Absence of Evidence or Evidence of Absence? Concurrent Decline in the Host Plant Onobrychis alba and the Butterfly Polyommatus orphicus in a Montane Habitat of Northern Greece

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

General comments

1. Section 2.2 is incomplete. It is entitled “Survey design and methods”; however, information about field survey methods is totally missing. How did the authors carry out butterfly, habitat and plant sampling? What methods were used? Standardized methods and protocols should be used in any survey.
2. How did the authors measure environmental degradation? All evidence provided in the Results comes for qualitative descriptions and observations, while no measurements of environmental parameters that can justify degradation are included such as vegetation and florescence measurements.
3. The butterfly species that was examined in the current study cannot be considered as a “specialized pollinator” or “associated pollinator” (lines 72-79, 207-209). European butterflies have only limited contribution to pollination. The larvae of some species may be bound to specific plants but in no case these butterflies are specialists to pollinate specific plants. Therefore, this kind of statements should be avoided.
4. The aim of the study should be described in the final part of the introduction and no results should be presented in this section, thus lines 81-83 should be reformulated accordingly.
5. The study focuses on a specific butterfly species and its host plant. Therefore, “montane community decline” in the title is not accurate, as the article does not examine any community. The title should be restructured accordingly (e.g. plant-host decline in a montane habitat in Northern Greece or relevant).

 

Specific comments

Keywords. “Local extinctions” is recommended to be removed from the keywords as the study was conducted only during one year.

Figure 1: The spatial reference system should be mentioned.

Table 1 should be removed from the manuscript. Species data is sensitive and localities should be obscure. Figure 1 depicts the sampling locations; thus table 1 is redundant. The habitat types of the six sampling sites can be added in the text.

102-104. As indicated in the Natura 2000 Viewer (https://natura2000.eea.europa.eu/), the study area is also designated as a Site of Community Interest (SCI) under the Habitats Directive (GR1140004), which protects non-bird species and is aligned with the study species. Lines 102-104 should thus be corrected accordingly.

106-108. Meteorological information is provided for the study region. The source of this information should also be provided.

124. This sentence should be added to the list with the selection criteria above.
125. Here “between 20 June and 25 July 2024” is redundant, it is already mentioned before (line 111).

130-132. This part should be moved to Section 2.2.

168. The phrase here is incomplete. Which species is P. eleniae synonym with? P. eleniae is not considered a taxonomically distinct species though.

Discussion. In many cases meteorological conditions create a harsh situation for plants and insects disrupting their life cycle. However, population fluctuations could also be plausible, and further research would be required to assess the future of this rare species.

204-206. “Rising temperatures may cause asynchrony between the life cycles of P. orphicus and the flowering period of its host plants, affecting reproductive success.”. There is evidence for this pattern from other studies, but it cannot be supported here as no such measurements were performed in this study.

207-209. “Pollination” sounds rather incongruous here and this kind of interrelationship between plants and butterflies should be avoided. Butterflies are not “specific pollinators”, and plants are not primarily dependent on them but on other pollinating insects (sensu stricto).

217. “effects of climate change impact” - effects sounds redundant here.

229-232. However, it is not clear whether this wildfire in the study area affected the habitat of the species and which way. As mentioned above a field methodology is missing from the article.

Author Response

General comments

Comment 1: Section 2.2 is incomplete. It is entitled “Survey design and methods”; however, information about field survey methods is totally missing. How did the authors carry out butterfly, habitat and plant sampling? What methods were used? Standardized methods and protocols should be used in any survey.

 

Response: We have fully rewritten Section 2.2 (“Survey Design and Methods”) to include detailed descriptions of butterfly, plant, and habitat surveys. This now specifies:

 

• search times and weather criteria,
• adapted Pollard & Yates-based thorough searches,
• alba phenology recording,
• qualitative habitat assessment and grazing pressure indicators,
• meteorological data source.

These details are now on pp. 4-5.

 

 

Comment 2: How did the authors measure environmental degradation? All evidence provided in the Results comes for qualitative descriptions and observations, while no measurements of environmental parameters that can justify degradation are included such as vegetation and florescence measurements.

 

Response: We acknowledge the limitation of not having quantitative vegetation data. The revised methods explicitly describe qualitative assessment protocols and livestock sign counts. This has been clarified in Section 2.2.

Comment 3: The butterfly species that was examined in the current study cannot be considered as a “specialized pollinator” or “associated pollinator” (lines 72-79, 207-209). European butterflies have only limited contribution to pollination. The larvae of some species may be bound to specific plants but in no case these butterflies are specialists to pollinate specific plants. Therefore, this kind of statements should be avoided.

 

Response: We removed references to P. orphicus as a “specialized” or “associated” pollinator. Wording was changed in the Introduction to focus on larval host plant specialization, not pollination dependency, while in Discussion it is kept general for pollinators.

 

Comment 4: The aim of the study should be described in the final part of the introduction and no results should be presented in this section, thus lines 81-83 should be reformulated accordingly.

 

Response: The aim of the study has been rewritten to present only the aim and scope, without preliminary results.

 

Comment 5: The study focuses on a specific butterfly species and its host plant. Therefore, “montane community decline” in the title is not accurate, as the article does not examine any community. The title should be restructured accordingly (e.g. plant-host decline in a montane habitat in Northern Greece or relevant).

Response: Title was changed to accurately reflect the study scope: Absence of Evidence or Evidence of Absence? Plant–Host Decline in a Montane Habitat of Northern Greece

Specific comments

Comment 6: Keywords. “Local extinctions” is recommended to be removed from the keywords as the study was conducted only during one year.

Response: Removed from keyword list.

Comment 7: Figure 1: The spatial reference system should be mentioned.

Response: Added WGS 84 (EPSG:4326)  as the spatial reference system in the figure caption.

Comment 8: Table 1 should be removed from the manuscript. Species data is sensitive and localities should be obscure. Figure 1 depicts the sampling locations; thus table 1 is redundant. The habitat types of the six sampling sites can be added in the text.

Response: Table 1 removed. Habitat descriptions for each site are now provided in the text.

Comment 9: 102-104. As indicated in the Natura 2000 Viewer (https://natura2000.eea.europa.eu/), the study area is also designated as a Site of Community Interest (SCI) under the Habitats Directive (GR1140004), which protects non-bird species and is aligned with the study species. Lines 102-104 should thus be corrected accordingly.

Response: Corrected to indicate that the study area is designated as SCI (GR1140004) under the Habitats Directive, in addition to SPA.

Comment 10: 106-108. Meteorological information is provided for the study region. The source of this information should also be provided.

Response: Added source: Hellenic National Meteorological Service.

Comment 11: Line 124. This sentence should be added to the list with the selection criteria above.

 

Response: fixed it

 

 

Comment 12: Line 125. Here “between 20 June and 25 July 2024” is redundant, it is already mentioned before (line 111).

 

Response: Removed the repeated phrase

Comment 13: Lines 130-132. This part should be moved to Section 2.2.

Response: Moved to Section 2.2.

Comment 14: Line 168 The phrase here is incomplete. Which species is P. eleniae synonym with? P. eleniae is not considered a taxonomically distinct species though.

 

Response: Clarified that P. eleniae is currently treated as a junior synonym of P. orphicus.

Comment 15: Discussion. In many cases meteorological conditions create a harsh situation for plants and insects disrupting their life cycle. However, population fluctuations could also be plausible, and further research would be required to assess the future of this rare species.

Response: Revised to acknowledge possible interannual variation, phenological mismatch, and detection limitations, with supporting references.

Comment 16: Lines 204-206. “Rising temperatures may cause asynchrony between the life cycles of P. orphicus and the flowering period of its host plants, affecting reproductive success.”. There is evidence for this pattern from other studies, but it cannot be supported here as no such measurements were performed in this study.

Response: Statement revised to be more cautious, noting that no such measurements were made in this study.

Comment 17: 207-209. “Pollination” sounds rather incongruous here and this kind of interrelationship between plants and butterflies should be avoided. Butterflies are not “specific pollinators”, and plants are not primarily dependent on them but on other pollinating insects (sensu stricto).

Response: Removed references to butterfly-specific pollination, replaced with general mention of plant–pollinator interactions.

Comment 18: Line 217 “effects of climate change impact” - effects sounds redundant here.

 

Response: Corrected wording to “climate change impacts.”

Comment 19: Lines 229-232. However, it is not clear whether this wildfire in the study area affected the habitat of the species and which way. As mentioned above a field methodology is missing from the article.

Response: Rewritten to state that potential wildfire effects on target habitat are uncertain and require further investigation.

Reviewer 2 Report

Comments and Suggestions for Authors

Thank you for this little, but highly relevant and kind of sad study of status of the butterfly endemic to Greek/Bulgarian/Albanian mountains, Polyommatus orphicus. And thank you also for rising the interest in Mt. Falakro, which happens to be my favourite spot (I had been there 3 times, once as a still young backpacker with a gilfriend, second time as a father, going with family by car, third timer with a busload of university students... But never saw P. orphicus ... and only the 2nd visit was at the right time, phenologically). 

But enough of nostalgia, lets go the review. 

I understgand all the limitations of searching for possibly extinct species, especially under time constraint (and in the sub-Mediterranean summer heaths). So, no problem here, but at lines 114-116, you are talking about spatial independence of the sites, and "permanent transects". Is this pseudo-statistical wizzardly really necessary, if you are searching for a species? I would skip the independence issue, and replace "transects" by "thorough searches", or something similar, just not to irritate those of readers, which would prefer "searching the entire mountain" over repeatedly walking transects. 

 

line 168: mising species name of the "P." thing (middle of the sentence). 

I have more reservations to Discussion, and general interpretation of the whole story. 

First, lines 182-183, you are mentioning factors contributing to detection failure ("..temporal mismatch, population fluctuations, or interannual variability..") - these are indeed quite likely, given that the year was exceptionally hot and dry, which could had shift the Onobrychis bloom, and P. orphicus flight, to earlier weeks of June. We all know such situations. It would be the optimistic case, of course, but it should be even more emphasized in the Discussion - and checked in field asap!

Second, repeatedly in Discussion, you are blaming grazing by "cattle and eqiunes" for habitat degradation, and contrasting it with "traditional grazing by sheep". Although I agree that overgrazing, or just badly executed grazing, often represents problem, we all should step out of the box, and view these issues in broader eco-evolutionary and historical perspectives. The butterfly had certainly evolved somewhere around Rodope mountain system, and sustained there all the climatic fluctuations of Pleistocene (how old is the species, btw.? please, check...), and for most of its evolution, there were wild equids, wild big bovids, and  wild Caprinae (C. ibex, most likely, plus possibly chamoix? as well) present in its habitats. Hence, it is the "traditional sheep" which is relatively novel (since the Neoliothic), as sheep ancestors arrived to the Balkans only with the Neolithic revolution. I know that this was long time ago, and the biota have had enough time to adapt to the sheep+goat system. But with decline of sheep+goat, would not it be better to adapt the cattle+horses pasture, so that it is less destructive to plants and insects? You are making some hints in this directin (lines 267-269), but may be it deserves some elaboration. 

Second, as side note, while both sheep and cattle likely prefer Onobrychis, horses more likely avoid it, preferring grasses over forbs and avoiding (fresh) Fabaceae plants. With dry plants, they do not care. 

 

Third, the fire issue. The droughts certainly do promote wildfires, but it is fact of life that wildfires had always been present in Mediterranean ecosystems - unless heavily grazed. What about using the fires (when they accidentally happen), grazing the post-fire vegetation (extremely nutritious for ungulates), and thus to in fact increase or restore the areas of calcareous grasslands? 

Finally, let me recommend two references, which touch the issue. Both are about Polyommatus (Agrodiaetus) damon, a relative of P. orphicus also developping on Onobrychis - and also struggling with droughts, and mismanagement of the sites. I hope they might help. 

Sucháčková Bartoňová, A., et al., 2021. Extremely endangered butterflies of scattered central European dry grasslands under current habitat alteration. Insect Systematics and Diversity, 5(5), p.6.

Šlancarová, J., et al.,  2012. How life history affects threat status: Requirements of two Onobrychis-feeding lycaenid butterflies, Polyommatus damon and Polyommatus thersites, in the Czech Republic. Biologia, 67(6), pp.1175-1185.

Author Response

Comment 1: Line 114–116 – “spatial independence” and “permanent transects”

Response: Replaced with “thorough searches of suitable habitat” to better reflect the actual search strategy. Removed unnecessary statistical terminology. Also, we added additional methodology in section 2.2

Comment 2: Line 168 – missing species name

Response: Corrected to include full species name.

Comment 3: Discussion – stronger emphasis on phenological mismatch

Response: Expanded discussion of phenological shifts in hot, dry years, citing Šlancarová et al. (2012) and Sucháčková Bartoňová et al. (2021), and recommending earlier-season surveys.

Comment 4: Discussion – grazing history and ecology

Response: Added context on historical grazing regimes (wild ungulates, introduction of domestic sheep/goats in Neolithic) and proposed adaptive cattle/horse grazing management. Included note on feeding preferences (horses vs cattle) for Onobrychis.

 

Comment 4: Fire management

Response: Included the ecological role of fire in Mediterranean grasslands and discussed potential controlled post-fire grazing as a management tool.

Lastly, I added the suggested references that I also believe will strengthen our manuscript

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript has been significantly improved after the revision. However, there are still some minor issues to be resolved, which would further improve content and increase clarity.

1. The title now depicts the content more accurately. However, it is still not very clear what the “plant-host” refers to; either to the decline of the larval host plant only or to the host-butterfly relationship (as stated for example in lines 73-81)? If the authors aim to acknowledge the concurrent decline of both host and butterfly populations (which I think they do), this should be formulated accordingly. If this is not the case, maybe “Host plant” would be more accurate?
2. Figure 1: As there is growing concern for threatened species, tools for geoprivacy have been developed to increase obscurity of species localities. Removing table 1 was aligned with this, however actual localities could be further obscured in fig. 1. Guidelines for obscuring localities are provided in different platforms; here is a widely used example: https://help.inaturalist.org/en/support/solutions/articles/151000169938-what-is-geoprivacy-what-does-it-mean-for-an-observation-to-be-obscured-
3. Lines 108-109: i. In accordance with the study system (i.e. butterflies and plants) SCI should be ahead of SPA. ii. “providing protection for non-bird species including invertebrates and plants associated with calcareous grasslands” is not completely relevant here and should be removed.
4. Line 143: “weather permitting” is redundant here, since favourable conditions are described in lines 147-148.
5. Line 150: “walking at a constant pace” should be deleted here as it has been already mentioned in line 147.
6. Line 152: What do “basic environmental descriptors” include? Is this related to lines 164-170?
7. Line 184: If precise precipitation data is available, this could be added inside a parenthesis here to further support this comparison.
8. I suggest authors review lines 200-211 as this part might be rather confusing (and includes repetitions). Line 207: Which species were the wild grazers before the Neolithic period? Line 208: “While the species likely adapted to moderate grazing” – Species refers to O. alba here? Line 209: Does this mean that now cattle and horse (it is mentioned in the Discussion) are the main or sole grazers in the study area?

9. Fig. 2: Does this legume pictured here has also signs of grazing (e.g. in its uppermost stems) or is it only a misunderstanding?
10. Fig. 4: I suggest the authors add in the legend of the figure the taxonomic relationship between P. eleniae and P. orphicus.
11. Line 222: Ref. 33 is outdated, as there is a more recent European assessment of this species. Since the species has been evaluated as Endangered in both the European and the national red lists, this is remarkably lacking and should certainly be added in the text to highlight the need for its conservation.
12. Line 252: “The complete desiccation of flowering plants and absence of Fabaceae inflorescences” – This may not be completely factual as this would lead to extirpation of the plants soon. Alternatively, the phenology of the plants and butterflies is asynchronous. What do the authors think about these two scenarios? Which of them do they support here?
13. Lines 258-273: This paragraph has substantially improved but again “pollinators” appear for the first time in line 268 and this looks rather mismatched here.
14. Line 278: impacts
15. Does this study comply with all ethical standards regarding animal handling and field work? Did this study have to be permitted by local authorities and bioethical committees? It is not mentioned anywhere in the manuscript (apart from releasing males after examining their genitalia).
16. References need to be checked again, for example: Some journal names are completely abbreviated, such as BR in ref. 1 and CCG in ref. 10 and 11, some are partially abbreviated, while others are not abbreviated at all. There is a mismatch in Pamperes and Pamperis in ref. 7. “Aufl.” should be deleted in ref. 7. The doi in ref. 10 is not working. Title in ref. 49 is partially capitalized. Etc.  

Author Response

Comment1: The title now depicts the content more accurately. However, it is still not very clear what the “plant-host” refers to; either to the decline of the larval host plant only or to the host-butterfly relationship (as stated for example in lines 73-81)? If the authors aim to acknowledge the concurrent decline of both host and butterfly populations (which I think they do), this should be formulated accordingly. If this is not the case, maybe “Host plant” would be more accurate?

 

Response: We agree and now make it explicit that we refer to the coupled decline of the host plant Onobrychis alba and the host-dependent butterfly Polyommatus orphicus.

New Title:

Absence of Evidence or Evidence of Absence? Concurrent Decline of the Host Plant Onobrychis alba and the Butterfly Polyommatus orphicus in a Montane Habitat of Northern Greece

 

Comment 2: Figure 1: As there is growing concern for threatened species, tools for geoprivacy have been developed to increase obscurity of species localities. Removing table 1 was aligned with this, however actual localities could be further obscured in fig. 1. Guidelines for obscuring localities are provided in different platforms; here is a widely used example: https://help.inaturalist.org/en/support/solutions/articles/151000169938-what-is-geoprivacy-what-does-it-mean-for-an-observation-to-be-obscured-

 

We thank the reviewer for this important comment. We agree with the principle of geoprivacy and the need to protect sensitive species. However, in our case, the majority of the localities of Polyommatus orphicus and its host plant Onobrychis alba are already publicly available from previously published sources. Furthermore, the map in Figure 1 does not provide sufficient resolution to allow extraction of precise coordinates, and therefore does not expose the species to additional risk. For these reasons, we have retained the current resolution of Figure 1 but have clarified in the caption that sensitive coordinates are not disclosed.

 

 

Comment 3: Lines 108-109: i. In accordance with the study system (i.e. butterflies and plants) SCI should be ahead of SPA. ii. “providing protection for non-bird species including invertebrates and plants associated with calcareous grasslands” is not completely relevant here and should be removed.

 

Response: Done.

 

 

Comment 4: Line 143: “weather permitting” is redundant here, since favourable conditions are described in lines 147-148.

 

Response: Deleted. No further change needed.

 

Comment 5: Line 150: “walking at a constant pace” should be deleted here as it has been already mentioned in line 147.

 

Response: Deleted to avoid repetition.

 

Comment 6: Line 152: What do “basic environmental descriptors” include? Is this related to lines 164-170?

 

Response: Clarified

 

Comment 7: Line 184: If precise precipitation data is available, this could be added inside a parenthesis here to further support this comparison.

 

Response: We now include a parenthetical range and the reference station.

 

Comment 8: I suggest authors review lines 200-211 as this part might be rather confusing (and includes repetitions). Line 207: Which species were the wild grazers before the Neolithic period? Line 208: “While the species likely adapted to moderate grazing” – Species refers to O. alba here? Line 209: Does this mean that now cattle and horse (it is mentioned in the Discussion) are the main or sole grazers in the study area?

 

Response: Rewritten for clarity, removed repetition, specified the referent of “species,” and stated current grazers.

 

Comment 9: Fig. 2: Does this legume pictured here has also signs of grazing (e.g. in its uppermost stems) or is it only a misunderstanding?

 

Response: Caption amended to state what is visible.

 

Comment 10: Fig. 4: I suggest the authors add in the legend of the figure the taxonomic relationship between P. eleniae and P. orphicus.

 

Response: Done.

 

Comment 11: Line 222: Ref. 33 is outdated, as there is a more recent European assessment of this species. Since the species has been evaluated as Endangered in both the European and the national red lists, this is remarkably lacking and should certainly be added in the text to highlight the need for its conservation.

 

Response: Updated text and references in the introduction to cite the European Red List of Butterflies (most recent assessment) and the national Red List for Greece, both listing P. orphicus as Endangered (EN).

 

Comment 12: Line 252: “The complete desiccation of flowering plants and absence of Fabaceae inflorescences” – This may not be completely factual as this would lead to extirpation of the plants soon. Alternatively, the phenology of the plants and butterflies is asynchronous. What do the authors think about these two scenarios? Which of them do they support here?

 

Response: Softened wording and stated our interpretation.

 

Comment 13: Lines 258-273: This paragraph has substantially improved but again “pollinators” appear for the first time in line 268 and this looks rather mismatched here.

 

Response: Response: Reframed entirely around larval host dependence; removed “pollinator” language.

 

Comment 14: Line 278: impacts

 

Response: Corrected to “impacts”.

 

Comment 15: Does this study comply with all ethical standards regarding animal handling and field work? Did this study have to be permitted by local authorities and bioethical committees? It is not mentioned anywhere in the manuscript (apart from releasing males after examining their genitalia).

 

Response: Added an Ethical Approval to the back matter and noted non-lethal handling in Methods. A formal research permit was obtained from the Ministry of Environment and Energy of Greece (Permit No. ΥΠΕΝ/ΔΔΔ/31412/1000, issued 22 March 2024; attached as Supplementary Material). We have now added a statement in the Methods section to clarify compliance with ethical requirements and permits.

 

Comment 16: References need to be checked again, for example: Some journal names are completely abbreviated, such as BR in ref. 1 and CCG in ref. 10 and 11, some are partially abbreviated, while others are not abbreviated at all. There is a mismatch in Pamperes and Pamperis in ref. 7. “Aufl.” should be deleted in ref. 7. The doi in ref. 10 is not working. Title in ref. 49 is partially capitalized. Etc.  

 

 

Response: We standardized all entries to the journal’s style: full journal names, sentence-case article titles, consistent author spelling, no “Aufl.”, working DOIs, and corrected capitalization.

Author Response File: Author Response.docx