Feeding Habits and Diet Overlap between Brown Trout Lineages from the Danube Basin of Croatia

Round 1

Reviewer 1 Report

I have read the manuscript with great interest. This is an original study that required a lot of work. The authors collected Salmo trutta from 10 different rivers in 3 different regions of Croatia. All captured individuals are subjected to genetic analysis to determine their origin. Further, the contents of the gut of all individuals were analyzed.

The material is small - only 80 fish, of which 8 fish with empty gut were excluded from the analysis. There are 7 AT, 20 DA and 45 hybrids left, i.e. a total of 72 fish from 10 rivers: AT were caught only in 4 rivers out of 10, DA - in 8 rivers, and only hybrids - in all rivers. The fish differed in length up to 3 times. The catch was carried out on different rivers in different years and in different months (April and May). To solve a scientific problem - comparing the diet of fish of different origins, it would be more correct to catch a sufficient number of fish in one river strictly at a certain time. But it is very difficult to do this in small rivers, where the population of brown trout is small.

The authors analyzed the obtained results quite carefully and came to the conclusion that there are no significant differences in feeding between brown trout of AT and DA lineages. The authors explain this by the fact that brown trout is an opportunistic euryphage fish and feeds on everything that is currently available in the water bodies. But such a conclusion would be more convincing if the authors would have offered another option for explaining such a similarity - an option related to the sensory physiology of brown trout. There is data that shows that the taste preferences of brown trout from different populations, even geographically distant ones, are very close! (see: Taste preferences of the brown trout Salmo trutta from three geographically isolated populations // J. Ichthyology. V. 45. No. 1. P. 111–123). All prey caught by fish are going through orosensory control, mainly by taste receptors, and only those prey that meet the fish requirements are swallowed by the fish. And if brown trout of different populations (=origins) have similar taste preferences, then food of brown trout in the same location should be the same. It should be noted that taste preferences in fishes are genetically determined and highly stable for environmental variables.

I don't really like the title of the article. The authors did not study food preferences, so "Feeding preferences" should be removed from the title. Since the material, as the authors themselves note, is small and may not accurately reflect reality, it was possible to change the title to something like that:  "Studies of diet overlap between brown trout lineages from the Danube Basin of Croatia".

 Some other remarks:

Lines 32-39: In this paragraph, it should be stated as clearly as possible whether or not the Black Sea line Salmo trutta lives in Croatia in the Danube basin. It is important. Now there is a lot of interesting information in this paragraph, but there is no information about the local brown trout.

Lines 40-47: It should be said about the origin of the Atlantic brown trout line, which was introduced into the rivers of Croatia (what populations of the Atlantic brown trout line were used for introduction?).

Line 53 and 63: phrase "food items of terrestrial origin captured near the surface" is unfortunate

95: Jankovac seems to be replaced by Jankovački potok, as indicated on line 77 and in Table 1.

Line 101: I consider the information about the classification of the fish into AT and DA lineages or their hybrids to be completely insufficient. It is strongly important to convincingly show how and by what criteria such a classification was made.

Lines 102-103: Bernatchez (1992) is not presented in references list

Line 171: since 2 fish were not classified, authors should talking about 80 fish analyzed, not 82.

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report

The Authors of the reviewed manuscript titled "Feeding preferences and diet overlap between brown trout lineages from the Danube Basin of Croatia" have put forth a relatively concise, but well-prepared paper which focuses on local riverine populations of the brown trout. The manuscript is of high quality, as there are basically no linguistic mistakes to be found throughout the entire text (which is truly a rarity) and there is almost nothing to complain about from a substantive standpoint. I only have pointed out one issue regarding the M&Ms section.

As the Editors will likely point out, the formatting of references within the text should be changed to a numerical arrangement.

Below, I have arranged a section-by-section commentary.

Abstract and Introduction: Excellently written, no complaints here. Everything is clear and comprehensive in both sections. I did not spot any linguistic mistakes, not even typos. Superb work.

Materials and Methods: This section is quite concise and I do not have any objections except for one important issue: the lack of any description of the genetic classification method, which allowed to separate the fish into the three studied lineages (DA, AT, HY). I believe that simply referring the Kanjuh et al. 2020 paper here is a too big of a shortcut. After all, this classification is a crucial component of the whole analysis. Therefore, at least a short paragraph (or subsection) could be added which would outline the major features of that lineage identification process, including the pickup of samples for genetic analyses and some main points of laboratory work, as well as software processing. Such addition would help much with the overall reception and interpretation of the manuscript. This final effect could also be achieved by preparing a supplementary file with a detailed description.

Results: It is highly commendable that the Authors kept this section relatively short, but have presented an abundance of data in appendices, maintaining an excellent readability of the manuscript even throughout this section.
The arrows and descriptions of prey types in Figure 2 are of the same color and are stacked closely one to another - is there some way to add clarity to this graph? Maybe use different colors? It is simply an advice, but certainly one worth considering.
There seems to be some issue with the interline of figure descriptions, but I believe that this issue will be resolved by the editors. Furthermore, Figures 3 and 4 partially disappeared due to improper positioning to the right, but they appear to be correct.

Discussion: Comprehensive and conclusive, no objections here. Good work.
Line 319: Correct style to "..., no sufficient samples of AT lineage have been found for further confirmation of this finding".

Author Response

Please see the attachment

Author Response File: Author Response.docx