Probiotics as Antibiotic Alternatives for Human and Animal Applications
Round 1
Reviewer 1 Report
General comments to the Authors
The manuscript provides a comprehensive review of the field of probiotics as alternatives to antibiotics. The abstract adequately described the subject area and the conclusions drawn by the authors.
The introduction gave a good background to the subject area and provided a good description of the areas of most interest in the subject of probiotics as alternatives to antibiotics.
The comprehensive list of prior literature was very good to see and I thank the authors for making the effort to gather this literature together.
I noted there was no reference in the text to any of the figures and only one in text reference to a table (Table 6). I presume these references should be where where the referencing errors but I strongly encourage the authors to check this accordingly. The tables and figures should be mentioned in the text otherwise they should not be included in the manuscript.
I have not cross-checked the referencing in the text against the references.
I find the manuscript needs a large number of minor edits to improve the use of language. My recommendations in this regard are listed in the specific comments below. I would strenuously recommend these specific comments be given attention by the authors and the edits made accordingly.
Specific comments:
Line 13 – replace ‘its’ with ‘their’
Line 16 – insert the word ‘the’ between ’within’ and ‘bacterial’
Line 17 – re-word the sentence to ‘… and might be the cause of millions of deaths each year in the future.’
Line 21 – re-word the sentence to ‘… is one approach in practicing …’
Line 22 - replace ‘evade’ with ‘avoid’
Line 35 – replace ‘that’ with ‘and’
Line 47 – start the sentence with ‘The”
Line 53 – replace ‘… in the opposite …’ with ‘… as opposed to …’
Line 68 – ‘human and animal’ should be plural ‘humans and animals’
Line 92 – a referencing error in the Word document needs to be fixed
Line 111 – better wording would be ‘… have been found to have a 25 times higher chance of exchanging genetic …’
Line 128 - a referencing error in the Word document needs to be fixed
Line 129 – ‘gene’ should be ‘genes’
Line 132 – ‘enzyme’ should be ‘enzymes’
Line 133 - a referencing error in the Word document needs to be fixed
Line 144 - a referencing error in the Word document needs to be fixed
Line 163 – ‘probiotic’ should be ‘ probiotics’
Line 166 - a referencing error in the Word document needs to be fixed
Line 183 – ‘aid’ should be ‘aids’
Line 196 – separate the words ’to’ and ‘pathogenic’ (they are joined)
Line 212 – remove ‘in’
Line 299 – ‘protein’ should be ‘proteins’
Line 310 – insert ‘the’ after ‘regulate’
Line 312 – remove the words ‘In a study research,’
Line 352 - a referencing error in the Word document needs to be fixed
Line 365 – ‘treatment duration’ is better than ‘duration treatment’
Line 371 - a referencing error in the Word document needs to be fixed
Line 382 – remove the word ‘thus’
Line 384 - a referencing error in the Word document needs to be fixed
Line 392 – replace ‘so on’ with ‘others’
Line 393 - a referencing error in the Word document needs to be fixed
Line 393 – add ‘supplemented’ after ‘feed’
Table 8 – the row regarding ‘Chickens’ has errors in the spelling of the species name
Line 404 – ‘strengths and assets’ should be replaced by ‘features and functions’
Line 406 – replace ‘develop’ with ‘possess’
Line 409 – remove the first instance of ‘as’
Line 410 – change ‘anti-microbes’ to ‘antimicrobials’
Line 419 – the formatting needs fixing in this line
Author Response
Dear Reviewer,
Thank you so much for your comments.
Please find below our responses point-by-point:
1) « I noted there was no reference in the text to any of the figures and only one in text reference to a table (Table 6). I presume these references should be where where the referencing errors but I strongly encourage the authors to check this accordingly. The tables and figures should be mentioned in the text otherwise they should not be included in the manuscript. I have not cross-checked the referencing in the text against the references »
Thank you so much for this remark.
All references have been carefully checked and inserted correctly in the revised manuscript. Please accept our apologize for the errors in the previous version that we didn’t sincerely understand.
2) « I find the manuscript needs a large number of minor edits to improve the use of language. My recommendations in this regard are listed in the specific comments below. I would strenuously recommend these specific comments be given attention by the authors and the edits made accordingly. »
Thank you so much for your time and recommendation.
All suggestions have been taken into account, and the manuscript has been corrected in this way.
Reviewer 2 Report
The review by Rabetafika et al. recapitulates the applications of probiotics in human and animal health keeping also the focus on the problem of antibiotic resistance emergence. Although the topic is of high interest, I find that the approach in some parts of the manuscript can be misleading in considering probiotics as a plain replacement alternative to antibiotic treatment and should be removed. As an example of this:
1- Section 2.3: Alternatives to Antibiotics. This section is supported by Figure 2. Along the section and Figure, the "modulation of gut microbiota" is mentioned. The way it is mentioned is somehow confusing since the modulation of gut microbiota is not an indication for antibiotic use (at least in most general cases) and consequently the use of probiotics to this aim is not really to replace antibiotic use. Oppositely, in the same section, other true replacement alternatives to antibiotic are mentioned such as phage therapy.
2-Section 4 probiotics vs. antibiotics pros and cons is written in the same way, as comparing two options for the same problem, whereas in most of the cases they can complement (eg. use of probiotic to reduce antibiotic caused dysbiosis)
Besides this major concern, I also suggest that the topics of antibiotic use in human health and animal production should be treated separately. In human health there are many situations where antibiotic use is the first choice and probiotics can complement (eg. as mentioned antibiotic used dysbiosis). There are other situations where there is no clear/controversial indication of antibiotic use, however probiotic may constitute an option (depending on trials, periodontal disease, acne, IBDs, recurrent infections with H. pylori or bacterial vaginosis) and there are other situations where antibiotic use is non indicated and probiotics appear as an appropriate option (C. dificcile associated diarrhea, acute diarrhea). In the present version all these situations are shown at the same level which can be also confusing.
On the other hand, in animal production the use of antibiotic as growth promoters is clearly an off label use that is forbidden and also an important cause of AMR propagation. In these cases, where there is not an specific infection to treat but health promotion/development is aimed, the use of probiotics may be and important option. Consequently I suggest to clearly differentiate the topic of antibiotics/probiotics in human health vs. animal production that in the present version are shown as almost parallel situations.
Finally, I suggest to eliminate all in vitro evidence from Table 6, since this is not comprehesive. I would suggest to focus on in vivo evidence besides clinical trials.
Minor changes:
- in the present version in many parts there is an error message: "(Error! Reference source not found.)." that should be removed
- line 196: "topathogenic"... should be replaced "to pathogenic".
Author Response
Dear Reviewer,
Thank you so much for your comments and suggestions, for which we address our sincere acknowledgment.
Please find below point-by-point our responses and revisions accordingly:
« Although the topic is of high interest, I find that the approach in some parts of the manuscript can be misleading in considering probiotics as a plain replacement alternative to antibiotic treatment and should be removed. As an example of this: »
1) Section 2.3: Alternatives to Antibiotics. This section is supported by Figure 2. Along the section and Figure, the "modulation of gut microbiota" is mentioned. The way it is mentioned is somehow confusing since the modulation of gut microbiota is not an indication for antibiotic use (at least in most general cases) and consequently the use of probiotics to this aim is not really to replace antibiotic use. Oppositely, in the same section, other true replacement alternatives to antibiotic are mentioned such as phage therapy.
Thank you so much for this comment.
The section 2.3 has been revised accordingly by replacing ‘‘Modulation Gut Microbiota’’ to ‘’Disease prevention’’ beside “Disease treatment” and distinguishing molecular and microbial action mechanisms while adding further examples in the new version of Figure 2. Moreover, additional explanations have been introduced in this section to precise the principal role of some potential antibiotic alternatives, including probiotics, their classification according to their functions with direct (e.g. inhibition, lysis) or indirect actions (e.g., gut microbiota and immune system modulation). The changes have been made from lines 149 to 167 in the revised manuscript, as follows:
“These alternatives aim at reducing in maximum the inappropriate and excessive use of antibiotics,… …Two alternative groups are distinguished according to their functions: (i) disease prevention through gut microbiota and immune system modulation (e.g. probiotics), and immune stimulation (e.g. vaccines), and (ii) disease treatment by reducing or suppressing bacterial infections (e.g. phage therapy, bacteriocins, nanoparticles, antibodies, and quorum sensing anti-virulence inhibitors) [50].”
We hope that this new version is appropriate for avoiding some confusing situation.
2) Section 4: Probiotics vs. antibiotics pros and cons is written in the same way, as comparing two options for the same problem, whereas in most of the cases they can complement (e.g. use of probiotic to reduce antibiotic caused dysbiosis).
Thank you so much for this comment and suggestions.
The section 4 has also revised accordingly by changing the title to a more appropriate one, as follows: “Advantages and disadvantages of probiotics as antibiotic alternatives”.
Moreover, the following phrase has been added from lines 380 to 382 to specify the complementary role of probiotics to reduce dysbiosis caused by antibiotics: “In addition, probiotics help to repopulate the gut with healthy microbiota and reduce dysbiosis caused by antibiotics. In this situation, probiotics can compensate for the antibiotic side effects”.
An example of probiotic disadvantage has been specified at lines 394-396, as follows:
“Among the probiotics’ disadvantages are their sensitivity under extreme stress conditions (e.g., temperature, acidity, moisture, etc.), which reduce their survival rate…”
3) Besides this major concern, I also suggest that the topics of antibiotic use in human health and animal production should be treated separately. In human health there are many situations where antibiotic use is the first choice and probiotics can complement (e.g. as mentioned antibiotic used dysbiosis). There are other situations where there is no clear/controversial indication of antibiotic use, however probiotic may constitute an option (depending on trials, periodontal disease, acne, IBDs, recurrent infections with H. pylori or bacterial vaginosis) and there are other situations where antibiotic use is non indicated and probiotics appear as an appropriate option (C. dificcile associated diarrhea, acute diarrhea). In the present version all these situations are shown at the same level which can be also confusing.
Thank you so much for these relevant suggestions.
In the section “Human applications”, the 3 cases of probiotic use, as support, possible and real option for antibiotic alternatives have been specified with some examples and cited references to avoid the confusing situations (Lines 409-418), as follows:
“However, it is important to distinguish different scenarios where probiotics are used for supporting antibiotics, or as really substitute options. For many situations in human health, the use of antibiotics remains the first choice in controlling bacterial infections, and probiotics are useful for repopulating the gut microbiota [115]. Other situations indicate no clear or controversial use of antibiotics, whereas the use of probiotics may constitute an alternative like in the cases of periodontal disease, acne, recurrent infections with Helicobacter pilori, and bacterial vaginosis [116–119]. Finally, there are other situations for which the antibiotic use is non indicated and probiotics appear as an appropriate option, such as in the case of acute and Clostridium difficile associated diarrhea [120,121]”.
4) On the other hand, in animal production the use of antibiotic as growth promoters is clearly an off label use that is forbidden and also an important cause of AMR propagation. In these cases, where there is not an specific infection to treat but health promotion/development is aimed, the use of probiotics may be and important option. Consequently I suggest to clearly differentiate the topic of antibiotics/probiotics in human health vs. animal production that in the present version are shown as almost parallel situations.
Thank you so much for these comment and suggestions.
The section “Animal applications” has been revised accordingly by 1) distinguishing the probiotic use as a real important option to substitute antibiotics for the animal growth and production, or as a potential alternative for disease controls and management; 2) adding a comprehensive summarizing figure (Fig. 4). The following changes have been made for this section at lines 426 - 439:
“Antibiotics are often used in animal farming as antimicrobial agents for enhancing animal growth and production, and controlling diseases [138]. An evident use of probiotics instead to antibiotics is supported in the case of promoting animal growth for which the goal consists of health development without specific infection targets [139,140]. A considerable number of probiotic strains have also the capacity to inhibit various animal pathogens, and may be potentially used as antibiotic alternatives in farming sectors of poultry, swine, cattle, and others for enhancing immune function, and disease prevention [10]. The benefits and inputs from probiotics as alternatives to antibiotics in animal health are outlined in Figure 4. Table 8 illustrates some recent examples of animals feed supplemented with lactic acid and soil-based bacteria, the form of administration, and the probiotic strain effects. The use of probiotics as feed supplements in animal farming allows, not only the reduction of AMR apparition, due to the excessive use of antibiotics, but also the diminution of the residue transfer risk to the animal products such as eggs, milks, and meats.”
5) Finally, I suggest to eliminate all in vitro evidence from Table 6, since this is not comprehensive. I would suggest to focus on in vivo evidence besides clinical trials.
All in vitro examples have been removed from Table 6.
6) Minor changes:
- in the present version in many parts there is an error message: "(Error! Reference source not found.)." that should be removed.
This general issue has been corrected.
- line 196: "topathogenic"... should be replaced "to pathogenic".
This has been corrected.
Reviewer 3 Report
I appreciate the effort carried out by the authors to provide as much information as possible regarding the potential use of probiotics as antibiotic alternatives with examples applications in human and animal health protection.
AMR is currently one of the most important research issues, but the review of the potential use of probiotics as alternatives has not been highlighted thoroughly. Consequently, I consider that the present paper is dealing with a very modern and sufficiently interesting issue for the readers.
The manuscript does well in dealing with the subject and all Tables and Figures are well organized and clear for the reader. The authors do also well in presenting the conclusions of the study.
However, a lot of references are missing as indicated in the text, for example in lines 92, 128-129, 133, 144, 166, 250-251, 352, 371-372, 384 etc. Please comment.
Author Response
Dear Reviewer,
Thank you so much for your comments.
Please find below our responses and explanation to your questions:
« A lot of references are missing as indicated in the text, for example, in lines 92, 128-129, 133, 144, 166, 250-251, 352, 371-372, 384 etc. Please comment »
We appreciate your remark on the missing references.
Please accept our apology for this issue. This was possibly due to the incorrect use of reference styles within the template.
All references in the present version have been carefully checked.
Round 2
Reviewer 1 Report
Dear Authors,
After reviewing the revised manuscript I am of the opinion that the issues raised in my first report have adequately addressed.
Thank you for taking the time to make the changes to the manuscript.
Reviewer 2 Report
All comments and concerns on previous version have been properly addressed
Reviewer 3 Report
I would like to thank the authors for revising the manuscript according to comments.
I suggest that the paper should accepted in its present form.
