Reviewer ’s report  
 
Title:  Effect of administration of a probiotic preparation on gut microbiota and immune response 
in healthy women in India  
 
Version: 1 Date: 04 Jan 2018 
 
Reviewer:  Alejandra Moreno  LeBlanc  
 
Reviewer’s report:  
 
Please include all comments for the authors in this box rather than uploading your report as an 
attachment. Please only upload as attachments annotat ed versions of manuscripts, graphs, 
supporting materials or other aspects of your report which cannot be included in a text format.  
 
Please overwrite this text when adding your comments to the authors.  
 
The article entitle "Effect of administration of a pro biotic preparation on gut microbiota and 
immune response in healthy volunteers" is an interesting work performed with 14 healthy 
women in a determined region of India. There is not the first work that studies the effect of 
probiotics on intestinal microbio ta and immune response; however, different results have been 
reported by different groups using different probiotics. These differences can be explained 
because it is known that probiotic effect is specific of strain, so the same effect could be not 
report ed for different microorganisms. Similarly, the population under study is another variable, 
as the authors discussed in the article.  
 
I have some specific comments that can help to improve the manuscript and I will explain in 
each section.  
 
Title: The title  should include more specification about the population under analysis, such as 
country (India) or women, or both.  
 
 
Abstract.  
 
1) Methods: There are other methodologies and samples studied that are not included in the 
abstract but they are in the results.  
 
2) Results: Line 31 -33, if the authors did not explain that the studies were performed in 
blood samples, they should clarify in this section the sample in which the results were 
obtained.  
 
3) Conclusions: I don't agree with the conclusion wrote by the autho rs. Their findings were 
obtained in healthy women. In my opinion a conclusion similar to one written at the end 
of the manuscript is according to the results obtained. The suggestions are not the 
conclusions obtained. It is important to consider the comple xity of each pathology and 
also the difference between probiotics or probiotic mixes that can be administrated. It is 
not correct to make a suggestion about the effect of probiotics on pathologies with an 
study with healthy volunteers, even more when there  are works that show, in certain 
pathologies, changes in the composition of the intestinal microbiota when certain 
probiotics are administered. The conclusion also should be for the probiotic mix under 
study and not as a conclusion for all probiotics  
 
 
Main text 
 
2.Methods:  
 
2.1. Subjects. Why did the authors not include a placebo group like it is usually included in this 
kind of study?  
 
2.3.2. Measurement of plasma cytokine levels. (Lines 2 to 5): Even when the authors analysed 
each cytokine as representative f or determined cell populations, the analysis of plasma levels 
does not warranty that the cytokine is produced by a single cell population. In my opinion, only 
the name of the cytokines should be included in the parenthesis. Why did the authors use ELISA 
for cytokine determinations in plasma and CBA for the determinations in culture supernatants? 
Was it random or do the authors have any explanation for those choices?  
 
 
Results:  
 
3.2. Composition of gut microbiota: The results show that probiotic administration did not 
induce any change in the diversity and composition of gut microbiota. However, there are 
important variations observed in some ranges. If the authors take the information from each 
woman included in the study, the results are still no significant? The lack of variation in the data 
was uniform for all the participants?  
 
3.3. Immune responses.  I have the same question for the immunological studies, even when 
there are some significant differences, the data (considering the individual baseline) varied  
similarly in all the participants? If the authors have any comments about these results, they can 
include in the discussion.  
 
3.3.2. Cytokine levels in plasma and culture supernatants. (Lines 48 to 51): Please clarify 
between what samples it was obtained a  reduction of values for these cytokines.  
 
 
Discussion  
 
Pg 14, lines 47 to 53. If the authors think that the consumption of a mix of probiotics explain the 
lack of effect on some cytokines because "the stimulatory effect of some organisms may be 
cancelled out  by the inhibitory effect of the other constituents", did they think that is better the 
consumption of a single probiotic? How do they explain that the anti -inflammatory effect of 
VSL#3 was observed against certain pathologies? Please discuss about this.  
 
 
Conclusion:  
 
Even when this conclusion is better that the one presented in the abstract because is about the 
results obtained in the work, there are some points that should be modified. Please clarify that T 
cells were analysed in blood.  
 
Pg. 16, lines 13 to 16. The authors wrote: "...it led to a reduction in the frequency of Th17 cells 
and in the production of pro -inflammatory cytokine from monocytes"; however IL -10 was also 
reduced, not only pro -inflammatory cytokines.  
 
At the end of the conclusion, it is imp ortant include again the name of the probiotic under study 
and not conclude for all probiotics.  
 
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
No 
 
Are the conclusions drawn adequately supported by the data shown?  
If not,  please explain in your comments to the authors.  
 
No 
 
Are you able to assess any statistics in the manuscript or would you recommend an 
additional statistical review?  
If an additional statistical review is recommended, please specify what aspects require further 
assessment in your comments to the editors.  
 
I recommend additional statistical review  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
1. Have you in the past five years received reimbursements, fees, funding, or salary from an 
organisation that may in any way gain or lose financially from the publication of this 
manuscript, either n ow or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose 
financially from the publication of this manuscript, either now or in the future?  
 
3. Do you hold or are you currently applying for any patents relating  to the content of the 
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I declare that I have no competing interests' 
below. If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests  
 
I agree to the open peer review policy of the journal. I understand that my name will be included 
on my report to the authors and, if the manuscript is accepted for publication, my named report 
including any attachments I upload wi ll be posted on the website along with the authors' 
responses. I agree for my report to be made available under an Open Access Creative Commons 
CC-BY license ( http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments 
which I do not wish to be included in my named report can be included as confidential comments 
to the editors, which will not be published.  
 
I agree to the open peer review policy  of the journal  
Reviewer ’s report  
 
Title:  Effect of administration of a probiotic preparation on gut microbiota and immune response 
in healthy women in India  
 
Version: 1 Date: 04 Jan 2018 
 
Reviewer:  Alejandra Moreno  LeBlanc  
 
Reviewer’s report:  
 
Please include all comments for the authors in this box rather than uploading your report as an 
attachment. Please only upload as attachments annotat ed versions of manuscripts, graphs, 
supporting materials or other aspects of your report which cannot be included in a text format.  
 
Please overwrite this text when adding your comments to the authors.  
 
The article entitle "Effect of administration of a pro biotic preparation on gut microbiota and 
immune response in healthy volunteers" is an interesting work performed with 14 healthy 
women in a determined region of India. There is not the first work that studies the effect of 
probiotics on intestinal microbio ta and immune response; however, different results have been 
reported by different groups using different probiotics. These differences can be explained 
because it is known that probiotic effect is specific of strain, so the same effect could be not 
report ed for different microorganisms. Similarly, the population under study is another variable, 
as the authors discussed in the article.  
 
I have some specific comments that can help to improve the manuscript and I will explain in 
each section.  
 
Title: The title  should include more specification about the population under analysis, such as 
country (India) or women, or both.  
 
 
Abstract.  
 
1) Methods: There are other methodologies and samples studied that are not included in the 
abstract but they are in the results.  
 
2) Results: Line 31 -33, if the authors did not explain that the studies were performed in 
blood samples, they should clarify in this section the sample in which the results were 
obtained.  
 
3) Conclusions: I don't agree with the conclusion wrote by the autho rs. Their findings were 
obtained in healthy women. In my opinion a conclusion similar to one written at the end 
of the manuscript is according to the results obtained. The suggestions are not the 
conclusions obtained. It is important to consider the comple xity of each pathology and 
also the difference between probiotics or probiotic mixes that can be administrated. It is 
not correct to make a suggestion about the effect of probiotics on pathologies with an 
study with healthy volunteers, even more when there  are works that show, in certain 
pathologies, changes in the composition of the intestinal microbiota when certain 
probiotics are administered. The conclusion also should be for the probiotic mix under 
study and not as a conclusion for all probiotics  
 
 
Main text 
 
2.Methods:  
 
2.1. Subjects. Why did the authors not include a placebo group like it is usually included in this 
kind of study?  
 
2.3.2. Measurement of plasma cytokine levels. (Lines 2 to 5): Even when the authors analysed 
each cytokine as representative f or determined cell populations, the analysis of plasma levels 
does not warranty that the cytokine is produced by a single cell population. In my opinion, only 
the name of the cytokines should be included in the parenthesis. Why did the authors use ELISA 
for cytokine determinations in plasma and CBA for the determinations in culture supernatants? 
Was it random or do the authors have any explanation for those choices?  
 
 
Results:  
 
3.2. Composition of gut microbiota: The results show that probiotic administration did not 
induce any change in the diversity and composition of gut microbiota. However, there are 
important variations observed in some ranges. If the authors take the information from each 
woman included in the study, the results are still no significant? The lack of variation in the data 
was uniform for all the participants?  
 
3.3. Immune responses.  I have the same question for the immunological studies, even when 
there are some significant differences, the data (considering the individual baseline) varied  
similarly in all the participants? If the authors have any comments about these results, they can 
include in the discussion.  
 
3.3.2. Cytokine levels in plasma and culture supernatants. (Lines 48 to 51): Please clarify 
between what samples it was obtained a  reduction of values for these cytokines.  
 
 
Discussion  
 
Pg 14, lines 47 to 53. If the authors think that the consumption of a mix of probiotics explain the 
lack of effect on some cytokines because "the stimulatory effect of some organisms may be 
cancelled out  by the inhibitory effect of the other constituents", did they think that is better the 
consumption of a single probiotic? How do they explain that the anti -inflammatory effect of 
VSL#3 was observed against certain pathologies? Please discuss about this.  
 
 
Conclusion:  
 
Even when this conclusion is better that the one presented in the abstract because is about the 
results obtained in the work, there are some points that should be modified. Please clarify that T 
cells were analysed in blood.  
 
Pg. 16, lines 13 to 16. The authors wrote: "...it led to a reduction in the frequency of Th17 cells 
and in the production of pro -inflammatory cytokine from monocytes"; however IL -10 was also 
reduced, not only pro -inflammatory cytokines.  
 
At the end of the conclusion, it is imp ortant include again the name of the probiotic under study 
and not conclude for all probiotics.  
 
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
No 
 
Are the conclusions drawn adequately supported by the data shown?  
If not,  please explain in your comments to the authors.  
 
No 
 
Are you able to assess any statistics in the manuscript or would you recommend an 
additional statistical review?  
If an additional statistical review is recommended, please specify what aspects require further 
assessment in your comments to the editors.  
 
I recommend additional statistical review  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
1. Have you in the past five years received reimbursements, fees, funding, or salary from an 
organisation that may in any way gain or lose financially from the publication of this 
manuscript, either n ow or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose 
financially from the publication of this manuscript, either now or in the future?  
 
3. Do you hold or are you currently applying for any patents relating  to the content of the 
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I declare that I have no competing interests' 
below. If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests  
 
I agree to the open peer review policy of the journal. I understand that my name will be included 
on my report to the authors and, if the manuscript is accepted for publication, my named report 
including any attachments I upload wi ll be posted on the website along with the authors' 
responses. I agree for my report to be made available under an Open Access Creative Commons 
CC-BY license ( http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments 
which I do not wish to be included in my named report can be included as confidential comments 
to the editors, which will not be published.  
 
I agree to the open peer review policy  of the journal  
Please review this paper.
user
Reviewer ’s report  
 
Title:  Effect of administration of a probiotic preparation on gut microbiota and immune response 
in healthy women in India  
 
Version: 1 Date: 04 Jan 2018 
 
Reviewer:  Alejandra Moreno  LeBlanc  
 
Reviewer’s report:  
 
Please include all comments for the authors in this box rather than uploading your report as an 
attachment. Please only upload as attachments annotat ed versions of manuscripts, graphs, 
supporting materials or other aspects of your report which cannot be included in a text format.  
 
Please overwrite this text when adding your comments to the authors.  
 
The article entitle "Effect of administration of a pro biotic preparation on gut microbiota and 
immune response in healthy volunteers" is an interesting work performed with 14 healthy 
women in a determined region of India. There is not the first work that studies the effect of 
probiotics on intestinal microbio ta and immune response; however, different results have been 
reported by different groups using different probiotics. These differences can be explained 
because it is known that probiotic effect is specific of strain, so the same effect could be not 
report ed for different microorganisms. Similarly, the population under study is another variable, 
as the authors discussed in the article.  
 
I have some specific comments that can help to improve the manuscript and I will explain in 
each section.  
 
Title: The title  should include more specification about the population under analysis, such as 
country (India) or women, or both.  
 
 
Abstract.  
 
1) Methods: There are other methodologies and samples studied that are not included in the 
abstract but they are in the results.  
 
2) Results: Line 31 -33, if the authors did not explain that the studies were performed in 
blood samples, they should clarify in this section the sample in which the results were 
obtained.  
 
3) Conclusions: I don't agree with the conclusion wrote by the autho rs. Their findings were 
obtained in healthy women. In my opinion a conclusion similar to one written at the end 
of the manuscript is according to the results obtained. The suggestions are not the 
conclusions obtained. It is important to consider the comple xity of each pathology and 
also the difference between probiotics or probiotic mixes that can be administrated. It is 
not correct to make a suggestion about the effect of probiotics on pathologies with an 
study with healthy volunteers, even more when there  are works that show, in certain 
pathologies, changes in the composition of the intestinal microbiota when certain 
probiotics are administered. The conclusion also should be for the probiotic mix under 
study and not as a conclusion for all probiotics  
 
 
Main text 
 
2.Methods:  
 
2.1. Subjects. Why did the authors not include a placebo group like it is usually included in this 
kind of study?  
 
2.3.2. Measurement of plasma cytokine levels. (Lines 2 to 5): Even when the authors analysed 
each cytokine as representative f or determined cell populations, the analysis of plasma levels 
does not warranty that the cytokine is produced by a single cell population. In my opinion, only 
the name of the cytokines should be included in the parenthesis. Why did the authors use ELISA 
for cytokine determinations in plasma and CBA for the determinations in culture supernatants? 
Was it random or do the authors have any explanation for those choices?  
 
 
Results:  
 
3.2. Composition of gut microbiota: The results show that probiotic administration did not 
induce any change in the diversity and composition of gut microbiota. However, there are 
important variations observed in some ranges. If the authors take the information from each 
woman included in the study, the results are still no significant? The lack of variation in the data 
was uniform for all the participants?  
 
3.3. Immune responses.  I have the same question for the immunological studies, even when 
there are some significant differences, the data (considering the individual baseline) varied  
similarly in all the participants? If the authors have any comments about these results, they can 
include in the discussion.  
 
3.3.2. Cytokine levels in plasma and culture supernatants. (Lines 48 to 51): Please clarify 
between what samples it was obtained a  reduction of values for these cytokines.  
 
 
Discussion  
 
Pg 14, lines 47 to 53. If the authors think that the consumption of a mix of probiotics explain the 
lack of effect on some cytokines because "the stimulatory effect of some organisms may be 
cancelled out  by the inhibitory effect of the other constituents", did they think that is better the 
consumption of a single probiotic? How do they explain that the anti -inflammatory effect of 
VSL#3 was observed against certain pathologies? Please discuss about this.  
 
 
Conclusion:  
 
Even when this conclusion is better that the one presented in the abstract because is about the 
results obtained in the work, there are some points that should be modified. Please clarify that T 
cells were analysed in blood.  
 
Pg. 16, lines 13 to 16. The authors wrote: "...it led to a reduction in the frequency of Th17 cells 
and in the production of pro -inflammatory cytokine from monocytes"; however IL -10 was also 
reduced, not only pro -inflammatory cytokines.  
 
At the end of the conclusion, it is imp ortant include again the name of the probiotic under study 
and not conclude for all probiotics.  
 
Are the methods appropriate and well described?  
If not, please specify what is required in your comments to the authors.  
 
Yes 
 
Does the work include the necessary controls?  
If not, please specify which controls are required in your comments to the authors.  
 
No 
 
Are the conclusions drawn adequately supported by the data shown?  
If not,  please explain in your comments to the authors.  
 
No 
 
Are you able to assess any statistics in the manuscript or would you recommend an 
additional statistical review?  
If an additional statistical review is recommended, please specify what aspects require further 
assessment in your comments to the editors.  
 
I recommend additional statistical review  
 
Quality of written English  
Please indicate the quality of language in the manuscript:  
 
Acceptable  
 
Declaration of competing interests  
Please complete a declaration of competing interests, considering the following questions:  
1. Have you in the past five years received reimbursements, fees, funding, or salary from an 
organisation that may in any way gain or lose financially from the publication of this 
manuscript, either n ow or in the future?  
 
2. Do you hold any stocks or shares in an organisation that may in any way gain or lose 
financially from the publication of this manuscript, either now or in the future?  
 
3. Do you hold or are you currently applying for any patents relating  to the content of the 
manuscript?  
 
4. Have you received reimbursements, fees, funding, or salary from an organization that 
holds or has applied for patents relating to the content of the manuscript?  
 
5. Do you have any other financial competing interests?  
 
6. Do you have any non -financial competing interests in relation to this paper?  
 
If you can answer no to all of the above, write 'I declare that I have no competing interests' 
below. If your reply is yes to any, please give details below.  
 
I declare that I have no competing interests  
 
I agree to the open peer review policy of the journal. I understand that my name will be included 
on my report to the authors and, if the manuscript is accepted for publication, my named report 
including any attachments I upload wi ll be posted on the website along with the authors' 
responses. I agree for my report to be made available under an Open Access Creative Commons 
CC-BY license ( http://creativecommons.org/licenses/by/4.0/ ). I understand that any comments 
which I do not wish to be included in my named report can be included as confidential comments 
to the editors, which will not be published.  
 
I agree to the open peer review policy  of the journal  
assistant