<xhtml:span xmlns:xhtml="http://www.w3.org/1999/xhtml" xml:lang="en">Antimicrobial Effect of Blue Light on Antibiotic-Sensitive and Drug-Resistant Escherichia coli: A Novel Isotropic Optical Fiber </xhtml:span>
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BACKGROUND Orthopaedic oncologic pelvic reconstructions have an elevated risk of infection with gram-negative bacteria. This study evaluates the bactericidal ability of a novel antimicrobial blue light (ABL)-emitting optical fiber on antibiotic-sensitive E. coli (AS-Ec) and ESBL-producing E. coli (ESBL-Ec). MATERIALS AND METHODS Time-to-kill assays used a 10 mL NaCl solution with a starting inocula of 1x105 CFU/mL for AS-Ec or ESBL-Ec; assays were repeated a minimum of 3 times per strain. Experimental tubes had either 1 optical fiber (20.1 mW/mm; low power [LP]) or 2 optical fibers (40.3 mW/mm; high power [HP]), which delivered 5 wavelengths of ABL over 60 minutes. Control tubes had no optical fibers. 50µL samples taken from each tube at 0, 10, 20, 30 and 60 min were streaked onto agar plates and incubated. CFU/mL was determined. Bactericidal reduction was defined as a 99.9% (≥3 log10) reduction in CFU/mL. One-way ANOVA were conducted. RESULTS Bactericidal effects were seen for AS-Ec under both LP-ABL and HP-ABL with a log10CFU/mL±SD difference of 3.44±0.35 (p=0.043) and 3.74±0.21 (p=0.048) at 30 min and 20 min, respectively. For ESBL-Ec, while there was a significant reduction in bacterial colony formation, the bactericidal threshold was not reached with a log10CFU/mL±SD difference of only 1.02±0.41 (p=0.034) and 2.53±0.22 (p=0.037) at 60 min for LP-ABL and HP-ABL, respectively. CONCLUSIONS A novel ABL-emitting optical fiber exhibited bactericidal effects in AS-Ec and a clinically meaningful reduction of ESBL-Ec, providing a promising avenue for the use of ABL as a potential therapy for gram-negative infections.
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All reviewer comments have been sufficiently addressed and the presented work is scientifically sound.
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The reviewers have highlighted minor concerns with the work presented. Please ensure that you address their comments.
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Comments to Author
The manuscript "Antimicrobial Effect of Blue Light on Antibiotic-Sensitive and Drug-Resistant E. coli: A Novel Isotropic Optical Fiber" addresses the critical issue of increasing antibiotic resistance among strains isolated from infections in orthopedic oncology patients and the necessity of developing novel, non-antibiotic methods for combating microorganisms. The authors have focused on creating a novel optical fiber that ensures equal light intensity over the entire active length of the fiber. Additionally, each optical fiber delivers five simultaneous and continuous wavelengths of antimicrobial blue light (aBL) at 405, 415, 435, 450, and 475 nm, which represents an innovative approach. Such research is essential for the broader clinical adoption of antimicrobial blue light. The manuscript is well …
Comments to Author
The manuscript "Antimicrobial Effect of Blue Light on Antibiotic-Sensitive and Drug-Resistant E. coli: A Novel Isotropic Optical Fiber" addresses the critical issue of increasing antibiotic resistance among strains isolated from infections in orthopedic oncology patients and the necessity of developing novel, non-antibiotic methods for combating microorganisms. The authors have focused on creating a novel optical fiber that ensures equal light intensity over the entire active length of the fiber. Additionally, each optical fiber delivers five simultaneous and continuous wavelengths of antimicrobial blue light (aBL) at 405, 415, 435, 450, and 475 nm, which represents an innovative approach. Such research is essential for the broader clinical adoption of antimicrobial blue light. The manuscript is well written and the experiments are carefully designed and conducted. However, I have a few minor comments regarding the manuscript: 1. In the title it is suggested to include the full species name - Escherichia coli, instead of the abbreviation 2. The authors do not mention the significant advantage of aBL—its low risk of developing resistance to this approach. 3. It appears that the authors should avoid claiming 100% bacterial eradication in the results (e.g., line 167) since the detection limit is between 1-2 log10, corresponding to approximately 99.99% reduction in viability. 4. What medium was used for the overnight cultures? The composition of the medium can influence the amount of endogenous porphyrins produced, making this information essential for the Materials and Methods section. Additionally, the authors do not provide a reference for the Wisplinghoff protocol, making it challenging to evaluate its appropriateness. 5. The authors irradiate bacterial samples in 0.9% NaCl. It is well-known that the type of medium can impact aBL efficacy. For example, Murdoch et al. (2012) reported that the presence of organic matter, such as proteins, can limit aBL penetration into bacteria, reducing its effectiveness. Therefore, irradiating bacteria in NaCl solution might yield higher effectiveness than irradiation in a growth medium, potentially offering a less accurate representation of aBL efficacy in live tissue. It may be worthwhile to consider this aspect in future studies. 6. While the authors observed differences in aBL response between sensitive E. coli strains and ESBL-producing strains, the Discussion section only briefly addresses the reasons for these differences. Prior studies (e.g., dos Anjos et al., 2020; 2024) may provide additional insights for this discussion. Since only two strains (one sensitive and one ESBL-producing) were tested, it is difficult to draw definitive conclusions about the reduced susceptibility of ESBL strains to aBL. Supporting this discussion with additional literature would strengthen the manuscript. 7. In the Discussion section (lines 233-238), the authors state, "there are other studies that have seen comparable success to our study," but authors do not cite any specific publications, making it unclear which studies are being referenced.
Please rate the manuscript for methodological rigour
Good
Please rate the quality of the presentation and structure of the manuscript
Satisfactory
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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Comments to Author
Design and methodology * Are the research objectives clearly stated and were they met? The objectives of this study were clearly stated- "to evaluate the bactericidal effect of this novel isotropic optical fiber on antibiotic-sensitive E. coli and ESBL-producing E. coli under clinically relevant in-vitro conditions." Although the bactericidal effects were successfully measured, the conditions were arguably not clinically relevant (see below). * Is there an adequate review of the published literature? Yes. * Are the methods described in enough detail to permit someone to reproduce the experiments? Somewhat. The authors should include (1) a description of how the output power of the Antimicrobial Blue Light (ABL) was measured and (2) the environmental conditions during the time to kill assays, …
Comments to Author
Design and methodology * Are the research objectives clearly stated and were they met? The objectives of this study were clearly stated- "to evaluate the bactericidal effect of this novel isotropic optical fiber on antibiotic-sensitive E. coli and ESBL-producing E. coli under clinically relevant in-vitro conditions." Although the bactericidal effects were successfully measured, the conditions were arguably not clinically relevant (see below). * Is there an adequate review of the published literature? Yes. * Are the methods described in enough detail to permit someone to reproduce the experiments? Somewhat. The authors should include (1) a description of how the output power of the Antimicrobial Blue Light (ABL) was measured and (2) the environmental conditions during the time to kill assays, particularly temperature. * Are the methods appropriate to the research? Do they address the problem, and do they include relevant controls? No. The 'problem' as it pertains to this paper is that single point illumination, the traditional delivery system of ABL, is not effective (or optimal) in certain anatomical areas such as pelvic reconstruction cavities. (1) Without the use of a single point illumination fiber as a control, the findings reported are not novel and do not directly address the performance of the POF as compared to other fiber types. (2) More effort could have been taken to replicate in vivo conditions (i.e. cavities and luminal spaces). While the time to kill assays with bacteria growing in suspension is a good first step in assessing fiber performance, additional experiments assessing the efficacy of the delivery methods (novel POF and single point illumination control) on bacteria growing on surfaces such as paired agar plates, with varying distance from the ABL source, would seem more appropriate. By varying the distance between the two opposing agar plates, a distance to kill graph could easily be generated. * Have the authors used appropriate statistical tools to address the findings? The authors should include asterisks in the figures to denote statistical significance. * Have the authors followed relevant ethical guidelines? We will leave this judgement up to the editor as we are not familiar with the companies involved. Results and conclusions * How significant are the findings described in the article, and do they represent an advance in knowledge and understanding? The authors make a compelling argument for the need of an optimized delivery system of ABL for use in certain anatomical spaces. The effects of ABL and the mechanism of action are well established in the literature. The novelty presented in this paper lies in the design of the Polymer Optical Fiber (POF), with the goal of more effectively delivering light into the pelvic cavity and other luminal spaces as compared to the traditional single point illumination type of fiber. While the authors indeed show that delivery of ABL with the POF attenuates bacterial growth, this is nothing new on its own. The authors claim to have designed an optical fiber that provides equal light intensity and power distribution over the entire active length of the fiber, yet this is not demonstrated with any data to back up the statement. Furthermore, the authors neglect to make any direct comparisons of the POF to single point illumination fibers regarding (1) power distribution, (2) killing efficacy, or (3) functionality in cavity-type spaces. Although the implications of an optimized ABL delivery system would make a significant clinical impact on patients suffering from periprosthetic joint infections, as it stands, this paper does not demonstrate a significant advancement in knowledge on the topic. * Have the results been interpreted properly? Yes. * Are the conclusions based on sound data, with reasonable reference to the published literature? The results for low power and high power are expressed as log reduction and percent reduction and control values are not reported. Including the raw data for low power, high power, and no power control would allow the readers to better come to their own conclusions regarding the soundness of data. General * Is the article well written and of appropriate length? Yes. The article is well written and uses clear language. The only part that was unclear is line 156: "NS" was not defined. * Is the number of tables and figures appropriate, or should some of the data be published as supplementary material? Authors should include more data to demonstrate or assess (1) the equal power intensity and distribution along the novel POF, (2) comparisons of killing efficacy between the novel POF and single point illumination fiber, (3) ability of the novel POF to kill bacteria in a cavity-type space (i.e. surface growth such as agar plates with an air interface), and (4) bacterial attenuation as a function of distance from the light sources. Although not completely necessary, the results from the temperature study (lines 180 - 182) could be nice to have in table format. * Is the supplementary material relevant to the article? N/A
Please rate the manuscript for methodological rigour
Satisfactory
Please rate the quality of the presentation and structure of the manuscript
Very good
To what extent are the conclusions supported by the data?
Not at all
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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