Collateral Sensitivity Associated with Antibiotic Resistance Plasmids
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Abstract
Collateral sensitivity (CS) is a promising alternative approach to counteract the rising problem of antibiotic resistance (ABR). CS occurs when the acquisition of resistance to one antibiotic produces increased susceptibility to a second antibiotic. For CS to be widely applicable in clinical practice, it would need to be effective against the different resistance mechanisms available to bacteria. Recent studies have focused on CS strategies designed against ABR mediated by chromosomal mutations. However, one of the main drivers of ABR in clinically relevant bacteria is the horizontal transfer of ABR genes mediated by plasmids. Here, we report the first analysis of CS associated with the acquisition of complete ABR plasmids, including the clinically important carbapenem-resistance conjugative plasmid pOXA-48. In addition, we describe the conservation of CS in clinical E. coli isolates and its application to the selective elimination of plasmid-carrying bacteria. Our results provide new insights that establish the basis for developing CS-informed treatment strategies to combat plasmid-mediated ABR.
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This manuscript is in revision at eLife
The decision letter after peer review, sent to the authors on December 29 2020, follows.
Summary
This paper describes very clearly a set of experiments to assess collateral sensitivity to certain antibiotics that is created by carriage of beta-lactam (incl carbapenenem) resistance plasmids in E. coli. This addresses one of the limitations of existing literature on CS, which typically focuses on the effects of resistance point mutations, which are clinically less significant. By documenting multiple ways that this CS is real and selectable and to a degree generalizable across genetic backgrounds, this is an important contribution in showing that CS is a real phenomoenon for clinically important resistance mechanisms.
Essential Revisions
The primary screen of 'antibiotics x plasmids' to identify …
This manuscript is in revision at eLife
The decision letter after peer review, sent to the authors on December 29 2020, follows.
Summary
This paper describes very clearly a set of experiments to assess collateral sensitivity to certain antibiotics that is created by carriage of beta-lactam (incl carbapenenem) resistance plasmids in E. coli. This addresses one of the limitations of existing literature on CS, which typically focuses on the effects of resistance point mutations, which are clinically less significant. By documenting multiple ways that this CS is real and selectable and to a degree generalizable across genetic backgrounds, this is an important contribution in showing that CS is a real phenomoenon for clinically important resistance mechanisms.
Essential Revisions
The primary screen of 'antibiotics x plasmids' to identify collateral sensitivity, presented in Figure 1B, lacks an analysis of the statistical significance of results. Supplementary data shows that measurements of MIC are a little too noisy to robustly identify 2-fold changes with only 4 or 5 replicates. Defining "significant" as "mean more than 2x" is not adequate. Using a power calculation derived from the data in the manuscript, a sample size should be determined to have a 90 (or other high) % chance of detecting a 2x difference given the variability observed between assays, and then they should be done. Ideally this would be for all organism-plasmid pairs, but at least for the ones that the preliminary screen found a mean of 2x for.
Recommendation (not required for acceptance, but please temper claims of clinical relevance if not done): The comparative killing data should be repeated in competition. This is technically more challenging but I believe not more so than the comparative growth curves. This would establish as proof of principle that a mixed population could be purified of plasmid-bearers by CS. Without this, the clinical relevance will still remain speculative. Also, two reviewers initially misread these as competition assays. The text and legend should emphasize that these are separate populations
(Not required for acceptance but suggestion for future work:) The presented work is solid but, as pointed out by the authors, there is no mechanistic explanations for the observations. It would be highly interesting to know if the collateral effects are due to specific genes (OXA-48 would be a good place to start) and/or if the observed effects are due to the plasmid backbone.
The experiment in Figure 3 demonstrates the exploitation of collateral sensitivity to preferentially inhibit plasmid-bearing bacteria. The terminology in this section refers to 'eradicate', 'mortality' etc, but in practice, the experiment defines survival as OD>0.2 after ~24 hours. It seems likely that in the 'non-surviving' conditions, waiting another day or two would show regrowth of some bacteria in these conditions.We don't think this requires any change to the experiment, only how the results are described: they show preferential inhibition of growth, not eradication. A more patient approach to identifying regrowth would be necessary to definitively state that these bacterial populations have been eradicated. Suggest tempering claims.
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