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Dear authors,
After confirming that you have address the revisions and corrections made by the reviewers, specifically to tone down the conclusions of the work since the findings are relevant but here may be other causes for the degeneration observed, I find the manuscript suitable for publication.
Also, I am communicating to the PeerJ staff your request for the email modification of one of the authors.
Thank you for choosing PeerJ for your manuscript.
All the best for your research moving forward.
Best regards,
Bernardo
[# PeerJ Staff Note - this decision was reviewed and approved by Konstantinos Kormas, a PeerJ Section Editor covering this Section #]
Dear authors,
Thank you for assessing the comments and concerns rise by the reviewers. The two experts that rise a number of concerns agree that the manuscript has now address most of them. The Reviewer 3 indicates that the manuscript needs few changes and please address the issue indicated in line 329 of the manuscript. I kindly request that the authors revise these issues and submit the manuscript with a rebuttal letter.
All the best for your research moving forward.
Best regards,
Bernardo
The authors addressed my previous concerns. Still, there is no solid link between degeneration and MAT type. However, the study addresses a significant subject.
The authors addressed my previous concerns. Still, there is no solid link between degeneration and MAT type. However, the study addresses a significant subject.
The authors addressed my previous concerns. Still, there is no solid link between degeneration and MAT type. However, the study addresses a significant subject.
The authors addressed my previous concerns. Still, there is no solid link between degeneration and MAT type. However, the study addresses a significant subject.
After carefully reviewing the changes made by the authors I’m confident that the revised version of this manuscript is of sufficient quality to be published with some minor corrections.
The authors have addressed all of my concerns regarding the intro, background, and literature cited, and incorporating those works into their now more robust discussion section.
They have also included a more thorough and detailed explanation of the structure of Cordyceps militaris mating-type locus, which aids on the interpretation of their results. Just as a minor correction, on line 78 of their corrected manuscript it reads “MAT1-1 1-3 gene” and it should be “MAT1-1-3 gene”.
The authors now present the sequences of the primers used as well the relevant information of the original sequences used to design the mentioned primers, allowing for reproduction of their results. Figure 6 has been corrected as suggested as well.
My only final suggestion to the authors would be to toned down their conclusions about the imbalance of mating-type genes. On line 329 they state “An imbalance or loss of mating-type genes has been identified as the primary cause of rapid degeneration in Cordyceps militaris strains during subculture”. Although the authors results show that this imbalance is very important for the rapid degeneration, since the authors did not test any other of the other possible causes (e.g., genetic mutations, methylation levels, etc), the original conclusion should be adjusted to something like “An imbalance or loss of mating-type genes has been identified as one of the primary causes of rapid degeneration in Cordyceps militaris strains during subculture”.
I want to highlight the importance of the presented work. As the authors state on their response letter and elsewhere on their manuscript, the purpose of this paper was to solve the problem of strain degeneration during subculture and their results are very useful on this regard. I believe this work provides important and applicable knowledge to a specific problem.
Minor typographical mistakes to correct:
Line 130: Remove the dash at the end of the line.
Line 304: It should be “subculture”.
Line 312: It should be “and has thus...”.
Line 321: the word “influence” is repeated.
Line 332: It should be “strains containing both mating types and insensitive to...” (remove “are”).
No further comments. The authors have addressed all of my concerns and suggestions.
No further comments. The authors have addressed all of my concerns and suggestions.
No further comments. The authors have addressed all of my concerns and suggestions.
Dear authors,
Three experts in the field have reviewed your manuscript and two have raised important issues. Reviewer 1 indicates that the authors should clarify the findings and provide clear experimental evidence of the mating type. Reviewer 3 indicates several aspects of the literature cited and methodological issues that need extensive revisions. Please address all the issues indicated by the reviewers for a second round of revisions.
All the best with your research moving forward.
Bernardo
**PeerJ Staff Note:** Please ensure that all review, editorial, and staff comments are addressed in a response letter and that any edits or clarifications mentioned in the letter are also inserted into the revised manuscript where appropriate.
**Language Note:** PeerJ staff have identified that the English language needs to be improved. When you prepare your next revision, please either (i) have a colleague who is proficient in English and familiar with the subject matter review your manuscript, or (ii) contact a professional editing service to review your manuscript. PeerJ can provide language editing services - you can contact us at [email protected] for pricing (be sure to provide your manuscript number and title). – PeerJ Staff
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The manuscript addresses a relevant subject that deserves attention, research, and data communication. Despite the appealing subject, I found the manuscript content in a preliminary stage that does not provide results to support the main conclusions, nor move forward our current knowledge on this subject.
One of the main problems of the manuscript is the fact that the authors jump from conidia to spores as synonyms and they are not. Conidia are generated by asexual reproduction, while spores come from sexual events. So, it is not clear whether the different generations/strains analyzed here are coming from asexual or sexual reproduction. If the latter, evidence of this should be included. The second main flaw in the experimental design is that the analysis of MAT type and fungal degeneration during subculturing are two disconnected observations that seem to be coincidental more than related ones. If sex reproduction or MAT types have a role in delaying fungal degeneration, then hypotheses should be generated and experimentally addressed. Regrettably, this is not the case and as a consequence, the main conclusion is not supported by facts. As a minor issue, the sequence of the ITS regions should be deposited in a repository, and the MAT types should be also sequenced and deposited.
The manuscript is well-structured, uses clear and professional English, and adequately references literature to contextualize the study within the current understanding of Cordyceps militaris cultivation challenges. Figures and tables are relevant, high quality, well-labelled, and described, providing essential support to the findings.
The research question is well defined, relevant, and fills an identified knowledge gap regarding the degeneration of Cordyceps militaris strains during subculture. The experimental design is rigorous, with methods described in sufficient detail for replication. The study represents original primary research within the scope of the journal.
Data appears robust, statistically sound, and supports the conclusions. The manuscript links its findings back to the original research question effectively, stating clearly how the imbalance or loss of mating-type genes contributes to strain degeneration. However, the impact and novelty of these findings are not directly assessed within the manuscript, and meaningful replication to confirm these results would be encouraged.
1. A more in-depth discussion on the potential molecular mechanisms underlying the observed effects of mating type ratio imbalance on strain degeneration could enrich the study.
2.Considering environmental factors or genetic drift in future studies could provide a more comprehensive understanding of the strain degeneration process.
3. Additional replication of the experiments with a broader range of strains might strengthen the conclusions drawn.
Reliable production of fruiting bodies of Cordyceps militaris is of great importance due to its numerous uses as a food supplement and medicinal properties, mainly in Asian countries. Usual systems of production relay in subculturing of selected strains with particular characteristics of interest. Unfortunately, after successive subculturing steps most strains suffer a degenerative process that ultimately greatly reduce the production of fruiting bodies. There are many hypothesized mechanisms or factors that are involved in such degeneration during the subculturing process. One of these factors could be the imbalance of the mating type locus on the mycelia during growth. The authors of this paper set to investigate the effect of different rates of mating type combinations as a possible solution to the obtain a more stable and reliable method of production.
The manuscript is, in general, written in professional English and is structured in a logical manner that is easy to follow.
Nevertheless, it is my opinion that the manuscript is lacking in some aspects, mainly on the rigor of its intro, background, and literature cited, as well as some key methodological information.
It is my opinion that the literature cited is too biased and ignores some very important publications needed to contextualize the proposed projected and in turn, this oversight from the authors reduces the effectivity of their intro and background. A key aspect of the proposed manuscript is based on the mating type locus of Cordycpes militaris, but the authors neglect to explain in detail the composition of this locus (made up of three main genes and not just two as they show) and its heterothallic nature also failing to provide important references such as the one describing the MAT locus in the sequenced strain of C. militaris (Zheng et al., 2011 10.1186/gb-2011-12-11-r116) or the one by Yokoyama et al., 2003 10.1128/AEM.69.8.5019-5022.2003. There also exists a recent previous work that looks too into the degeneration due to subculturing and the effect of mating type imbalance that the authors omit in their literature (Vu et al., 2023 10.3390/jof9100971). As a key point of their work, the determination of the mating types is done through a PCR test but their findings are not reproducible since the authors do not present the sequences of the used primers and only refer to a thesis that is not accessible. In line 118 they say that “MAT1-1F, MAT1-1R, MAT1-2F, and MAT1-2R genes were amplified” but those are not the genes but the names of the primers used.
The authors also fail to use the correct terminology for the presence of one or two mating types. These should be referred as homokarion and heterokarion, respectively, and not to be confused with homothallic and heterothallic.
Finally, in lines 85-87 the authors state: “This study investigated a single mating-type strain of Cordyceps militaris, focusing on subculture impacts, mating-type combinations, and fruiting to determine the primary causes of strain degeneration.” Although their results show the effects of the mating types mixes, these do not determine conclusively “the primary causes of strain degeneration”.
As stated above, the methodology is not reproducible since the authors fail to disclose the sequences of their PCR primers.
In figure 6 authors should use the same scale so it is possible to compare across the different graphs. Even with the statistical tests I'm not sure that their conclusions are actually correct since the standard deviations seem to greatly vary. I encourage the authors to use a different sets of statistical tests to corroborate their findings.
Same as the experimental design.
Also, as stated above, their findings do not sustain their stated main objective of determining the primary causes of strain degeneration.
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