Acinetobacter baumannii Co-Resistant to Extended Spectrum Beta-Lactamases and Carbapenemases in Six Peruvian Hospital Centers

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Review Report

General Summary

This study provides valuable insights into MDR A. baumannii in Peruvian hospitals. Addressing the limitations, particularly through a larger, geographically diverse sample and incorporating additional resistance mechanisms, could strengthen the research and inform broader public health strategies. Advantage

• Describe the phenotypic and molecular characteristics of Acinetobacter baumannii isolates carrying resistance genes to beta-lactams and carbapenems.
• Focus on isolates from six public Peruvian hospital health centers.
• Utilized the automated MicroScan system.
• Interpreted results according to M100 S30 CLSI 2020 guidelines.
• Employed conventional polymerase chain reaction (PCR).
• Visualized results using 1% agarose gel electrophoresis.

Important of Results

• Strain Reactivation:
• Out of 21 total strains, 9 (TRU1, PM1, PM2, CUS1, CUS2, CUS3, CAL1, CAL2, CAL3) were reactivated.
• Resistance Patterns:
• 77.8% resistance observed to Imipenem, Ciprofloxacin, and Cefepime.
• 66.7% resistance to Meropenem and Ceftazidime, indicating high levels of multidrug resistance.
• Gene Expression:
• Group A beta-lactamase genes (blactx-m and blatem) expressed in CUS1, CUS2, and CUS3 strains.
• Group D carbapenemase gene (blaoxa) identified in CUS3, which uniquely presented all three resistance gene groups.

Disadvantages

1. The study is confined to six public health centers in Peru, which may not be representative of the entire country or other regions. This limits the generalizability of the findings.
2. With only 21 strains analyzed, the sample size is relatively small. A larger sample would provide more robust data and might reveal additional patterns or resistance mechanisms.
3. Dependence on the MicroScan system for susceptibility testing could introduce biases or inaccuracies compared to manual methods or other automated systems.
4. While PCR is effective for detecting known resistance genes, it may not identify novel or less common genes. This could result in an incomplete understanding of the resistance mechanisms present.
5. Visualizing PCR results with 1% agarose gel electrophoresis is a relatively low-resolution method. More advanced techniques, such as sequencing, could provide more detailed information about the genetic characteristics of the isolates.
6. The study does not account for temporal changes in resistance patterns, which can evolve rapidly. Longitudinal studies would be needed to monitor trends over time.
7. The study mentions the need for rigorous surveillance and infection control but does not evaluate current practices or provide specific recommendations for improvements in these areas.
8. Although the conclusion calls for the development of alternative therapies, the study does not explore potential options or provide a framework for their development and implementation.
9.  The study does not consider environmental factors that might contribute to the spread of resistant strains, such as hospital hygiene practices, antibiotic usage policies, or patient demographics.
10. Focusing primarily on beta-lactamases and carbapenemases may overlook other important resistance mechanisms, such as efflux pumps or porin mutations, that could contribute to multidrug resistance.

Comments

• The study provides a comprehensive overview of the resistance profiles of Acinetobacter baumannii in selected Peruvian hospitals.
• The methodology is sound, employing both automated susceptibility testing and PCR for gene identification.
• The findings underscore the importance of addressing multidrug resistance through improved clinical practices and research into new treatment options.
• The study's conclusions are well-supported by the data and highlight the pressing need for public health interventions to mitigate the spread of resistant strains.

 

Comments on the Quality of English Language

 Minor editing 

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Title: Emergency of Infections by Acinetobacter baumannii co-resistant to Extended Spectrum Beta-lactamases and Carbapenemases in Peruvian Hospitals Health Centers

This manuscript is well-written by the authors. I do believe that if they can improve the manuscripts following all comments. It might have a chance to publish in the journal.

Comments

1. Topic: Please rewrite. It is complicate. No need full stop in the title.  

2. Please edit or correct the information on the abbreviation (a-e is not necessary). It would be better if the authors used English in the abbreviation.

3. Line 27: Please replace the word “is” by “was”.

4. Line 32 and Line 34: Please remove a full stop in the sentence.

5. Please write the name of the gene “bla_ctx-m” and other genes in not bold. Please use the same type of alphabet.

6. For the scientific name of the bacteria, the first time is written as Acinetobacter baumannii, the next time is written as A. baumannii. Please edit.

7. In this study, the authors used the word “Emergence”. I suggest to use the word occurrence or prevalence.

8. Line 65-67: Please edit. What is the fact in this sentence? Actually, Covid-19 is virus but the authors are talking about the resistance in bacteria.

9. Line 75-78: Why does this sentence highlight as green color?

10. Line 92-84: “These samples 92 were stored in vials with Trypticase Soy Agar at -20 °C” This sentence is incorrect. Actually, the bacterial samples are kept at -20 °C using Trypticase Soy broth plus 10%glycerol. Please edit.

11. Line 102: Nine isolates…

12. Line 102 and 121: Why did the authors select 9 isolates?

13. Line 122: An incubation at 24 h is not exponential phase of A. baumannii. Please edit.

14. Line 123: 1 x 10⁹ cfu/ml, please edit.

15. Do the authors perform statistical analysis? Please describe.

16. Please increase the magnification of the Figure.

17. Why does the error bar in the Figure 1 is high?

18. I suggest the authors to present the antimicrobial susceptibility in Table. 

19. Table 2: Please write the name of the genes in the same style as the text.

20. Please add the information of the discussion. Try to compare the results (the author’s hypothesis) with other finding by other researchers.

21. Please delete some introduction and result sentences in the discussion part.

22. Please modify the conclusion. It should be summarized on the key finding.

 

 

 

 

 

 

Comments for author File: Comments.docx

Comments on the Quality of English Language

Please correct the grammar. 

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

Dear Prof. Dr. Editor;

You will find enclosed my comments about the revision of the Manuscript ID : microbiolres-3050646 entitled " Emergency of Infections by Acinetobacter baumannii co-resistant to Extended Spectrum Beta-lactamases and Car-bapenemases in Peruvian Hospitals Health Centers”.

In this study, the authors assess the phenotypic and molecular multidrug resistance of several strains of A. baumannii to beta-lactams and carbapenems in six public Peruvian hospitals health centers in Peruvian public hospitals. The authors used different microbiological and molecular methods to reveal the resistance patterns.

Overall the work is well written. The methodological approaches lead to results that are fairly well exploited in interpreting and discussing the obtained results.

Despite this effort, the redaction of the paper contains some unclear points; and still requires amelioration in order to valorize this work.  

 

My comments are listed as follows:

Abstract:

-        Line 32: replace “ visualized by 1% agarose gel electrophoresis” with “PCR products were visualized by 1% agarose gel electrophoresis”

Introduction

Lines 81-83:  The authors do not give enough arguments; how can decision-makers and/or researchers use their results of simple identification and demonstration of resistance of A. baumannii in combating this problem?

Materials and Methods

Linse 89-92: Are the strains collected by the authors of this work during the period mentioned (between 2017 and 2019), or are they used from a collection? it must be clarified!

Lie 102: How do you explain the non-reactivation of the other strains? Could this be related to the storage conditions? if, yes, do you have an idea about a possible degradation and/or mutation the 9 strains recovered, which can thus influence their resistance?!

 

Results

Line 141 : It would be important to add a figure demonstrating the PCR products visualized.

Line 174: The results summarized in Figure 1 show very significant deviations. Therefore, some isolates may be interesting by showing low resistance, especially for GM and TM?? Explain?

 

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Reviewer 4 Report

Comments and Suggestions for Authors

This manuscript has shown a study that characterizes Acinetobacter baumannii isolates from six public Peruvian hospitals, revealing high multidrug resistance, particularly to beta-lactams and carbapenems. Multiple key resistance genes, including blactx-m, blatem, and blaoxa, were identified using PCR. The findings highlight an urgent need for enhanced surveillance, infection control, and alternative treatment strategies to combat rising bacterial resistance.

Please find my comments below,

Line 75-78. please remove the green highlight.

Line 121-134. Please provide the complete details of the PCR cycle.

Table 2. Please provide references for each resistance gene.

Figure 1 has a table in it. Please fix it.

Quality of Figure 2 is low. Please redraw it.

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Reviewer 5 Report

Comments and Suggestions for Authors

After a careful reading, this reviewer found several weaknesses of this study. Please follow: 

Title: "Emergency of infections" does not sound the right title. Please improve the title. 

Methods: Please briefly mention the methods of sample collection. 

Methods: L108: Please provide the citation for the Beckman protocol used.

Methods: Were the bacterial isolates used for phenotypic and molecular characterization cultured in separate experiments? This is not clear in the current draft of the manuscript. 

Methods: L129-131: Please also provide the volumes used in microliters, for PCR programs. 

Results: L151: "bacterial isolates reactivated"? Not sure what it means.

Results: Please explain the figure legends in more details, for Figures 1 and 2. 

Results: Please provide agarose gel images for representation of PCR detection. Data for molecular detection/characterization is not provided.

Results: To this reviewer it appears that the authors have repeatedly written the word "expression" for the markers they detected using PCR. In my understanding, the authors did not investigate the expression of markers, rather they simply detected the presence or absence of certain genes using the conventional PCR assays. If that stands correct, I suggest authors modify the text and must mention that the presence of bacterial isolate(s) in patients' samples was detected using a PCR program, without using the word "expression".  

Discussion: L222: "gene expression profiles". I failed to understand this claim. It was a simple PCR based detection and not the gene expression profiling. This must be corrected throughout the manuscript. 

Discussion: L263-265: "The differential expression of these genes in the strains studied reinforces the notion that A. baumannii employs a variety of mechanisms to adapt and counteract the action of anti-biotics". Authors should provide data and citations to support such statements. Otherwise it provides no information. 

Discussion: L267: "; highlights the unusual presence of a class A ESBL in;;;". Should be re-written for clarity, with details. What is ESBL, there is no information on that.

Discussion: L272-275: "The expression of blactx-min a non-fermenting bacillus such as A. baumannii is striking and its presence could be explained because blactx-m has the ability to distribute cosmopolitanly as well as the ease of invading human and animal cellular structures." Needs to be re-written for improvement and clarity. 

L275-276: ", Furthermore, the presence of ESBL can promote the prevalence of these markers in nosocomial pathogens, as indicated by the first report of MBL (NDM-1)". Needs significant improvements in writing. 

L304: "believed to be located on a small fragment of genetic material". The entire paragraph needs rewriting. 

Overall, the results and discussion sections need significant re-writing. Also, please provide in-text citations. 

Comments on the Quality of English Language

I suggest authors use a professional editor for English improvement. The current version has grammatical and other English errors which must be addressed. 

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Reviewer 6 Report

Comments and Suggestions for Authors

This manuscript is well written and well documented article. The importance of detecting the AMR and its geographical distribution is necessary to have an overall knowledge to see the present situation with the microbial resistance to the available treatments. It is also important to diagnose and treat the infections properly to minimize the effect of AMR. The manuscript is showing the Peruvian hospital acquired A.baumannii infection strains are developing AMR through beta-lactamase and carbapenemase gene expressions, of which developing carbapenem resistance should be a real concern in clinical aspect and it is showing a need to develop comprehensive selection of antimicrobial therapies. The representations in the table 1 and table 3 are also very helpful to see the distribution of the bacterial isolates which is directly associated with the nosocomial infection and the clinical care management.

There are two minor comments I would like to mention:

1. A green highlighted texts persist in the line no 75 to 78. I believe it is a proof-reading error which should be corrected. 

2. In Table 2 the size of the genome could be expressed as 'bp' rather than 'pb' as it is mentioned presently.

Overall the theme and the data of the manuscript are relevant to clinical studies, public health, and basic science.

Author Response

It was placed in the added documents

Author Response File: Author Response.pdf

Round 2

Reviewer 5 Report

Comments and Suggestions for Authors

Thanks for revising and considering my previous comments. No more comments from me.