Benthic Diatoms on Macrophytes of the Israeli Mediterranean Coast

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript offers findings into the species composition, ecology, and phytogeography of epiphytic diatoms inhabiting macrophytes along the Israeli Mediterranean coast. In general, the manuscript is well constructed, but there are several aspects that warrant improvement in the manuscript to enhance its overall quality and clarity.

 Major concerns

1. The sampling time was limited between March to May of 2021, which may potentially restrict the understanding of seasonal dynamics throughout the entire year. It is recommended that the authors describe the rationale behind this specific time selection.

 2. I suggest the author to keep the “Bray-Curtis” related method and descriptions in the manuscript to be clear and consistent.

 3. I am curious about why the environmental variables only measured by one sample in each sampling site?

 4. Why did not adopt the molecular technique (e.g. based on 18S rRNA) for taxonomic identification of collected diatoms.

Figures and Tables

1. I suggest the authors to improve the quality and the readability of all figures in the manuscript, by using some professional graphing software to plot figures instead of using EXCEL.

 2. check the reference ([2]) in the figure legend of Fig. 3.

 3.Table 4 is the repeated data of Fig. 3, please delete one of the data. Besides, why connected different sites in Fig.3? I suggest the author to use bars instead of using dot-line plot.

 4.Table 5: The descriptions of abbreviations of “Hab” and “Sap” were lost.

Comments on the Quality of English Language

Minor editing of English language needed.

Author Response

Dear Editor,

Thank you and the Reviewer 1 comments. Please find the responses below.

With best regards,

Prof Sophia Barinova

Corresponding author

 

Responses to the Reviewer 1 comments

Comments and Suggestions for Authors

The manuscript offers findings into the species composition, ecology, and phytogeography of epiphytic diatoms inhabiting macrophytes along the Israeli Mediterranean coast. In general, the manuscript is well constructed, but there are several aspects that warrant improvement in the manuscript to enhance its overall quality and clarity.

Major concerns

1. The sampling time was limited between March to May of 2021, which may potentially restrict the understanding of seasonal dynamics throughout the entire year. It is recommended that the authors describe the rationale behind this specific time selection.

Response: We did not set the task of studying seasonal dynamics at the first stage of studying the diversity of diatoms but focused on identifying the species composition. However, in the MM section it is added that the selected season is the most favorable for the development of autotrophs in these climatic conditions.

2. I suggest the author to keep the “Bray-Curtis” related method and descriptions in the manuscript to be clear and consistent.

Response: The program is cited [41] and is publicly available with a detailed description of the methods. There are all clear for the users.

3. I am curious about why the environmental variables only measured by one sample in each sampling site?

Response: This is at least 5 measurements of each specified parameter in scooped water, the results are statistically processed, and the average is given. Variations are extremely low for all parameters, excluding nitrate nitrogen, but everything is within the error limits of the specified HANNA instruments.

4. Why did not adopt the molecular technique (e.g. based on 18S rRNA) for taxonomic identification of collected diatoms.

Response: Because at the first stage we set the task of identifying the diversity of diatoms for the first time. Molecular methods require a more nuanced study, which is planned to be done from populations isolated from crops, since general 18S analysis does not provide species identification. That is, it is beyond the scope of the task.

Figures and Tables

1. I suggest the authors to improve the quality and the readability of all figures in the manuscript, by using some professional graphing software to plot figures instead of using EXCEL.

Response: We have improved the readability of the figures by removing the connection of the columns, however, the text format of the figures exported from EXCEL is quite satisfactory for the technical editor when the manuscript is not in the draft format, but in the published format.

2. check the reference ([2]) in the figure legend of Fig. 3.

   Response: the figure reconstructed as you recommended.

3.Table 4 is the repeated data of Fig. 3, please delete one of the data. Besides, why connected different sites in Fig.3? I suggest the author to use bars instead of using dot-line plot.

Response: done. We reconstructed Figure 3 as the histogram as you recommend but left it because there can be seen the trend lines, that are used for conclusion.

 4.Table 5: The descriptions of abbreviations of “Hab” and “Sap” were lost.

Response: corrected

Comments on the Quality of English Language

Minor editing of English language needed.

Response: Thank you, done

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The paper reports on the diatom assemblages of macrophytic algae along the Israeli coast from the West Bank to Gaza. 25 sites (I could only count 23 on Fig. 1: improve detail please) were sampled once. A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast. The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis. Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

 

Abstract

Delete superfluous information.

Introduction

Delete irrelevant sections.

M & M

 

Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me. Some parts seem to be missing (where are the Pearson correlations?). There seems to be no purpose for the cluster analysis. The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

 

Discussion & conclusions

Structure and contents need considerable improvement.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Author Response

Dear Editor,

Thank you and the Reviewer 1 comments. Please find the responses below.

With best regards,

Prof Sophia Barinova

Corresponding author

 

Responses to the Reviewer 2 comments

 

Comments and Suggestions for Authors

The paper reports on the diatom assemblages of macrophytic algae along the Israeli coast from the West Bank to Gaza. 25 sites (I could only count 23 on Fig. 1: improve detail please) were sampled once.

Response: Table 1 gives the response for this comment. Sites not only named but also geographical coordinates are given for each sampling point. Let’s us please to correct your understanding the study area – sa it can be seen on the Figure 1 and Table 1, the studied points were located not in the continental part of this territory but only on the coastline between Rosh-HaNiqra and Ashkelon and in no case affected other continental territory, as you write “along the Israeli coast from the West Bank to Gaza”

 

A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast.

Response: As stated in the article, this is not a relatively unexplored coastline, but a completely unexplored coastline in terms of diatom diversity. Thank you for appreciating the floristic aspect of the article.

 

The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis.

Response: Thank you. Undoubtedly, the article provides only the first data; this aspect is expected to be studied later.

 

Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

Response: Thank you very much for your detailed comments. We will follow it.

 

Abstract

Delete superfluous information.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

 

Introduction

Delete irrelevant sections.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

 

M & M

 Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Response: Thank you, the article provides only primary data on the diversity of diatoms that have not previously been studied in this region, and the experimental approaches you propose will be applied later in detailed studies.

 

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me.

Response: Most likely, you do not divide our data into the number of species on the site and the rate of each species or we don’t clearly understand you. However, these are two different series of numbers. The first talks about the number of species on the site, the second about their abundance of their cells. The first reflects taxonomic diversity, and the second can be used to identify dominant species and calculate saprobity indices.

 

Some parts seem to be missing (where are the Pearson correlations?).

Response: Pearson correlations have been calculated and presented in the article for some parameters, reference cited under number 42.

 

There seems to be no purpose for the cluster analysis.

Response: Since cluster analysis was carried out for the entire species composition, but the tree turned out to have no sharp differences in the clusters, we consider this construction useful, since it allows us to draw a conclusion about the continuity of the species composition of communities along the studied coast. At the same time, if we include the number of species, abundance scores and chemical indicators in the analysis, then in Figure 8 we can see two main clusters, the defining indicator of which is species richness and abundance, and two sites - Mikmoret (harbor) and Ashkelon (harbor), remain individual.

 

The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

Response: Most likely, you are referring to the calculated saprobity indices in Table 6. This calculation includes all species that have the species-specific saprobity indices shown in Table 5. Thus, you can see exactly which and how many species are included in the calculation by looking at Appendix Table A 1. All data is open, but we do not seek to multiply repetitions of information by placing the main table in the Appendix.

 

Discussion & conclusions

Structure and contents need considerable improvement.

Response: We try to do it, thank you. We replaced some of the figures, rearranged them in a clearer format, added trend lines for parameter changes, and made corrections to the text.

 

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Response: Thank you very much for your detailed comments. We will follow it.

Comments in Attachment:

Unfortunately, the rationale for using a saprobic index for marine conditions is entirely based on studies that I cannot assess because they are in Russian and not easy to come by. So I will not comment on this.

Response: It is in English also, please give your attention to the references number 17 and 43.

Personally, I would be very wary to do so, but assuming that this would work, this still necessitates an estimation of relative abundance (perhaps also weighed by biovolumes, if you like) to apply weighted averaging of scores reliably - as in formula 1 - in a reproducible way. The traditional way to estimate hi requires counting a fairly large number of valves (in your case cells). Your abundance scores are only a crude reflection or relative abundance.

Response: The semi-quantitative scores of the cell’s abundance is represented for each species on the each sampling site in the Appendix Table A 1.

For instance, assume that you have only 2 taxa and both are 'frequent', does this mean their relative abundance is equal (e.g. 50 %) and that they carry 'the same amount of information', therefore should receive the same weight?

Response: This method does not use percentage; it is giving the scores of abundances only. You are right, the percentage is not usable because we can calculate the sum of scores for the sample, but never – the sum of percentage.

Note that scores 1 to 3 have undefined intervals. What with 7, 12 or 20 cells?

Response: as above, it is not relevant.

I assume that your ordinal scoring starts from probability-based reasoning, but then some additional explanation is required because the decision rules are not clear to me. To assess all taxa on a slide you have to scan it completely (which usually takes more time than actual counting a fixed number, so I cannot see much reason to use a much cruder approximation, anyway). The area of transects and fields of view varies with magnification... Is score 6 given for all the views, together covering the slide or only for the first one inspected (not plausible it seems, the first cell would always lead to 6)? The same for 4 and 5. How do you separate circular views from rectangular transects?

Response: we added x40 for the last line in Table 2. All other scores are as is.

The abundance scores also depend primarily on the amount of sample (i.e. total surface area of the thalli) used, the initial density of cells on the substrate, as well as the final dilution.. Without complete standardization of procedures you cannot add them up decently and make comparisons between samples regarding densities. A better way would be to work really quantitatively, e.g. using the number of cells on a fixed surface area  of the substrate, say 1 square cm.

Response: We avoid these problems when use the semi-quantitative method with the scores assessment. But in Future it possible to calculate the quantitative variables such as cells number per area and biomass per area if the experimental research will be done.

A sentence giving the most important taxa would be more informative than the nr of classes, etc.. Now it is necessary to consult the supplementary table to get an idea.

Response: Yes, we added the full list of diatoms for each sampling site because all researchers can see our advances. By the way, the mentioned taxonomic structure is a part of floristic analysis to which you recommended to concentrate our paper. The comparison to the similar diatom flora found in the north from our study area in Tartus is represent also part of floristic analysis – comparative floristic.

It seems to me that the only (somewhat) useful measure you have to compare with with regard to pollution or nutrients would be nitrate. I didn't read anything about this.

Response: Please read papers no 17 and 43 as well as: Barinova, S. 2017. On the Classification of Water Quality from an Ecological Point of View. International Journal of Environmental Sciences & Natural Resources, 2(2): 1-8. https://juniperpublishers.com/ijesnr/pdf/IJESNR.MS.ID.555581.pdf

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

While I agree with the authors' assessment that benthic marine diatoms have potential uses as bioindicators and that more extensive floristic work must be conducted in order to unlock this potential, there were several aspects of this manuscript that confounded me. Some of the conclusions and inferences appeared overly broad considering the only the epiphyton was sampled, and the utility of comparisons drawn in the Discussion were not always clear. The language impeded clarity in many places (which can be fixed), but I am also concerned about some valuable resources going unmentioned in this study, in particular Alvarez-Blanco & Blanco 2014 "Benthic diatoms from Mediterranean coasts" (Bibliotheca Diatomologica 60). I believe this, and other suggested resources, would greatly aid the authors in their diatom identification, as some of their illustrations were not always unambiguously identified.

I think the science overall is sound and useful, however, and I would encourage resubmission after significant edits and taxonomic evaluation.

Additional comments and suggested edits:

Abstract

“In this work, the composition, ecology, and phytogeography of diatoms of the macrophytes epiphyton are of epiphyton diatoms of the macrophytes is presented for the first time.”

“Altogether 85 diatom taxa were found among the epiphyton of 25 species of green brown and red macroalgae from the Israeli coast between March and May of 2021. These diatoms represent 3 classes, 17 orders, 26 families and 41 genera.”

“The distribution of diatoms are compared to that of other macrophytes and…”

“Ecological characteristics, relative abundance and dominant species were reported and statistically analyzed. Of all species recorded, 55% were considered marine species, 30% marine-brackish, 22% arctic-boreal-tropical, 29% cosmopolites and 12% widespread in their ranges.” Delete previous sentence.

“…arctic-boreal-tropical” This is an odd combination. I assume this means the proposed distribution was specific to either an arctic, boreal OR tropical habitat…not a species with a presence in all three.

“The Index of toxic influence on photosynthesis (WESI) and Index saprobity (S = 1.69-2.71) were compared to species richness and anthropogenic loads.” Delete previous sentence.

“The influence of the anthropogenic loads on the coastal territories…”

“…it was concluded that the section of the Israeli coast documented here is oligo-mesotrophic compared to…” Where is the Gulf of Tartus?

Introduction

“Mentioned that research has begun on important but little-studied groups: diatoms, cyanobacteria, fungi, and lichens.” I do not understand what the authors mean by this; are they stating that the bibliographic review specifically states that diatoms, cyanobacteria, fungi and lichens are understudied in the Mediterranean?

“…green (76) algae, with taxonomical and ecological approaches [2], but diatom microalgae were not included in this work.”

“…Mediterranean basin are given for specific areas: the port of Ischia is a volcanic island in the Tyrrhenian Sea, part of the Mediterranean Sea off the western coast of Italy..”

“…and ecology of algae in the phytobenthos, not only in fresh and brackish, but also marine waters, which are poorly studied in comparison.”

“In addressing this problem, the primary objective is to create marine macroalgae databases and checklists in different areas.” But the authors were just discussing the importance of microalgae (benthic diatoms)—shouldn’t that be the primary objective?

“At the same time, seaweed populations found in the eastern Mediterranean are known to have low biodiversity due to their oligotrophic environment [2], as evidenced by the chlorophyll content of the water [2425].” So, the assumption is that the benthic diatoms will follow the same pattern? What is the relevance of this statement to benthic diatoms as bioindicators?

Materials and Methods

“…at a depth of 0.3-0.5 m at the coordinates provided in Table 1. The collection of…”

“…measured at the sampling time using a HANNA HI 9813 Waterproof…”

“The GPS coordinates of sampling sites were determined with a GARNMIN GISMAP 64.”

Table 1: GPS coordinates should have units (degrees, minutes)

“…plastic tubes, fixed in 4% neutral formaldehyde, the same samples were not fixed for direct study and moved to the Institute of Evolution…” I’m not sure what the authors mean here—is it that an aliquot of the samples were taken and stored separately without fixative?

“Metadata associated with each sample was inserted into our database in Microsoft Access.”

“For each collected sample, the diatoms present were documented by microphotograph, while the macroalgae were measured and photographed using an OMAX 3D-500 digital camera Leica DM2500 (Leica Microsystems EMEA, United Kingdom) light microscope under 400–1000x magnification and photographed by Omax 9.0 MP USB Digital Camera.”

“Species were identified whenever possible, down to the genus or species level, based on established morphological features.” Please provide citations for the resources used to identify, at the very least, the diatoms presented here.

“To study the species composition of microalgae fouling of the different macrophytes from the Israeli coast of the Mediterranean Sea, dry samples were delivered by mail to the Department of Aquaculture and Marine Pharmacology Kovalevsky Institute of Biology of Southern Seas of Russian Academy of Sciences.” Presumably, this does not refer to the SEM surveys mentioned in the previous sentence, but rather, the protocol mentioned in the following paragraph?

“Algae samples were placed in containers and filled with distilled water in the Petri dish.” What Petri dish? The Petri dishes were not the containers in which algal samples were placed?

“The resulting washings were poured…”

“To identify diatom species literary sources were used, including Handbooks and Atlas [2829-3738].” An illustrated flora of diatoms from Britain and Japan hardly seems appropriate for the Israel coastline. I would recommend Witkowski et al. 2000 “Diatom Flora of Marine Coasts” (Iconographia Diatomologica 7), Alvarez-Blanco & Blanco 2014 “Benthic Diatoms from Mediterranean Coasts” (Bibliotheca Diatomologica 60).

Results

“In our sampling sites of the Israeli coast, the values of salinity ranged from 39 psu to 41 psu during the study period, whereas salinity in the Mediterranean Sea increased from west to east from 36 to 39.5 psu.”

“…period in all sites, while pH fluctuated from 7.1 to 8.0 depending on the place of study, and nitrate N-NO3 ranged from 0 – 3.9 mg/L (Table 4).”

“Can be seen that even the variable values are fluctuated from site to site, the trend lines directions are the same.” To what does this refer—the salinity?

“Altogether 85 species of pennate diatoms were found in the green, brown, and red algae epiphyton in the Israeli coastal waters of the Mediterranean Sea (Table 5, Appendix Table A 1, Appendix Figures B 1-B 4), representing 3 classes, 17 orders, 26 families and 41 genera. 94% of these were benthic forms.”

“…prevailed from all macrophytes…” What do the authors mean by “prevailed”? The green algae were the most abundant?

“In terms of phytogeographical distribution, diatoms identified here are found in almost all zones of the World Ocean, with 30% of species are cosmopolites and 21% were arctic-boreal-tropical (Table 5).” I presume this is what the authors mean, rather than that diatoms, in general, are found across the world’s oceans. I would be careful with any biogeographical assessment, however, as the resources used to identify diatoms were limited and might have biased the identification of species to those found in both the UK and Japan—the sources provided by the authors.

“Table 6 indicates that green algae macrophytes were the most common substrate for diatoms, hosting the highest number of species.” Word choice.

“Diatom species richness fluctuated from one in Bet Yanai (site 17) to 27 in Hadera Tal Afar (site 14) with the dominant species at each site listed in Table 7.”

“…Dan Panorama were also species rich, with 25 diatom species each.”

“…communities correlated with the activity of diatom growth, expressed by abundance scores…” I’m not sure what the authors mean by this. Diatom growth rates were not measured; how are the authors correlating this to abundance?

“The results are visible in Figure 4 where cannot be seen so large correlations such as species richness and abundance scores, but visually the Index saprobity often fluctuated similarly.” Again, I’m uncertain of what the authors are trying to express here.

Figure 4—I presume the “no species” line is an abbreviation for “number of species”, not “no species present.” Perhaps change the legend to “# species” or “no. species”

Table 7—for site 20, does “all sporadically” mean that there was no clear dominant taxon?

“The role of diatom species richness in known algae…” Nothing about Figure 5 says anything about the “role” diatoms play in these communities, only their species richness

“…where macrophyte Rhodophyta species were dominated and other macrophytes were presented really well but diatoms take only small part of algae communities because study of diatoms just started.” Again, I’m not sure what the authors mean by this. Presumably they’re stating that the Rhodophyte algae had the most number of species in the vast majority of the sites in comparison to the diatom species richness. It could be argued that this result has less to do with the paucity of diatom studies in the region and more about the fact that diatoms were only counted from the epiphyton at each site.

“In the same histogram, fluctuation of the Index saprobity (S) and anthropogenic load scores are visualized along with the species richness valves. Both variables appear correlated and tend to increase from North to South. We feel it is particularly remarkable that Index S was higher near the harbors on the coast, up to 3.0 on the sites at Haifa, Mikmoret, and Palmahim.” Please verify that these are the points that were intended to be made here.

“Index WESI, which reflects photosynthetic stress (Figure 6), was indicated as “below normal” in only a few sites; these sites were Rosh HaNiqra, Haifa Bat Galim Casino, Yam – Dor Habonim, Hadera, and Yam Evtah, most of which are located near harbors or touristic attractions. All these sites contain low species number and abundance, which might suggest that some toxic substances were present and stressed the diatom community. One of these substances may be an oil spill that occurred during the sampling period. The influence of oil as the main impact factor is supported by the fact that suppression of diatom communities was observed even at the Rosh HaNiqra site, where there is no anthropogenic impact, but we noted clots of oil on the coastal rocks.”

“The Bray-Curtis similarity tree of the species composition…” “…represents a certain continuum…” What do the authors mean by “certain continuum”? “…conditions in which epiphytic diatom communities are formed.” Remember that only the epiphyton was quantified in this study.

The authors keep referring to “abundance”. The location of these values (Appendix A) should be cited somewhere in the Results. This was quantified as categories of relative abundance mentioned in Table 2, correct?

“…Mikmoret (harbor) and Ashkelon (harbor), remain unlike the rest of the sites.”

Discussion

“This is especially true for benthic species leading a substrate-attached lifestyle.” The authors need to remember that there are many different substrate types in these habitats, and that they only quantified the epiphyton. Be careful making broad characterizations…

“During a cruise on the R/V «Akademik Kovalevsky» along the banks of the Aegean Sea, samples of the epiphyton of the green algae…”

I am curious why the authors are making so many references to depth and deeper collections in the Mediterranean, when the samples collected in this study were consistently shallow.

“When comparing the species composition of diatom benthos in deep and shallow water areas of the Egypt coast, 20 common species were noted. The difference in the composition and number of species can be explained by different habitats (season, depth, type of substrate, etc.) and sampling methods.” This line is repeated in two paragraphs.

“Diatom species Achnanthes brevipes, Licmophora abbreviata, Rhaphoneis nitida, found in the Israeli coastline were also found near the Suez Canal [9].” Please verify that these are the points that were intended to be made here.

“Assemblage 11 represents the Mediterranean samples…” Assemblage 11 from what? Zalat 2002?

“…are infrequently in some samples from the other sites.” Other sites where? In this study? In Zalat 2002? “Other sites” not in the Mediterranean?

“The Egyptian sector near Port Said of the Mediterranean Sea showed that dominated species of Bacillariophyta.” What do the authors mean by this?

“It has been established that the similarity in the composition of the diatom flora between the banks of the Aegean Sea and the Egyptian sector of the Mediterranean Sea is 31%” Percent similarity in species present? Genera present? Relative abundance?

The last paragraph of the Discussion is a seemingly random list of statements apparently summarizing the results of three studies, with very little apparent context or comparison to the results of this study.

Appendix Figure B1. b) there appears to be a stauros on the raphid valve (the consistent dot set against the raphe-bearing valve), which would suggest a species other than brevipes. Difficult to identify the other micrographs to species from these images.

Appendix Figure B2. Increase contrast in images. b-c) the sternum on the rapheless valve is eccentric, rather than central as it would be in A. longipes.

Appendix Figure B3. b-c) These are Grammatophora spp, not Hyalosira. The slit-like areolae on the valvocopula are diagnostic for Grammatophora. h) It is difficult to see the valve areolae in this specimen, which should be linear for L. abbreviata.

 

 

 

 

 

Comments on the Quality of English Language

See "Comments and Suggestions for Authors"

Author Response

Responses to Reviewer 3 Report 1

Dear Editor,

Thank you and the Reviewer 3 for comments.

Please find our responses to each comment and comment of comment below.

With best regards,

Prof Sophia Barinova,

Corresponding author.

 

Comments and Suggestions for Authors

While I agree with the authors' assessment that benthic marine diatoms have potential uses as bioindicators and that more extensive floristic work must be conducted in order to unlock this potential, there were several aspects of this manuscript that confounded me. Some of the conclusions and inferences appeared overly broad considering the only the epiphyton was sampled, and the utility of comparisons drawn in the Discussion were not always clear. The language impeded clarity in many places (which can be fixed), but I am also concerned about some valuable resources going unmentioned in this study, in particular Alvarez-Blanco & Blanco 2014 "Benthic diatoms from Mediterranean coasts" (Bibliotheca Diatomologica 60). I believe this, and other suggested resources, would greatly aid the authors in their diatom identification, as some of their illustrations were not always unambiguously identified.

I think the science overall is sound and useful, however, and I would encourage resubmission after significant edits and taxonomic evaluation.

Response: The authors are deeply grateful to Reviewer 3 for hard work on the our manuscript. All comments were taken into account and corrections were made in the text of the manuscript. In addition, we used the Alvarez-Blanco & Blanco 2014 "Benthic diatoms from Mediterranean coasts" and revised the identification of some diatoms as well as replaced and edited some microphotographs. For which we are very grateful for the valuable help of Reviewer 3.

Additional comments and suggested edits:

Abstract

In this work for the first-time species composition, ecology, and phytogeography of diatoms of the macrophytes epiphyton are presented

Response: corrected.

“In this work, the composition, ecology, and phytogeography of diatoms of the macrophytes epiphyton are presented for the first time.”

Response: corrected.

“Altogether 85 diatom taxa were found among the epiphyton of 25 species of green brown and red macroalgae from the Israeli coast between March and May of 2021. These diatoms represent 3 classes, 17 orders, 26 families and 41 genera.”

Response: corrected.

“The distribution of diatoms are compared to that of other macrophytes and…”

Response: corrected.

“Ecological characteristics, relative abundance and dominant species were reported and statistically analyzed. Of all species recorded, 55% were considered marine species, 30% marine-brackish, 22% arctic-boreal-tropical, 29% cosmopolites and 12% widespread in their ranges.” Delete previous sentence.

Response: corrected.

“…arctic-boreal-tropical” This is an odd combination. I assume this means the proposed distribution was specific to either an arctic, boreal OR tropical habitat…not a species with a presence in all three.

Response. An arctic-boreal-tropical species is a species that is listed in both tropical, temperate, and arctic climate zones, i.e., from the southern tropic to the north pole, according to the littoral data.

“The Index of toxic influence on photosynthesis (WESI) and Index saprobity (S = 1.69-2.71) were compared to species richness and anthropogenic loads.” Delete previous sentence.

Response: corrected.

“The influence of the anthropogenic loads on the coastal territories…”

Response: corrected.

“…it was concluded that the section of the Israeli coast documented here is oligo-mesotrophic compared to…” Where is the Gulf of Tartus?

Response: Because the diatom communities in the east side of Mediterranean Sea were studied in Tartus only. This is only 150 km north of our points 1 and 2.

Introduction

“Mentioned that research has begun on important but little-studied groups: diatoms, cyanobacteria, fungi, and lichens.” I do not understand what the authors mean by this; are they stating that the bibliographic review specifically states that diatoms, cyanobacteria, fungi and lichens are understudied in the Mediterranean?

Response: Yes, we cite information from this review that indicates references to initial studies of these groups, which are poorly studied and just started in the 80s on the Mediterranean Sea.

“…green (76) algae, with taxonomical and ecological approaches [2], but diatom microalgae were not included in this work.”

Response: corrected.

Mediterranean basin are given for the different its areas: port of Ischia is a volcanic island in the Tyrrhenian Sea is part of the Mediterranean Sea off the western coast of Italy

Response: corrected.

“…and ecology of algae in the phytobenthos, not only in fresh and brackish, but also marine waters, which are poorly studied in comparison.”

Response: corrected.

“In addressing this problem, the primary objective is to create marine macroalgae databases and checklists in different areas.” But the authors were just discussing the importance of microalgae (benthic diatoms)—shouldn’t that be the primary objective?

Response: You're absolutely right. It was a typo. We've corrected it. We apologise for that.

“At the same time, seaweed populations found in the eastern Mediterranean are known to have low biodiversity due to their oligotrophic environment [2], as evidenced by the chlorophyll content of the water [2425].” So, the assumption is that the benthic diatoms will follow the same pattern? What is the relevance of this statement to benthic diatoms as bioindicators?

Response: benthic diatoms as bioindicators can show the trophic state

Materials and Methods

“…at a depth of 0.3-0.5 m at the coordinates provided in Table 1. The collection of…”

Response: corrected.

“…measured at the sampling time using a HANNA HI 9813 Waterproof…”

Response: corrected.

“The GPS coordinates of sampling sites were determined with a GARNMIN GISMAP 64.”

Response: corrected.

Table 1: GPS coordinates should have units (degrees, minutes)

Response: corrected.

 

“…plastic tubes, fixed in 4% neutral formaldehyde, the same samples were not fixed for direct study and moved to the Institute of Evolution…” I’m not sure what the authors mean here—is it that an aliquot of the samples were taken and stored separately without fixative?

Response: An aliquots of the samples were taken without fixative for direct live algae study and for. visual diversity assessment.

“Metadata associated with each sample was inserted into our database in Microsoft Access.”

Response: corrected.

“For each collected sample, the diatoms present were documented by microphotograph, while the macroalgae were measured and photographed using an OMAX 3D-500 digital camera Leica DM2500 (Leica Microsystems EMEA, United Kingdom) light microscope under 400–1000x magnification and photographed by Omax 9.0 MP USB Digital Camera.”

 Response: corrected.

“Species were identified whenever possible, down to the genus or species level, based on established morphological features.” Please provide citations for the resources used to identify, at the very least, the diatoms presented here.

Response: The sentence has moved below, where the identifiers are referenced.

“To study the species composition of microalgae fouling of the different macrophytes from the Israeli coast of the Mediterranean Sea, dry samples were delivered by mail to the Department of Aquaculture and Marine Pharmacology Kovalevsky Institute of Biology of Southern Seas of Russian Academy of Sciences.” Presumably, this does not refer to the SEM surveys mentioned in the previous sentence, but rather, the protocol mentioned in the following paragraph?

Response: This sentence is paraphrased.

“The resulting washings were poured…”“Algae samples were placed in containers and filled with distilled water in the Petri dish.” What Petri dish? The Petri dishes were not the containers in which algal samples were placed?

Response: This sentence is paraphrased.

“To identify diatom species literary sources were used, including Handbooks and Atlas [2829-3738].” An illustrated flora of diatoms from Britain and Japan hardly seems appropriate for the Israel coastline. I would recommend Witkowski et al. 2000 “Diatom Flora of Marine Coasts” (Iconographia Diatomologica 7), Alvarez-Blanco & Blanco 2014 “Benthic Diatoms from Mediterranean Coasts” (Bibliotheca Diatomologica 60).

Response: We've added references to the following sources: Witkowski et al. 2000, Alvarez-Blanco & Blanco 2014. We apologise for this omission.

Results

“In our sampling sites of the Israeli coast, the values of salinity ranged from 39 psu to 41 psu during the study period, whereas salinity in the Mediterranean Sea increased from west to east from 36 to 39.5 psu.”

Response: corrected.

“…period in all sites, while pH fluctuated from 7.1 to 8.0 depending on the place of study, and nitrate N-NO3 ranged from 0 – 3.9 mg/L (Table 4).”

Response: corrected.

“Can be seen that even the variable values are fluctuated from site to site, the trend lines directions are the same.” To what does this refer—the salinity?

Response: It applies to all variables mentioned on Figure 3 (nitrates, pH, salinity, and conductivity) The sentence corrected.

“Altogether 85 species of pennate diatoms were found in the green, brown, and red algae epiphyton in the Israeli coastal waters of the Mediterranean Sea (Table 5, Appendix Table A 1, Appendix Figures B 1-B 4), representing 3 classes, 17 orders, 26 families and 41 genera. 94% of these were benthic forms.”

Response: corrected.

“…prevailed from all macrophytes…” What do the authors mean by “prevailed”? The green algae were the most abundant?

Response: The green algae were the most abundant in the Israeli coastal waters. This sentence is paraphrased.

“In terms of phytogeographical distribution, diatoms identified here are found in almost all zones of the World Ocean, with 30% of species are cosmopolites and 21% were arctic-boreal-tropical (Table 5).” I presume this is what the authors mean, rather than that diatoms, in general, are found across the world’s oceans. I would be careful with any biogeographical assessment, however, as the resources used to identify diatoms were limited and might have biased the identification of species to those found in both the UK and Japan—the sources provided by the authors.

Response: This sentence is paraphrased. Here we present data on phytogeographical characteristics of species taken from literature and from Section Detailed Distribution with Sources in Algaebase.

“Table 6 indicates that green algae macrophytes were the most common substrate for diatoms, hosting the highest number of species.” Word choice.

Response: corrected.

“Diatom species richness fluctuated from one in Bet Yanai (site 17) to 27 in Hadera Tal Afar (site 14) with the dominant species at each site listed in Table 7.”

Response: corrected.

“…Dan Panorama were also species rich, with 25 diatom species each.”

Response: corrected.

“…communities correlated with the activity of diatom growth, expressed by abundance scores…” I’m not sure what the authors mean by this. Diatom growth rates were not measured; how are the authors correlating this to abundance?

Response: corrected, “Diatom growth rates” were deleted.

 

“The results are visible in Figure 4 where cannot be seen so large correlations such as species richness and abundance scores, but visually the Index saprobity often fluctuated similarly.” Again, I’m uncertain of what the authors are trying to express here.

Response: This sentence is rephrased.

Figure 4—I presume the “no species” line is an abbreviation for “number of species”, not “no species present.” Perhaps change the legend to “# species” or “no. species”

Response: corrected

Table 7—for site 20, does “all sporadically” mean that there was no clear dominant taxon?

Response: Yes, there was no clear dominant taxon and all species were found sporadically.

“The role of diatom species richness in known algae…” Nothing about Figure 5 says anything about the “role” diatoms play in these communities, only their species richness

Response: Word “role” is replaced by “contribution”.

“…where macrophyte Rhodophyta species were dominated and other macrophytes were presented really well but diatoms take only small part of algae communities because study of diatoms just started.” Again, I’m not sure what the authors mean by this. Presumably they’re stating that the Rhodophyte algae had the most number of species in the vast majority of the sites in comparison to the diatom species richness. It could be argued that this result has less to do with the paucity of diatom studies in the region and more about the fact that diatoms were only counted from the epiphyton at each site.

Response: corrected

“In the same histogram, fluctuation of the Index saprobity (S) and anthropogenic load scores are visualized along with the species richness valves. Both variables appear correlated and tend to increase from North to South. We feel it is particularly remarkable that Index S was higher near the harbors on the coast, up to 3.0 on the sites at Haifa, Mikmoret, and Palmahim.” Please verify that these are the points that were intended to be made here.

Response: added, corrected

“Index WESI, which reflects photosynthetic stress (Figure 6), was indicated as “below normal” in only a few sites; these sites were Rosh HaNiqra, Haifa Bat Galim Casino, Yam – Dor Habonim, Hadera, and Yam Evtah, most of which are located near harbors or touristic attractions. All these sites contain low species number and abundance, which might suggest that some toxic substances were present and stressed the diatom community. One of these substances may be an oil spill that occurred during the sampling period. The influence of oil as the main impact factor is supported by the fact that suppression of diatom communities was observed even at the Rosh HaNiqra site, where there is no anthropogenic impact, but we noted clots of oil on the coastal rocks.”

Response: corrected.

“The Bray-Curtis similarity tree of the species composition…” “…represents a certain continuum…” What do the authors mean by “certain continuum”? “…conditions in which epiphytic diatom communities are formed.” Remember that only the epiphyton was quantified in this study.

Response: Clarification is made in this sentence that it is the epiphyton communities that are being referred to.

The authors keep referring to “abundance”. The location of these values (Appendix A) should be cited somewhere in the Results. This was quantified as categories of relative abundance mentioned in Table 2, correct?

Response: Reference to Appendix Table A 1 added. Abundance scores were quantified using categories of relative abundance from Table 2.

“…Mikmoret (harbor) and Ashkelon (harbor), remain unlike the rest of the sites.”

Response: corrected.

Discussion

“This is especially true for benthic species leading a substrate-attached lifestyle.” The authors need to remember that there are many different substrate types in these habitats, and that they only quantified the epiphyton. Be careful making broad characterizations…

Response: This statement applies to all benthic diatoms, especially those leading an attached lifestyle on any substrate, including epiphyton.

“During a cruise on the R/V «Akademik Kovalevsky» along the banks of the Aegean Sea, samples of the epiphyton of the green algae…”

Response: corrected.

I am curious why the authors are making so many references to depth and deeper collections in the Mediterranean, when the samples collected in this study were consistently shallow.

Response: We wanted to point to common species in different areas and depths of the Mediterranean Sea, which may be indicative of the typicality of these species in the sea.

“When comparing the species composition of diatom benthos in deep and shallow water areas of the Egypt coast, 20 common species were noted. The difference in the composition and number of species can be explained by different habitats (season, depth, type of substrate, etc.) and sampling methods.” This line is repeated in two paragraphs.

Response: corrected.

“Diatom species Achnanthes brevipes, Licmophora abbreviata, Rhaphoneis nitida, found in the Israeli coastline were also found near the Suez Canal [9].” Please verify that these are the points that were intended to be made here.

Response: This sentence is paraphrased. Rhaphoneis nitida is deleted.

“Assemblage 11 represents the Mediterranean samples…” Assemblage 11 from what? Zalat 2002?

Response: Yes, it is from Zalat 2002. This sentence is rephrased.

“…are infrequently in some samples from the other sites.” Other sites where? In this study? In Zalat 2002? “Other sites” not in the Mediterranean?

Response: That phrase is deleted.

“The Egyptian sector near Port Said of the Mediterranean Sea showed that dominated species of Bacillariophyta.” What do the authors mean by this?

Response: This sentence is deleted.

“It has been established that the similarity in the composition of the diatom flora between the banks of the Aegean Sea and the Egyptian sector of the Mediterranean Sea is 31%” Percent similarity in species present? Genera present? Relative abundance?

Response: Here is a comparison of species composition by presence/absence of species.

The last paragraph of the Discussion is a seemingly random list of statements apparently summarizing the results of three studies, with very little apparent context or comparison to the results of this study.

Response: This paragraph is edited.

Appendix Figure B1. b) there appears to be a stauros on the raphid valve (the consistent dot set against the raphe-bearing valve), which would suggest a species other than brevipes. Difficult to identify the other micrographs to species from these images.

Response: The light photos are presented in the manuscript mainly to give the reader an image of diatom inhabiting of macroalgae (photos a-c) and of their life state. In addition, as it is known, the shape of cells and the number and arrangement of chloroplasts are important for the identification of diatom species, in addition to the structure of their frustules. As for photo b), the consistent dot set against the raphe-bearing valve is a central fascia, which is characteristic of the species Achnanthes brevipes

Appendix Figure B2. Increase contrast in images. b-c) the sternum on the rapheless valve is eccentric, rather than central as it would be in A. longipes.

Response: Images edited. You're absolutely right about Achnanthes images. We apologize for this mistake. Now, only Achnantes brevipes is presented in images a-c.

Appendix Figure B3. b-c) These are Grammatophora spp, not Hyalosira. The slit-like areolae on the valvocopula are diagnostic for Grammatophora. h) It is difficult to see the valve areolae in this specimen, which should be linear for L. abbreviata.

Response: As for Grammatophora, we agree with you completely and have corrected our mistake. For L. abbreviata we added image of valve fragment, where the areolae are clearly visible.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

I am satisfied with the revisions. 

Author Response

Dear Editor,

Thank you and the Reviewer 1 for comments.

Please find our responses to each comment and comment of comment below.

With best regards,

Prof Sophia Barinova,

Corresponding author.

 

Comments and Suggestions for Authors

I am satisfied with the revisions. 

RESPONSE: Thank you very much.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Prof. Barinova, herewith (in bold) my replies to your responses, ordered as previously.

The paper reports on the diatom assemblages of macrophytic algae along the Israeli coast from the West Bank to Gaza. 25 sites (I could only count 23 on Fig. 1: improve detail please) were sampled once.

Response: Table 1 gives the response for this comment. Sites not only named but also geographical coordinates are given for each sampling point. Let’s us please to correct your understanding the study area – sa it can be seen on the Figure 1 and Table 1, the studied points were located not in the continental part of this territory but only on the coastline between Rosh-HaNiqra and Ashkelon and in no case affected other continental territory, as you write “along the Israeli coast from the West Bank to Gaza”

Sorry, a geographic slip, apparently. I was fully aware that these are only coastal samples; not West Bank, meant Lebanon of course… (that’s what you get with all the news). The main point is the quality of the map, only 23 dots visible not 25.

A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast.

Response: As stated in the article, this is not a relatively unexplored coastline, but a completely unexplored coastline in terms of diatom diversity. Thank you for appreciating the floristic aspect of the article.

'Relative' with regard to other parts of the Med. and in general.

 

The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis.

Response: Thank you. Undoubtedly, the article provides only the first data; this aspect is expected to be studied later.

If I read 'to identify indicator properties of Israeli aquatic flora that can be used to monitor the quality of coastal waters', that means more than just an inventory of taxa.

Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I stand with this comment.

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

Response: Thank you very much for your detailed comments. We will follow it.

Abstract

Delete superfluous information.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Still includes unsubstantiated conclusions. Decision to be made by editor as I have no access to other reviews.

 

Introduction

 

Delete irrelevant sections.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

 

Sorry, I can’t agree. Partly irrelevant in my opinion. Decision to be made by editor.

M & M

Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Response: Thank you, the article provides only primary data on the diversity of diatoms that have not previously been studied in this region, and the experimental approaches you propose will be applied later in detailed studies.

There’s nothing experimental proposed, only more reliable and reproducible methodology. Identification of live diatom cells at species level, particularly without prior knowledge of assemblage composition, is not recommendable for floristic studies. Only permanent slides of oxidised material allow adequate observation of diagnostic features and additionally serve as a repository for later study and reference.

 

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me.

Response: Most likely, you do not divide our data into the number of species on the site and the rate of each species or we don’t clearly understand you. However, these are two different series of numbers. The first talks about the number of species on the site, the second about their abundance of their cells. The first reflects taxonomic diversity, and the second can be used to identify dominant species and calculate saprobity indices.

Sorry, there was no misunderstanding involved. I just don’t trust your species numbers.

 

Some parts seem to be missing (where are the Pearson correlations?).

Response: Pearson correlations have been calculated and presented in the article for some parameters, reference cited under number 42.

I expected (at the least) correlations for other pairs of variables than those in figure 3 and sample order. I cannot corroborate your 'visual' interpretations.

 

The only relation evident from alternative figure 3 is for EC (and still unexpected to me – and remaining unexplained - not for salinity), which is actually even more complex than just linear.

There seems to be no purpose for the cluster analysis.

 

Response: Since cluster analysis was carried out for the entire species composition, but the tree turned out to have no sharp differences in the clusters, we consider this construction useful, since it allows us to draw a conclusion about the continuity of the species composition of communities along the studied coast. At the same time, if we include the number of species, abundance scores and chemical indicators in the analysis, then in Figure 8 we can see two main clusters, the defining indicator of which is species richness and abundance, and two sites - Mikmoret (harbor) and Ashkelon (harbor), remain individual.

I have read nothing about this in your discussion. Figure 7: chaining is typical for single linkage clustering. Figure 8: it’s really not done to mix variables that are inherently different and with different scales in this type of analysis; there are appropriate methods, however.

The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

Response: Most likely, you are referring to the calculated saprobity indices in Table 6. This calculation includes all species that have the species-specific saprobity indices shown in Table 5. Thus, you can see exactly which and how many species are included in the calculation by looking at Appendix Table A 1. All data is open, but we do not seek to multiply repetitions of information by placing the main table in the Appendix.

 

Sorry, you cannot ask the reader to do these basic calculations themselves.

Discussion & conclusions

Structure and contents need considerable improvement.

Response: We try to do it, thank you. We replaced some of the figures, rearranged them in a clearer format, added trend lines for parameter changes, and made corrections to the text.

I cannot discern appreciable improvement.

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Response: Thank you very much for your detailed comments. We will follow it.

 

Comments in Attachment:

Unfortunately, the rationale for using a saprobic index for marine conditions is entirely based on studies that I cannot assess because they are in Russian and not easy to come by. So I will not comment on this.

Response: It is in English also, please give your attention to the references number 17 and 43.

Sorry, these publications offer no insight into the basis of saprobity scores for marine taxa, or the validation of these metrics for marine conditions – that is, do they relate significantly to whatever they are assumed to indicate. Again no comment on the method itself, but why calculate something which may not be relevant (and for which relevance is not indicated by results presented here, either).

Personally, I would be very wary to do so, but assuming that this would work, this still necessitates an estimation of relative abundance (perhaps also weighed by biovolumes, if you like) to apply weighted averaging of scores reliably - as in formula 1 - in a reproducible way. The traditional way to estimate hi requires counting a fairly large number of valves (in your case cells). Your abundance scores are only a crude reflection or relative abundance.

Response: The semi-quantitative scores of the cell’s abundance is represented for each species on the each sampling site in the Appendix Table A 1.

Although it can be done, it is not customary to use ordinal data in WA.

For instance, assume that you have only 2 taxa and both are 'frequent', does this mean their relative abundance is equal (e.g. 50 %) and that they carry 'the same amount of information', therefore should receive the same weight?

Response: This method does not use percentage; it is giving the scores of abundances only. You are right, the percentage is not usable because we can calculate the sum of scores for the sample, but never – the sum of percentage.

Indeed, it’s much more crude. I assume that you don’t want to count because it takes some time, but in that case why not use ‘simultaneous observation’, instead of mixing slides, transects and fields?

 

Note that scores 1 to 3 have undefined intervals. What with 7, 12 or 20 cells?

 

Don’t understand. There are numbers that cannot be classified (6 to 9, 16 to 24 cells)… Seems relevant to me.

I assume that your ordinal scoring starts from probability-based reasoning, but then some additional explanation is required because the decision rules are not clear to me. To assess all taxa on a slide you have to scan it completely (which usually takes more time than actual counting a fixed number, so I cannot see much reason to use a much cruder approximation, anyway). The area of transects and fields of view varies with magnification... Is score 6 given for all the views, together covering the slide or only for the first one inspected (not plausible it seems, the first cell would always lead to 6)? The same for 4 and 5. How do you separate circular views from rectangular transects?

Response: we added x40 for the last line in Table 2. All other scores are as is.

Doesn't clarify much to me. Even more surprising that you are able to identify more than a few uncleaned diatoms at 400x in water – I couldn’t.

 

The abundance scores also depend primarily on the amount of sample (i.e. total surface area of the thalli) used, the initial density of cells on the substrate, as well as the final dilution.. Without complete standardization of procedures you cannot add them up decently and make comparisons between samples regarding densities. A better way would be to work really quantitatively, e.g. using the number of cells on a fixed surface area of the substrate, say 1 square cm.

Response: We avoid these problems when use the semi-quantitative method with the scores assessment. But in Future it possible to calculate the quantitative variables such as cells number per area and biomass per area if the experimental research will be done.

Your method ranks taxa from less to more abundant relative to each other, but the density of cells on a slide and therefore the individual scores (as well as their sum) depends on their concentration in the suspension you apply. Without standardisation, the concentrations are not comparable between samples. Why would one ‘six’ at a site be equivalent to six ‘ones’ at another?

A sentence giving the most important taxa would be more informative than the nr of classes, etc.. Now it is necessary to consult the supplementary table to get an idea.

Response: Yes, we added the full list of diatoms for each sampling site because all researchers can see our advances. By the way, the mentioned taxonomic structure is a part of floristic analysis to which you recommended to concentrate our paper. The comparison to the similar diatom flora found in the north from our study area in Tartus is represent also part of floristic analysis – comparative floristic.

There is no need for an additional table (info on prevalence could btw also be easily included in Table 5).

It seems to me that the only (somewhat) useful measure you have to compare with with regard to pollution or nutrients would be nitrate. I didn't read anything about this.

Response: Please read papers no 17 and 43 as well as: Barinova, S. 2017. On the Classification of Water Quality from an Ecological Point of View. International Journal of Environmental Sciences & Natural Resources, 2(2): 1-8. https://juniperpublishers.com/ijesnr/pdf/IJESNR.MS.ID.555581.pdf

I meant that this is not covered at all in results or discussion.

Also, still unclear how you arrive at ‘scores for anthropogenic loading’ following Einav and Israel 2008 Checklist of seaweeds from the Israeli Mediterranean: Taxonomical and ecological approaches. Israel Journal of Plant Sciences 56: 127–184 DOI: 10.1560/IJPS.57.1–2.127. They do not mention anything like this.

Also, I missed replies on some comments.

An additional question. You added this was 'when water temperature and solar activity increase'. Suggests that assemblages might just be particularly unstable at this time of year.

Superfluous keywords still present and why vertical lines in figure 4?

 

 

 

Comments on the Quality of English Language

Not improved adequately.

Author Response

Dear Editor,

Thank you and the Reviewer 2 for comments.

Please find our responses to each comment and comment of comment below.

With best regards,

Prof Sophia Barinova,

Corresponding author.

 

Comments and Suggestions for Authors

Dear Prof. Barinova, herewith (in bold) my replies to your responses, ordered as previously.

The paper reports on the diatom assemblages of macrophytic algae along the Israeli coast from the West Bank to Gaza. 25 sites (I could only count 23 on Fig. 1: improve detail please) were sampled once.

Response: Table 1 gives the response for this comment. Sites not only named but also geographical coordinates are given for each sampling point. Let’s us please to correct your understanding the study area – sa it can be seen on the Figure 1 and Table 1, the studied points were located not in the continental part of this territory but only on the coastline between Rosh-HaNiqra and Ashkelon and in no case affected other continental territory, as you write “along the Israeli coast from the West Bank to Gaza”

Sorry, a geographic slip, apparently. I was fully aware that these are only coastal samples; not West Bank, meant Lebanon of course… (that’s what you get with all the news). The main point is the quality of the map, only 23 dots visible not 25.

RESPONSE: I understood your geographical mistakes. I understand your bias, but I am not guided by the news. Let me also ask you to give your attention to Table 1 where are mentioned the GIS coordinates of each sampling point and you can see that some of them are located very close to each other, so same points are doubled on the map of this resolution. But there are 25 points in any case. However, to clear up any confusion, we are replacing Figure 1 with one with more resolution and sampling points numbered.

 

A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast.

Response: As stated in the article, this is not a relatively unexplored coastline, but a completely unexplored coastline in terms of diatom diversity. Thank you for appreciating the floristic aspect of the article.

'Relative' with regard to other parts of the Med. and in general.

RESPONSE: OK I understood. The other parts of the Mediterranean diatom diversity are mentioned in the Discussion part.

 

The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis.

Response: Thank you. Undoubtedly, the article provides only the first data; this aspect is expected to be studied later.

If I read 'to identify indicator properties of Israeli aquatic flora that can be used to monitor the quality of coastal waters', that means more than just an inventory of taxa.

RESPONSE: Yes, of course, it means that in the revealed diversity will be found an indicator species to diverse of environmental variables and we only on the first step of this way, expected to be studied later.

 

Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I stand with this comment.

RESPONSE: The rationale for the conclusions follows from many details. We believe it is the scientific method to look at the details and summarize them into conclusions. English was subject to improvement.

 

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

Response: Thank you very much for your detailed comments. We will follow it.

Abstract

Delete superfluous information.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Still includes unsubstantiated conclusions. Decision to be made by editor as I have no access to other reviews.

RESPONSE: Unfortunately, I did not find any unfounded conclusions. It is detailed handling of the material that provides the basis for conclusions both in the taxonomic and floristic aspect, and from the point of view of interaction with the environment, which is given in the Conclusions and in the Abstract

 

Introduction

 

Delete irrelevant sections.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

 

Sorry, I can’t agree. Partly irrelevant in my opinion. Decision to be made by editor.

RESPONSE: We edited the text, taking into account your advice

M & M

Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Response: Thank you, the article provides only primary data on the diversity of diatoms that have not previously been studied in this region, and the experimental approaches you propose will be applied later in detailed studies.

There’s nothing experimental proposed, only more reliable and reproducible methodology. Identification of live diatom cells at species level, particularly without prior knowledge of assemblage composition, is not recommendable for floristic studies. Only permanent slides of oxidised material allow adequate observation of diagnostic features and additionally serve as a repository for later study and reference.

RESPONSE: As described in the MM, diatoms were identified as living material only at the first stage of the study, then the material was fixed, species identification was made in permanent preparations, then the cleaned valves were examined in an electron microscope, the results are given in the Appendix. It was not difficult for us to make this determination, since this is our many years of professional practice. On the other hand, the process of establishing the species composition in a community goes exactly like this, first identifying species, then generalizing them as the species composition of the community, but not vice versa, as you recommend. We even wonder where knowledge about the initial composition of a community can come from if its species are not determined. Permanent slide surveys are an experimental work and then, according to our many years of experience, species that are not typical for natural communities can gain an advantage. Here we present only natural communities, as all florists usually do, and not by doing an experiment.

 

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me.

Response: Most likely, you do not divide our data into the number of species on the site and the rate of each species or we don’t clearly understand you. However, these are two different series of numbers. The first talks about the number of species on the site, the second about their abundance of their cells. The first reflects taxonomic diversity, and the second can be used to identify dominant species and calculate saprobity indices.

Sorry, there was no misunderstanding involved. I just don’t trust your species numbers.

RESPONSE: Our material is identified by basic, detailed, and generally accepted methods, handbooks are cited, and photographs of species are provided. If you do not trust our definitions, you can indicate where exactly, which photos show erroneous definitions.

 

Some parts seem to be missing (where are the Pearson correlations?).

Response: Pearson correlations have been calculated and presented in the article for some parameters, reference cited under number 42.

I expected (at the least) correlations for other pairs of variables than those in figure 3 and sample order. I cannot corroborate your 'visual' interpretations.

RESPONSE: Correlation coefficients for other pairs of parameters were calculated (the table is available in added Appendix A Table A 2), however, they turned out to be not significant, so they were not presented in the article. It is proposed to only look at general trends.

 

The only relation evident from alternative figure 3 is for EC (and still unexpected to me – and remaining unexplained - not for salinity), which is actually even more complex than just linear.

RESPONSE: Yes, you are right. More significant correlation looks like with EC, correlarion Table added.

 

There seems to be no purpose for the cluster analysis.

 

Response: Since cluster analysis was carried out for the entire species composition, but the tree turned out to have no sharp differences in the clusters, we consider this construction useful, since it allows us to draw a conclusion about the continuity of the species composition of communities along the studied coast. At the same time, if we include the number of species, abundance scores and chemical indicators in the analysis, then in Figure 8 we can see two main clusters, the defining indicator of which is species richness and abundance, and two sites - Mikmoret (harbor) and Ashkelon (harbor), remain individual.

I have read nothing about this in your discussion. Figure 7: chaining is typical for single linkage clustering. Figure 8: it’s really not done to mix variables that are inherently different and with different scales in this type of analysis; there are appropriate methods, however.

RESPONSE: Two analyzes using the same method were presented on a different basis - community composition in Figure 7 and indicator group composition and environmental parameters in Figure 8. Significant differences are visible and are described below.

 

The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

Response: Most likely, you are referring to the calculated saprobity indices in Table 6. This calculation includes all species that have the species-specific saprobity indices shown in Table 5. Thus, you can see exactly which and how many species are included in the calculation by looking at Appendix Table A 1. All data is open, but we do not seek to multiply repetitions of information by placing the main table in the Appendix.

 

Sorry, you cannot ask the reader to do these basic calculations themselves.

RESPONSE: We have already performed these basic calculations and presented the results. The specific phrase in the previous answer was intended solely for you, so that you could see the transparency of the analysis done.

 

Discussion & conclusions

Structure and contents need considerable improvement.

Response: We try to do it, thank you. We replaced some of the figures, rearranged them in a clearer format, added trend lines for parameter changes, and made corrections to the text.

I cannot discern appreciable improvement.

RESPONSE: If you can follow the changes in the track changes, it will become clear what exactly has been improved or replaced. A lot of them.

 

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Response: Thank you very much for your detailed comments. We will follow it.

 

Comments in Attachment:

Unfortunately, the rationale for using a saprobic index for marine conditions is entirely based on studies that I cannot assess because they are in Russian and not easy to come by. So I will not comment on this.

Response: It is in English also, please give your attention to the references number 17 and 43.

Sorry, these publications offer no insight into the basis of saprobity scores for marine taxa, or the validation of these metrics for marine conditions – that is, do they relate significantly to whatever they are assumed to indicate. Again no comment on the method itself, but why calculate something which may not be relevant (and for which relevance is not indicated by results presented here, either).

RESPONSE: Since saprobity indices show only the degree of organic pollution, this has nothing to do with the salinity of the water, but only with organic pollution. Therefore, in the article and in the additional publication we provided you with links to this generally accepted method, obviously, after reading which you could understand why and on what basis this is done. Yes, you are right, for species from the marine environment, species-specific saprobity indices are not yet sufficiently known, however, we consider our work useful, since we supplement the ecological properties of the identified diatom species using data known for indicators of organic pollution.

 

Personally, I would be very wary to do so, but assuming that this would work, this still necessitates an estimation of relative abundance (perhaps also weighed by biovolumes, if you like) to apply weighted averaging of scores reliably - as in formula 1 - in a reproducible way. The traditional way to estimate hi requires counting a fairly large number of valves (in your case cells). Your abundance scores are only a crude reflection or relative abundance.

Response: The semi-quantitative scores of the cell’s abundance is represented for each species on the each sampling site in the Appendix Table A 1.

Although it can be done, it is not customary to use ordinal data in WA.

RESPONSE: We can cite hundreds of publications where semi-quantitative methods for calculating indices are used. However, the main ones have already been cited in the article and, apparently, there is no need to multiply the citations, thereby simply increasing the list of references used.

 

For instance, assume that you have only 2 taxa and both are 'frequent', does this mean their relative abundance is equal (e.g. 50 %) and that they carry 'the same amount of information', therefore should receive the same weight?

Response: This method does not use percentage; it is giving the scores of abundances only. You are right, the percentage is not usable because we can calculate the sum of scores for the sample, but never – the sum of percentage.

Indeed, it’s much more crude. I assume that you don’t want to count because it takes some time, but in that case why not use ‘simultaneous observation’, instead of mixing slides, transects and fields?

RESPONSE: There is no mixing. We replaced Table 2 where were some misprints, so it can be clear for the assessment of scores. The given method of semi-quantitative analysis is widely used. We have added here one more reference, however, we consider it unproductive to multiply quotes:

Alster, A., Kaplan-Levy, R.N., Barinova, S.S., Zohary, T. Analyzing semiquantitative phytoplankton counts. Hydrobiologia, 2023. https://doi.org/10.1007/s10750-023-05391-4.

 

Note that scores 1 to 3 have undefined intervals. What with 7, 12 or 20 cells?

 

Don’t understand. There are numbers that cannot be classified (6 to 9, 16 to 24 cells)… Seems relevant to me.

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18.

 

I assume that your ordinal scoring starts from probability-based reasoning, but then some additional explanation is required because the decision rules are not clear to me. To assess all taxa on a slide you have to scan it completely (which usually takes more time than actual counting a fixed number, so I cannot see much reason to use a much cruder approximation, anyway). The area of transects and fields of view varies with magnification... Is score 6 given for all the views, together covering the slide or only for the first one inspected (not plausible it seems, the first cell would always lead to 6)? The same for 4 and 5. How do you separate circular views from rectangular transects?

Response: we added x40 for the last line in Table 2. All other scores are as is.

Doesn't clarify much to me. Even more surprising that you are able to identify more than a few uncleaned diatoms at 400x in water – I couldn’t.

RESPONSE: We can because we used the permanent slide methods as well as SEM method. The calculation of the cells number is followed by species definition as given in MM but not vice versa.

 

The abundance scores also depend primarily on the amount of sample (i.e. total surface area of the thalli) used, the initial density of cells on the substrate, as well as the final dilution.. Without complete standardization of procedures you cannot add them up decently and make comparisons between samples regarding densities. A better way would be to work really quantitatively, e.g. using the number of cells on a fixed surface area of the substrate, say 1 square cm.

Response: We avoid these problems when use the semi-quantitative method with the scores assessment. But in Future it possible to calculate the quantitative variables such as cells number per area and biomass per area if the experimental research will be done.

Your method ranks taxa from less to more abundant relative to each other, but the density of cells on a slide and therefore the individual scores (as well as their sum) depends on their concentration in the suspension you apply. Without standardisation, the concentrations are not comparable between samples. Why would one ‘six’ at a site be equivalent to six ‘ones’ at another?

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18. It is standard – one drop of the sediment from fixed sample as given in addition ref 18.

 

A sentence giving the most important taxa would be more informative than the nr of classes, etc.. Now it is necessary to consult the supplementary table to get an idea.

Response: Yes, we added the full list of diatoms for each sampling site because all researchers can see our advances. By the way, the mentioned taxonomic structure is a part of floristic analysis to which you recommended to concentrate our paper. The comparison to the similar diatom flora found in the north from our study area in Tartus is represent also part of floristic analysis – comparative floristic.

There is no need for an additional table (info on prevalence could btw also be easily included in Table 5).

RESPONSE: Table 5 represents not only pecies composition for all studied sites, but also ecological and phytogeographical characteristics. It is basic floristic and ecological material of this first studied diatoms in the region. But Table 5 does not include the species in communities of each of 25 studied sites. It is presented in Appendix A Table 1, so each can be see the species content that was found in first when the research will be continued.

 

It seems to me that the only (somewhat) useful measure you have to compare with with regard to pollution or nutrients would be nitrate. I didn't read anything about this.

Response: Please read papers no 17 and 43 as well as: Barinova, S. 2017. On the Classification of Water Quality from an Ecological Point of View. International Journal of Environmental Sciences & Natural Resources, 2(2): 1-8. https://juniperpublishers.com/ijesnr/pdf/IJESNR.MS.ID.555581.pdf

I meant that this is not covered at all in results or discussion.

Also, still unclear how you arrive at ‘scores for anthropogenic loading’ following Einav and Israel 2008 Checklist of seaweeds from the Israeli Mediterranean: Taxonomical and ecological approaches. Israel Journal of Plant Sciences 56: 127–184 DOI: 10.1560/IJPS.57.1–2.127. They do not mention anything like this.

RESPONSE: It is your mistake. When Prof Einav did the site description it mentioned what anthropogenic load was for found in each studied site, we only scored it. For example: Site 1 Rosh HaNiqra – “low levels of deterioration caused by human activities”, page 128; Site “Natanya were dramatically changed from natural rocky beaches fit for seaweed growth into “bathing beaches””; Yafo Site – “this site has been turned into bathing beaches and the natural topography largely changed”; Site Tel Taninim – “site is made of abrasion platforms and likely receives significant amounts of nutrients from the close by Taninim River effluents”, page 144, etc.

 

Also, I missed replies on some comments.

An additional question. You added this was 'when water temperature and solar activity increase'.

RESPONSE: This is wrong. Our experience in the region shows that the waters are gradually warming up and insolation is gradually increasing. We also noticed that it is in spring that macrophytes actively develop in the coastal photic zone, as in the continental reservoirs of the region. Your experience and assumptions may be from a different region.

 

Suggests that assemblages might just be particularly unstable at this time of year.

RESPONSE: reduced

 

 

Superfluous keywords still present and why vertical lines in figure 4?

RESPONSE: For better focus of attention to the highly correlated variables.

 

Comments on the Quality of English Language

Not improved adequately.

RESPONSE: Hope now is better.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

I appreciate the authors' considerations for my concerns and suggested edits. Aside from some lingering issues with the text and grammar, I am satisfied.

Comments on the Quality of English Language

There are still a couple of rough spots in places where the text has been edited with "Track changes"--particularly passages at the  end of Page 10 and on Page 18.

Round 3

Reviewer 2 Report

Comments and Suggestions for Authors

Overall, if this paper would merely present a first account of assemblage composition, without any of the ecological assumptions and less conjecture, and still with substantial re-editing, it could possibly obtain my approval, but as it stands I have too many doubts and unanswered questions. Additional comments marked v3 in bold. My suggestion would be to elaborate the documentation of all the listed taxa (by LM and, as much as possible SEM) to increase its value as a guide for further study (as stated in the intro to be principal aim), summarize the environmental data, remove all the rest and only present a brief discussion of composition and geographical distributions. This would improve the focus of the paper and avoid all points of preliminary nature or open to discussion. I have not replied to every response of the authors to previous comments (if ever given), except where indicated by v3 in bold.

Response: Table 1 gives the response for this comment. Sites not only named but also geographical coordinates are given for each sampling point. Let’s us please to correct your understanding the study area – sa it can be seen on the Figure 1 and Table 1, the studied points were located not in the continental part of this territory but only on the coastline between Rosh-HaNiqra and Ashkelon and in no case affected other continental territory, as you write “along the Israeli coast from the West Bank to Gaza”

Sorry, a geographic slip, apparently. I was fully aware that these are only coastal samples; not West Bank, meant Lebanon of course… (that’s what you get with all the news). The main point is the quality of the map, only 23 dots visible not 25.

RESPONSE: I understood your geographical mistakes. I understand your bias, but I am not guided by the news. Let me also ask you to give your attention to Table 1 where are mentioned the GIS coordinates of each sampling point and you can see that some of them are located very close to each other, so same points are doubled on the map of this resolution. But there are 25 points in any case. However, to clear up any confusion, we are replacing Figure 1 with one with more resolution and sampling points numbered.

v3 Better.

A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast.

Response: As stated in the article, this is not a relatively unexplored coastline, but a completely unexplored coastline in terms of diatom diversity. Thank you for appreciating the floristic aspect of the article.

'Relative' with regard to other parts of the Med. and in general.

RESPONSE: OK I understood. The other parts of the Mediterranean diatom diversity are mentioned in the Discussion part.

The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis.

Response: Thank you. Undoubtedly, the article provides only the first data; this aspect is expected to be studied later.

If I read 'to identify indicator properties of Israeli aquatic flora that can be used to monitor the quality of coastal waters', that means more than just an inventory of taxa.

RESPONSE: Yes, of course, it means that in the revealed diversity will be found an indicator species to diverse of environmental variables and we only on the first step of this way, expected to be studied later.

v3 The paper does not present the necessary data to substantiate this aim '... to determine ... the main environmental factors influencing their richness and diversity, and to identify indicator species that, together with macroalgal data, can serve as an auxiliary tool in coastal zone monitoring.' It is a first and still partial account of the diatom flora present along this coast. If, for instance, the same sites had been sampled three months later, very different assemblages might even have occurred. You can do the taxonomy/floristics, if necessary - which is a legitimate aim in itself - or you collect species data and environmental proxies together. Your contribution fits in the first category, whilst making no contribution to taxonomy. In my opinion, you have not clarified anything about '... the main environmental factors influencing their richness and diversity...'. At present, there is no such analysis (or the necessary data to support such) in the paper. This should, however, preceed any tentative interpretations of assemblage composition in terms of ambient environmental conditions.

Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I stand with this comment.

RESPONSE: The rationale for the conclusions follows from many details. We believe it is the scientific method to look at the details and summarize them into conclusions. English was subject to improvement.

v3 Readability remains problematic.

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

Response: Thank you very much for your detailed comments. We will follow it.

Abstract

Delete superfluous information.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Still includes unsubstantiated conclusions. Decision to be made by editor as I have no access to other reviews.

v3 See further down.

RESPONSE: Unfortunately, I did not find any unfounded conclusions. It is detailed handling of the material that provides the basis for conclusions both in the taxonomic and floristic aspect, and from the point of view of interaction with the environment, which is given in the Conclusions and in the

v3 It remains a mystery to me how one can claim that 'The study ... clearly demonstrates the possibility ... for water quality indication' when at the same time 'we observed a lack of correlation between species richness, saprobity index, WESI, and anthropogenic loads, and environmental variables.' etc. (and why periphyton, in particular? the diatom studies I know of concerning oil spills and pollution in the marine littoral are mostly on sediments). You conclude that '...to solve these problems, it is necessary to create checklists of species depending on salinity, water depth, temperature, and the type of substrate.' How is this going to solve anything? Check lists are not the answer to this. Still unclear to me what (which) point(s) the discussion wants to make with regard to species composition in different parts of the eastern Med., Black Sea, etc. .

Abstract

Introduction

Delete irrelevant sections.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Partly irrelevant in my opinion. Decision to be made by editor.

RESPONSE: We edited the text, taking into account your advice

v3 The first paragraph concerns a bibliography from 1981. That's almost half a century ago. Relevance questionable. Your overview of diatom epiphyton studies in the Med. is not complete, so why bother. I also have no idea what, for example, the Spanish Emporda (a completely different setting), primary production of epipelic microphytobenthos (the Westerschelde does not even connect with the Med., btw) or the number of phyla in the rest of Israel have to do with this paper.

M & M

v3 Relation of first paragr. to the rest of the paper remains obscure to me.

Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Response: Thank you, the article provides only primary data on the diversity of diatoms that have not previously been studied in this region, and the experimental approaches you propose will be applied later in detailed studies.

There’s nothing experimental proposed, only more reliable and reproducible methodology. Identification of live diatom cells at species level, particularly without prior knowledge of assemblage composition, is not recommendable for floristic studies. Only permanent slides of oxidised material allow adequate observation of diagnostic features and additionally serve as a repository for later study and reference.

RESPONSE: As described in the MM, diatoms were identified as living material only at the first stage of the study, then the material was fixed, species identification was made in permanent preparations, then the cleaned valves were examined in an electron microscope, the results are given in the Appendix. It was not difficult for us to make this determination, since this is our many years of professional practice. On the other hand, the process of establishing the species composition in a community goes exactly like this, first identifying species, then generalizing them as the species composition of the community, but not vice versa, as you recommend. We even wonder where knowledge about the initial composition of a community can come from if its species are not determined. Permanent slide surveys are an experimental work and then, according to our many years of experience, species that are not typical for natural communities can gain an advantage. Here we present only natural communities, as all florists usually do, and not by doing an experiment.

v3 Initially, I could not find anything on 'permanent preparations' for LM in your MM. Now I can trace something with some difficulty but not in the MM (see further; numbering of refs here is a mess, btw). With 'permanent' I mean permanent mounts of oxidized valves embedded in a highly refractive mountant (not substrates that are deployed at sampling sites for colonisation experiments, which seems to be what's hinted at in the reply). What I do read is that taxa were abundance-ranked based on observations of cells in water. 'Then microalgae were washed off macrophytes with a scraper, and the suspension was placed in glass containers for further processing of these samples. Further, the resulting washings were poured into containers, thoroughly mixed, and aqueous preparations of microalgae were used for observation in the light microscopes.' In this case, details are insufficiently clear for many taxa to allow reliable identification (or even separation of taxa with similar general morphology). I also wonder how you could relate SEM-based identifications (often more difficult than with the LM) with the LM observations in some cases, particularly for less abundant taxa.

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me.

Response: Most likely, you do not divide our data into the number of species on the site and the rate of each species or we don’t clearly understand you. However, these are two different series of numbers. The first talks about the number of species on the site, the second about their abundance of their cells. The first reflects taxonomic diversity, and the second can be used to identify dominant species and calculate saprobity indices.

Sorry, there was no misunderstanding involved. I just don’t trust your species numbers.

RESPONSE: Our material is identified by basic, detailed, and generally accepted methods, handbooks are cited, and photographs of species are provided. If you do not trust our definitions, you can indicate where exactly, which photos show erroneous definitions.

v3 Since only 19 taxa are illustrated (<25%), I cannot evaluate the quality of your identifications and I have no great concerns about most of them, but at least one of your abundant species definitely does not extend into marine conditions (Encyonema perpusilla). It is also highly unlikely to find every taxon observed in a sample with LM also in the SEM (and sometimes visa versa) – no problem for abundant ones, but the rest? As there was no standardized way of obtaining species numbers (cf. previous remark on stopping rules, etc), the 'totals' cannot be compared reliably between samples.

Some parts seem to be missing (where are the Pearson correlations?).

Response: Pearson correlations have been calculated and presented in the article for some parameters, reference cited under number 42.

I expected (at the least) correlations for other pairs of variables than those in figure 3 and sample order. I cannot corroborate your 'visual' interpretations.

RESPONSE: Correlation coefficients for other pairs of parameters were calculated (the table is available in added Appendix A Table A 2), however, they turned out to be not significant, so they were not presented in the article. It is proposed to only look at general trends.

The only relation evident from alternative figure 3 is for EC (and still unexpected to me – and remaining unexplained - not for salinity), which is actually even more complex than just linear.

RESPONSE: Yes, you are right. More significant correlation looks like with EC, correlarion Table added.

v3 Since no use of these data is made in relation to assemblage composition, any 'trends' are irrelevant with respect to the rest of the paper. Water samples were collected over a period of 2 months (6/03, 23/04 and 6/05), so who is to say that differences are independent of sampling date. Also does not explain why EC weirdly varies unrelated to salinity.

There seems to be no purpose for the cluster analysis.

Response: Since cluster analysis was carried out for the entire species composition, but the tree turned out to have no sharp differences in the clusters, we consider this construction useful, since it allows us to draw a conclusion about the continuity of the species composition of communities along the studied coast. At the same time, if we include the number of species, abundance scores and chemical indicators in the analysis, then in Figure 8 we can see two main clusters, the defining indicator of which is species richness and abundance, and two sites - Mikmoret (harbor) and Ashkelon (harbor), remain individual.

I have read nothing about this in your discussion. Figure 7: chaining is typical for single linkage clustering. Figure 8: it’s really not done to mix variables that are inherently different and with different scales in this type of analysis; there are appropriate methods, however.

RESPONSE: Two analyzes using the same method were presented on a different basis - community composition in Figure 7 and indicator group composition and environmental parameters in Figure 8. Significant differences are visible and are described below.

v3 Reply does not address the previous remarks.

The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

Response: Most likely, you are referring to the calculated saprobity indices in Table 6. This calculation includes all species that have the species-specific saprobity indices shown in Table 5. Thus, you can see exactly which and how many species are included in the calculation by looking at Appendix Table A 1. All data is open, but we do not seek to multiply repetitions of information by placing the main table in the Appendix.

Sorry, you cannot ask the reader to do these basic calculations themselves.

RESPONSE: We have already performed these basic calculations and presented the results. The specific phrase in the previous answer was intended solely for you, so that you could see the transparency of the analysis done.

v3 I cannot imagine why you are wary to present this info more directly.

Discussion & conclusions

Structure and contents need considerable improvement.

Response: We try to do it, thank you. We replaced some of the figures, rearranged them in a clearer format, added trend lines for parameter changes, and made corrections to the text.

I cannot discern appreciable improvement.

RESPONSE: If you can follow the changes in the track changes, it will become clear what exactly has been improved or replaced. A lot of them.

v3 A matter of opinion, apparently.

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Response: Thank you very much for your detailed comments. We will follow it.

Comments in Attachment:

Unfortunately, the rationale for using a saprobic index for marine conditions is entirely based on studies that I cannot assess because they are in Russian and not easy to come by. So I will not comment on this.

Response: It is in English also, please give your attention to the references number 17 and 43.

Sorry, these publications offer no insight into the basis of saprobity scores for marine taxa, or the validation of these metrics for marine conditions – that is, do they relate significantly to whatever they are assumed to indicate. Again no comment on the method itself, but why calculate something which may not be relevant (and for which relevance is not indicated by results presented here, either).

RESPONSE: Since saprobity indices show only the degree of organic pollution, this has nothing to do with the salinity of the water, but only with organic pollution. Therefore, in the article and in the additional publication we provided you with links to this generally accepted method, obviously, after reading which you could understand why and on what basis this is done. Yes, you are right, for species from the marine environment, species-specific saprobity indices are not yet sufficiently known, however, we consider our work useful, since we supplement the ecological properties of the identified diatom species using data known for indicators of organic pollution.

v3 Personally, I would question the saprobity concept for well-oxygenated marine littoral waters, but I also could not find saprobity values for many of the marine taxa in refs 39 or 45 (both by Barinova) as indicated, so where do they actually come from?

Personally, I would be very wary to do so, but assuming that this would work, this still necessitates an estimation of relative abundance (perhaps also weighed by biovolumes, if you like) to apply weighted averaging of scores reliably - as in formula 1 - in a reproducible way. The traditional way to estimate hi requires counting a fairly large number of valves (in your case cells). Your abundance scores are only a crude reflection or relative abundance.

Response: The semi-quantitative scores of the cell’s abundance is represented for each species on the each sampling site in the Appendix Table A 1.

Although it can be done, it is not customary to use ordinal data in WA.

RESPONSE: We can cite hundreds of publications where semi-quantitative methods for calculating indices are used. However, the main ones have already been cited in the article and, apparently, there is no need to multiply the citations, thereby simply increasing the list of references used.

For instance, assume that you have only 2 taxa and both are 'frequent', does this mean their relative abundance is equal (e.g. 50 %) and that they carry 'the same amount of information', therefore should receive the same weight?

Response: This method does not use percentage; it is giving the scores of abundances only. You are right, the percentage is not usable because we can calculate the sum of scores for the sample, but never – the sum of percentage.

Indeed, it’s much more crude. I assume that you don’t want to count because it takes some time, but in that case why not use ‘simultaneous observation’, instead of mixing slides, transects and fields?

v3 The problem is not with 'percentages', as such, but with constancy of proportional abundance.

RESPONSE: There is no mixing. We replaced Table 2 where were some misprints, so it can be clear for the assessment of scores. The given method of semi-quantitative analysis is widely used. We have added here one more reference, however, we consider it unproductive to multiply quotes:

Alster, A., Kaplan-Levy, R.N., Barinova, S.S., Zohary, T. Analyzing semiquantitative phytoplankton counts. Hydrobiologia, 2023. https://doi.org/10.1007/s10750-023-05391-4.

v3 In contrast to your analyses, the phytoplankton procedure starts from a known equal volume which makes things comparable and it's also much more logical as it only uses 'field's of view'.

Note that scores 1 to 3 have undefined intervals. What with 7, 12 or 20 cells?

Don’t understand. There are numbers that cannot be classified (6 to 9, 16 to 24 cells)… Seems relevant to me.

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18.

vs3 Better.

I assume that your ordinal scoring starts from probability-based reasoning, but then some additional explanation is required because the decision rules are not clear to me. To assess all taxa on a slide you have to scan it completely (which usually takes more time than actual counting a fixed number, so I cannot see much reason to use a much cruder approximation, anyway). The area of transects and fields of view varies with magnification... Is score 6 given for all the views, together covering the slide or only for the first one inspected (not plausible it seems, the first cell would always lead to 6)? The same for 4 and 5. How do you separate circular views from rectangular transects?

Response: we added x40 for the last line in Table 2. All other scores are as is.

Doesn't clarify much to me. Even more surprising that you are able to identify more than a few uncleaned diatoms at 400x in water – I couldn’t.

RESPONSE: We can because we used the permanent slide methods as well as SEM method. The calculation of the cells number is followed by species definition as given in MM but not vice versa.

v3 Suddenly, permanent slides have appeared, so what was it? I'm puzzled now.

The abundance scores also depend primarily on the amount of sample (i.e. total surface area of the thalli) used, the initial density of cells on the substrate, as well as the final dilution.. Without complete standardization of procedures you cannot add them up decently and make comparisons between samples regarding densities. A better way would be to work really quantitatively, e.g. using the number of cells on a fixed surface area of the substrate, say 1 square cm.

Response: We avoid these problems when use the semi-quantitative method with the scores assessment. But in Future it possible to calculate the quantitative variables such as cells number per area and biomass per area if the experimental research will be done.

Your method ranks taxa from less to more abundant relative to each other, but the density of cells on a slide and therefore the individual scores (as well as their sum) depends on their concentration in the suspension you apply. Without standardisation, the concentrations are not comparable between samples. Why would one ‘six’ at a site be equivalent to six ‘ones’ at another?

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18. It is standard – one drop of the sediment from fixed sample as given in addition ref 18.

v3 Table is ok now but this does not solve the problem. The 'drops' are not comparable. Simple solution: forget the abundance scores (which reflect the dominance structure, not the 'activity of diatom growth' which is something different) because it inherently depends on the number of taxa.

A sentence giving the most important taxa would be more informative than the nr of classes, etc.. Now it is necessary to consult the supplementary table to get an idea.

Response: Yes, we added the full list of diatoms for each sampling site because all researchers can see our advances. By the way, the mentioned taxonomic structure is a part of floristic analysis to which you recommended to concentrate our paper. The comparison to the similar diatom flora found in the north from our study area in Tartus is represent also part of floristic analysis – comparative floristic.

There is no need for an additional table (info on prevalence could btw also be easily included in Table 5).

RESPONSE: Table 5 represents not only pecies composition for all studied sites, but also ecological and phytogeographical characteristics. It is basic floristic and ecological material of this first studied diatoms in the region. But Table 5 does not include the species in communities of each of 25 studied sites. It is presented in Appendix A Table 1, so each can be see the species content that was found in first when the research will be continued.

It seems to me that the only (somewhat) useful measure you have to compare with with regard to pollution or nutrients would be nitrate. I didn't read anything about this.

Response: Please read papers no 17 and 43 as well as: Barinova, S. 2017. On the Classification of Water Quality from an Ecological Point of View. International Journal of Environmental Sciences & Natural Resources, 2(2): 1-8. https://juniperpublishers.com/ijesnr/pdf/IJESNR.MS.ID.555581.pdf

I meant that this is not covered at all in results or discussion.

v3 And still this remains a compilation of separate, unrelated data.

Also, still unclear how you arrive at ‘scores for anthropogenic loading’ following Einav and Israel 2008 Checklist of seaweeds from the Israeli Mediterranean: Taxonomical and ecological approaches. Israel Journal of Plant Sciences 56: 127–184 DOI: 10.1560/IJPS.57.1–2.127. They do not mention anything like this.

RESPONSE: It is your mistake. When Prof Einav did the site description it mentioned what anthropogenic load was for found in each studied site, we only scored it. For example: Site 1 Rosh HaNiqra – “low levels of deterioration caused by human activities”, page 128; Site “Natanya were dramatically changed from natural rocky beaches fit for seaweed growth into “bathing beaches””; Yafo Site – “this site has been turned into bathing beaches and the natural topography largely changed”; Site Tel Taninim – “site is made of abrasion platforms and likely receives significant amounts of nutrients from the close by Taninim River effluents”, page 144, etc.

v3 No sorry, this is not 'my mistake'. I read these very general descriptions of mainly physical coastline alteration (useful for macroalgae as they determine availability of their substrates, but much less for their epiphytes) and find them quite unsuitable to translate them into meaningful 'scores for anthropogenic loading'. This simply lacks an objective methodology.

Also, I missed replies on some comments.

v3 Remains. See previous comments.

An additional question. You added this was 'when water temperature and solar activity increase'. Suggests that assemblages might just be particularly unstable at this time of year.

RESPONSE: This is wrong. Our experience in the region shows that the waters are gradually warming up and insolation is gradually increasing. We also noticed that it is in spring that macrophytes actively develop in the coastal photic zone, as in the continental reservoirs of the region. Your experience and assumptions may be from a different region.

v3 The rate of assemblage change is likely to follow the rate of change in the ambient environment rather closely, so why would this be wrong? If you look for the more stable 'mature' and species-rich assemblages you should pick the season which is most favorable as well as environmentally stable.

RESPONSE: reduced

Superfluous keywords still present and why vertical lines in figure 4?

RESPONSE: For better focus of attention to the highly correlated variables.

Comments on the Quality of English Language

Not improved adequately.

RESPONSE: Hope now is better.

v3 Remains.

Comments on the Quality of English Language

no

Author Response

Responses to Reviewer 2 Report 3

Dear Editor,

Thank you and the Reviewer 2 for comments.

Please find our responses to each comment and comment of comment below.

With best regards,

Prof Sophia Barinova,

Corresponding author.

 

Comments and Suggestions for Authors

Overall, if this paper would merely present a first account of assemblage composition, without any of the ecological assumptions and less conjecture, and still with substantial re-editing, it could possibly obtain my approval, but as it stands I have too many doubts and unanswered questions. Additional comments marked v3 in bold. My suggestion would be to elaborate the documentation of all the listed taxa (by LM and, as much as possible SEM) to increase its value as a guide for further study (as stated in the intro to be principal aim), summarize the environmental data, remove all the rest and only present a brief discussion of composition and geographical distributions. This would improve the focus of the paper and avoid all points of preliminary nature or open to discussion. I have not replied to every response of the authors to previous comments (if ever given), except where indicated by v3 in bold.

Response 3: The authors are very grateful to Reviewer 2 for his careful attention to our manuscript and his willingness to help. We would like to note that at this stage we have done a lot of work to correct the manuscript. We have carefully corrected the text, clarified the identification of some species, re-corrected references and the accuracy of their citation.

Response: Table 1 gives the response for this comment. Sites not only named but also geographical coordinates are given for each sampling point. Let’s us please to correct your understanding the study area – sa it can be seen on the Figure 1 and Table 1, the studied points were located not in the continental part of this territory but only on the coastline between Rosh-HaNiqra and Ashkelon and in no case affected other continental territory, as you write “along the Israeli coast from the West Bank to Gaza”

Sorry, a geographic slip, apparently. I was fully aware that these are only coastal samples; not West Bank, meant Lebanon of course… (that’s what you get with all the news). The main point is the quality of the map, only 23 dots visible not 25.

RESPONSE: I understood your geographical mistakes. I understand your bias, but I am not guided by the news. Let me also ask you to give your attention to Table 1 where are mentioned the GIS coordinates of each sampling point and you can see that some of them are located very close to each other, so same points are doubled on the map of this resolution. But there are 25 points in any case. However, to clear up any confusion, we are replacing Figure 1 with one with more resolution and sampling points numbered.

v3 Better.

A few variables (salinity, pH, nitrate) were measured, as well. Some taxa are documented by SEM (unfortunately not always the most interesting ones). In my opinion, the most important contribution of this ms is to the diatom floristics (85 taxa) of this relatively unexplored part of the Mediterranean coast.

Response: As stated in the article, this is not a relatively unexplored coastline, but a completely unexplored coastline in terms of diatom diversity. Thank you for appreciating the floristic aspect of the article.

'Relative' with regard to other parts of the Med. and in general.

RESPONSE: OK I understood. The other parts of the Mediterranean diatom diversity are mentioned in the Discussion part.

The authors aim to relate assemblage diversity and composition to (assumed) pollution but this is not convincingly demonstrated and would require more extensive data, consideration of alternative sources of variation and more rigorous (multivariate) analysis.

Response: Thank you. Undoubtedly, the article provides only the first data; this aspect is expected to be studied later.

If I read 'to identify indicator properties of Israeli aquatic flora that can be used to monitor the quality of coastal waters', that means more than just an inventory of taxa.

RESPONSE: Yes, of course, it means that in the revealed diversity will be found an indicator species to diverse of environmental variables and we only on the first step of this way, expected to be studied later.

v3 The paper does not present the necessary data to substantiate this aim '... to determine ... the main environmental factors influencing their richness and diversity, and to identify indicator species that, together with macroalgal data, can serve as an auxiliary tool in coastal zone monitoring.' It is a first and still partial account of the diatom flora present along this coast. If, for instance, the same sites had been sampled three months later, very different assemblages might even have occurred. You can do the taxonomy/floristics, if necessary - which is a legitimate aim in itself - or you collect species data and environmental proxies together. Your contribution fits in the first category, whilst making no contribution to taxonomy. In my opinion, you have not clarified anything about '... the main environmental factors influencing their richness and diversity...'. At present, there is no such analysis (or the necessary data to support such) in the paper. This should, however, preceed any tentative interpretations of assemblage composition in terms of ambient environmental conditions.

Too much consideration of details and too little structure and clear reasoning do not make for pleasant reading. Language and readability also need considerable attention throughout.

I stand with this comment.

RESPONSE: The rationale for the conclusions follows from many details. We believe it is the scientific method to look at the details and summarize them into conclusions. English was subject to improvement.

Response 3: We did just that, determined the species composition and identified indicators for 6 environmental indicators, and using statistical methods, described the interaction of the species composition of diatoms and the main environmental indicators, and also identified the intensity of organic water pollution and correlated this with the strength of anthropogenic load. That is: “to determine ... the main environmental factors influencing their richness and diversity, and to identify indicator species that, together with macroalgal data, can serve as an auxiliary tool in coastal zone monitoring”

v3 Readability remains problematic.

Response 3: We made many corrections to the text in the manuscript in terms of language. We hope, that the manuscript is now more comprehensible

I have made many more detailed comments and suggestions on the pdf, the following are a few general ones.

Response: Thank you very much for your detailed comments. We will follow it.

Abstract

Delete superfluous information.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Still includes unsubstantiated conclusions. Decision to be made by editor as I have no access to other reviews.

v3 See further down.

Response 3: Abstract is edited.

RESPONSE: Unfortunately, I did not find any unfounded conclusions. It is detailed handling of the material that provides the basis for conclusions both in the taxonomic and floristic aspect, and from the point of view of interaction with the environment, which is given in the Conclusions and in the

v3 It remains a mystery to me how one can claim that 'The study ... clearly demonstrates the possibility ... for water quality indication' when at the same time 'we observed a lack of correlation between species richness, saprobity index, WESI, and anthropogenic loads, and environmental variables.' etc. (and why periphyton, in particular? the diatom studies I know of concerning oil spills and pollution in the marine littoral are mostly on sediments). You conclude that '...to solve these problems, it is necessary to create checklists of species depending on salinity, water depth, temperature, and the type of substrate.' How is this going to solve anything? Check lists are not the answer to this. Still unclear to me what (which) point(s) the discussion wants to make with regard to species composition in different parts of the eastern Med., Black Sea, etc. .

Response 3: In this article, our aim was to study for the first time the species composition of epiphyton diatoms  in the Israeli sector of the Mediterranean Sea and to compare it with other Mediterranean waters. In order to calculate the saprobic index, it is necessary to cover the flora of the study area as widely as possible, that is why species checklists for the region are so important.  In addition, it is important to identify the presence of indicator species in the region at the initial stage of research, that's why we write that our work shows the possibility of using it for further bioindication work. We did not seek to carry out taxonomic work specifically but made efforts to identify the species composition. But the recommendation to eliminate all data analysis, leaving only the floristic part, turns out to be unacceptable, since only a list of species will remain, and this is not published in journals without data analysis. Moreover, using bioindication methods, we identified indicators such as organic pollution with help. saprobity indices, which is impossible to do in a simple floristic list. Bioindication helps in cases where data on the habitat is insufficient or absent, determining the amplitude of each of them according to the community living within these limits.

Abstract

Introduction

Delete irrelevant sections.

Response: We follow the advice of other reviewers who indicate that these passages are satisfactory. We nevertheless edited the text a little, taking into account your advice.

Sorry, I can’t agree. Partly irrelevant in my opinion. Decision to be made by editor.

Response 3: We edited the text, taking into account your advice

v3 The first paragraph concerns a bibliography from 1981. That's almost half a century ago. Relevance questionable. Your overview of diatom epiphyton studies in the Med. is not complete, so why bother. I also have no idea what, for example, the Spanish Emporda (a completely different setting), primary production of epipelic microphytobenthos (the Westerschelde does not even connect with the Med., btw) or the number of phyla in the rest of Israel have to do with this paper.

Response 3: Here we want to show the lack of study of the region. In addition, we wanted to cover the geography of benthic diatom studies of the Mediterranean Sea.

M & M

v3 Relation of first paragr. to the rest of the paper remains obscure to me.

Response 3:  A description of the study area...

Consider a more reliable way to estimate species diversity and abundance using permanent mounts, or just stick to the floristics (see remarks on ms).

Response: Thank you, the article provides only primary data on the diversity of diatoms that have not previously been studied in this region, and the experimental approaches you propose will be applied later in detailed studies.

There’s nothing experimental proposed, only more reliable and reproducible methodology. Identification of live diatom cells at species level, particularly without prior knowledge of assemblage composition, is not recommendable for floristic studies. Only permanent slides of oxidised material allow adequate observation of diagnostic features and additionally serve as a repository for later study and reference.

RESPONSE: As described in the MM, diatoms were identified as living material only at the first stage of the study, then the material was fixed, species identification was made in permanent preparations, then the cleaned valves were examined in an electron microscope, the results are given in the Appendix. It was not difficult for us to make this determination, since this is our many years of professional practice. On the other hand, the process of establishing the species composition in a community goes exactly like this, first identifying species, then generalizing them as the species composition of the community, but not vice versa, as you recommend. We even wonder where knowledge about the initial composition of a community can come from if its species are not determined. Permanent slide surveys are an experimental work and then, according to our many years of experience, species that are not typical for natural communities can gain an advantage. Here we present only natural communities, as all florists usually do, and not by doing an experiment.

v3 Initially, I could not find anything on 'permanent preparations' for LM in your MM. Now I can trace something with some difficulty but not in the MM (see further; numbering of refs here is a mess, btw). With 'permanent' I mean permanent mounts of oxidized valves embedded in a highly refractive mountant (not substrates that are deployed at sampling sites for colonisation experiments, which seems to be what's hinted at in the reply). What I do read is that taxa were abundance-ranked based on observations of cells in water. 'Then microalgae were washed off macrophytes with a scraper, and the suspension was placed in glass containers for further processing of these samples. Further, the resulting washings were poured into containers, thoroughly mixed, and aqueous preparations of microalgae were used for observation in the light microscopes.' In this case, details are insufficiently clear for many taxa to allow reliable identification (or even separation of taxa with similar general morphology). I also wonder how you could relate SEM-based identifications (often more difficult than with the LM) with the LM observations in some cases, particularly for less abundant taxa.

Response 3:  Corrected in ms. Each sample was studied in parallel in SM and SEM, so it was possible to associate identified species based on the photographs obtained. The methodology of sample preparation is described in Section 2.1. Sampling and material processing. Sample preparation of epiphyton diatoms was carried out according to Simonsen, 1962 with our own additions (Blaginina, A.; Ryabushko, L, 2021). Our methods refer to standard algological methods.

Results

I have substantial doubts on the possibility to estimate comparable species numbers from the observations. Relations are mainly inferred ‘visually’, but not transparent to me.

Response: Most likely, you do not divide our data into the number of species on the site and the rate of each species or we don’t clearly understand you. However, these are two different series of numbers. The first talks about the number of species on the site, the second about their abundance of their cells. The first reflects taxonomic diversity, and the second can be used to identify dominant species and calculate saprobity indices.

Sorry, there was no misunderstanding involved. I just don’t trust your species numbers.

RESPONSE: Our material is identified by basic, detailed, and generally accepted methods, handbooks are cited, and photographs of species are provided. If you do not trust our definitions, you can indicate where exactly, which photos show erroneous definitions.

v3 Since only 19 taxa are illustrated (<25%), I cannot evaluate the quality of your identifications and I have no great concerns about most of them, but at least one of your abundant species definitely does not extend into marine conditions (Encyonema perpusilla). It is also highly unlikely to find every taxon observed in a sample with LM also in the SEM (and sometimes visa versa) – no problem for abundant ones, but the rest? As there was no standardized way of obtaining species numbers (cf. previous remark on stopping rules, etc), the 'totals' cannot be compared reliably between samples.

Response 3:  Encyonema perpusillum was mostly sporadic in the samples, except some sites where strong coastal spreading is observed. It can be assumed that this species entered the sea with river discharges and took habitat at those stations, where spreading is observed.

Some parts seem to be missing (where are the Pearson correlations?).

Response: Pearson correlations have been calculated and presented in the article for some parameters, reference cited under number 42.

I expected (at the least) correlations for other pairs of variables than those in figure 3 and sample order. I cannot corroborate your 'visual' interpretations.

RESPONSE: Correlation coefficients for other pairs of parameters were calculated (the table is available in added Appendix A Table A 2), however, they turned out to be not significant, so they were not presented in the article. It is proposed to only look at general trends.

The only relation evident from alternative figure 3 is for EC (and still unexpected to me – and remaining unexplained - not for salinity), which is actually even more complex than just linear.

RESPONSE: Yes, you are right. More significant correlation looks like with EC, correlarion Table added.

v3 Since no use of these data is made in relation to assemblage composition, any 'trends' are irrelevant with respect to the rest of the paper. Water samples were collected over a period of 2 months (6/03, 23/04 and 6/05), so who is to say that differences are independent of sampling date. Also does not explain why EC weirdly varies unrelated to salinity.

Response 3:  The absence of a clear correlation between salinity and conductivity is not a criterion for the reliability of the data. Naturally, you assume that these indicators should correlate, but this is not always the case, since the measured values ​​indicate different things - the number of ions in the water and the resistance of the water. In any case, the device that measured the indicators is certified; data on the devices is indicated in the MM section. The period of data collection is also not a criterion of reliability, since in one second no data on a 150 km section and 25 points can be obtained in any case, however, both of these indicators were determined in 3 repetitions and simultaneously, from which the average values ​​were calculated.

There seems to be no purpose for the cluster analysis.

Response: Since cluster analysis was carried out for the entire species composition, but the tree turned out to have no sharp differences in the clusters, we consider this construction useful, since it allows us to draw a conclusion about the continuity of the species composition of communities along the studied coast. At the same time, if we include the number of species, abundance scores and chemical indicators in the analysis, then in Figure 8 we can see two main clusters, the defining indicator of which is species richness and abundance, and two sites - Mikmoret (harbor) and Ashkelon (harbor), remain individual.

I have read nothing about this in your discussion. Figure 7: chaining is typical for single linkage clustering. Figure 8: it’s really not done to mix variables that are inherently different and with different scales in this type of analysis; there are appropriate methods, however.

RESPONSE: Two analyzes using the same method were presented on a different basis - community composition in Figure 7 and indicator group composition and environmental parameters in Figure 8. Significant differences are visible and are described below.

v3 Reply does not address the previous remarks.

Response 3:  Two analyzes using the same method were presented on a different basis - community composition in Figure 7 and indicator group composition and environmental parameters in Figure 8. Significant differences are visible and are described below.

 

The relevance of calculated index values cannot be assessed without knowing which part of the assemblage is included.

Response: Most likely, you are referring to the calculated saprobity indices in Table 6. This calculation includes all species that have the species-specific saprobity indices shown in Table 5. Thus, you can see exactly which and how many species are included in the calculation by looking at Appendix Table A 1. All data is open, but we do not seek to multiply repetitions of information by placing the main table in the Appendix.

Sorry, you cannot ask the reader to do these basic calculations themselves.

RESPONSE: We have already performed these basic calculations and presented the results. The specific phrase in the previous answer was intended solely for you, so that you could see the transparency of the analysis done.

v3 I cannot image ne why you are wary to present this info more directly.

Response 3:  We are not afraid of anything, it was strange to even suggest this. We go step by step in the analysis and clearly present the results of each step, mentioning on the basis of what and by what methods this stage of analysis was carried out. But if you recommend keeping only the list of species and eliminating the entire analysis without discussion, then your comments are unclear.

Discussion & conclusions

Structure and contents need considerable improvement.

Response: We try to do it, thank you. We replaced some of the figures, rearranged them in a clearer format, added trend lines for parameter changes, and made corrections to the text.

I cannot discern appreciable improvement.

RESPONSE: If you can follow the changes in the track changes, it will become clear what exactly has been improved or replaced. A lot of them.

v3 A matter of opinion, apparently.

Response 3:  can be

Comments on the Quality of English Language

I have made no specific comments in the text but thorough revision is absolutely necessary. Some parts are quite difficult to comprehend.

Response: Thank you very much for your detailed comments. We will follow it.

Comments in Attachment:

Unfortunately, the rationale for using a saprobic index for marine conditions is entirely based on studies that I cannot assess because they are in Russian and not easy to come by. So I will not comment on this.

Response: It is in English also, please give your attention to the references number 17 and 43.

Sorry, these publications offer no insight into the basis of saprobity scores for marine taxa, or the validation of these metrics for marine conditions – that is, do they relate significantly to whatever they are assumed to indicate. Again no comment on the method itself, but why calculate something which may not be relevant (and for which relevance is not indicated by results presented here, either).

RESPONSE: Since saprobity indices show only the degree of organic pollution, this has nothing to do with the salinity of the water, but only with organic pollution. Therefore, in the article and in the additional publication we provided you with links to this generally accepted method, obviously, after reading which you could understand why and on what basis this is done. Yes, you are right, for species from the marine environment, species-specific saprobity indices are not yet sufficiently known, however, we consider our work useful, since we supplement the ecological properties of the identified diatom species using data known for indicators of organic pollution.

v3 Personally, I would question the saprobity concept for well-oxygenated marine littoral waters, but I also could not find saprobity values for many of the marine taxa in refs 39 or 45 (both by Barinova) as indicated, so where do they actually come from?

Response 3:  The cited works collected literature information on species-specific saprobity indices, the most part of which comes from cited paper of Sladecek [16] with addition of some other.

Personally, I would be very wary to do so, but assuming that this would work, this still necessitates an estimation of relative abundance (perhaps also weighed by biovolumes, if you like) to apply weighted averaging of scores reliably - as in formula 1 - in a reproducible way. The traditional way to estimate hi requires counting a fairly large number of valves (in your case cells). Your abundance scores are only a crude reflection or relative abundance.

Response: The semi-quantitative scores of the cell’s abundance is represented for each species on the each sampling site in the Appendix Table A 1.

Although it can be done, it is not customary to use ordinal data in WA.

RESPONSE: We can cite hundreds of publications where semi-quantitative methods for calculating indices are used. However, the main ones have already been cited in the article and, apparently, there is no need to multiply the citations, thereby simply increasing the list of references used.

For instance, assume that you have only 2 taxa and both are 'frequent', does this mean their relative abundance is equal (e.g. 50 %) and that they carry 'the same amount of information', therefore should receive the same weight?

Response: This method does not use percentage; it is giving the scores of abundances only. You are right, the percentage is not usable because we can calculate the sum of scores for the sample, but never – the sum of percentage.

Indeed, it’s much more crude. I assume that you don’t want to count because it takes some time, but in that case why not use ‘simultaneous observation’, instead of mixing slides, transects and fields?

v3 The problem is not with 'percentages', as such, but with constancy of proportional abundance.

RESPONSE: There is no mixing. We replaced Table 2 where were some misprints, so it can be clear for the assessment of scores. The given method of semi-quantitative analysis is widely used. We have added here one more reference, however, we consider it unproductive to multiply quotes:

Alster, A., Kaplan-Levy, R.N., Barinova, S.S., Zohary, T. Analyzing semiquantitative phytoplankton counts. Hydrobiologia, 2023. https://doi.org/10.1007/s10750-023-05391-4.

Response 3:  This part with Table 2 corrected

v3 In contrast to your analyses, the phytoplankton procedure starts from a known equal volume which makes things comparable and it's also much more logical as it only uses 'field's of view'.

Note that scores 1 to 3 have undefined intervals. What with 7, 12 or 20 cells?

Don’t understand. There are numbers that cannot be classified (6 to 9, 16 to 24 cells)… Seems relevant to me.

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18.

Response 3:  Our subject is periphyton and we detailed described the procedure in MM part with relevant corrections.

vs3 Better.

I assume that your ordinal scoring starts from probability-based reasoning, but then some additional explanation is required because the decision rules are not clear to me. To assess all taxa on a slide you have to scan it completely (which usually takes more time than actual counting a fixed number, so I cannot see much reason to use a much cruder approximation, anyway). The area of transects and fields of view varies with magnification... Is score 6 given for all the views, together covering the slide or only for the first one inspected (not plausible it seems, the first cell would always lead to 6)? The same for 4 and 5. How do you separate circular views from rectangular transects?

Response: we added x40 for the last line in Table 2. All other scores are as is.

Doesn't clarify much to me. Even more surprising that you are able to identify more than a few uncleaned diatoms at 400x in water – I couldn’t.

RESPONSE: We can because we used the permanent slide methods as well as SEM method. The calculation of the cells number is followed by species definition as given in MM but not vice versa.

v3 Suddenly, permanent slides have appeared, so what was it? I'm puzzled now.

Response 3:  Information on the preparation of permanent preparations has been entered in the Materials and Methods section. We apologise for this omission.

The abundance scores also depend primarily on the amount of sample (i.e. total surface area of the thalli) used, the initial density of cells on the substrate, as well as the final dilution.. Without complete standardization of procedures you cannot add them up decently and make comparisons between samples regarding densities. A better way would be to work really quantitatively, e.g. using the number of cells on a fixed surface area of the substrate, say 1 square cm.

Response: We avoid these problems when use the semi-quantitative method with the scores assessment. But in Future it possible to calculate the quantitative variables such as cells number per area and biomass per area if the experimental research will be done.

Your method ranks taxa from less to more abundant relative to each other, but the density of cells on a slide and therefore the individual scores (as well as their sum) depends on their concentration in the suspension you apply. Without standardisation, the concentrations are not comparable between samples. Why would one ‘six’ at a site be equivalent to six ‘ones’ at another?

RESPONSE: We replaced Table 2 where were some misprints, so it can be clearly for the assessment of scores and added reference 18. It is standard – one drop of the sediment from fixed sample as given in addition ref 18.

v3 Table is ok now but this does not solve the problem. The 'drops' are not comparable. Simple solution: forget the abundance scores (which reflect the dominance structure, not the 'activity of diatom growth' which is something different) because it inherently depends on the number of taxa.

Response 3:  The appropriate correction has been made, however, the method is so widely known and has been used for more than a hundred years (since 1908) that we see no reason to exclude its use in our work.

A sentence giving the most important taxa would be more informative than the nr of classes, etc.. Now it is necessary to consult the supplementary table to get an idea.

Response: Yes, we added the full list of diatoms for each sampling site because all researchers can see our advances. By the way, the mentioned taxonomic structure is a part of floristic analysis to which you recommended to concentrate our paper. The comparison to the similar diatom flora found in the north from our study area in Tartus is represent also part of floristic analysis – comparative floristic.

There is no need for an additional table (info on prevalence could btw also be easily included in Table 5).

RESPONSE: Table 5 represents not only pecies composition for all studied sites, but also ecological and phytogeographical characteristics. It is basic floristic and ecological material of this first studied diatoms in the region. But Table 5 does not include the species in communities of each of 25 studied sites. It is presented in Appendix A Table 1, so each can be see the species content that was found in first when the research will be continued.

It seems to me that the only (somewhat) useful measure you have to compare with with regard to pollution or nutrients would be nitrate. I didn't read anything about this.

Response: Please read papers no 17 and 43 as well as: Barinova, S. 2017. On the Classification of Water Quality from an Ecological Point of View. International Journal of Environmental Sciences & Natural Resources, 2(2): 1-8. https://juniperpublishers.com/ijesnr/pdf/IJESNR.MS.ID.555581.pdf

I meant that this is not covered at all in results or discussion.

v3 And still this remains a compilation of separate, unrelated data.

Response 3:  These are separate parts of the analysis, carried out step by step, to highlight different aspects of the interaction between environments and communities.

Also, still unclear how you arrive at ‘scores for anthropogenic loading’ following Einav and Israel 2008 Checklist of seaweeds from the Israeli Mediterranean: Taxonomical and ecological approaches. Israel Journal of Plant Sciences 56: 127–184 DOI: 10.1560/IJPS.57.1–2.127. They do not mention anything like this.

RESPONSE: It is your mistake. When Prof Einav did the site description it mentioned what anthropogenic load was for found in each studied site, we only scored it. For example: Site 1 Rosh HaNiqra – “low levels of deterioration caused by human activities”, page 128; Site “Natanya were dramatically changed from natural rocky beaches fit for seaweed growth into “bathing beaches””; Yafo Site – “this site has been turned into bathing beaches and the natural topography largely changed”; Site Tel Taninim – “site is made of abrasion platforms and likely receives significant amounts of nutrients from the close by Taninim River effluents”, page 144, etc.

v3 No sorry, this is not 'my mistake'. I read these very general descriptions of mainly physical coastline alteration (useful for macroalgae as they determine availability of their substrates, but much less for their epiphytes) and find them quite unsuitable to translate them into meaningful 'scores for anthropogenic loading'. This simply lacks an objective methodology.

Нет, извините, это не «моя ошибка». Я прочитал эти очень общие описания главным образом физических изменений береговой линии (полезные для макроводорослей, поскольку они определяют доступность своих субстратов, но гораздо меньше для их эпифитов) и нашел их совершенно непригодными для перевода в значимые «оценки антропогенной нагрузки». Здесь просто не хватает объективной методологии.

Also, I missed replies on some comments.

Response 3:  But we analyzed the texts available in work 2 and we were able to identify information about anthropogenic changes in relation to 23 of the mentioned sites, but in the text we added that also according to our observations.

v3 Remains. See previous comments.

Response 3:  But we analyzed the texts available in work 2 and we were able to identify information about anthropogenic changes in relation to 23 of the mentioned sites, but in the text we added that also according to our observations.

 

An additional question. You added this was 'when water temperature and solar activity increase'. Suggests that assemblages might just be particularly unstable at this time of year.

RESPONSE: This is wrong. Our experience in the region shows that the waters are gradually warming up and insolation is gradually increasing. We also noticed that it is in spring that macrophytes actively develop in the coastal photic zone, as in the continental reservoirs of the region. Your experience and assumptions may be from a different region.

v3 The rate of assemblage change is likely to follow the rate of change in the ambient environment rather closely, so why would this be wrong? If you look for the more stable 'mature' and species-rich assemblages you should pick the season which is most favorable as well as environmentally stable.

Response 3: We do not make assumptions. We were not looking for seasons that were convenient for us. We report the first study of diatoms in coastal communities of this coastal site. But, as indicated in the answer above, in the communities of continental waters of this region there really is an outbreak of diatom diversity precisely during the spring period, when insulation increases and the water begins to warm up.

RESPONSE: reduced

Superfluous keywords still present and why vertical lines in figure 4?

RESPONSE: For better focus of attention to the highly correlated variables.

Response 3: words are shortened. lines are shown to focus the reader's attention on correlating variables.

Comments on the Quality of English Language

Not improved adequately.

RESPONSE: Hope now is better.

v3 Remains.

Response 3: We have made additional revisions to the manuscript and corrections in terms of the language.

Author Response File: Author Response.pdf