Leaf Traits and Fluctuating Asymmetry as Stress Indicators in a Mangrove Species After an Extreme Rainfall Event

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Authors, 

Thanks much for introducing this outstanding work .I have carefully reviewed the manuscript throughly and found some parts needs additional enhancements and corrections. All these keypoints addresses in my report attached below. 

Comments for author File: Comments.pdf

Comments on the Quality of English Language

The provided manuscript lacks some slight English improvements to better readability. 

Author Response

Dear Authors, 

Thanks much for introducing this outstanding work. I have carefully reviewed the manuscript throughly and found some parts needs additional enhancements and corrections. All these keypoints addresses in my report attached below. 

Thank you for taking the time to review our manuscript and for your thoughtful comments. We appreciate your positive remarks regarding our work and your detailed suggestions for improvement. We have carefully addressed the key points outlined in your report and made necessary enhancements to strengthen the manuscript. We provided a detailed response to each comment and have clarified the modifications made. 

 

Title and Abstract 

The abstract does not possess some results that would be particularly important such as the percent increase in leaf area (LA) and specific leaf area (SLA) or any of the significance levels of asymmetry (FA). This absence certainly lessens the power and clarity of one’s findings. 

Furthermore, the background provided in the abstract is vague as this section looks to be better focused on describing the research gap succinctly and more pointedly.

We appreciate the reviewer’s insightful comments on the abstract. Following the suggestions, we have revised the abstract to include specific results for leaf area (LA), specific leaf area (SLA), and significance levels of fluctuating asymmetry (FA). We now report that FA significantly increased only after the extreme rainfall event (t = 1.759, df = 149, p = 0.080), whereas LA and SLA increased by 28% (p < 0.01) and 33% (p < 0.01), respectively (Lines 23-25). Additionally, we have clarified the research gap by emphasizing the need to understand the impact of extreme rainfall on the developmental instability on mangroves (Lines 19-21). We trust that these adjustments have enhanced the power and clarity of our findings, as requested.

 

Introduction 

 

There appears to be no explicit hypothesis encapsulating the relationship between FA and stress due to extreme rainfall, and therefore the expected outcomes remain undefined. Along with this, the purpose of the study is never directly articulated.

We express our gratitude and would like to draw attention to the fact that we have implemented the necessary modifications to the text to make the hypothesis more explicit (Lines 109-115).

 

There seems to be an absence of up-to-date sources on the studies focused on directional or fluctuating asymmetry of mangroves under climate change stress which would compromise the contextual framework of the study

We appreciate the reviewer's comment. To the best of our knowledge, there have been no additional studies on directional or fluctuating asymmetry in mangroves under climate change stress beyond those cited in our manuscript. We recognize that this may narrow the breadth of the contextual framework, but it also highlights the novelty and relevance of our study in addressing this gap.

 

Materials and Methods 

 

The study framework is usually clear, yet the justification for the sampling sizes is vague (10 participants during the dry season and 30 during the wet season). This is questionable, as without proper justification, doubts regarding the statistical validity may arise.

Thank you for your comment. We understand the importance of justifying sample sizes and appreciate the opportunity to clarify this. Initially, the study was designed to focus on herbivory, and based on a standardized global protocol, five individuals per sampling point were considered adequate for estimating herbivory effects. This approach is commonly used in ecological studies to ensure a reasonable estimate. However, after the extreme rain event, we realized that this climatic disturbance could introduce significant variability in plant responses. Given the unpredictable nature of the event, we expanded the sampling size to 30 individuals in the wet season, to better capture the variability and ensure that the data represented the full range of responses. However, we took advantage of this unpredictable weather event to investigate how such drastic climatic changes might affect stress indicators in mangrove plants, particularly leaf area and specific leaf area. This shift in focus allowed us to explore the broader impact of extreme climate events on mangrove health. We have added this rationale to the Materials and Methods (Lines 329-337) section for clarity, as follows: "The initial sampling design focused on herbivory, with 5 individuals per sampling point based on a standardized global protocol. However, following an extreme climatic event, we expanded the sample size to 30 individuals during the wet season to better capture the variability induced by the event. While stress measurement was not the primary aim of the sampling, we leveraged this unexpected weather event to investigate how drastic climatic changes could affect stress indicators in mangrove plants." We hope that this explanation provides more context for the sampling design and our rationale for adjusting it to account for the impacts of extreme events.

 

The extreme rainfall event chosen as a natural experiment does not fully describe how would one consider this event as an adequate representation of other climate events. 

We appreciate the reviewer’s valuable feedback. To address this concern, we have expanded our description of the extreme rainfall event and provided additional references on its social and economic impacts. According to McPhilips et al. (2018), extreme events are typically defined based on their intensity, frequency, duration, magnitude, and impacts. (Lines 311-314). Many studies have defined extreme events using magnitude alone—typically surpassing the 90th percentile of historical means—and our observed rainfall event exceeded ~97% of the historical mean, a threshold that strongly supports its classification as extreme (Lines 318-323). Furthermore, this rainfall event produced significant environmental impacts, including widespread flooding, landslides, and sediment displacement, which caused substantial damage to the infrastructure and led to the displacement of local populations (lines 325-326). These social and economic consequences validate the classification of the event as extreme in the region. We believe these added details clarify why our chosen event is a suitable representation of extreme climate conditions, and highlight the broader ramifications of such events in both the environment and the affected communities.

 

The text does not explain and recognize the presence of potential confounding factors like differences in soil salinity and tides which could influence the measured traits. 

Thank you for your valuable comment. We agree that factors such as soil salinity and tidal variation can influence plant traits and should be considered when interpreting the results. Unfortunately, we did not specifically measure or control for these variables in this study. Thus, they may represent potential confounding factors that could have affected the measured traits, such as leaf area and specific leaf area. We recognize that soil salinity can influence water uptake and nutrient availability, and tidal changes can impact the physical environment of mangrove plants, influencing their growth and stress responses. Because these factors were not directly accounted for in our experimental design, we acknowledge that they may contribute to some of the observed variability in plant traits, along wiith the effects of seasonality and climatic changes. We have added a limitation statement in the Discussion section (lines 187-193) to reflect this consideration, as follows: "One limitation of this study is the lack of control over potential confounding factors such as soil salinity and tidal variations, which can significantly influence plant traits in L. racemosa. These factors may have contributed to the observed variation in leaf area and specific leaf area and should be considered when interpreting our findings. Future studies could benefit from explicitly measuring and controlling for these variables to better isolate the effects of seasonal and climatic factors on plant traits." We hope that this addresses this concern and adds clarity to the potential limitations of this study.

 

The research does not describe how a measurement error (ME) of 0.19 percent was derived nor does it consider how it would affect reliability of the observed asymmetry. 

Thank you for your comment. We acknowledge the need to clarify the derivation and implications of the measurement error (ME). As described in the manuscript (Lines 357-363), ME was calculated using the formula ME = MSwithin / (sA² + MSwithin) × 100%, where MSwithin represents intra-individual variance, and sA² is the added variance component (Palmer and Stroebek, 2003). The sA² component was derived from MSamong - MSwithin/n, with MSamong and MSwithin obtained from an ANOVA using leaf identity as a predictor and repeated leaf measurements as the response variable. This approach ensures that the ME represents the proportion of within-individual variation relative to total variation. We also added a rationale that supports our reported ME of 0.19%, indicating that measurement imprecision contributed minimally to the observed asymmetry, falling well below the 1% threshold for high precision (Májeková et al., 2024) (Lines 363-364). Furthermore, while ME can be more consequential for species with very small leaves (Graham et al., 2015) (Lines 364-366), Laguncularia racemosa has relatively large leaves, further minimizing concerns regarding measurement reliability. Given these points, we are confident that ME does not compromise the reliability of the asymmetry patterns observed in our study.

 

Results 

 

Noteworthy differences with regards to leaf area and SLA are significant, however, the reporting is missing critical commentary on inter-individual variation or site specific variation that may have resulted in the observed differences. 

Thank you for your valuable comment. We agree that considering inter-individual and site-specific variation is essential for interpreting the observed differences in leaf area (LA) and specific leaf area (SLA). To address this, we clarify that our analyses were conducted using Generalized Linear Mixed Models (GLMMs), where season was included as a fixed predictor variable. Additionally, we accounted for individual variation by nesting individual plants within sampling points as random effects to capture site-specific influences. Our variance estimates indicated that sampling points contribute minimally to the variation in both traits. For LA, the variance associated with sampling points alone was 4.89 × 10⁻¹⁰, while the variance for individual plants within sampling points was 0.053, suggesting that LA variation is more strongly structured at the individual level rather than by site. In contrast, for SLA, the variance for individual plants was 0.005, and the variance for sampling points alone was 0.0003, indicating that both inter-individual and site-level variations were minor contributors to the observed SLA differences. To improve transparency, we explicitly incorporated these variance estimates into the Results section (Lines 136-143), ensuring that the contribution of inter-individual and site-specific variation is clearly reported. Additionally, we have added the following clarification to the Materials and Methods section (Lines 381-386): "To verify whether leaf area and specific leaf area varied between the dry season and after extreme rainfall (ERE), we constructed two Generalized Linear Mixed Models (GLMMs) using a Gaussian distribution. In these models, season was included as a fixed predictor variable, while plant individuals were nested within the sampling point as a random effect, and traits were used as response variables. This approach allowed us to account for inter-individual and site-specific variation in the analysis." We believe that these additions strengthened the interpretation of our results and enhanced the transparency of our analytical framework.

 

The reporting of p-values and some secondary findings is scant and by extension, some results appear under validated.

Thank you for your thoughtful comment. We have carefully reviewed all the statistical analyses presented in the manuscript to ensure that the reporting is comprehensive. We have included all the necessary details, including p-values, to support our results and ensure transparency. We believe that the current reporting provides a clear and robust foundation for our findings. 

 

The authors state that the nitrogen retention is indeed greater but the relative increase in asymmetry is greater still. Their linking remarks and no direct relationships are offered to support this claim. 

Thank you for your comment. We apologize for the confusion caused by the statement regarding nitrogen retention. As you pointed out, we do not have direct data on nitrogen retention in this study, and this was intended as a possible explanation for the observed patterns in fluctuating asymmetry (FA), rather than a confirmed relationship. We recognize the importance of providing stronger evidence for any claims made and have softened the discourse to clarify that the link between nitrogen retention and FA is a suggestion at this stage. Instead of stating it as a definitive finding, we framed it as a potential hypothesis to explain the greater increase in asymmetry observed after the extreme rainfall event (Lines 178-186). 

 

Discussion 

 

In regards to the increase in fluctuating asymmetry, the discussion offers little on the anticipated biological explanation of nutrients during exceedingly heavy rainfall and its impacts on developmental stability. 

Thank you for your observation. We agree that the link between nutrient dynamics and fluctuating asymmetry (FA) could be better elaborated. In response, we have expanded the discussion (Lines 175-186) to include a more detailed explanation of how the extreme rainfall event led to increased freshwater input, thereby enhancing the availability of nutrients such as nitrogen, which are crucial for growth. As we now explain, excess nitrogen can trigger rapid growth in mangroves, potentially leading to instability in plant development due to an inability to control development under high nutrient availability. This instability, in turn, may increase FA as a result of developmental noise. We have referenced relevant studies that highlight how high nutrient availability can disturb normal plant development and contribute to FA. We hope that this provides a more comprehensive understanding of the biological mechanisms involved.

 

Wind speed, mechanical stress, and the role of other stressors are mentioned in passing without adequate exploration of their contribution to the observed effects.

Multiple factors, including wind speed and mechanical stress, likely contributed to the observed effects. In this study, we highlighted the potential role of these variables, particularly in relation to extreme rainfall and its impact on plant stability and gas exchange. However, due to data limitations, we were unable to disentangle their individual contributions. Future research will aim to assess these variables more comprehensively by monitoring conditions before, during, and after extreme weather events. Nonetheless, our findings provide important preliminary evidence that advances our understanding of how extreme climatic events affect flora and fauna in the Neotropics.

 

The authors speculate that antisymmetry and some form of fluctuating asymmetry occurs simultaneously in response to different types of stress. However, they have not properly examined whether this observation is typical, or atypical, in relation to different types of plants.

We appreciate this comment and acknowledge that a broader taxonomic and ecological perspective would strengthen our conclusions. Our interpretation was based on a comparative analysis of data from two distinct periods—before and after an extreme weather event—suggesting that both antisymmetry and fluctuating asymmetry may arise under different stress conditions. However, further research is required to determine whether this pattern is consistent across different plant species and functional groups. Future studies incorporating multiple taxa and long-term monitoring are essential to better contextualize these findings. We inserted this suggestion in lines 237-240.

 

Little work has been done relating the results obtained for Laguncularia racemosa to other mangrove species or varying conditions of the environment.

We acknowledge that further comparisons with other mangrove species and environmental conditions would provide a more comprehensive understanding of the observed effects (Lines 244-246). This study serves as an important first step, offering preliminary evidence that extreme weather events influence the growth stability of L. racemosa. These findings will inform the design of long-term research, where we aim to collect robust data across multiple species and environmental contexts before and after extreme climatic events. We believe that this study lays the groundwork for future investigations into the broader ecological impacts of extreme weather in Neotropical mangrove ecosystems.

 

Figures and Tables 

 

Particularly for some of the figures on the distributions of fluctuating asymmetry, particularly those that display fluctuation asymmetry distributions, there is inadequate additional information explaining important findings and trends in the data that have been gathered.

Thank you for your insightful comment. In response to your suggestion, we have revised the figures displaying fluctuating asymmetry distributions. Previously, we included statistical values (such as t and p-values) directly on the figures. We have now removed these values from the figures (Line 125) and have instead retained them only in the main text (Lines 117-123) to maintain clarity and improve the overall readability of the visuals. We believe that this adjustment enhances the focus on the key trends and findings in the figures while still providing the necessary statistical details in the text.

 

Box plots for LA and SLA might be more informative if they were presented together with confidence intervals or other summary measures for a better explanation of results.

Thank you for this suggestion. We would like to clarify that the current box plots already provide a robust summary of the data distribution, including the interquartile range (boxes), median (black line), and data spread (whiskers). Additionally, we included the mean as a red dot for direct comparison and supplemented the figure with 95% confidence intervals around the mean to highlight the differences (Lines 148-153).

 

References 

 

This section of the document contains references that in all probability might be discussing the new concepts on the extremes of weather conditions and fluctuation asymmetry being one of plant stress indicators. Such references might provide or enhance the scope of the discussion in this thesis.

The suggestion is commendable. The requested actions were implemented, and several references were appropriately used, such as in lines 167, 171-172, 203-206, and 232-234. 

 

The references section contains some of the more recently published studies that may pertaining to the concept of multi extreme weather patterns, however, this does not enhance the depth of discussion as much as other concepts have the potential to do.

Thank you for your feedback. In response to your comment, we have carefully reviewed the references we included in the manuscript and reconsidered how they can enhance the breadth and depth of our discussion. We aimed to broaden our interpretation of our results by integrating additional insights from the literature on extreme weather conditions and plant stress indicators, particularly in relation to the broader ecological impacts of climatic extremes. We have expanded the Discussion section to better connect our findings to these references, thus providing a more comprehensive context for our results. This includes incorporating relevant studies on multi-extreme weather patterns and exploring how they might influence the physiological responses observed in mangrove species such as Laguncularia racemosa. By re-evaluating the literature and refining the connections to our own findings, we hope to offer a more nuanced interpretation that strengthens the manuscript's contribution to the field.

Reviewer 2 Report

Comments and Suggestions for Authors

Article Title: Leaf Traits and Fluctuating Asymmetry as Stress Indicators in a Mangrove Species After an Extreme Rainfall Event

The manuscript presents a study on the impact of extreme rainfall on the physiological and abiotic characteristics of Laguncularia racemosa. However, several sections require improvements to enhance clarity, conciseness, and informativeness.

 

Abstract

• Lines 30-32: Improve readability and ensure that the conclusion effectively summarizes the key findings while maintaining coherence with the study’s objectives.

Introduction

• The section is too lengthy. It should be more concise while retaining key ideas. Consider restructuring to improve readability.
• Lines 110-115: Clearly state the main research goal and hypotheses to provide a stronger foundation for the study.

Results

• The section needs substantial enhancement. Currently, it lacks informative data and clear statements about the obtained results. Ensure that findings are explicitly described and linked to the study’s objectives.

Discussion

• The discussion requires further refinement to provide a deeper interpretation of the results. Strengthen the connection between findings and relevant stress-response mechanisms.
• Consider incorporating the following references to enhance the discussion on plant stress responses:
• Harnessing GABA Pathways to Improve Plant Resilience Against Salt Stress. Horticulturae, 10(12), 1296. https://doi.org/10.3390/horticulturae10121296
• Optimization of Progesterone Treatment via Response Surface Methodology to Maintain Quality and Mitigate Chilling Injury in Prunus persica L. New Zealand Journal of Crop and Horticultural Science, 1-19. https://doi.org/10.1080/01140671.2025.2458819

Materials and Methods

• Lines 234-242: Move this section to the Introduction to provide a clearer background and justification for the methodology.
• Line 251: Delete "see" to improve clarity and readability.

Author Response

The manuscript presents a study on the impact of extreme rainfall on the physiological and abiotic characteristics of Laguncularia racemosa. However, several sections require improvements to enhance clarity, conciseness, and informativeness.

Thank you for your thoughtful comments. We appreciate your comments and agree that improving clarity, conciseness, and informativeness in certain sections of the manuscript has enhanced its overall quality. We have addressed these points and made revisions to the following areas:

• We have reviewed the introduction to ensure a clearer and more concise presentation of the study's objectives and the significance of the research.
• In the methods section, we streamlined the explanation of the experimental design and statistical analyses to improve readability and ensure that the approach is fully transparent and understandable.
• We have refined the results section by emphasizing key findings and ensuring that all data are clearly presented and discussed.
• We have also made adjustments in the discussion section, ensuring that we provide an adequate context for our findings and acknowledge relevant limitations.

We believe that these revisions will address the concerns regarding clarity and conciseness and improve the overall informativeness of the manuscript. Please, find below point-by-point answers to each of your specific comments.

 

Abstract

 

Lines 30-32: Improve readability and ensure that the conclusion effectively summarizes the key findings while maintaining coherence with the study’s objectives.

Thank you for your suggestion. We revised the conclusion to improve readability and ensure that it effectively summarizes our key findings while maintaining coherence with the objectives of the study. The updated text now states: "These findings demonstrate the differential effects of rainfall extremes on leaf traits and asymmetry, positioning FA, LA, and SLA as mangrove stress indicators." (Lines 26-28). This revision highlights the main outcomes of our study, emphasizing the role of leaf traits and asymmetry in detecting stress responses in L. racemosa.

 

Introduction

 

The section is too lengthy. It should be more concise while retaining key ideas. Consider restructuring to improve readability.

Thank you for your comment. We respectfully disagree with the suggestion to shorten the introduction, as we believe that the current length is essential to provide a comprehensive understanding of the background and context of the study. The introduction covers several important aspects, including the physiological effects of extreme rainfall, the role of fluctuating asymmetry (FA) as a stress indicator, and the potential effects of excessive nitrogen retention due to increased precipitation. These elements are crucial for framing the objectives of the study. The introduction also highlights the complexity of mangrove ecosystems, where extreme weather events such as heavy rainfall can lead to both positive and negative consequences for plant growth. By addressing these complexities, we ensure that readers have a clear understanding of the theoretical background before exploring the specific aims and hypotheses of our study. Additionally, we aimed to present a broad view of the environmental stressors in mangrove systems to help contextualize our research within a wider field. While we understand the concern for conciseness, we feel that shortening this section would risk losing the critical background information that is necessary for a clear understanding of the study's context and rationale. However, we reviewed the introduction for any area where we could improve readability without sacrificing essential details.

 

Lines 110-115: Clearly state the main research goal and hypotheses to provide a stronger foundation for the study.

We agree with your comment. We have clearly stated our main research goal and hypotheses (Lines 108-115) to guide the study's direction. This provided a stronger foundation for understanding the expected outcomes and how they align with the research focus.

•  

Results

 

The section needs substantial enhancement. Currently, it lacks informative data and clear statements about the obtained results. Ensure that findings are explicitly described and linked to the study’s objectives.

Thank you for your comment. We agree that the section needed enhancement. To address this, we have revised it to provide clearer, more detailed descriptions of the obtained results and explicitly linked them to the objectives of the study. We have ensured that the key findings are highlighted and discussed in relation to our research questions, offering a more informative and cohesive interpretation of the data.

 

Discussion

 

The discussion requires further refinement to provide a deeper interpretation of the results. Strengthen the connection between findings and relevant stress-response mechanisms.

Thank you for your suggestion. We agree that the discussion could benefit from a deeper interpretation of the results. We have refined this section to better connect our findings with the relevant stress-response mechanisms in plants. We now highlight how the observed changes in leaf traits, such as fluctuating asymmetry, leaf area, and specific leaf area, are consistent with the known physiological responses to environmental stress, particularly in relation to extreme rainfall events. This revision aims to provide a clearer understanding of the mechanisms underlying the stress-induced changes in L. racemosa and strengthen the overall interpretation of our results.

•  

Consider incorporating the following references to enhance the discussion on plant stress responses:

• Harnessing GABA Pathways to Improve Plant Resilience Against Salt Stress. Horticulturae, 10(12), 1296. https://doi.org/10.3390/horticulturae10121296

• Optimization of Progesterone Treatment via Response Surface Methodology to Maintain Quality and Mitigate Chilling Injury in Prunus persica L. New Zealand Journal of Crop and Horticultural Science, 1-19. https://doi.org/10.1080/01140671.2025.2458819

Thank you for your valuable feedback on our discussion. We have refined this section to provide a deeper interpretation of our results and strengthen the link between our findings and relevant stress-response mechanisms. Specifically, we have expanded our discussion on plant stress responses by incorporating insights from the suggested reference: Harnessing GABA Pathways to Improve Plant Resilience Against Salt Stress (Horticulturae, 10(12), 1296). This addition allowed us to better contextualize the potential role of γ-aminobutyric acid (GABA) in mitigating salinity stress and enhancing plant resilience in our system (Lines 222-226). However, we opted not to incorporate the second suggested reference (Optimization of Progesterone Treatment via Response Surface Methodology to Maintain Quality and Mitigate Chilling Injury in Prunus persica L., New Zealand Journal of Crop and Horticultural Science), as it focuses on chilling stress and a phylogenetically distant plant group (Rosaceae, order Rosales), which differs significantly from our study system (Combretaceae, order Myrtales, APG IV). Given these differences, we believe that the mechanisms discussed in that paper may not be directly applicable to our study.

 

Materials and Methods

 

Lines 234-242: Move this section to the Introduction to provide a clearer background and justification for the methodology.

Thank you for your suggestion regarding the relocation of this section to the Introduction. We understand that it is important to provide a clearer background and justification for the methodology. However, we believe that the details regarding L. racemosa—such as its characteristics, distribution, and specific traits like salt excretion mechanisms—are more appropriate for the Study Species section, as they provide context specific to our study organism rather than abroader theoretical background. We also agree with your previous comment that the Introduction is almost too lengthy, and we wanted to avoid overloading it with species-specific information that may not be as relevant to the general theoretical framework we are presenting. Additionally, we think that the ecological patterns mentioned (e.g., salt excretion and leaf turnover) may apply to a variety of mangrove species, as such, we chose to focus on the theoretical background in the Introduction. For these reasons, we have retained the species-specific information in the Study Species section, where it directly supports the rationale for choosing L. racemosa as the model species for this study. We hope this approach clarifies our decision, and we believe it maintains a balance between providing the necessary species-specific context and keeping the Introduction focused on a broader theoretical background.

 

Line 251: Delete "see" to improve clarity and readability.

Thank you for pointing this out to us! Changes made as requested (Line 311).

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript entitled „Leaf traits and fluctuating asymmetry as stress indicators in a mangrove species after an extreme rainfall event“ investigates, whether Fluctuating Asymmetry (FA), leaf area (LA) and specific leaf area (SLA) can be used as biotic indicators of environmental stress in the mangrove tree species Laguncularia racemosa, in this case of stress by flooding. The authors compare traits of leaves collected both, under dry conditions, and after an exceptional rain fall event. Results indeed show significant levels of FA in L. racemosa after the extreme event. As well as higher values of LA and SLA. In addition, during the dry season plants showed signs of antisymmetry. This is a nice little study highlighting that both genetically determined and environmentally triggered responses of leaf traits occur within the same plant species. I have only a couple of straight forward recommendation for improvement which qualify for a moderate revision. As this was not an available choice I opted for major revision, which it is not, from my point of view.

In general, please better discuss/explain in the discussion section the reason for the behavior of a plant, to develop FA, and what is the benefit for the plant? Is there anyone? (LL143-145, 164-172). In addition, why does precipitation tend to increase nitrogen retention in mangroves and how does extreme precipitation situations lead to increased leaf asymmetry.

L59-62: show the three types (Fluctuating Asymmetry (FA), leaf area (LA) and specific leaf area (SLA)) in a schematic figure/sketch.

L123-127: Figure caption of Figure 1: I recommend to change colors??? Conventionally red indicates (hot and) dry conditions, while blue better symbolizes wet conditions.

L172-174: unclear, what is meant

L222-232: Study area section: include a map and a climate chart showing the mean climatic conditions, i.e. monthly mean temperature and precipitation values

L233-243: Study species: where does L. r. occur along the environmental gradient form sea to land, i.e. within the common zonation of Atlantic mangroves.

Author Response

The manuscript entitled “Leaf traits and fluctuating asymmetry as stress indicators in a mangrove species after an extreme rainfall event” investigates, whether Fluctuating Asymmetry (FA), leaf area (LA) and specific leaf area (SLA) can be used as biotic indicators of environmental stress in the mangrove tree species Laguncularia racemosa, in this case of stress by flooding. The authors compare traits of leaves collected both, under dry conditions, and after an exceptional rainfall event. Results indeed show significant levels of FA in L. racemosa after the extreme event. As well as higher values of LA and SLA. In addition, during the dry season plants showed signs of antisymmetry. This is a nice little study highlighting that both genetically determined and environmentally triggered responses of leaf traits occur within the same plant species. I have only a couple of straight forward recommendation for improvement which qualify for a moderate revision. As this was not an available choice I opted for major revision, which it is not, from my point of view.

Thank you for your positive and constructive feedback regarding our manuscript. We greatly appreciate your recognition of the value of our study and the insights it provides regarding the potential use of fluctuating asymmetry (FA), leaf area (LA), and specific leaf area (SLA) as indicators of stress in Laguncularia racemosa. We also appreciate your recommendation for further improvements and have revised the manuscript based on your suggestions. We have addressed the recommended changes in a way that enhances the clarity and depth of the study, as we believe that these revisions will further strengthen the manuscript. Thank you again for your thoughtful review. 

 

In general, please better discuss/explain in the discussion section the reason for the behavior of a plant, to develop FA, and what is the benefit for the plant? Is there anyone? (LL143-145, 164-172). In addition, why does precipitation tend to increase nitrogen retention in mangroves and how does extreme precipitation situations lead to increased leaf asymmetry.

Thank you for your insightful comment. We appreciate your request for a deeper discussion on the potential benefits of fluctuating asymmetry (FA) and the mechanisms underlying the observed patterns in leaf traits. In response, we have expanded the Discussion section to provide a clearer explanation of why plants may develop FA. However, we acknowledge that FA is generally considered a marker of developmental instability, and while it typically reflects environmental stress or genetic instability, it is not usually thought to have direct benefits for the plant. Instead, FA can be seen as a consequence of the plant’s inability to achieve perfect symmetry in response to environmental stressors, such as extreme rainfall. The development of FA could indicate a disturbance in the plant’s developmental processes, particularly when facing factors such as nutrient imbalance or environmental stress. As we have added in the revised manuscript (lines 175-186), high precipitation leads to increased freshwater influx in mangrove systems, particularly in riverine mangroves. This influx of freshwater often results in an increase in nutrients, especially nitrogen, which plays a critical role in leaf growth. Higher nitrogen availability under intense rainfall can lead to accelerated growth of mangrove species, potentially contributing to developmental instability. We hypothesized that in conditions of nitrogen excess, the plant may experience a form of "growth overload", where it invests heavily in rapid leaf production at the expense of structural stability, thereby increasing susceptibility to stress and leading to greater FA. In particular, excessive nitrogen may disrupt the plant's ability to allocate resources effectively, as rapid leaf growth could result in less investment in the structural and defensive traits that typically protect the plant. This could explain why extreme rainfall events, through their impact on nitrogen availability, lead to increased FA: the plants are growing faster than they can properly manage their resources, leading to imbalanced development. We have revised the manuscript to include these points and to clarify the mechanisms underlying FA development in mangroves under extreme weather conditions. We hope that this additional explanation addresses your concerns and enhances the understanding of our results.

 

 

L59-62: show the three types (Fluctuating Asymmetry (FA), leaf area (LA) and specific leaf area (SLA)) in a schematic figure/sketch.

We appreciate the reviewer's suggestion. We have now included a schematic figure (Figure 5) in the Methods section that illustrates the three types of asymmetry (including Fluctuating Asymmetry) as well as SLA and LA variables.

 

L123-127: Figure caption of Figure 1: I recommend to change colors??? Conventionally red indicates (hot and) dry conditions, while blue better symbolizes wet conditions.

Thank you for your valuable suggestion. We agree with your observation that initial colors may have been counterintuitive. As you pointed out, red is often associated with hot and dry conditions, whereas blue is typically used to symbolize wet conditions. In response to your comment, we have revised the color scheme to use orange for the dry period and blue for the rainy period, which aligns more closely with these conventional associations. Additionally, to enhance cohesiveness between figures, we applied the same color scheme to both Figure 1 (Line 125) and Figure 2 (Line 149). We believe this change improves clarity and strengthens the visual consistency across the figures.

 

L172-174: unclear, what is meant

Thank you for pointing this out to us. We have revised lines 194-198 to improve clarity and to ensure the meaning is more easily understood. We believe that the rewording addresses ambiguity and enhances the overall readability of the section.

 

L222-232: Study area section: include a map and a climate chart showing the mean climatic conditions, i.e. monthly mean temperature and precipitation 

values

Thank you for your suggestion. We added a map of the study area (Figure 3A) and a climate chart  (Figure 4) showing the mean climatic conditions, including monthly mean temperature and precipitation values. These additions have now been included in the manuscript to provide a clearer context for the study area and its climate.

 

L233-243: Study species: where does L. r. occur along the environmental gradient from sea to land, i.e. within the common zonation of Atlantic mangroves.

Thank you for your comment. We have added the requested information regarding the occurrence of Laguncularia racemosa along the environmental gradient from sea to land within the common zonation of the Atlantic mangroves (Lines 291-294). This clarification has been included in the revised manuscript to provide a more comprehensive understanding of the species' distribution along the gradient.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Authors, 

I'm truly grateful for considering my comments positively this resulted in elevating manuscript quality. 

Comments on the Quality of English Language

The English writing style could benefit from additional minor grammatical correction. 

Author Response

Dear Authors, 

I'm truly grateful for considering my comments positively this resulted in elevating manuscript quality.

We sincerely appreciate your thoughtful feedback and constructive suggestions. Your insights have significantly contributed to improving the quality of our manuscript.

 

The English writing style could benefit from additional minor grammatical correction. 

We appreciate your suggestion regarding the English writing style. To ensure the highest quality of language, we carefully reviewed the entire manuscript and made minor grammatical and stylistic improvements where necessary, such as in lines 24 (in which we replaced compared to by compared with, indicating that we are comparing similar things - i.e. seasons), line 50 (in which we inserted a comma after the word conductance), line 58 (in which we replaced the word symmetric by symmetrical), and line 95 (in which we replaced the word The by An). Other additional grammatical corrections are highlighted in yellow.

Reviewer 2 Report

Comments and Suggestions for Authors

The author has made several enhancements; however, they disagree with condensing the introduction. While the introduction contains many sentences that could be streamlined.

Author Response

The author has made several enhancements; however, they disagree with condensing the introduction. While the introduction contains many sentences that could be streamlined.

We appreciate your feedback on the introduction. We carefully reviewed this section and streamlined sentences where possible while ensuring that the necessary context and background information were preserved, such as in lines 32-39,  and in lines 40-42 and 48-52 (in which we divide the phrases in two, to enhance clarity). We also identified areas where we could reduce wordiness by removing unnecessary phrases. For instance, the sentence “However, some studies have shown that antisymmetry may be an indicator of developmental instability, and may represent a transitional stage of symmetry breaking in organisms under significant stress, generating nonlinear developmental processes [18].” was revised to “However, antisymmetry may be an indicator of developmental instability, and may represent a transitional stage of symmetry breaking in organisms under significant stress, generating nonlinear developmental processes [18]” (Lines 65-67). In the same sense, the sentence “For example, previous studies have shown plants under water limitation or droughts can exhibit more leaf FA than plants under adequate water availability [23], reflecting developmental instabilities due to water restriction.” was revised to “For example, plants under water limitation or droughts can exhibit more leaf FA than plants under adequate water availability [23], reflecting developmental instabilities due to water restriction.” (Lines 72–75). We also tried to simplify some phrases, such as the case of the sentence “Therefore, it is important to verify whether plants in extreme weather conditions undergo variations in their characteristics linked to growth and resource acquisition, as abrupt abiotic changes caused by extreme climatic events can have multifaceted effects on plant physiology.”, which now reads as “It is important to assess whether extreme weather conditions alter plant traits related to growth and resource acquisition, as abrupt abiotic changes can have complex effects on plant physiology.” (Lines 95-97). Additionally, we also standardized terminology for various concepts, such as replacing “precipitation” with “rainfall”.

Reviewer 3 Report

Comments and Suggestions for Authors

Dear authors, dear editors,

thank you very much, authors, for the efforts you made to improve the manuscript. I find you perfectly addressed all the issues raised by me in the first round of review and I am very satisfied with the revised version. From my point of view the manuscript can be accepted for publication in the current form.

Author Response

Dear authors, dear editors, thank you very much, authors, for the efforts you made to improve the manuscript. I find you perfectly addressed all the issues raised by me in the first round of review and I am very satisfied with the revised version. From my point of view the manuscript can be accepted for publication in the current form.

Thank you for your positive feedback and for recognizing our efforts in improving the manuscript. We truly appreciate your thoughtful comments and are grateful for your support throughout the review process.