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Article
Peer-Review Record

Experimental Study of the Quantitative Impact of Flow Turbulence on Algal Growth

Water 2021, 13(5), 659; https://doi.org/10.3390/w13050659
Reviewer 1: O. R. Anderson
Reviewer 2: Anonymous
Water 2021, 13(5), 659; https://doi.org/10.3390/w13050659
Received: 12 January 2021 / Revised: 4 February 2021 / Accepted: 8 February 2021 / Published: 28 February 2021
(This article belongs to the Special Issue Algae: Indices of Water and Ecological Quality)

Round 1

Reviewer 1 Report

The revisions that I recommended have been adequately addressed by the authors.

Author Response

We greatly appreciate your comments. Thank you very much.

Reviewer 2 Report

The Manuscript “Experimental study of the quantitative impact of flow turbulence on algal growth” requires revision before accepted for publication. The specific comments are given below.

  1. Remove abbreviations from the summary. Explain in the text.
  2. Define the research hypothesis.
  3. Ln 38 Remove numbering.
  4. Ln 79 Describe exactly why Microcystis aeruginosa was tested.
  5. Please indicate the manufacturer, city, country when mentioning the equipment.
  6. Ln 105-113 Remove numbering.
  7. Table 2 Center Maximum and Minimum.
  8. If you wrote “significant”, did you mean statistically significant? E.g Ln 177 “lower significantly”, Ln 182 “increased significantly”. Give significance level.
  9. Add the standard deviations in the results listed in the text.
  10. Expand the discussion. Compare and describe the accurate results other researchers have obtained.
  11. Remove the numbering in the Conclusions chapter.

Author Response

Response to Reviewer 2 Comments

Manuscript ID: water-1090928

 

30 January 2021

 

Dear Editor and reviewers,

Thank you very much for your letter of 24 January 2020, which informed us that our manuscript "Experimental study of the quantitative impact of flow turbulence on algal growth" (Manuscript ID: water-1090928) can be considered for publication in Water after major revisions. We appreciate both your help and the referees' concerning the improvement of this paper.

We have revised this paper and prepared a detailed reply to reviewers' and editors' comments in the attached file of "Responses to reviewers_water_1090928". We have read all questions carefully and made corresponding revisions as suggested. Besides carefully considering the comments and making corresponding revisions, we have read the manuscript several times and made some revisions based on our understanding. We hope that you find these revisions acceptable and thank you for your kind consideration of our manuscript.

 

Point 1: Remove abbreviations from the summary. Explain in the text.

 

Response 1: The abbreviation (CFD) has been removed from the abstract and explained in Line 183 of the revised manuscript.

 

Point 2: Define the research hypothesis.

 

Response 2: We revised the Introduction chapter (Line 89-99 in the revised manuscript). The previous research results showed poor comparability because of different parameters selected to characterize the turbulence intensity. And a lack of functions to describe the effect of flow turbulence on algal growth.

And we reworded the research hypothesis (Line 94-99 in the revised manuscript).

 

Point 3: Ln 38 Remove numbering.

 

Response 3: Revised as requested.

 

Point 4: Ln 79 Describe exactly why Microcystis aeruginosa was tested.

 

Response 4: Microcystis aeruginosa is the most typical bloom all over the world. And Microcystis blooms can initiate severe environmental and ecological events. However, the effect mechanisms of hydrodynamics on Microcystis aeruginosa remain unclear. Therefore, hydrodynamics (characterized by turbulence intensity) on the growth of Microcystis aeruginosa was studied in this study. (Line 35-40 in the revised manuscript)

 

Point 5: Please indicate the manufacturer, city, country when mentioning the equipment.

 

Response 5: We have made corrections (Line174-178 in the revised manuscript).

 

Point 6: Ln 105-113 Remove numbering.

 

Response 6: Revised as requested.

 

Point 7: Table 2 Center Maximum and Minimum.

 

Response 7: Revised as requested.

 

Point 8: If you wrote "significant", did you mean statistically significant? E.g Ln 177 "lower significantly", Ln 182 "increased significantly". Give significance level.

 

Response 8: The level of significance level was set at p< 0.05. We have made corrections (Line 234, 279, 296, and 340 in the revised manuscript).

 

Point 9: Add the standard deviations in the results listed in the text.

 

Response 9: We added Figure 2b, 4b (the standard deviations of cell density and concentration of Chl-a) and redrew Figure 3, 5.

 

Point 10: Expand the discussion. Compare and describe the accurate results other researchers have obtained.

 

Response 10: We revised the discussion (Line 391-470 in the revised manuscript).

 

Point 11: Remove the numbering in the Conclusions chapter.

 

Response 11: Revised as requested.

 

In addition:

 

We added "3.4 Turbulence effect function parameters calibration and model validation" in the Result chapter.  In the section, we proposed an exponential function to incorporate the effect of flow turbulence, calibrated the function parameters, and validated the function (Line 349-389 in the revised manuscript).

Author Response File: Author Response.docx

Round 2

Reviewer 2 Report

Thank you for considering my comments.

Author Response

Dear editor,

  Thanks for your comments. We have revised the English language and style of the manuscript.

Author Response File: Author Response.docx

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.


Round 1

Reviewer 1 Report

Review: Experimental study of the quantitative impact of flow turbulence on algal growth

by Zhang et al.

 

Summary: Experimental work linking turbulence conditions to growth of bloom producing algae; recreating turbulence in a beaker as energy dissipation rate (the amount of energy lost by the viscous forces in the turbulent flow) is unique but is not representing what Cyanobacteria will experience; turbulence in aquatic habitats mixes particles, as a result reduces light availability and particles are damaging cell membrane covers.

 

I am a little bit confused if the photosynthetic efficiency was highest at ε was 0.19 m2s-3 
why that did not coincide with highest growth rate, that was not addressed clearly. The growth rate of Microcystis aeruginosa (Kützing) Kützing was the largest when ε was 6.44×10-2 m2s-3, that was the second lowest turbulence in the experiment,

 

The experiment was done on one cyanobacterium species, so generalizations for all algae are unacceptable.

 

My biggest problem is understanding how the authors counted cells in considering Microcystis aeruginosa exhibits complex irregular colonies of different size. Were the colonies broken? then where were the cells in the beakers, as a cyanobacterium this species gathers on the surface to access higher light intensity. Where were the samples collected from? surface of the beaker or below surface? What amount was collected? How many cells were counted?

 

what software was used for comparisons? what results are significantly different?

 

were there replicas of the treatments?

 

Specific suggestions:

None of the bar graphs have error bars or any means for comparison (see fig. 5), so I have hard time comparing between treatments and getting to the same conclusions as the authors.

 

When introducing a Latin binomial name, you have to provide author’s name; for example, you should state that publication in 2003 of

Anabaena circinalis Rabenhorst ex Bornet & Flahault should related with the current taxonomic designation of this taxon as Dolichospermum circinale (Rabenhorst ex Bornet & Flahault) P. Wacklin, L. Hoffmann & J. Komárek

Other suggestions:

-the end of introduction has no period at the end?

-there are many missing punctuations (see ‘the mean maximum and minimum values ‘ should be ’the mean, maximum, and minimum values) 


- labels like in fig. 1 to the top left that are confusing.  

Fig. 7 is impossible to view.

All figure captions have to be re written and informative

Reviewer 2 Report

This is an interesting study, and with a focus on a cyanobacterium it adds additional data on the effects of hydrodynamics on algal blooms beyond some of the eukaryotic data previously published. I have a few comments and suggestions to make.

Line.  Comment

93-94.  The authors report that RPM was set at 0, 100, 150, 200 RPM, the corresponding value in Table 2 for 200 RPM is 0.19 for the average ε. However, in lines 124 to 125 the range is stated as ε = 7.4 x 10-3 – 0.62 m2/s3. The latter in Table 2 is for 300 RPM, not 200 RPM. The 200 RPM is listed in Table 2 as 0.19. Can authors clarify?

  1. Need to clarify how is the chlorophyll a concentration expressed, is it per cell or per unit volume of culture medium or what? I see in Figure 4, that the units are µg/L which suggests it is per L of culture medium. But, this should be made explicit in the Methods section.

Table 2.  The title wording and the wording of column 1 should be “Rotor speed”  OR if the reference is to the motion of the fluid it should be “Rotational speed” not Rotate speed.

To be clear, in the table column headings, use whole words for Average, Maximum and Minimum, there is sufficient space for the whole words.

133 reword “---- the growth curves of Microcystis aeruginosa began----"

134-135           Was there a statistical test for significance, or is this a qualitative judgment?  If the experiments were repeated only twice, then I do not see how a statistical test would be applied for the pairwise data, but if the data were compiled across the time span from day 7 forward it may be possible.

I am not particularly concerned if no statistical test was done for the data in this graph, the differences in the treatment and control curves for algal biomass are at least twice or higher. However, we also need to know if the plotted values are the means of two or more replications (as per line 101), or what?

Line 142-143.  It is clear that the growth trace for 150 RPM is higher than the other velocity curves, but again, unless we know what the standard error bars are on the data points, it is difficult to know if this is statistically significant. The ratio of the 150 RPM growth rate to the other two is on the order of 1.2 x.  I do believe that the column graphs require the addition of an error bar for each column, including comments in the text about the magnitude of the errors and implications for interpretations.

Figure 3. This is a plot of the average growth rate and is consistent with the biomass curve data, but again we need standard error bars on the column graph data to make some judgment about the significance of the differences in the column graph data.  At least, some mention of how many measurements (replicates) were made to obtain the averages is also recommended.

152-154.  The authors state “After six days of culture, all experimental groups entered the logarithmic growth phase, and Chlorophyll-a concentration increased logarithmically. However, there were differences among experimental groups.”

What statistical evidence confirms the conclusion that there were differences among the experimental groups. The coordinate points for  most of the trend lines are identical or almost identical.  There are no error bars on the coordinate points, so no more exact judgment can be made.  At best, the authors can only claim the trend in the experimental groups is greater than in the control group, unless some additional statistical evidence is presented to back up their claims for differences within the experimental treatments.

166  recommend rewording

“Furthermore, the maximum increase was found at 200 RPM, up to 175.53% compared to the control group, which indicated --------"

168-169.  Rewording suggestion “On the 6th day, algae entered the logarithmic growth phase. All of the groups’ photosynthetic rates, as measured by O2 production, increased significantly; and there was little difference among them.”

  1. The authors need to modify this sentence to be more discriminating in the scope of the claim. First, there is only minimal (no statistical) evidence that 150 RPM produces greater algal biomass, with the exception possibly of Fig. 3 that provides more synoptic evidence that the 150 RPM treatment does yield a higher average growth rate. Other data provide less convincing statistical evidence that there are major differences among the RPM treatments.

That is, instead of the vague statement of this sentence in line 193, a more nuanced wording is recommended. “In the experiment, the flow turbulence promoted Microcystis aeruginosa, particularly the growth rate (e.g., Fig. 3), notably at 150 RPM. This is probably due to two mechanisms: (1) ---------.

203-204. Here also, the authors need to recognize that nutrient flux is only one of the likely limiting conditions. CO2 flux (which is  technically not a ‘nutrient’) also could contribute and needs to be recognized. That is the static boundary layer could limit CO2 and or HCO3- available to the cell due to their depletion in the boundary layer.  Recommended rewording as follows – or some variation of it.

“This is consistent with the research results of our team. We discovered more substantial turbulence promoted Microcystis aeruginosa growth that may be attributed to greater availability and  uptake of essential dissolved resources such as nutrients and/or CO2, due to disruption of the static boundary layer.”

218   small rewording “------- the rate decreased noticeably [10].”

220-221. This sentence is correct that in single-celled organisms the growth of the population is produced by binary division. However, this is true for most eukaryotic single-celled algae as well. So, the authors need to clarify this sentence, because the prior sentences were comparing this cyanobacteria to eukaryotic algal cells.  They need to recognize this difference first before launching into a statement about cell division and population growth that is characteristic of most single-celled biota.

That is in Line 220 add some additional text such as

“However, as a cyanobacterium, the single-celled Microcystis aeruginosa may exhibit some particular response patterns different from single-celled eukaryotic algae that were studied in the preceding reviewed research. Nonetheless, as with many single-celled organisms, the growth of the algal population is a result of binary division, and it is difficult to distinguish growth and proliferation from cell division.”

  1. reword. “Parameters b and c affect the sensitivity of the function.”
  2. “---turbulence on ?(?) becomes more sensitive, b=0.01344 ± 0.00770 m4/s6.”

Further suggestions for the Discussion

Laboratory simulations and microcosms do provide a more controlled analysis and evaluation of phenomena in the natural environment that may not be possible in-situ; but of course lab simulations introduce conditions and circumstances that are not equivalent to the natural environment. For example, the vortices in a fluid-filled beaker generated by a stirring bar, may not be equivalent to more straight-line currents in a larger volume in the natural environment. Some laboratory generated vortices may partially simulate natural spiral eddies that set up in an aquatic natural environment; but there are also other forces in the lab experimental apparatus (such as the impact forces of the impeller stir bar on the suspended algae) that would not necessarily be present in the natural environment.  Some of these aspects of the laboratory experiment should at least be recognized.

 

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