Occurrence and Seasonal Variation of Picoplankton at Saiysad Freshwater in Taif City, Saudi Arabia
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThis excellent manuscript is an investigation of tiny photosynthetic organisms in fragile freshwater ecosystems of Saudi Arabia. The work was carried out using modern methods, the data were statistically verified and appear reliable. Only minor typographical errors listed below prevent this manuscript from being published immediately, but these can be easily corrected.
Line 97. The Methods section does not list the manufacturers of the devices used by the author.
Figure 3: It is better to decipher the fractions of plankton directly in the legend. This will shorten the caption to the figure and make it more understandable; abbreviations seem unnecessary.
Line 153. Gasol and Morán (2015). This reference is not formatted according to the rules and is not included in the list of references.
Line 211-232. In this section, please check the formatting of superscript characters.
Line 434-436. Some items in the Abbreviations section look strange. For example, PE is phycoerythrin, not orange fluorescence. The explanation of PerCP and Cy5-5 items also does not match the abbreviations.
Line 514. The reference to the book should look something like this: Aminot A., Rey F. Determination of chlorophyll a by spectroscopic methods // ICES Techniques in Marine Environmental Sciences. 1999. P. 1-15.
Author Response
Comment 1: This excellent manuscript is an investigation of tiny photosynthetic organisms in fragile freshwater ecosystems of Saudi Arabia. The work was carried out using modern methods, the data were statistically verified and appear reliable. Only minor typographical errors listed below prevent this manuscript from being published immediately, but these can be easily corrected.
Response 1:
Dear Reviewer,
I sincerely appreciate the reviewer’s positive and encouraging comments on this study and thank you for the opportunity to resubmit my revised manuscript entitled “Occurrence and seasonal variation of picoplankton at Saiysad freshwater in Taif city, Saudi Arabia” to Water. I am grateful for the time and effort you have dedicated to reviewing this paper and for recognizing the reliability of the methods and results presented. Regarding the minor typographical errors noted, I have carefully revised the manuscript and corrected them. A detailed point-by-point response to the reviewer’s comments is provided below.
Comment 2: Line 97. The Methods section does not list the manufacturers of the devices used by the author.
Response 2: As suggested by the reviewer, the manufacturers of the devices have been included in the revised manuscript and highlighted in yellow. Please see lines 118, 119, 141, and 154.
Comment 3: Figure 3: It is better to decipher the fractions of plankton directly in the legend. This will shorten the caption to the figure and make it more understandable; abbreviations seem unnecessary.
Response 3: Thank you for your insightful suggestion to improve clarity and readability. I have revised Figure 3 to explicitly label picoplankton, nanoplankton and microplankton. All abbreviations have been removed from both the figure and its caption. The updated version is included in the revised manuscript (see Figure 3, lines 211–216.
Comment 4: Line 153. Gasol and Morán (2015). This reference is not formatted according to the rules and is not included in the list of references.
Response 4: I apologize for this error. The reference has now been properly formatted and included in the reference list. Please see the line # 155.
Comment 5: Line 211-232. In this section, please check the formatting of superscript characters.
Response 5: Thank you for your comment. The formatting of superscript characters in lines 211–232 has been carefully checked and revised as requested.
Comment 6: Line 434-436. Some items in the Abbreviations section look strange. For example, PE is phycoerythrin, not orange fluorescence. The explanation of PerCP and Cy5-5 items also does not match the abbreviations.
Response 6: I apologize for the errors in the Abbreviations section. The entries have been corrected as follows: PE: Phycoerythrin; PerCP-Cy5-5: Peridinin chlorophyll protein-Cyanine 5.5. The revised manuscript reflects these corrections.
Comment 7: Line 514. The reference to the book should look something like this: Aminot A., Rey F. Determination of chlorophyll a by spectroscopic methods // ICES Techniques in Marine Environmental Sciences. 1999. P. 1-15.
Response 7: The reference has now been corrected to "Aminot, A., & Rey, F. (2002). Standard procedure for the determination of chlorophyll a by spectroscopic methods. ICES Techniques in Marine Environmental Sciences, 1–25."
Please find the revised manuscript attached.
Author Response File: 
Author Response.pdf
Reviewer 2 Report
Comments and Suggestions for AuthorsThis manuscript describes the occurrence and seasonal variation of picoplankton in Saiyasad freshwater, Taif City, Saudi Arabia. The study was mainly conducted through analyses of water quality, chlorophyll, and picoplankton abundance using flow cytometry. Overall, the manuscript is well written and well structured. However, some point requires clarification. The minor comments are listed below:
L74–77: Please provide a reference for this statement.
L90–96: This section should clearly state the objective of the study. What is the main objective? Please clarify.
L107: What is the distance between each sampling station? The stations appear to be close to one another, which may explain why the results did not differ significantly.
L167: Please provide data comparing nitrogen and phosphorus levels across the sampling stations for each season in Supplementary Table 1.
L203: It seems that in some seasons, the data may show significant differences. For example, pChl-a in summer and fall varies among stations. Particular attention should also be given to these differences.
L211: The authors mentioned that the picoplankton were classified as Synechococcus and Prochlorococcus. However, only Synechococcus was reported in this study. What about Prochlorococcus?
L240: What is the relationship between environmental variables and the sampling stations? Please provide this information, as it is valuable for water resource management, as discussed.
L294: This study was conducted in a freshwater ecosystem. Why did the authors measure salinity instead of electrical conductivity? Please clarify.
L312: The discussion of water quality does not clearly explain why this water resource is considered eutrophic. Please provide clarification.
L365: The authors stated that this freshwater resource is used for multiple purposes. Therefore, the ecological conditions at each sampling location should be compared.
Author Response
Comment 1: This manuscript describes the occurrence and seasonal variation of picoplankton in Saiyasad freshwater, Taif City, Saudi Arabia. The study was mainly conducted through analyses of water quality, chlorophyll, and picoplankton abundance using flow cytometry. Overall, the manuscript is well written and well structured. However, some point requires clarification. The minor comments are listed below:
Response 1:
Dear Reviewer,
I sincerely thank the reviewer for careful evaluation and positive comments regarding the clarity and structure of the manuscript. I also appreciate the constructive feedback and have addressed all the minor points listed below in the revised manuscript, with the changes highlighted in yellow.
Comment 2: L74–77: Please provide a reference for this statement.
Response 2: Thank you for the comment. References have been added to support this statement:
- McQuatters-Gollop A, Stern RF, Atkinson A, Best M, Bresnan E, Creach V, et al. The silent majority: Pico- and nanoplankton as ecosystem health indicators for marine policy. Ecol Indic 2024;159:111650.
- Li P, Luo Z, Zhu X, Dang Z, Zhang D, Sui X. Picoplankton Groups and Their Responses to Environmental Factors in Small Cascade Hydropower Stations. Water (Basel) 2025;17:903.
Comment 3: L90–96: This section should clearly state the objective of the study. What is the main objective? Please clarify.
Response 3: I have clarified the main objective of the study in the revised manuscript “The main objectives of this study are to (1) investigate the seasonal and spatial variability of both autotrophic and heterotrophic picoplankton in a freshwater ecosystem, and (2) evaluate the influence of environmental changes and nutrient inputs from human activities, particularly agriculture and irrigation, on their abundance, distribution, and community structure.” Please see the line 91-95.
Comment 4: L107: What is the distance between each sampling station? The stations appear to be close to one another, which may explain why the results did not differ significantly.
Response 4: I thank the reviewer for raising this important point. The three sampling stations were selected relatively close to one another to ensure the presence of water throughout the year, as sediment accumulation can substantially reduce water levels in some months. I acknowledge that the proximity of the stations may contribute to the lack of significant spatial differences; however, varying environmental conditions were observed, as described in the manuscript. This clarification has been added to the Methods section of the revised manuscript as follows: ‘The three sampling stations were at relatively short distances to ensure water was consistently present throughout the year, as sediment accumulation can occasionally limit water availability.’ Please see lines 115–117.
Comment 5: L167: Please provide data comparing nitrogen and phosphorus levels across the sampling stations for each season in Supplementary Table 1.
Response 5: If I understand correctly, the nitrogen (NO₃⁻) and phosphorus (PO₄³⁻) values across the sampling stations for each season are already presented in Figure 2 of the manuscript. I believe this figure provides a clear comparison of seasonal and spatial variations of these nutrients.
Comment 6: L203: It seems that in some seasons, the data may show significant differences. For example, pChl-a in summer and fall varies among stations. Particular attention should also be given to these differences.
Response 6: pChl-a was compared across all seasons using the Kruskal–Wallis test to assess seasonal variation. The analysis showed no significant differences among seasons (p = 0.484), indicating that overall pChl-a concentrations remained relatively stable throughout the year. Although small fluctuations were observed, with slightly higher values in Spring and Summer and lower values in Fall, these variations were not statistically significant.
Comment 7: L211: The authors mentioned that the picoplankton were classified as Synechococcus and Prochlorococcus. However, only Synechococcus was reported in this study. What about Prochlorococcus?
Response 7: Prochlorococcus was not detected in this study because it is typically found in oligotrophic ecosystems, whereas Wadi Saiysad is a highly eutrophic ecosystem. This explanation has already been provided in the Discussion section (see lines 350–351).
Comment 8: L240: What is the relationship between environmental variables and the sampling stations? Please provide this information, as it is valuable for water resource management, as discussed.
Response 8: The mean seasonal and spatial variations in environmental parameters are presented in Figure 2 and Table S1. The results show that while significant seasonal differences were detected for temperature, pH, nitrate, phosphate and N:P ratios, no significant differences were observed among the three sampling stations. This indicates that seasonal changes exert a stronger influence on water quality than spatial variation, likely due to the close proximity of the stations. This clarification has already been included in the revised manuscript (lines 293-297).
Comment 9: L294: This study was conducted in a freshwater ecosystem. Why did the authors measure salinity instead of electrical conductivity? Please clarify.
Response 9: The reviewer has raised an important point. Although electrical conductivity is often used in freshwater studies, salinity and EC are strongly correlated, as both reflect the concentration of dissolved ions. In this study, salinity values remained consistently low and stable across all stations and seasons, making it a reliable indicator of ionic strength in the freshwater ecosystem. I also acknowledge that salinity can serve as an indicator of anthropogenic and agricultural influences, particularly given the presence of farms adjacent to Wadi Saiysad. However, the persistently low and stable salinity values suggest that nutrient enrichment, rather than ionic accumulation, was the dominant signal of anthropogenic impact.
Comment 10: L312: The discussion of water quality does not clearly explain why this water resource is considered eutrophic. Please provide clarification.
Response 10: Wadi Saiysad is considered eutrophic because it exhibits persistent nutrient enrichment, particularly elevated nitrate (NO₃⁻) concentrations and consistently high nitrogen-to-phosphorus (N:P) ratios, indicating phosphorus limitation. In this study, NO₃⁻ concentrations were markedly high, whereas phosphate (PO₄³⁻) levels remained relatively low, resulting in elevated N:P ratios. These findings, together with evidence of nutrient inputs from agricultural runoff and treated wastewater, confirm that the Wadi functions as a eutrophic freshwater ecosystem. All of this information is already included in the manuscript (lines 304–321).
Comment 11: L365: The authors stated that this freshwater resource is used for multiple purposes. Therefore, the ecological conditions at each sampling location should be compared.
Response 11: The ecological conditions at each sampling location are described in the Materials and Methods section: “Station 1 (St. 1) was situated in a heavily used area with evident human activity and visible nutrient enrichment (Fig. 1C), Station 2 (St. 2) was dominated by aquatic vegetation and supported diverse populations of zooplankton and small fish (Fig. 1D), and Station 3 (St. 3) was located in a stretch of actively flowing freshwater (Fig. 1E)”. Please see lines 110–114.
Author Response File: Author Response.pdf
Round 2
Reviewer 1 Report
Comments and Suggestions for AuthorsI am very grateful for your positive evaluation of my modest contribution. I have one final request: please could you adjust the references in lines 303, 311, 322, and 382-383 to comply with the journal’s guidelines. I wish you continued success in your research and look forward to your future publications.
Reviewer 2 Report
Comments and Suggestions for AuthorsThank you very much for your careful revisions and for your efforts in addressing the reviewers’ comments.
