Assessment of the Diversity, Distinctiveness and Conservation of Australia’s Central Queensland Coastal Rainforests Using DNA Barcoding

Round 1

Reviewer 1 Report

Dear Author

This is a well-designed study; With reliable sample collection procedures, the methods used to assess genetic diversity based on phylogeny (DNA barcoding) are appropriate. The results are therefore highly reliable. The author designs the manuscript well; reasonable structure, presents it clearly, and the style attracts readers.

Minor suggestion:

The introduction is too long. There are too much on issues related to rainforests in Australia, and The Central Queensland Coast (CQC) rainforest. The author can cleverly truncate to bring down into the discussion part, with little information.

The Phylogenetic Diversity in the introduction section is too short. The author should introduce the background and tendency of this approach to make it clear to the readers.  In addition, the author needs to further analyse the weakness of the Phylogenetic Diversity approach in this study in comparison with traditional ones. 

What is the most significant outcome in the assessment of the diversity, and distinctiveness, of Australia's Central Queensland Coastal rainforests obtained in this study by the Phylogenetic Diversity method?

Best wishes

Author Response

Response to Reviewer 1 Comments

Manuscript ID: diversity-2200657

Thank you for your comments and opportunity to revise the manuscript.

Point 1: The introduction is too long. There is too much on issues related to rainforests in Australia and the central Queensland coast (CQC) rainforest. The author can cleverly truncate to bring down into the discussion part, with little information.

Response 1: We have amended the introduction to reduce its length and reduce the amount of repetition. However Reviewer 2 has requested clarification for some points, so we have added a small number of sentences to provide this.

Point 2: The Phylogenetic Diversity in the introduction section is too short. The author should introduce the background and tendency of this approach to make it clear to the readers. In addition, the author needs to further analyse the weakness of the Phylogenetic Diversity approach in this study in comparison with traditional ones.

Response 2: Thank you for drawing our attention to this. We have altered the text of lines 105-111, page 3, in order to explain the Phylogenetic Diversity approach more clearly.

Point 3: What is the most significant outcome in the assessment of the diversity, and distinctiveness of Australia’s Central Queensland coastal rainforest obtained in this study by the Phylogenetic diversity method?

Response 3: We have altered the text in the Conclusions to explain the most significant outcome in line 730 of the original manuscript (now 851 in the track changes copy).

Reviewer 2 Report

Dear authors,

Congratulations on excellent research and valuable and perfect results. Suggestions for clarification and potential improvement of the manuscript follow in the attached document.

Best Wishes.

Comments for author File: Comments.pdf

Author Response

Response to Reviewer 2 Comments

Manuscript ID: diversity-2200657

Thank you for your comments and opportunity to revise and enhance the manuscript. Thank you also for your positive comments, they were greatly appreciated.

Point 1: I suggest adding the main patterns that drive species distribution between and within the studied areas. Also highlight the correlations between the latter.

Response 1: We have ammended the Abstract as suggested. (line 4 of the original manuscript)

Point 2: I suggest highlighting the main selection pressures that affect the similarity and phylogenetic diversity of the sampled areas

Response 2: We have ammended line 27 of the original manuscript to reflect this.

Point 3: In what sense are these areas little known? In terms of structure, dynamics or richness?

Response 3: In response to Reviewer 1 we have reorganised the introduction. This sentence has been ammended and moved from line 45 in the original text to line 77 in the second paragraph of the tracked changes copy.

Point 4: In what sense do the results of the proposed research help in the conservation of the studied areas?

Response 4: We have ammended this sentence highlight the conservation value of this research. (line 142 of the original manuscript, now 165 in tracked changes copy).

Point 5: Is it important to insert a question about the relationship of phylogenetic diversity between the areas studied and what factors drive them?

Response 5: Thank you for this suggestion. We have added a question in the Aims (question 3.)

Point 6: Highlight the number of taxa sampled?

Response 6: We have ammended the first sentence of the Materials and Methods section to include the number of taxa sampled in lines 87, 163, and added a new paragraph in line 183 to make this more clear to the reader.

Point 7: Separate extraction from amplification. Why use these markers?

Response 7: We have separated the extraction and PCR amplifictation and ammended this paragraph to make the use of these markers more clear for the reader, in line 175 of the original manuscript.

Point 8: Inform the sequence number for each marker. Also the taxa number

Response 8: We have ammended this section (lines 173 -184 of the original manuscript) for more clarity about the sequence number and we have added the number of contigs for each marker in the next section (Queensland Rainforest Phylogeny).

Point 9: Methods that generated phylogenetic trees? Or transfer to the phylogenetic analysis session

Response 9: We have ammended this sentence to include the options used in Geneious. (line 195 of the original manuscript and line 246 of the revised copy)

Point 10: Why not APG IV?

Response 10: To assemble our phylogeny, we used the Phylomatic V3 online tool which used APG III. There is not an option to change this aspect of the program.

Point 11: Where do we find the list? Add a call to it.

Response 11: This list can be accessed via the Herbrecs database online. We have added the web address (line 231 of the original manuscript, now line 291 of the revised text) Also a full list of species will be added in the supplimentary materials.

Point 12: Considering the same markers used?

Response 12: We have ammended the text to make this more clear for the reader. (line 281 in the original manuscript and line 341 of the revised text)

Point 13: Families and Genera?

Response 13: This paragraph has been moved to the methods section and we have included a call to the table showing species, family and genus richness.

Point 14: From which publications did these come?

Response 14: These sequences are freely available from Genbank and BOLD online databases. The list of species will contain the Accession numbers where possible to enable the reader to find them easily.

Point 15: Part of the methodology

Response 15: Thank you for this suggestion, we have move these sentences to the methodology section (line 281 of the revised text).

Point 16: How was the conservation status calculated?

Response 16: We have ammended this paragraph to include the government websites where the conservation status of threatened plant species can be found in line 486 of the revised text.

Point 17: What does this suggest?

Response 17: We have ammended the text in the first paragraph of the discussion to more clearly explain this. (line 685 of the revised text).

Point 18: Could it be that the rainfall volumes of such regions direct their similarities and richness.

Response 18: We have ammended this sentence to clarify our explanation (line 582 of the original manuscript and 696 of the revised text)

Point 19: Would they have a relationship with the endemism present in such areas? Their conservation status?

Response 19: We have ammended this sentence also to clarify this point. (line 600 of the original manuscript and line 719 of the revised text).

Point 20: Could the results of this study support areas for the conservation of species found in places of intense anthropic pressure?

Are the phylogenetic diversity corrections found between the areas due to the sharing of widely distributed species within them? Your disturbance levels?

Response 20: We have ammended the final sentence to touch on the conservation near areas of anthopic pressure, however we find that we are unable to comment on disturbance here as this was not included in this study. This will be the focus of the final Thesis chapter of the lead author.

Reviewer 3 Report

I think the manuscript is interesting for Diversity journal as the authors use the DNA barcode sequences (some identified by them) to establish the phylogenetic distances from species belonging to different Australian regions. Specifically, they establish the need for conservation actions. However, I believe that some revisions must be made to complete the manuscript.

- First of all it is important to indicate in the methods section  the primers used for the three marker genes and also to specify the length of the amplified fragment used in the multi-alignments with the sequences obtained from Genbank and BOLD databases.

- It would be important to add the list of species examined in supplementary materials. - Furthermore, it would be important to establish what is the percentage of species in these forests that do not  have DNA barcodes.

Minor comment: the quality of the figures 3 and 6 must be improved.

Author Response

Response to Reviewer 3 Comments

Manuscript ID: diversity-2200657

We appreciate your comments and opportunity to revise the manuscript. Thank you.

Point 1: It is important to specify the primers used for the three gene markers and to specify the length of the amplified fragment used in the multi-alignments with the sequences obtained from Genbank and BOLD databases.

Response 1: We have included a table in the Appendix containing the PCR primers used and referred to this in the text (line 179 in the original manuscript, 212 in the tracked changes copy). We have clarified the quality controls used for selecting the sequences used. (lines 181 and 198 of the original manuscript, now 235 and 251 of the track changes copy).

Point 2: It would be important to add the list of species examined in the supplementary materials.

Response 2: We are constructing a table of species examined and where possible the Accession numbers for each of the 3 markers. This will be completed within the next couple of days.

Point 3: It would be important to establish what is the percentage of species in these forests that do not have DNA barcodes.

Response 3: We have clarified the percentage of species barcoded for this study from our list of species known from the Central Queensland Coastal region (line 200 of the original manuscript or 253 of the tracked changes copy).

Point 4: The quality of figures 3 and 6 must be improved.

Response 4: We have improved the quality of figures 3 and 6 by using the .jpeg images and removing the individual titles from each NMDS.