Aminoglycoside Antibiotics Inhibit Phage Infection by Blocking an Early Step of the Infection Cycle
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Curated by eLife
Evaluation Summary:
Earlier work demonstrated the inhibition of phage infection by aminoglycosides in Mycobacteria. Following up on this prior work, the authors demonstrate that the acetylated form of apramycin retains its anti-phage activity while blocking its antibacterial activity. The authors observed that MgCl abrogated the anti-phage effects of aminoglycosides and that MgCl can inhibit aminoglycoside uptake leading to the conclusion that antibiotic uptake was likely important for the anti-phage effects. Consistent with this conclusion, the initial stages of phage infection (adsorption and DNA injection) were not impacted by the antibiotics.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Abstract
Predation by phages is a major driver of bacterial evolution. As a result, elucidating antiphage strategies is crucial from both fundamental and therapeutic standpoints.
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Evaluation Summary:
Earlier work demonstrated the inhibition of phage infection by aminoglycosides in Mycobacteria. Following up on this prior work, the authors demonstrate that the acetylated form of apramycin retains its anti-phage activity while blocking its antibacterial activity. The authors observed that MgCl abrogated the anti-phage effects of aminoglycosides and that MgCl can inhibit aminoglycoside uptake leading to the conclusion that antibiotic uptake was likely important for the anti-phage effects. Consistent with this conclusion, the initial stages of phage infection (adsorption and DNA injection) were not impacted by the antibiotics.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The …
Evaluation Summary:
Earlier work demonstrated the inhibition of phage infection by aminoglycosides in Mycobacteria. Following up on this prior work, the authors demonstrate that the acetylated form of apramycin retains its anti-phage activity while blocking its antibacterial activity. The authors observed that MgCl abrogated the anti-phage effects of aminoglycosides and that MgCl can inhibit aminoglycoside uptake leading to the conclusion that antibiotic uptake was likely important for the anti-phage effects. Consistent with this conclusion, the initial stages of phage infection (adsorption and DNA injection) were not impacted by the antibiotics.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Reviewer #1 (Public Review):
This manuscript is based on prior work, which demonstrated the inhibition of phage infection by aminoglycosides in Mycobacteria. Following up, the authors demonstrate in this manuscript that the acetylated form of apramycin retains its anti-phage activity while blocking its antibacterial activity. Along these lines, the authors describe that MgCl2 diminished the anti-phage effects of aminoglycosides, presumably due to inhibition of aminoglycoside uptake. Then the authors investigated if apramycin impacts the initial stages of phage infection, and found that DNA injection was not impacted leading to the important question of the apramycin target and if phages would actually encounter apramycin before or after infection, and which effects other aminoglycosides might have on the infection process.
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Reviewer #2 (Public Review):
The primary observations are very interesting. Their impacted is lessened by the prior demonstration of the inhibition of phage infection by aminoglycosides in Mycobacteria (Reference 18). This prior work also suggests a similar stage of phage infection is inhibited. As a consequence, perhaps the most novel and striking aspect of the paper is the demonstration that the acetylated form of apramycin retains its anti-phage activity while blocking its antibacterial activity. Further development of this aspect of the paper, or greater mechanistic insight into how aminoglycosides block phage infection would clearly strengthen the work.
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Reviewer #3 (Public Review):
The authors effectively show that aminoglycosides can protect against phage infection - and that this is by inhibiting lytic infections and not by promoting lysogeny.
It was notable that not all aminoglycosides had a protective effect. It would be helpful if the authors could comment on the differences between those that had an effect versus those that didn't in their discussion.
It was observed that MgCl abrogated the anti-phage effects of aminoglycosides, and noted that MgCl can inhibit aminoglycoside uptake. From this, the authors concluded that antibiotic uptake was likely important for the anti-phage effects - and this was nicely (but indirectly) confirmed by showing that the initial stages of phage infection (adsorption and DNA injection) were not impacted by the antibiotics. The observation did, …
Reviewer #3 (Public Review):
The authors effectively show that aminoglycosides can protect against phage infection - and that this is by inhibiting lytic infections and not by promoting lysogeny.
It was notable that not all aminoglycosides had a protective effect. It would be helpful if the authors could comment on the differences between those that had an effect versus those that didn't in their discussion.
It was observed that MgCl abrogated the anti-phage effects of aminoglycosides, and noted that MgCl can inhibit aminoglycoside uptake. From this, the authors concluded that antibiotic uptake was likely important for the anti-phage effects - and this was nicely (but indirectly) confirmed by showing that the initial stages of phage infection (adsorption and DNA injection) were not impacted by the antibiotics. The observation did, however, raise the question of whether bacteria whose antibiotic resistance is conferred by efflux would be more susceptible to phage infection than those which modify the antibiotic itself? This would be a point worth addressing in the discussion. Is antibiotic efflux more commonly associated with other antibiotic classes?
Testing the phage resistance of natural aminoglycoside producers effectively showed that these Streptomyces species could produce potent anti-phage compounds. For S. tenebrarius, an apramycin producer, spent medium was observed to have strong protective effects. Are there S. tenebrarius strains available that are defective in their ability to make apramycin? This would be a useful control to include to ensure that these strains aren't making another metabolite/releasing another compound with anti-phage activity. Similarly with the kasugamycin-producing strain - purified kasugamycin didn't have a profound effect on phage defense, based on the data presented in Fig. 1c or 2a, yet the addition of culture supernatant from the producing strain was as effective as apramycin in protecting against phage predation. It would be useful to be able to separate the aminoglycoside-specific effects, from the effect of other compounds produced by these strains (it would also be useful to know if they make anti-phage adjuvants that enhance the effects of the aminoglycosides).
The demonstration of distinct anti-bacterial versus anti-phage properties for apramycin is fascinating. Given that many of the initial experiments presented in the manuscript were done using host strains expressing antibiotic-modifying resistance determinants, might this suggest that all results obtained to this point were with modified antibiotics? (despite unmodified variants being added to the cultures) This could suggest a trade-off between anti-phage effects versus anti-bacterial effects (for microbes that are not resistant - or that do not encode antibiotic-modifying resistance determinants). Which effects are more potent? This would be an important point to discuss/address (in some cases, the concentration of antibiotic needed to protect against phage infection seems quite high).
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