Description of a Novel Fish Pathogen, Plesiomonas shigelloides subsp. oncorhynchi, Isolated from Rainbow Trout (Oncorhynchus mykiss): First Genome Analysis and Comparative Genomics

Round 1

Reviewer 1 Report

Authors wrote an interesting manuscript dealing with an actual concern in aquaculture, with a specific address to public health.

The manuscript is well written, and Authors expose clearly the concepts.

Hereby, some minor flaws:

-          L57 -58: correct the number of the references and write them according to the style of the journal.

-          L58: please, provide a short explanation about “post-natural disaster infections”

-          I encourage to underline the differences form the previous study (ref. 21) àdifferent method, more specific protocol, enrollment of different species…

-          L163/164: May be the web sites could be removed from here and placed only in the references list? This is a question for Editors, also.

-          L190: I would change “inoculated” with “seeded” (it is more appropriate here)

-          Paragraph 2.5: why Authors chose these molecules? A lot of them could not be used to treat fishes according to the current European Legislation. Lease, state the reason of this selection (experimental I think… but please provide a reason and then discuss a bit this point)

-          L321: not “against” but “testing”.

-          L322: A. salmonicida is not listed as control strain in M&M section. Please, provide a correction.

-          Paragraph 3.6: it is true that all these drugs are not listed in EUCAST (actually Plesiomonas shigelloides is not listed… so probably authors considered Enterococcus spp??) but these data are very important. So please, implement a little bit more this part. Moreover, it is necessary to also implement the explanation of table 4 (and please, provide the unit of the inhibition zone). Moreover, update the access date because it is really far from now.

-          L375/378/385/386/387410/411/428/438/442/444/447/457/462: names of the bacteria in italics please

-          L433: why capital letter for trimethoprim/sulfamethoxazole and not for ciprofloxacin? ? please, check and uniform the style.

Author Response

Dear Reviewer,

Thank you for your valuable evaluation. We have completed the manuscript edition and all suggestions were edited according to your comments. In addition, we also provided additional information based on the questions. The manuscript file includes changes (tracking change format) and a clean file.

Author Response File: Author Response.docx

Reviewer 2 Report

General comments

The present article is an interesting study. The wording of the introduction section needs to be improved. The analytical part of the methodology is standard and well described in general, but it would have been desirable to understand the phenotypic pathogenicity of V78 directly in fish by performing an in vivo challenge assay. The results should be shown in a simpler and more summarized way, trying not to repeat in the text what the reader sees in the graphs.

Lastly, it would be essential for the authors to confirm the biological guidelines necessary to taxonomically classify new bacterial species or subspecies: International Journal of Systematic and Evolutionary Microbiology. Published by the Microbiology Society and owned by the International Committee on Systematics of Prokaryotes (ICSP), a committee of the Bacteriology and Applied Microbiology Division of the International Union of Microbiological Societies, International Journal of Systematic and Evolutionary Microbiology is the leading forum for the publication of novel microbial taxa and the ICSP’s official journal of record for prokaryotic names.

Specific comments

L46-L52: Considering the more direct and concrete description, with adequate references, detailed between L53 and L69, I consider that these lines are superfluous (they repeat the idea).

L46-L48: You are talking about human clinical findings in this statement, right?

L54: It should indicate the scientific names of the fish species, especially that of rainbow trout because it is the first time it is mentioned after the abstract.

L54-L63: I suggest revising the wording. In general, they are repetitive ideas that could be better expressed (more concretely).

L62-L67: I suggest using "farmed" instead "cultured" 

L70-L88: It is a bit heavy to read this paragraph. I suggest being more concise.

L91-L93: I suggest including a line regarding what the previous study is about and briefly state what the difference is in what the authors are now describing.

L95: You talk about "pathogenicity" of a bacterium isolated from sick rainbow trout raised in Turkey that experienced outbreaks presumably caused by Plesiomonas. Why didn't you confirm Koch's postulates and the level of pathogenicity by an experimental challenge in vivo? Could this information be an important input for this genomic study?

L193: It would have been interesting to know the phenotypic pathogenicity level of the isolate by in vivo experimental challenge in naïve trout.

L319: It would have been interesting to know the minimum inhibitory concentration (MIC) of V78 to the antibiotics for which it showed susceptibility in the antibiogram.




L204-L344: I suggest finding a better and simpler way to display the results, perhaps using a comparative table between the groups and describing the innate, humoral and cellular immune response genes that were observed to be overexpressed or downregulated.

L363-L364: Wording should be improved. In general, the discussion is very light, it needs a more analytical than descriptive focus especially on the only novelty of what is being reported: the RNA-seq results in vaccinated fish. It does not take advantage of very important data such as salmon strain, sex, time of challenge, etc.

Tables and figures: Perhaps the authors could be a bit more informative in describing each one.

Author Response

Dear Reviewer,

Thank you for your valuable evaluation. We have completed the manuscript edition and all suggestions were edited according to your comments. In addition, we also provided additional information based on the questions. The manuscript file includes changes (tracking change format) and a clean file.

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Thanks to the authors for considering suggestions. The manuscript has been significantly improved. I would just like suggest you please to include in the discussion some lines regarding the actions that will follow with regard to the formal IJSEM guidelines and to project the work that should be done at phenotypic and clinical level with experimental challenges in naive rainbow trout. You could discuss about that. 

Author Response

Dear Author,

We appreciate your contributions. Your suggestions improve manuscript quality and impact in addition to validation of our strains to be novel species. According to the comments, we have evaluated International Committee on Systematics of Prokaryotes and the Bacteriology and Applied Microbiology Division of the International Union of Microbiological Societies guidelines for validating a novel bacteria species. Similar to the reviewer's suggestions, we added a phylogenetic tree 16S rRNA gene-based for both isolates, 16S rRNA sequence accessions, ANI and dDDH values, isolation, habitat and sample description, morphology and growth conditions, physiology, functional and ecological analysis and species descriptions in the manuscript. We also discussed our results to be a novel bacteria species in the discussion part and added a suggestion for determining the pathological effects of the presented species by experimental analysis. The manuscript has been edited in English language and style.

We would like to present our editions in lines 411-416 and 453-463 in the discussion part and in lines 553-561 in the description of novel species.

We hope that our revisions meet your suggestions.

On behalf of the Authors

Best wishes

Author Response File: Author Response.docx