dfrA5 on R388 plasmid backbone | |
| Recombinant DNA reagent | WTA3 | This study | Custom integron array blaVEB1–dfrA5–aadB on R388 plasmid backbone | |
| Recombinant DNA reagent | WTA4 | This study | Custom integron array dfrA5–aadB–blaVEB1 on R388 plasmid backbone | |
| Recombinant DNA reagent | WTA5 | This study | Custom integron array aadB–blaVEB1– dfrA5 on R388 plasmid backbone | |
| Recombinant DNA reagent | WTA6 | This study | Custom integron array aadB–dfrA5–blaVEB1 on R388 plasmid backbone | |
| Recombinant DNA reagent | Δint1A3 | This study | Array WTA3 with a 948 bp deletion of the integrase intI1 | |
| Recombinant DNA reagent | pAMBL1 | San Millan et al., 2015a | KP873172.1 | Clinical plasmid containing a blaVIM-1–aadB integron array |
| Software, algorithm | breseq | Barrick et al., 2014 | RRID:SCR_010810 | Version 0.33.2 |
| Software, algorithm | CNOGpro | Brynildsrud, 2018 |
Additional files
-
Supplementary file 1
Primers used in this study.
- https://cdn.elifesciences.org/articles/62474/elife-62474-supp1-v2.xlsx
-
Supplementary file 2
List of mutations / recombinations at x1024 MIC.
- https://cdn.elifesciences.org/articles/62474/elife-62474-supp2-v2.xlsx
-
Supplementary file 3
List of mutations / recombinations at x4 MIC.
- https://cdn.elifesciences.org/articles/62474/elife-62474-supp3-v2.xlsx
-
Supplementary file 4
List of duplicated cassettes from the INTEGRALL database.
- https://cdn.elifesciences.org/articles/62474/elife-62474-supp4-v2.doc
-
Transparent reporting form
- https://cdn.elifesciences.org/articles/62474/elife-62474-transrepform-v2.pdf
