Autochthonous Conifers of Family Pinaceae in Europe: Broad Review of Morpho-Anatomical and Phytochemical Properties of Needles and Genetic Investigations

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Authors,

The work is great and just some commenets were prepared to make it better to understand.

Comments for author File: Comments.pdf

Author Response

Response to Reviewer 1

• Reviewer 1, Abstract, Comment: 17: In this part, first the aims of the study should be defined. Why are the authors going to prepare this article?

Authors: Lines 17-22, Sentences corrected or added: The aim of this review is providing cumulative information about variability of needle mor-pho-anatomy, terpenes and n-alkanes as well as genetics of Pinaceae species, native in Europe. The first morpho-anatomical examinations of needles were conducted in 19th century. The composition and abundance of terpenes differ between genera and families but also within the same genus pointed to their taxonomic importance.

• Reviewer 1, Abstract, Comment, 19: Why did the authors directly talk about the alkanes with no introductions before? Some sentences should be stated first.

Authors: L.23-24, Sentences to be added:  n-Alkanes on the needle wax surfaces of conifers are sometimes very useful markers of species and population variability.

• Reviewer 1, Abstract, Comment, L. 20: What are the C29, C31, or C27? What are their importance?

Authors: L.24-25, Sentence to be repaired:  The most abundant n-alkanes in Abies species were nonacosane (C29), hentriacontane (C31) or heptacosane (C27), whereas ...

• Reviewer 1, Introduction, Comment: Authors just collected and presented some info about the conifers. The aim and the frame of the article is not defined. How the authors collected the information/ What were the selected articles’ features? What are the main purposes of the article? What are you looking for? And some related questions must be answered to understand the authors’ reasons to prepare this work.

Authors: L. 54-58, Sentences to be added: The aims of this study were to investigate how many Pinaceae species are native to Europe, to quote them and to collect and analyze articles in terms of their morpho-anatomy, terpenes and n-alkanes variability and their genetic properties. Based of these informations one could find unexplored species in terms of analyzed properties. According to our knowledge this type of review has not been published so far.

• Reviewer 1, 297-311, Comment: The para need to be supported by some references

 

Authors: L. 311, references in L. 320, 323 and 327-328 were added.

 

• Reviewer 1, Conclusion. This review paper provides a detailed examination of the terpene composition of European Pinaceae species. Why the authors focus on this features? This must be explained with other examined features.

Authors: L. 383-386, Sentences to be added: It is well known that examined features are very variable between species. This was confirmed in tabular displays. On the other side, one could see from tables that some species have not been examined yet in some features and this fact is very important information for further investigations.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The authors have reviewed Morpho-anatomical and phytochemical properties of needles and genetic investigations in autochthonous conifers in Europe. However, before publication, there are some major issues needed to be improved.

1. The first and most important thing is that the authors should significantly improve the Tables in the manuscript. As presented, it is very difficult for me (I think for other readers as well) to understand.

2. Lot of references have been widely listed and well summarized, but this review still lacks some valuable works, such as Dr. Xiao-Ru Wang’s group (Department of Ecology and Environmental Science, Umea University, https://www.umu.se/en/staff/xiao-ru-wang/?flik=publikationer).

3. As for molecular markers (cpDNA etc.), pay attention to taxonomy based on molecular systematics (Brief history of the gymnosperms：classification，biodiversity，phytogeography，and ecology [M]．Pretoria：South African National Biodiversity Institute，2007; DOI:  10.11646/phytotaxa.19.1.3; DOI: 10.1016/j.pld.2022.05.003; DOI:  10.1371/journal.pone.0119248).

Author Response

Response to Reviewer 2

• Reviewer 2, 1. The first and most important thing is that the authors should significantly improve the Tables in the manuscript. As presented, it is very difficult for me (I think for other readers as well) to understand.

 

Authors: We have no idea how to improve Tables. Since other three reviewers didn’t comment table formatting, we left them the same as before reviewing.

 

• Reviewer 2, 2. Lot of references have been widely listed and well summarized, but this review still lacks some valuable works, such as Dr. Xiao-Ru Wang’s group (Department of Ecology and Environmental Science, Umea University, https://www.umu.se/en/staff/xiao-ru-wang/?flik=publikationer).

 

Authors: According to Your suggestion, we added two references [271] and [272] in Table 4 as well as in Reference list, L. 1057-1058 and L. 1059-1061, respectively.

 

• Reviewer 2, 3. As for molecular markers (cpDNA etc.), pay attention to taxonomy based on molecular systematics (Brief history of the gymnosperms：classification，biodiversity，phytogeography, and ecology [M]．Pretoria：South African National Biodiversity Institute，2007; DOI:  10.11646/phytotaxa.19.1.3; DOI: 10.1016/j.pld.2022.05.003; DOI:  10.1371/journal.pone.0119248).

 

Authors: According to Your suggestion, we added one sentence in the Introduction part, L. 51-53, as well as reference 3 in Reference list (L. 397-398).

 

Reviewer 3 Report

Comments and Suggestions for Authors

 

The authors collected a large amount of material on the structure of needles, chemical components and molecular markers of European species of the Pinaceae family and presented it in a convenient compact form.

Notes:

(1) lines 17-18 "The first morpho-anatomical examinations of needles were conducted in the 1920s." This phrase should be deleted because it is incorrect. The study of the anatomy of conifers was intensively carried out already in the second half of the 19th century, many of the works were carried out carefully and contained interesting details. For example, this 1893 work is a review of the anatomy of conifers and contains references to earlier works:

  MASTERS, M. T. Notes on the Genera of Taxaceae and Coniferae. Journ. Lynn. Soc., vol. xxx(1893), pp. 1-42 [https://ia800708.us.archive.org/view_archive.php?archive=/22/items/crossref-pre-1909-scholarly-works/10.1111%252Fj.1095-8339.1877.tb02314.x.zip&file =10.1111%252Fj.1095-8339.1893.tb02386.x.pdf]

For the article being reviewed, it is not necessary to cite 19th century publications.

(2) Line 87 “the cones fall apart” In Abies, the cones grow upward; after the seeds ripen, the scales and seeds fall off, and the axis of the cone remains on the tree.

(3) Table 2. The explanations at the end of the table need to be expanded, reference to Petrakis is not enough. “0.1-1.0% (=); 1.1 – 5.0% (>); 5.1-15.0% (>>); more than 15.0% (>>>)”. What do percentages mean? Is this a percentage of the needle mass? Some components do not have such marks, which needs to be explained. What does the order in component list mean for each sample? Does the largest one go first? Then it is not clear why in Abies alba Atrificial [96] “3 –Carene =” is located before “Tricyclene >”; in Abies nordmanniana Turkey [100] “Camphene =” and “α -Chumullene =” are located before “α -Pinene >”, etc.

(4) Petrakis et al. 2001 in the Table 2 – no number in the list of references; is it [122] ?

(5) The authors do cite a lot of their publications. These articles mostly contain factual material about the chemical composition of different conifer species. In the field of studying small molecule plant substances and testing their biological activity, small publications are traditionally practiced and they are appropriate in a thematic review. The high rate of self-citation is due to the fact that the authors did not use numerous publications of other authors on the same topic. It would be correct to cite those works where a certain substance was first discovered in a given species, in order to show respect to previous researchers.

But I cannot advise authors to choose older publications, because there is a requirement from publishers to place primarily works of the last 5-10 years in the bibliography. In my opinion, this is a bad practice. The social image of taxonomists, who are required to cite priority publications starting with Carl Linnaeus, as well as plant anatomy, where some interesting data were obtained 50-100 years ago and have not lost their relevance, suffers from these formal requirements.

Author Response

Response to Reviewer 3

• Reviewer 3, (1) lines 17-18 "The first morpho-anatomical examinations of needles were conducted in the 1920s." This phrase should be deleted because it is incorrect. The study of the anatomy of conifers was intensively carried out already in the second half of the 19th century, many of the works were carried out carefully and contained interesting details. For example, this 1893 work is a review of the anatomy of conifers and contains references to earlier works:

 

MASTERS, M. T. Notes on the Genera of Taxaceae and Coniferae. Bot. Journ. Linn. Soc., vol. 30 (1893), pp. 1-42 [https://ia800708.us.archive.org/view_archive.php?archive=/22/items/crossref-pre-1909-scholarly-works/10.1111%252Fj.1095-8339.1877.tb02314.x.zip&file =10.1111%252Fj.1095-8339.1893.tb02386.x.pdf]

 

For the article being reviewed, it is not necessary to cite 19th century publications.

Authors: Lines 17-18. The sentence "The first morpho-anatomical examinations of needles were conducted in the 1920s." was corrected as follow: The first compressive morpho-anatomical examinations of needles were conducted in 19^th century [16]. We also added this reference in References, L. 421.

• Reviewer 3, (2) Line 87 “the cones fall apart” In Abies, the cones grow upward; after the seeds ripen, the scales and seeds fall off, and the axis of the cone remains on the tree.

Authors: Thank You very much for this warning, You are wright. The part of sentence “the cones fall apart” was reconstructed as “the scales and seeds fall apart” L. 101.

• Reviewer 3, (3) Table 2. The explanations at the end of the table need to be expanded, reference to Petrakis is not enough. “0.1-1.0% (=); 1.1 – 5.0% (>); 5.1-15.0% (>>); more than 15.0% (>>>)”. What do percentages mean? Is this a percentage of the needle mass?

Authors, Table 2, Footnote.  Thank You very much for this comment. New part of sentence was corrected in footnote: (Symbols refere to Petrakis et al. 2001) [120] are used to denote differences in compound amounts presented as percentages of total terpene amount:

• Reviewer 3, Some components do not have such marks, which needs to be explained.

 

Authors, Table 2, Explanation: for the last compounds symbols are not suitable, because terpene profiles end with terpene compound without symbols.

 

• Reviewer 3, What does the order in component list mean for each sample? Does the largest one go first? Then it is not clear why in Abies alba Atrificial [96] “3 –Carene =” is located before “Tricyclene >”; in Abies nordmanniana Turkey [100] “Camphene =” and “α -Chumullene =” are located before “α -Pinene >”, etc.

Authors, Table 2, Explanation: Yes, compound with the largest amount always goes first. When the symbol = is used that means that difference is between 0.1 and 1.0%.

• Reviewer 3, (4) Petrakis et al. 2001 in the Table 2 – no number in the list of references; is it [122]?

Authors, Table 2, Footnote, Answer: Thank You for detecting our mistake. Yes, it was 122, but after corrections it obtained number 120.

• Reviewer 3, (5) The authors do cite a lot of their publications. These articles mostly contain factual material about the chemical composition of different conifer species. In the field of studying small molecule plant substances and testing their biological activity, small publications are traditionally practiced and they are appropriate in a thematic review. The high rate of self-citation is due to the fact that the authors did not use numerous publications of other authors on the same topic. It would be correct to cite those works where a certain substance was first discovered in a given species, in order to show respect to previous researchers.

 

But I cannot advise authors to choose older publications, because there is a requirement from publishers to place primarily works of the last 5-10 years in the bibliography. In my opinion, this is a bad practice. The social image of taxonomists, who are required to cite priority publications starting with Carl Linnaeus, as well as plant anatomy, where some interesting data were obtained 50-100 years ago and have not lost their relevance, suffers from these formal requirements.

Authors, Answer: You are wright for all. It is also very hard to find old articles. Since we have a lot of self-citations, we must reduce their number. We deleted our 18 references and consequently, shortened table rows.

Reviewer 4 Report

Comments and Suggestions for Authors

The manuscript “Autochthonous conifers of family Pinaceae in Europe: Morpho-anatomical and phytochemical properties of needles and genetic investigations” from Nikolić and collaborators brings an importante review with extensive collection of literature data on different aspects of 23 species of Pinaceace extant in Europe.

The data collected forms a pool of very rich information to understand various aspects of the Pinaceae family, including evolutionary ones. However, the data could be explored more deeply, especially in terms of morphoanatomy, which is the most superficially discussed topic.

Please see my comments in the pdf.

Comments for author File: Comments.pdf

Author Response

Reviewer 4

Comments and Suggestions for Authors

The manuscript “Autochthonous conifers of family Pinaceae in Europe: Morpho-anatomical and phytochemical properties of needles and genetic investigations” from Nikolić and collaborators brings an importante review with extensive collection of literature data on different aspects of 23 species of Pinaceace extant in Europe.

The data collected forms a pool of very rich information to understand various aspects of the Pinaceae family, including evolutionary ones. However, the data could be explored more deeply, especially in terms of morphoanatomy, which is the most superficially discussed topic.

Please see my comments in the pdf.

Suggestions of Reviewer 4 in the manuscript

• Reviewer 4, Title,“I suggest that the title conveys the idea that this manuscript is a broad review of these aspects”.

Authors, Title of review, Answer: Thank You for Your suggestion. New title of the review is: Autochthonous conifers of family Pinaceae in Europe: Broad review of morpho-anatomical and phytochemical properties of needles and genetic investigations

• Reviewer 4, Abstract, Line 18. - And? Please, insert some more informative sentence about this just like you did for chemicals.

Authors, Answer, L.19-21.: Thank You for Your suggestion. According to comment of Reviewer 3 and 4 this sentence was repaired as follow: The first morpho-anatomical examinations of needles were conducted in the 19th century .A lot of species were investigated up to now, but population variability of many conifer species is still not known.

• Reviewer 4, Abstract, Line 23. – Pinus? Picea?

Authors, Answer, L.28.: It was corrected as Pinus.

• Reviewer 4, Abstract, Line 24. – Which were the main conclusions of these studies?

Authors, Answer, L.33-34. We presented the main conclusions in the next sentence. We added them this sentence: These investigations are of great value not only on biodiversity level, but also in the levels of ecology, physiology, taxonomy and evolution.

• Reviewer 4, Introduction, Line 40: Please, see journal guidance for this citation (Ogg et al. (2008) and cited by [1]).

Authors, Answer, Line 47. Thank You for this suggestion, but authors decided to delete reference Ogg et al. (2008), so corrected part of sentence is: According to [1] and references cited therein, the Pinaceae …...

• Reviewer 4, page 3, Line 114: “Pinus” in italic

Authors, Answer, page 3, Line 127: Thank You for notice this mistake. We corrected it as “Pinus”. We also corrected word “Picea“in L. 113, and word ”found” in L. 132.

Page 4, Line 168. According to comment of Referee 3 we had to change this sentence with another one: The first compressive morpho-anatomical investigations of needles were published in 19^th century [16].

• Reviewer 4, Page 4. Sentence from line 160-161 have to be incorporated in line 159 before the sentence beginning with: By incorporating….

 

Authors, Answer, Page 4, Lines 159-161. Thank You for suggestion. We rotated places of these two sentences, L. 172-175: By incorporating anatomical tests, it becomes possible to identify hybrids or lower taxonomic units with a relatively high level of certainty. Needle morphology is in-strumental in species identification and their classification within plant systematics [17].

 

• Reviewer 4, 4^th Caption of Table 1. I didn't understand. Cuticle is a constituent of epidermal cells. Would these values be the thickness of the epidermis as a whole (including cuticle)?

Authors, Answer, 4th Caption of Table 1. Thank You for the suggestion. It will be corrected as Epidermis thickness.

• Reviewer 4, 5^th Caption of Table 1. Please, provide the legend for this abbreviation.

 

Authors, Answer, 5^th Caption of Table 1. Thank You for the suggestion, but we decided to put whole word Hypodermis instead of Hypoder.

 

• Reviewer 4, 6^th Caption of Table 1. Do all the works cited refer to the same way of collecting this data (in all of them is it the total number of ducts or in all of them is it the density of ducts)? This can be a conflicting point. and would make comparisons impossible.

Authors, Answer, 6^th Caption of Table 1. We decided to put mean values in parenthesis.

• Reviewer 4, page 9, Line 184: A deeper discussion of these data could be provided (explore the influence of different environmental conditions of the different collection sites on morphoanatomical characteristics).

Authors, Answer, page 9, L.199-201: Thank You for the suggestion. The sentence: The greatest variation in Serbia could be the consequence of its very variable environmental conditions, too. was inserted.

• Reviewer 4, page 9, Line 187: What about comments on the wide variation in the number of ducts? However, consider my previous comments about duct number and density.

 

Authors, Answer, page 9, Line 187: Authors think that changed 6^th Caption of Table 1: Inserted parenthesis in table could solve the reviewer’s dilemma about number of resin ducts variability.

• Reviewer 4, page 9, Line 188: What explains? influence of the environment? constitutive character?

 

Authors, Answer, page 9, L. 207-209.  Influence of genetic and environmental factors to such variation surely exists, but their part in overall variation were not examined.

 

• Reviewer 4, page 27, Line 345: And what does the data collected from literature suggest? I see that you have a very informative set of anatomical data in your hands that deserve to be explored in more depth.

 

Authors, Answer, page 27, L.366-367: These variable sets of morpho-anatomical data suggest that new varieties or forms could be selected in future.

All other corrections in text were signed in green.

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

No comments.