Review Reports - Effects of Single or Combined Administration of Dietary Synbiotic and Sodium Propionate on Humoral Immunity and Oxidative Defense, Digestive Enzymes and Growth Performances of African Cichlid (<i>Labidochromis lividus</i>) Challenged with <i>Aeromonas hydrophila</i>

Round 1

Reviewer 1 Report

The manuscript evaluated the single and combined supplementation of SYN and SP. The paper is interesting, but it needs revision before further processing.

Abstract: where is the aim of the study? What are the biological responses assessed? Where are the design and methods of the experiment?
The authors just said the supplemented groups are better than the control. I see many groups which group gives the best result in each parameter measured?
Keywords are too much. Only mention the most important and significant words and don’t repeat the words in the title.
All Latin names are italic, please correct the whole manuscript.
L51: Synbiotic
Table 1: please add the corresponding indication of the number 4 in the footnote. No. 3 in the footnote is repeated.
The fat in the diet is too high. According to what the authors formulate the diets. Please insert the reference.
How was the feeding done? How were fish fed ad libitum?
Please insert a reference for water parameters.
L110: what are the health indicators measured?
L128: add intake after crude protein
Tables: please transfer the definition of the groups from the title to the footnote. If there is no significance of a parameter between groups, don’t put superscripts on these data. Put superscripts only on the significant data.
It is better to obtain the difference between mean by Tukey’s test instead of Duncan and change the significance accordingly.
Results: don’t restrict on saying the supplemented groups are better than the control. Please mention the best groups as there is significance between groups.
Please revise the results in the text with their corresponding in the tables as there are some errors. For example, the authors said, “the alternative hemolytic complement activity (ACH50) increased significantly in fish fed on the supplemented diets compared to the control” while the opposite is present in the figure.
Discussion should be improved. The order of the parameters in the discussion should be the same in the result section. Provide interpretation to each result and focus on the most significant results. Please don’t repeat the results again.
L478: correct the number of days
L482: Did the authors calculate the economic efficiency?

Author Response

Response to reviewers on Fishes-1453964

Manuscript Title: Effects of single or combined administration of dietary synbiotic and sodium propionate on humoral immunity and oxidative defense, digestive enzymes and growth performances of African cichlid (Labidochromis lividus) challenged with Aeromonas hydrophila

The authors gratefully thank to the editor and referees for their constructive comments and recommendations which definitely help to improve the readability and quality of the paper. All the comments that need to be incorporated to the revised manuscript have been done and the others that only need to be clarified for the referees are explained here only. Detailed responses to the comments and recommendations are as follows. Please note that we replied to all the comments immediately below the comments.

Reviewer#1:

Dear honorable Reviewer#1: Thank you very much for your constructive recommendations. Here, we responded to your comments. Your requested changes are highlighted in green color. We hope the following descriptions are acceptable.

Comment 1: Abstract: where is the aim of the study? What are the biological responses assessed? Where are the design and methods of the experiment?

Response 1: Abstract was edited based on the comments.

Comment 2: The authors just said the supplemented groups are better than the control. I see many groups which group gives the best result in each parameter measured?

Response 2: Abstract was edited based on the comments.

Comment 3: Keywords are too much. Only mention the most important and significant words and don’t repeat the words in the title.

Response 3: Thanks of your suggestion. We revised it.

Comment 4: All Latin names are italic, please correct the whole manuscript

Response 4: Thanks of your attention. We revised it.

Comment 5: L51: Synbiotic.

Response 5: Thanks of your attention. We revised it.

Comment 6: Table 1: please add the corresponding indication of the number 4 in the footnote. No. 3 in the footnote is repeated.

Response 6: Thanks of your attention. We revised it.

Comment 7: The fat in the diet is too high. According to what the authors formulate the diets. Please insert the reference.

Response 7: Thanks for your kind comments. As you know, the test fish is regarded as a carnivorous fish. The test diets were produced with a twin-screw extruder. The nutrient requirements of ornamental fish especially African cichlid (Labidochromis lividus) did not actually determine. Based on the experience and laboratory observations, the reported diet formulation is highly acceptable with African cichlid (Labidochromis lividus).

Sarkheil, M., Ameri, M., Safari, O., (2021). Application of alginate-immobilized microalgae beads as biosorbent for removal of total ammonia and phosphorus from water of African cichlid (Labidochromis lividus) recirculating aquaculture system. Environmental Science and Pollution Research. doi: 10.1007/s11356-021-16564-w

Comment 8: How was the feeding done? How were fish fed ad libitum?

Response 8: The test fish was fed manually to satiety.

Comment 9: Please insert a reference for water parameters.

Response 9: Thanks of your kind attention. We added it. In the present study, the cultured fish was an ornamental fish belong to Cichlidae family. African cichlids require a warmer environment, with an ideal range of 23-28°C. Water temperature adjusted for culture of different species of Cichlidae was 23-28°C in different studies as below:

Erdogan, F., Erdogan, M., Gümüş, E. 2012. Effects of Dietary Protein and Lipid Levels on Growth Performances of Two African Cichlids (Pseudotropheus socolofi and Haplochromis ahli). Turkish Journal of Fisheries and Aquatic Sciences 12: 635-640.

Ribbink, A.J. The Behaviour of Hemihaplochromis Philander, A. South African Cichlid Fish. Zoologica Africana. 6 (2): 263-288 (1971).

https://www.petwave.com.au/assets/files/PetWave-African-Cichlid-Care-Sheet.pdf

https://ornamentalfish.org/wp-content/uploads/2013/07/30-African-Malawi-Cichlids.pdf

Comment 10: L110: what are the health indicators measured?

Response 10: Thanks of your kind attention. We purchased five hundred fish from a local ornamental fish supplier, transferred them to four glass aquariums (200 L), and checked appearance characteristics of fish for two weeks. We did not measure health indicators of fish.

Comment 11: L128: add intake after crude protein

Response 11: Thanks of your attention. We revised it.

Comment 12: Tables: please transfer the definition of the groups from the title to the footnote. If there is no significance of a parameter between groups, don’t put superscripts on these data. Put superscripts only on the significant data.

Response 12: Thanks of your attention. We revised it in the Tables.

Comment 13: It is better to obtain the difference between mean by Tukey’s test instead of Duncan and change the significance accordingly.

Response 13: Thanks for your attention. Tukey’s test was run and the results were similar to Duncan’s multiple range test.

Comment 14: Results: don’t restrict on saying the supplemented groups are better than the control. Please mention the best groups as there is significance between groups.

Response 14: Thanks of your suggestion. We added the highest and lowest values of different parameters between dietary groups that had significant differences (result section).

Comment 15: Please revise the results in the text with their corresponding in the tables as there are some errors. For example, the authors said, “the alternative hemolytic complement activity (ACH50) increased significantly in fish fed on the supplemented diets compared to the control” while the opposite is present in the figure.

Response 15: Thanks of your attention. We revised it.

Comment 16: Discussion should be improved. The order of the parameters in the discussion should be the same in the result section. Provide interpretation to each result and focus on the most significant results. Please don’t repeat the results again.

Response 16: Thanks for your kind attention. Discussion was re-written.

Comment 17: L478: correct the number of days.

Response 17: Thanks of your attention. We revised it.

Comment 18: L482: Did the authors calculate the economic?

Response 18: Thanks for your kind attention. The aim of present study is to compare the singular and combination effects of SYN and SP on some biological indices of test fish. We would like to focus on the possibility of using these compounds. The economical indices were not measured.

Author Response File: Author Response.docx

Reviewer 2 Report

The paper investigates n aquaculture area that is of interest worldwide, which is improving fish health and consequently increasing production. The MS is well written The objectives are very clear and concise, the methodology is clearly written so that it can be reproduced and the discussion clarifies the results very well.

There are few points to clarify:
- Throughout the text the Latin names of the species or most of them are without italics form.
- Was the concentration of bacteria in the feed investigated? and the SP component? That is, did the microbiological analysis of the feed to confirm that what had been added really was what the fish were eating?

Author Response

Response to reviewers on Fishes-1453964

Reviewer#2

Dear honorable Reviewer#2: Thank you very much for your constructive comments. We tried to amend the MS according to your comments. Your requested changes are highlighted in blue color.

Comment 19: throughout the text, the Latin names of the species or most of them are without italics form.

Response 19: Thanks of your attention. We revised it in the manuscript.

Comment 20: Was the concentration of bacteria in the feed investigated? and the SP component? That is, did the microbiological analysis of the feed to confirm that what had been added really was what the fish were eating?

Response 20: Thanks of your attention. We run a pre-test before the main study. The microbiological analysis of Bactocell^® (Lallemand Inc., Montreal, QC, Canada; 7.59 log CFU g^-1) was checked in ACECR. It was 7.59±0.15 log CFU g^-1. So, we decided to report the microbiological activity of company and confirmed it. But, the SP concentration was not measured because of some technical problems.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The manuscript can be accepted in its current form.