Bioacoustics Reveal Species-Rich Avian Communities Exposed to Organophosphate Insecticides in Macadamia Orchards
Round 1
Reviewer 1 Report
The paper used new way to explore the relastionship between avifauna and pesticide. The expeimental design is reasonalble, and the result is reliable. I recommend the acceptance after minor modification.
The suggestion is
Is it possible to give a table about the taxa composition(i.e. order) of anthrpods?
Author Response
Please note that all reviewer's comments were answered in a single file (attached).
Responses to Reviewer #1
The paper used new way to explore the relastionship between avifauna and pesticide. The expeimental design is reasonalble, and the result is reliable. I recommend the acceptance after minor modification. The suggestion is: Is it possible to give a table about the taxa composition(i.e. order) of anthrpods?
Response: Following the reviewer’s suggestion, we have included a new Table (3) with all related information.
Author Response File: 
Author Response.docx
Reviewer 2 Report
This is a very interesting and topical project, and I applaud the combination of ecotoxicological themes and a bioacoustic/ecological protocol; it's an integrative approach I'd like to see more people taking. The article itself is well written and coherent. I have only a few notes: - a more thorough description of the plant distribution and biome of the study sites would be a useful addition - I can make a guess, but I don't think it's entirely clear what a "water spray card" is - Figure 2's use of a "u" and an "s" seems like a confusing choice. I'd suggest coding all unsprayed sites in a similar pattern or color theme (for instance slashed or yellow/orange) and sprayed in a different pattern/color theme (for instance solid or blue/green). To illustrate site you could easily keep the same color or style of outline. That would make it easier to glean differences (or lack thereof)Author Response
Please note that all reviewer's comments were answered in a single file (attached).
Responses to Reviewer #2
This is a very interesting and topical project, and I applaud the combination of ecotoxicological themes and a bioacoustic/ecological protocol; it's an integrative approach I'd like to see more people taking. The article itself is well written and coherent.
I have only a few notes: - a more thorough description of the plant distribution and biome of the study sites would be a useful addition - I can make a guess, but I don't think it's entirely clear what a "water spray card" is - Figure 2's use of a "u" and an "s" seems like a confusing choice. I'd suggest coding all unsprayed sites in a similar pattern or color theme (for instance slashed or yellow/orange) and sprayed in a different pattern/color theme (for instance solid or blue/green). To illustrate site you could easily keep the same color or style of outline. That would make it easier to glean differences (or lack thereof)
Response: We appreciate the reviewer’s suggestions. As requested, a through description of plants and biome was added to the manuscript. (See L 112-124, 141-154 & 188-191). Spray cards are commonly used in insecticide-related field experiments to ensure that any potential drift does not impact control sites. Also, we have edited and improved Figure 2 with highly contrasting colors, but have kept the letters, to allow visualization for red-green blind people (following the E-in-C and other reviewer’s requests).
Author Response File: Author Response.docx
Reviewer 3 Report
Manuscript presents important issues related to the chemization of crops and the impact of these treatments on various groups of organisms. Before publication, the article requires some improvements as there is much confusion regarding the research methodology. The abstract and the results also require some corrections. Below I present my comments; I hope they will be helpful.
Comments:
I would consider the title of the manuscript. It suggests more the number and species composition of birds. However, research is more related to bird activity and arthropod abundance
I think that the abstract is very much part of a scientific publication. The abstract could more precisely refer to the obtained research results. Now, there are manly generalities that are not very exploratory and do not encourage you to read the entire article. There are results in the work that are interesting and should be exposed. You should emphasize changes more, before and after spraying.
L89 now iii and iii - schould be iii, iv
L90 - whether changes in the arthropod community following trichlorfon applications affect bird activity. - Are you sure you show this dependence in your research, or rather only the number of birds and arthropods? And the answer to this question is mainly part of the discussion section? (In line L-109 you write: "the short-term impacts")
L93-103 - The habitat description in the study area is too short. Essentially limited to the dominant species. It should be extended with information about the area covered by the study area by trees, shrubs, height structure, dimensional structure, etc. If such data are available, I suggest enriching this part, it will be easier to understand the species composition of bird and arthropod communities.
L108-110 You use the terms: location and sites. The size of these areas should be indicated. If they were small, also describe their surroundings, if it could greatly influence the groups of animals inside them.
L111 " not recently sprayed (henceforth 'unsprayed')" - requires clarification, because I do not fully understand what period has passed since the previous chemical spraying
L112-115 - Were any other treatments not described that could affect birds or arthropods? Also before the breeding season begins.
L112-115 - Lepidex is not used in many countries. Describe this organophosphate insecticide in more detail. How does this chemical affect arthropods? Does the producer mention any effects on birds, mammals etc.
L 125 - You can better describe the beginning and end of the breeding season for birds in these communities. Not everyone knows the bird assemblages and these habitats in Australia
L108-127 - Describe how the spraying was done, the time of day, the equipment used.
L129-133 - How acoustic recorders were placed inside the sites. Centrally? What distance range does this device have according to these settings. How did you solve the problem of linking the data of the birds recorded and only observed visually?
Table 1 Pre, Post1 and Post3 - All abbreviations should be explained below the table, or indicated where they are explained in the text.
L215-218 Add References
L219 - I can't find /3/ in the table. How did you solve the problem of non-recorded but watched species in the% column?
L221-222 - "Different letters indicate different groups based on Tukey’s post-hoc test results". Lichmera indistincta - Post1 ab?
Can you explain in methods what values (range) acoustic activity ratios can take and what the maximum values and what the minimum values mean? The same for arthropod abundance ratios. It is not entirely understandable.
Figure 3A. There are no statistically significant differences? Please check again.
L260 -Should be figure 2.
L272 Figure, Table? Where is this information presented?
Author Response
Please note that all reviewer's comments were answered in a single file (attached).
Responses to Reviewer #3
Manuscript presents important issues related to the chemization of crops and the impact of these treatments on various groups of organisms. Before publication, the article requires some improvements as there is much confusion regarding the research methodology. The abstract and the results also require some corrections. Below I present my comments; I hope they will be helpful.
Response: We appreciate the reviewer’s comments since we think that it has greatly improved the quality of our study. We have answered to each and every comment below.
I would consider the title of the manuscript. It suggests more the number and species composition of birds. However, research is more related to bird activity and arthropod abundance.
Response: We have changed the title in response to the reviewer’s comment by more directly addressing insecticide exposure among birds in the orchards. Because there were over 62 species of bird, we have maintained the words species-rich, but the new title does not highlight that our records of species diversity are novel (although we do believe they are high) or that they are the main focus of the study. Our study shows that 62 species in total were found in macadamia stands; we have also conducted previous research in the Bundaberg area, where we found similar species counts in eucalyptus patches nearby macadamia orchards. Indeed, we have checked data collected in the Bundaberg area uploaded to eBird (ebird.org). We did check particularly eucalyptus woodland areas nearby where the surveys for this study were carried out, and have found: 65 species in the Meadowvale Nature Park, 56 species in Miara, 60 species in Lake Gregory, and 52 species in Burrum Coast National Park.
I think that the abstract is very much part of a scientific publication. The abstract could more precisely refer to the obtained research results. Now, there are manly generalities that are not very exploratory and do not encourage you to read the entire article. There are results in the work that are interesting and should be exposed. You should emphasize changes more, before and after spraying.
Response: We appreciate the reviewer’s suggestion and have rewritten the abstract accordingly (L 34-63).
L89 now iii and iii - schould be iii, iv
Response: Corrected.
L90 - whether changes in the arthropod community following trichlorfon applications affect bird activity. - Are you sure you show this dependence in your research, or rather only the number of birds and arthropods? And the answer to this question is mainly part of the discussion section? (In line L-109 you write: "the short-term impacts")
Response: This point has been clarified in the manuscript, which now reads “to determine whether changes in the arthropod community following trichlorfon applications are associated with bird activity”.
L93-103 - The habitat description in the study area is too short. Essentially limited to the dominant species. It should be extended with information about the area covered by the study area by trees, shrubs, height structure, dimensional structure, etc. If such data are available, I suggest enriching this part, it will be easier to understand the species composition of bird and arthropod communities.
Response: We have expanded the information, following this and other comments made by other reviewers (L 141-144 & 160-174).
L108-110 You use the terms: location and sites. The size of these areas should be indicated. If they were small, also describe their surroundings, if it could greatly influence the groups of animals inside them.
Response: The size of the locations, though they were all macadamia orchards, differed from one another, but we chose paired sites with similar characteristics to ensure a robust experimental design. We have expanded the information regarding the characteristics of the studied orchards (L 159-165), and details on distances between paired sites and locations (L 170-174).
L111 " not recently sprayed (henceforth 'unsprayed')" - requires clarification, because I do not fully understand what period has passed since the previous chemical spraying
Response: We have specified the information on ‘unsprayed’ sites. Now it reads “for at least 45 days”.
L112-115 - Were any other treatments not described that could affect birds or arthropods? Also before the breeding season begins.
Response: There were not any other pesticide treatments carried out.
L112-115 - Lepidex is not used in many countries. Describe this organophosphate insecticide in more detail. How does this chemical affect arthropods? Does the producer mention any effects on birds, mammals etc.
Response: Trichlorfon is an organophosphate insecticide which is widely used in agriculture worldwide since the 50s [3]. According to the Encyclopedia of Toxicology by Karanth (2014), trichlorfon is currently used on several field crops and foods, including also several developing countries. Trichlorfon is sold under various product names, including – but not limited to - Anthon®, Bovinos®, Briten®, Chlorophos®, Dylox®, Dyrex®, Leivasom®, Neguvon®, Proxol®, Totalene®, and Trinex®. In Australia, during the period when the study was conducted, Lepidex ® was a commercial insecticide product (500 g/l trichlorfon) commonly used in macadamia orchards – as well as in other crops [30]. Nowadays, Lepidex ® use has been discontinued. However, Dipterex ®, containing exactly the same formulation as the previous product (trichlorfon 500 g/l), is commercialized and its use is recommended against “a wide variety of insect pests in various situations”, including three pests of macadamia, according to the producer information sheet (see https://cdn.nufarm.com/wp-content/uploads/sites/22/2018/05/14075246/Dipterex_500SL_label.pdf). This information has been added to Appendix A (L 568-580).
L 125 - You can better describe the beginning and end of the breeding season for birds in these communities. Not everyone knows the bird assemblages and these habitats in Australia
Response: Information has been added to the manuscript (L 188-189).
L108-127 - Describe how the spraying was done, the time of day, the equipment used.
Response: Insecticide spraying was conducted with air blast sprayers, normally during the afternoon. Information has been added to the manuscript (L 163-165).
L129-133 - How acoustic recorders were placed inside the sites. Centrally? What distance range does this device have according to these settings. How did you solve the problem of linking the data of the birds recorded and only observed visually?
Response: Acoustic recorders were located in the center of each site. We conducted preliminary tests to ensure that acoustic recorders could record sounds (vocalizations made by common species in the area) within a priori defined areas, ensuring no overlap was produced between paired sites. Based on the results we obtained, we then defined the distances between paired sites to ensure no overlap was produced (L 171-174). Bird point-counts were carried out to visually ratify the identification of some species recorded with acoustic recorders. We have clarified this in the manuscript (L 109-111).
Table 1 Pre, Post1 and Post3 - All abbreviations should be explained below the table, or indicated where they are explained in the text.
Response: Please note that the information is provided in the Materials and Methods section (L 178-182).
L215-218 Add References
Response: Added to the text.
L219 - I can't find /3/ in the table. How did you solve the problem of non-recorded but watched species in the% column?
Response: Fixed. Please find that ‘3’ refers to bird vocalizations recorder with audio recorders. All species observed during point counts were recorded with bioacoustics too. Also, please note that we have now included a new supplementary table (S1) with all the observations made using 10-min point counts.
L221-222 - "Different letters indicate different groups based on Tukey’s post-hoc test results". Lichmera indistincta - Post1 ab?
Response: Corrected. Now it reads “Groups with the same letter are not significantly indicate different…”.
Can you explain in methods what values (range) acoustic activity ratios can take and what the maximum values and what the minimum values mean? The same for arthropod abundance ratios. It is not entirely understandable.
Response: Corrected. Now it reads “Ratios had values between 0 and 1, with 1 indicating no activity or abundance recorded at unsprayed sites.”
Figure 3A. There are no statistically significant differences? Please check again.
Response: We have double-checked the analyses, and we confirm that there are no statistical significances (P > 0.05).
L260 -Should be figure 2.
Response: Corrected.
L272 Figure, Table? Where is this information presented?
Response: Fixed.
Author Response File: Author Response.docx
Round 2
Reviewer 3 Report
The Authors revised the manuscript according to my comments. In my opinion (and not only ) tables and figures should be self-explanatory, so all abbreviations used in them should be re-explained without having to look for them in the text (or at least a given subsection where to look for them). This improves readability and makes the article easier to use. However, this is a comment that does not affect the rating of the article. The substantive issues have been corrected. The research and manuscrypt are interesting, congratulations.
Author Response
Dear Reviewer,
Thank you very much for your comments and feedback. Following your advice, we have added information to explain all abbreviations in each figure and table in the manuscript.
On behalf of all authors,
Regards,
Eduardo
