|As I also reviewed the original submission, I am omitting a summary of the manuscript here. |
Although the authors changed the title of their manuscript, the interpretation of the presented data hasn’t changed much (or at least it doesn’t come across to the reader). At the moment it is relatively hard to judge the presented manuscript/data as there are still many cases where the statements/conclusions/claims by the authors are not supported by the results. Unfortunately, the authors do not present any further analyses or statistics to back up their statements (I have given a few examples below; however, this list is not comprehensive!). An overall suggestion to the authors would be to look for someone who would be able and willing to aid with the manuscript re-structuring and re-writing, but who is himself or herself not involved in these experiments (someone in their departments maybe? I think it would help to get an outside-the-box opinion here for the interpretation of the data). At the current state, there seem to be too many misinterpretations in the manuscript in order to recommend publication (even with some revisions). I have made some more specific comments below.
In general, the authors try to make (big) claims about the effect of CO2 on the microbial community; however, basically provide neither analyses/statistics that support these claims nor is it visible in the figures. Further, the authors have not given any type of error on their measurements making it hard to see actual differences. Overall, in many places it seems that the authors see what they want to see (i.e. an effect of CO2 concentrations on the microbial community).
Again, I would recommend to add an overview of the experimental setup to the methods section. I understand that the experimental setup is described in a different manuscript for the special issue. However, as a reader of this manuscript, I actually do not want to have to read another manuscript first in order to find out what the authors did here; at least a summary would be needed here.
Many places in the manuscript are still not written to the extent that they can be easily followed, for example: “The higher loss rates (days 5 to 9; Fig. 3e) resulted in a decrease in abundance, which was stronger for the low fCO2 mesocosms (as illustrated by M1) due to the significantly higher gross growth rates for the high fCO2 mesocosm (represented by M3; Fig. 3b). The positive correlation of Pico I peak abundance with fCO2 on day 13 (Fig. 3h, R2=0.94) was lost upon another decline in abundance. Significantly higher losses at high fCO2, a combination of grazing and lysis, resulted in a more dramatic crash at high fCO2 and abundances becoming similar again around day 17 (Fig. 3a).” This paragraph is simply not understandable for someone not working on these experiments without having to read the sentences multiple times (which disrupts the flow reading dramatically).
The figures were not labeled at the end of the manuscript (no matter whether this is because of forgetting to label them or whether the submission software omitted to do so); so, for the purpose of the review I assumed that the figures were 1-11 in the way they came out of the printer and I am referring to them in that way.
In general, I think that the authors should try to improve the readability of their manuscript by increasing the quality of their figures, e.g., not having figures that span nearly two full pages (see Figure 3) or omitting repetitive data.
Showing the ambient data together with the mesocosm is essential to judge whether changes in the mesocosms were, for example, induced by enclosement or were naturally happening outside as well. This does not mean that, in case changes were induced by enclosement, that differences between mesocosms are not useful; they are valuable data. Not presenting the ambient microbial community leaves the reader with guessing what happened and potentially suspecting that the data was omitted because they would alter the conclusions of the paper. If the authors would like to increase the understandability of their manuscript, I highly recommend adding the ambient data to the current main manuscript figures.
For example, Suppl. Figure 1 shows that changes in the Synechococcus population are pretty much the result of ‘bottle effects’ (I agree that a mesocosm is a large bottle, but it is an enclosement). Is this possibly related to cleaning procedures or stirring of the mesocosms as outlined in Paul et al.? While other groups are coherent with the ambient water.
After looking into Paul et al. for the experimental setup, it seemed that the CO2 concentrations were actually measured during the mesocosms, why aren’t the actual measured concentrations used for any of the plots instead of the targeted concentrations (which were only achieved in the first few days but weren’t maintained)? This seems such an obvious thing to do and a fact that is completely ignored by the authors.
Abstract: “Of these groups 2 picoeukaryotic groups increased in abundance whilst the other groups, including prokaryotic Synechococcus spp., decreased with increasing fCO2.” Looking at both Figure 2a and Suppl. Figure 1a, I seriously do not see where this statement comes from! There is a short period of time (~ 7-16 days) where Syn. are lower in the low fCO2 than in the higher ones but for most of the experiment it is the other way around. Unfortunately, the authors also do not supply any other analysis or statistics that could possibly back up their claim. Unfortunately, these kind of statements and interpretation is occurring throughout the manuscript, which leads me to the suggestion that the current manuscript is far from being acceptable for publication at this time.
Figure 1: The authors claim that the decline in total phytoplankton around the end of phase 2 and throughout phase 3 is due to a decrease in Synechococcus. This is supported by Figure 2a. However, Suppl Figure 1a shows Synechococcus populations staying up rather than declining to concentrations in Figure 1. After doing some research through this manuscript, I found that the difference between the figures is the depth reference here, which is 0.3-10 m for Figure 1 and 0-17 m for Suppl. Figure 1. This difference actually suggests that either the distribution of Synechococcus is not equal throughout the water column or that there is a sinking out of Synechococcus? I don’t know why this is, maybe the authors should think about this deviation which is currently not acknowledged anywhere.
Figure 1: The authors say that the total phytoplankton abundance at the end of phase 1 is significantly higher in the high fCO2 treatment than the low fCO2 treatments. Again, I am really sorry, but I cannot see this in the plots and the authors again don’t provide any analysis or statistics for this. (see p 10 lines 18-20)
Figure 2: Panel c is absolutely unnecessary here (and in following figures); it is the same data as in panel a and its addition here suggests that it is indeed different data and is making the figure unnecessarily large. Panel d: What are the errors on these measurements? I wonder whether these differences are actual changes or just variation? Further, this is comparing growth rates from time point 10 days to growth and loss rates of time points 4-7. In my opinion, this is totally misleading as day 10 is obviously different than 4-7 as can be seen in panel a. (see p 11, l 10-13).
Figure 2a legend: It is not necessary to present 4 (!) different significance levels here. For most claims in the manuscript no analyses or statistics are provided while here it is over the top. Either s.th. is significantly different or not.
p 10, l 21 and following: If CO2 has a strong positive effect on Pico 1 and 2, how can they then be comparable in abundances to the surrounding water? Maybe I don’t understand this sentence.
Suppl. Figure 1: This is (probably) the most informative figure of the entire manuscript.
p 11, l3-5: This claim is not supported by any analysis. Any time series statistics, PCA, similarity analysis, community composition comparisons?
p 17, l 17-18: this is an odd comparison in size fractions and the results are not exclusive of each other.
p 18, l 20-22: There is consistency among the mesocosms maybe, but there is already a deviation to the surrounding water indicating an effect of enclosement, at least on some groups.
Figure S3 is missing!