Riverine Inputs Impact the Diversity and Population Structure of Heterotrophic Fungus-like Protists and Bacterioplankton in the Coastal Waters of the South China Sea
Round 1
Reviewer 1 Report
For me, there are not enough microphotographs of the objects under study.
Author Response
Point 1: For me, there are not enough microphotographs of the objects under study.
Response 1: Thank you for your suggestion. In this study, the abundance and diversity of LP were all determined by their DNA, but not directly counting. Therefore, there were not enough microphotographs in our study. Moreover, the DNA-based methods were widely used to detect LP in different marine environments (such as ref. 25, 45 and 67). Thus, the method of this study was common and reliable.
- Xie, N.; Sen, B.; Song, Z.; Zhao, Y.; Chen, Z.; Shi, W.; Zhang, Y.; Zhang, J.; Johnson, Z.I.; Wang, G. High phylogenetic diversity and abundance pattern of Labyrinthulomycete protists in the coastal waters of the Bohai Sea. Environ. Microbiol. 2018, 20, 3042-3056.
- Bai, M.; Sen, B.; Wang, Q.; Xie, Y.; He, Y.; Wang, G. Molecular detection and spatiotemporal characterization of Labyrinthu-lomycete protist diversity in the coastal waters along the Pearl River Delta. Microb. Ecol. 2019, 77, 394-405.
- Liu, Y.; Singh, P.; Sun, Y.; Luan, S.; Wang, G. Culturable diversity and biochemical features of thraustochytrids from coastal waters of Southern China. Appl. Microbiol. Biotechnol. 2014, 98, 3241-3255.
Author Response File: 
Author Response.docx
Reviewer 2 Report
Review
Paper title: Riverine inputs impact the diversity and population structure of heterotrophic fungus-like protists in the coastal waters of the South China Sea
The authors conducted a study to reveal the effects of a salinity gradient in the coastal zone of the South China Sea on the abundance and community structure of Labyrinthulomycetes protists. The authors used two datasets obtained in cruises conducted in the summer period of 2016 and 2018. The authors found that the highest abundance occurred in the plume zone while the lowest in the pelagic zone. The authors found a species-specific distribution of protists and attributed it to the salinity gradient in the study area. These results may have important implications for further studies and provide a basis for expanding our knowledge of the role of marine protistan micro-eukaryotes in marine environments.
All these reasons explain the relevance of the paper by Suisui Wang and co-authors submitted to "Water".
General scores.
The data presented by the authors are original and significant. The study is correctly designed and the authors used appropriate collecting methods. In general, the statistical analyses are performed with good technical standards. The authors conducted careful work that may attract the attention of a wide range of specialists focused on marine microbial communities.
Recommendations.
L 170-172. Delete this sentence. “To evaluate the impacts of salinity on the LP abundance, qPCR method was used to determine the LP molecular abundance of water samples collected along a plume, off-shore and pelagic gradient during 2 summer SCS cruises”.
Figure 1. The authors should define the letters used on histograms.
L 205. The authors noted that “Statistical significance was analyzed by Kruskal test” but they did not provide a description of this method in the “Materials and Methods” section.
References should be formatted according to Rules for Authors.
Specific comments.
L 24. Change “was dominant” to “dominated”
L 27. Change “salinity gradient” to “the salinity gradient”
L 28. Change “of in interesting” to “of interesting”
L 32. Change “Changes of marine environmental factors influences” to “Changes in marine environmental factors influence”
L 35. Change “changing” to “change”
L 42. Change “such like” to “such as”
L 43. Change “further salinity has been reported to display” to “has displayed”
L 45. Change “estuary” to “estuarine”
L 48. Change “coastal area of East China Sea” to “the coastal area of the East China Sea”
L 52. Change “at brackish waters” to “in brackish waters”
L 56. Change “high level” to “high”
L 59. Change “in Pearl River-influenced region” to “in the Pearl River-influenced region”
L 69. Change “herbirory” to “herbivory”
L 72. Change “coastal” to “coastal zone”
L 80. Change “and however,” to “and”
L 82. Change “level” to “levels”
L 85. Change “distribution” to “the distribution”
L 92. Change “distinct salinity gradient [42,43], which provide us” to “a distinct salinity gradient [42,43], which provide us with”
L 93. Change “a salinity gradient” to “the salinity gradient”
L 94. Change “waters was” to “waters were”
L 98. Change “distribution” to “the distribution”
L 104. Change “Two liters of water in individual station were collected by” to “At each station, two liters of water were collected by a”
L 110. Change “salinity of each station” to “salinity”
L 111. Change “equipped with” to “equipped with the”
L 112. Change “method defined by the previous reports” to “the method defined by previous reports”
L 133. Change “previous study” to “a previous study”
L 149. Change “were used the following protocol” to “were used in the following protocol”
L 179. Change “displayed the” to “displayed a”
L 180. Change “lowest” to “lowest values”
L 186. Change “salinity gradient” to “the salinity gradient”
L 191. Change “difference” to “differences”
L 194. Change “plume region” to “the plume region”
L 202. Change “salinity gradient” to “the salinity gradient”
L 209. Change “High correlation” to “High correlations”
L 212. Change “strong relationship” to “a strong relationship”
L 213. Change “bacterioplankton community structure” to “the bacterioplankton community structure”
L 216. Change “gradients” to “gradient”
L 218. Change “gradient” to “gradients”
L 223. Change “with two” to “into two”
L 232. Change “identical with” to “identical to”
L 233. Change “that plume” to “that the plume”
L 237. Change “than those of the oceanic detected in” to “in comparison to”
L 246. Change “on genera level” to “on the genera level”
L 247. Change “gradients” to “gradient”
L 249. Change “accounted” to “accounted for”
L 251. Change “accounted” to “accounted for”
L 252. Change “was exceeded” to “exceeded”
L 256. Change “were largely varied” to “largely varied”
L 265. Change “abundance of offshore were all nearly” to “abundances in the offshore zone were nearly”
L 266. Change “increasing” to “increase”
L 269. Change “in 2015 summer” to “in the summer of 2015”
L 272. Change “The water salinity” to “Water salinity”
L 277. Change “increased” to “increase”
L 281. Change “formed” to “form”
L 282. Change “significantly positive” to “significant positive”
L 285. Change “generally is associated” to “is generally associated”
L 287. Change “In another word” to “In other words”
L 287. Change “salinity gradient” to “the salinity gradient”
L 294. Change “maybe one reason of” to “may be one reason for”
L 301. Change “sensitive” to “sensitivity”
L 303. Change “In genus level, plume LP communities were charactered” to “At the genus level, plume LP communities were characterized”
L 307. Change “to find high abundance” to “to find a high abundance”
L 310. Change “were intensities associated” to “were associated”
L 312. Change “river” to “the river”
L 316. Change “offshore and pelagic ocean” to “the offshore and pelagic ocean”
L 321. Change “description on” to “description of”
L 323. Change “South China Sea” to “the South China Sea”
Author Response
Please see the attachment
Author Response File: Author Response.docx
Reviewer 3 Report
Review for the paper "Riverine inputs impact the diversity and population structure of heterotrophic fungus-like protists in the coastal waters of the South China Sea" by Suisui Wang, Kalyani Sen, Yaodong He, Mohan Bai, Guangyi Wang submitted to "Water".
General comment.
The authors studied the distribution and diversity of labyrinthulomycetes protists along a salinity gradient in the South China Sea. Protists are major sources of mortality for both phytoplankton and bacteria in marine environments and are known to be important consumers in the World’s Ocean. Therefore, their role in marine ecosystems is crucial. However, many aspects of protistan ecology remain less-studied. The present study revealed that a peak of LP density was found in the plume while a minimum occurred in offshore waters. The authors stated that the spatial variation in LP community was clearly associated with the salinity gradient although LP diversity tended to be lower in the brackish waters. Different taxa were found to prevail in the offshore and inshore communities. I think that this report expands our knowledge regarding the functioning of pelagic ecosystems in the estuarine regions and may be interesting for scientists dealing with ecology of aquatic ecosystems in coastal waters worldwide. The authors used an adequate dataset and methods to collect the samples. Statistical methods seem to be valid and correctly used. Main results are illustrated with relevant Figures and Tables. Discussion is short and focused on the main findings. I have some suggestions to improve the ms.
Major concerns.
- The aim of the study was confined to salinity only. However, temperature may also be an important factor affecting plankton communities in the coastal waters studied. I suggest the authors to conduct an additional analysis to test the possible impact of water temperature on the microbes. The authors must prove that temperature did not influence the microplankton in their study. At least, a simple correlation between LP/bacterioplankton and water temperature would help a reader to understand the role of water temperature in spatial variations of LP and bacterioplankton.
- Environmental background is a very important part of any plankton study but the authors gave no description regarding oceanography of the study area and during their cruises. These sections must be included in the ms.
Specific remarks.
Title. The authors also presented data on bacterioplankton. Therefore, I recommend providing a more relevant title. For example, Riverine inputs impact the heterotrophic fungus-like protists and bacterioplankton in the coastal waters of the South China Sea.
Keyword. Should be updated with DNA sequencing.
L25. Consider replacing "specialists of" with "common in".
L28. Delete "in".
L32. Consider replacing "influences" with "influence".
L35. Consider replacing "largely changing the environmental factors" with "largely determine changing in the environmental factors".
L35-36. Delete ‘changing environmental’.
L38. Insert ‘biological’ before ‘aspects’.
L42-44. Consider replacing ‘some previous studies further salinity has been reported to display a strong relationship (p < 0.001, r > 0.7) with bacterioplankton community structures’ with ‘some previous studies have been reported strong relationships (p < 0.001, r > 0.7) between salinity and bacterioplankton communities’.
L47. Please, specify both populations. Above you mentioned three different groups.
L50-51. Delete ‘in the microbial communities’.
L56. Delete ‘level'.
L62. Consider replacing ‘low-salt’ with ‘less-saline’.
L72. Consider replacing ‘coastal of Bohai Sea, Osaka Bay and pelagic of South China Sea’ with ‘coastal waters of the Bohai Sea, Osaka Bay and pelagic waters of the South China Sea’.
L92. Consider replacing ‘provide’ with ‘provides’.
L95, 103. Consider replacing ‘2 with ‘two’.
L110. Consider replacing ‘of each station‘ with ‘at each station‘.
L112. Consider replacing ‘method defined‘ with ‘method proposed‘.
L113-118. This section should be moved in the Results.
L133. Consider replacing ‘in previous study‘ with ‘in a previous study‘.
L161. The Shannon index may be calculated based on log2 or ln. Please, specify which logarithmic base was used in your calculations.
Results. The authors mentioned that they measured temperature and salinity during their cruises. However, there are no data regarding environmental conditions. I strongly recommend including a section to describe spatial variation in the environmental conditions in the study area in 2016 and 2018. In this section the authors must clarify the criteria to delineate three different habitats – plume, offshore and pelagic waters. A map showing the location of the three different habitats would be desirable to illustrate the main patterns. Therefore, the map presented as supplementary material (Fig. S1) must be included in the main ms.
L171-172. Consider replacing ‘along a plume, offshore and pelagic gradient’ with ‘along plume, offshore and pelagic gradients’.
L175, 192. Consider replacing ‘3’ with ‘three’.
L176. Consider replacing ‘LP average’ with ‘average LP’.
L178. Consider replacing ‘in the waters of offshore’ with ‘in the offshore waters’.
L178-179. Delete ‘trend patterns of’.
L181. Insert ‘LP abundance’ before ‘occurred’.
Fig. 1 caption. I suggest replacing with ‘Figure 1. Average abundances of LP (a) and bacterioplankton (b) along a plume to pelagic gradient in the South China Sea in 2016 and 2018.
L198-200. This section is more appropriated for the Discussion and should be moved in this part of the ms.
L205. Consider replacing ‘Kruskal test’ with ‘Kruskal-Wallis test’.
L251. Consider replacing ‘accounted 21.8% and 32.9% of the total sequences of offshore and pelagic’ with ‘accounted for 21.8% and 32.9% of the total sequences in offshore and pelagic waters’.
L256. Delete ‘were largely’.
L259. Insert blank after ‘habitat’.
L263. Consider replacing ‘had been reported’ with ‘has been reported’.
L265. Consider replacing ‘of offshore’ with ‘in offshore waters’.
L267. Consider replacing ‘offshore’ ‘offshore waters’.
L276. Consider replacing ‘brackish salinity’ with ‘lowered salinity’.
L278. Delete ‘amount’.
L279. Delete ‘take place’.
L280. Consider replacing ‘the stimulus’ with ‘a stimulus’.
L284. Delete ‘reported to be’.
L289. Consider replacing ‘reported LP growth rate decreased’ with ‘reported decreased LP growth rate’.
L293. Consider replacing ‘, which conflicted’ with ‘that contradicts’.
L309-310. Consider replacing ‘Ulkenia zoospores were intensities associated’ with ‘that Ulkenia zoospores were associated’.
L317. Consider replacing ‘As Aplanochytrium was charactered’ with ‘Aplanochytrium was characterized’.
L318. Consider replacing ‘It suggested’ with ‘It suggested that’.
L322. Insert comma after ‘(LP)’.
Author Response
Please see the attachment.
Author Response File: Author Response.docx
Round 2
Reviewer 3 Report
Second review for the paper "Riverine inputs impact the diversity and population structure of heterotrophic fungus-like protists in the coastal waters of the South China Sea" by Suisui Wang, Kalyani Sen, Yaodong He, Mohan Bai, Guangyi Wang submitted to "Water".
The authors have addressed my concerns and revised the manuscript.
Some text revisions are still required.
L36. Consider replacing ‘salinity gradient’ with ‘a salinity gradient’
L43. Consider replacing ‘some previous studies have been reported’ with ‘some previous studies have reported’
L49. Consider replacing ‘were also found different’ with ‘were also found to be different’
L125. Consider replacing ‘Map of sampling stations in South China Sea’ with ‘Map of sampling stations in the South China Sea’
L170. Consider replacing ‘calculated based ln’ with ‘based on natural logarithm’
L179. Consider replacing ‘in range’ with ‘in ranges’
L235. Consider replacing ‘no significantly’ with ‘no significant’
L238. Consider replacing ‘did not the main factor affecting’ with ‘did not affect’
L320. Consider replacing ‘diversity changing’ with ‘diversity patterns’
L323. Consider replacing ‘decreased LP growth rate’ with ‘a decrease in an LP growth rate’
Author Response
Please see the attachment.
Author Response File: Author Response.docx
