Effect of a Corn Silage-Based Finishing Diet on Growth, Carcass Composition, Meat Quality, Methane Emissions and Carbon Footprint of Crossbred Angus Young Bulls

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The general idea of this study is based on the use of agricultural by-products (spent grains from brewing) and corn silage as available and inexpensive feedstuffs for fattening or raising calves, instead of relying on expensive concentrated feedstuffs (imported grains). This idea is not new and is already being applied in many countries around the world, particularly in some African countries. What new features does this study offer, from the author's perspective?

The English language of the text is acceptable but could be improved. For example, lines 92-94 are unclear and need to be rephrased.

The flow of ideas is good, although I, as a reader, felt bored due to the lengthy and prolix explanation of some points. On the other hand, some points (the in vitro experiment, for example) were not adequately explained, and I did not understand their purpose.

Some of the research methods are very old (the in vitro experiment), but most of the research methods are acceptable in terms of modernity.

The effect of the treatments on animal health was not studied, and no blood samples were taken to verify this.

I believe that the environmental impact of the treatments (methane and carbon dioxide production) could have been supported through less expensive and more accurate in vitro experiments than those used in this research. I am unsure why the researcher did not conduct the required laboratory measurements in this regard, since he conducted a laboratory in vitro experiment.

What is meant by "crude fat" in Tables 1 and 2?

In Table 3, the reader may mistakenly understand that the animals consumed more feed in the TMR treatment than in the control. It is well known that feed intake is only measured based on dry matter, so please delete the first row from the table.

The high moisture content in the spent grains left over from beer production makes them a suitable medium for fungal growth. Here, the author did not explain anything regarding fungal growth or any measurements of mycotoxins, which have a significant negative impact on animal health and the quality of the resulting meat.

Meat quality factors vary depending on consumer taste, but in my country, relying on less concentrated feedstuff in bull’s feeding results in lower meat quality and, consequently, a lower meat price. I believe the selling price of the meat should also be mentioned to accurately assess production efficiency from an economic perspective.

The author stated that using TMR saved 40% of the feed cost compared to the control group. When I calculated the cost of producing a kilogram of meat, I found that it did indeed decrease, but only by approximately 28%, not 40%. Please check.

The conclusion is acceptable, but I would have liked it to be more in-depth. Overall, the research team has made a sufficient effort. Therefore, I recommend publishing it after making some of the modifications I mentioned earlier.

Author Response

R1 - The general idea of this study is based on the use of agricultural by-products (spent grains from brewing) and corn silage as available and inexpensive feedstuffs for fattening or raising calves, instead of relying on expensive concentrated feedstuffs (imported grains). This idea is not new and is already being applied in many countries around the world, particularly in some African countries. What new features does this study offer, from the author's perspective?

AU: We recognize that the main idea is not new and has been applied in several regions, but even so, it is important to confirm it in more intensified production system, as those used in the South-western Europe where the finishing of beef cattle is commonly done based on concentrate compound feeds. Moreover, we measured the in vivo methane emissions, estimated the carcass composition of animals, and calculated the carbon footprint of both nutritional strategies. These techniques are not commonly applied together in most studies addressing the general issue of replacing imported grains with locally produced forages.

Thus, the new features involve a comprehensive approach that combines productive performance, carcass composition, in vivo methane and carbon dioxide emissions, meat quality, carbon footprint, and feed costs in a trial that replicated the practical conditions used by producers.

Moreover, this is an extension of previous studies of our team where we used haylages where in the present study we used maize silage. A commercial company is currently applying this approach in production, and we hope that this paper and the example of the company will encourage the farmers from countries dependent from imported grains to finish the beef cattle with alternative feeding strategies. 

 

R1 - The English language of the text is acceptable but could be improved. For example, lines 92-94 are unclear and need to be rephrased.

AU: Thanks for pointing that out. We rephrased the sentence.

R1 - The flow of ideas is good, although I, as a reader, felt bored due to the lengthy and prolix explanation of some points. On the other hand, some points (the in vitro experiment, for example) were not adequately explained, and I did not understand their purpose.
AU: We tried to explain clearly all the methodology used. In fact, we did not conduct an in vitro experiment per se. The only use of an in vitro measurement was for the estimation of the digestibility of diets. That value is critical to estimate the metabolizable energy of diets. The Tilley & Terry in vitro digestibility is a classic method used in most Animal Nutrition lab a routine method in the feed evaluation. It is an in vitro method where the feed samples are incubated with rumen fluid in the first stage and then treated with acid pepsin.  The Animal Nutrition Lab of the INIAV provides that service for their researchers, and we used it. Thus, the In vitro digestibility values presented should be interpreted just as one more “parameter” characterizing the diets nutritional value and not an “in vitro experiment” conducted by us.

R1 - Some of the research methods are very old (the in vitro experiment), but most of the research methods are acceptable in terms of modernity.

AU: Yes, some of the methods used, despite being already classic, are still the currently widely used and valuable.

R1 - The effect of the treatments on animal health was not studied, and no blood samples were taken to verify this.

AU: The animals remained all apparently healthy and thus we did not monitor their blood clinical parameters. We acknowledge that this would be important complementary data and will be aware of that in future experiments.

 

R1 - I believe that the environmental impact of the treatments (methane and carbon dioxide production) could have been supported through less expensive and more accurate in vitro experiments than those used in this research. I am unsure why the researcher did not conduct the required laboratory measurements in this regard, since he conducted a laboratory in vitro experiment.

AU: We understand the reviewers’ suggestion. We indeed have access to equipment to measure methane production in vitro and have been conducting several in vitro experiments in previous years. In vitro experiments are less expensive and ideal to screen several treatments (i.e. additives, plants, etc) or to test in a well-controlled condition some hypothesis difficult to test in vivo. However, the in vitro method only provides us with an first indication, and it is not possible to translated quantitatively the emissions to in vivo real conditions. One of our objectives was to quantify the methane production (g/d) in our feeding and housing system and for that in vivo methods are required.

 

R1 - What is meant by "crude fat" in Tables 1 and 2?

AU – Crude fat (also known as ether extract) is one of the basic Wende analytical parameters used in food and feed evaluation (such as dry matter, ashes, crude protein and crude fiber). The crude fat is obtained after extraction of the feed samples with petroleum ether and comprise lipids, pigments, waxes and every material that can be extracted with that solvent.

R1 - In Table 3, the reader may mistakenly understand that the animals consumed more feed in the TMR treatment than in the control. It is well known that feed intake is only measured based on dry matter, so please delete the first row from the table.

AU: The reviewer is right that dry matter (DM) intake should be considered for most uses. We provide both the “as fed” intake and the dry matter intake because total DM can in special situations be compromised by the bulk of the feed ingested “as fed”. Also, if you are feeding bovines, it is not irrelevant if you have to feed 12 kg or 24 kg per animal per day.

However, to avoid confusion we accept the reviewer suggestion and deleted the “as fed” intake values from the table and results section.

 

R1 - The high moisture content in the spent grains left over from beer production makes them a suitable medium for fungal growth. Here, the author did not explain anything regarding fungal growth or any measurements of mycotoxins, which have a significant negative impact on animal health and the quality of the resulting meat.

AU: Indeed, we did not address this issue, as we found no evidence of major notable fungal growth in our practical test, although we detected an odor alteration. Unfortunately, we did not monitored blood health indicators neither the mycotoxin content. Nevertheless, we address these type of concerns and clearly state that we can not exclude the negative effects of using  Brewer's spent wet grains on diet palatability, potential health issues and growth on the discussion and added a new reference supporting that. (lines 469–473)

 

R1 - Meat quality factors vary depending on consumer taste, but in my country, relying on less concentrated feedstuff in bull’s feeding results in lower meat quality and, consequently, a lower meat price. I believe the selling price of the meat should also be mentioned to accurately assess production efficiency from an economic perspective.

AU: We appreciate the comment, and it is true that consumers preference varies among regions and countries. The sensory quality of the meat was assessed by a panel of trained tasters and even if the “overall acceptability” and “flavour” evaluation might bias by regional preferences, other sensory parameters evaluated   like tenderness and juiceness are more universal. Moreover, the meat colour, pH, shear force and fatty acid profile are all objective quality traits. In the European Union, these factors do not determine directly the selling price. The officially the selling price of meat is determined by the ‘SEUROP’ carcass grading classification – that is based only on the conformation and fat cover of the carcass. Nevertheless, in practice, if a batch animals are fairly homogenous, a common price is often applied to the all batch. That was the case of these animals where carcass were paid at the same price (5.8 euros/kg) irrespective of the fact that 70% of the carcass were graded as R2 and 30% were graded as U2.

 

R1 - The author stated that using TMR saved 40% of the feed cost compared to the control group. When I calculated the cost of producing a kilogram of meat, I found that it did indeed decrease, but only by approximately 28%, not 40%. Please check.

AU: Yes, you are correct, but that is exactly what is described in lines 298-300, i.e., in feed cost in €/day, the decrease is 40%, and when we convert feed cost to kg Live Weight gain, the decrease is 28%.

 

R1 - The conclusion is acceptable, but I would have liked it to be more in-depth. Overall, the research team has made a sufficient effort. Therefore, I recommend publishing it after making some of the modifications I mentioned earlier.

AU: We appreciate all comments and have made most of the suggested modifications. We tried to elaborate more on the potential implications of our data on the improvement of the sustainability of the cattle finishing systems commonly used in south-western Europe.

Reviewer 2 Report

Comments and Suggestions for Authors

The topic is highly relevant, as it addresses the sustainability of beef cattle production by linking animal performance, meat quality, and greenhouse gas emissions.

The experimental design is clearly described, and the results have practical implications.

The description of methane measurement using the GreenFeed system is appropriate; however, the high individual variation observed should be more thoroughly discussed as a potential source of uncertainty. It is also suggested to provide more detail on how social dominance among animals was managed, since this factor could bias data collection.

The life cycle assessment (LCA) considered only the finishing phase (12 weeks), disregarding earlier stages (breeding, rearing), which may account for up to 80% of total emissions. Although this limitation is acknowledged, it is important to emphasize to the reader that the results are partial and do not represent the full carbon footprint of beef production.

Regarding meat lipid composition, it would be valuable to further discuss the potential implications for human health, given the article’s focus on sustainability and meat quality.

Some tables (e.g., fatty acid composition) are overly detailed. It is recommended to summarize the key findings within the main text and move the full datasets to the supplementary material.

The discussion could be made more concise by avoiding duplication between the presentation of results and their interpretation.

Recommendation

I recommend publication after minor revisions, particularly with respect to:

Highlighting the partial nature of the LCA (finishing phase only).

Improving the clarity of results presentation by summarizing extensive tables.

Expanding the discussion on the nutritional implications of the meat lipid profile for consumers.

Author Response

Reviewer 2

R2 - The description of methane measurement using the GreenFeed system is appropriate; however, the high individual variation observed should be more thoroughly discussed as a potential source of uncertainty. It is also suggested to provide more detail on how social dominance among animals was managed, since this factor could bias data collection.

AU: Thank you for the pertinent comment. Indeed, we agree that this deserves to be further discussed and revise this section accordingly (Lines 529-536). The individual variation is probably due to the genetics of the animals and the rumen microbiota of each animal. This individual variation is normal, and to prevent it from affecting the results, the groups of animals in each pen were distributed considering the farm of origin in an attempt to avoid either genetic and environmental factor bias that might affect animal responses.

About the social dominance, the reviewer is also right, and we added new information about this in the M&M (lines 145 - 147). However, we would like to explain that in this specific trial, we did not feel that dominance among animals was a problem during visits to the GF, perhaps because they were a calm breed (Angus) and also because they were of the same age group and live weight. In fact, we had experienced, in other experiments, a serious dominance problem in the access to GF with grazing cows of different ages.

 

R2 - The life cycle assessment (LCA) considered only the finishing phase (12 weeks), disregarding earlier stages (breeding, rearing), which may account for up to 80% of total emissions. Although this limitation is acknowledged, it is important to emphasize to the reader that the results are partial and do not represent the full carbon footprint of beef production.

AU: Yes, the reviewer is correct, and we point this out at various stages of the work (Materials and Methods, Results, and Discussion). Also, in the title of the table showing these results, we have ‘life cycle analysis for the finishing period (12 weeks)’, but in accordance with the comment, we have added the word ‘partial’ to this title, in MM and in Conclusion, when referring to the LCA, to make it clearer to the reader.

 

R2 - Regarding meat lipid composition, it would be valuable to further discuss the potential implications for human health, given the article’s focus on sustainability and meat quality.  Expanding the discussion on the nutritional implications of the meat lipid profile for consumers.

AU: Thanks for the suggestion. We added a paragraph to the discussion about this important topic.

 

16 - Some tables (e.g., fatty acid composition) are overly detailed. It is recommended to summarize the key findings within the main text and move the full datasets to the supplementary material.

AU: Thank you for the comment. We have taken the recommendation into account and reduced Tables 7 and 8 to only the most important results, moving the detailed and complete tables to ‘Supplementary Material’ (Tables S1 and S2).

17 - The discussion could be made more concise by avoiding duplication between the presentation of results and their interpretation.

AU: Thank you for your comment. We have reviewed and tried to identify and delete the repeated content.

Reviewer 3 Report

Comments and Suggestions for Authors

This manuscript is very interesting and this work is huge.

However, I have same suggestion as following:

 

Line 37 lower in the control group than in the TMR group? Need improve.

 

 

Table 1 added the full name of the abbreviate.

 

Table 2 metabolizable energy measured or calculate value.

 

 

Line 343 CH₄, subscript. Double checked.

 

Line 396 to 397, add the p values.

 

Line 413, add “,”

 

Line 415, space.

 

4.1, 4.2, and 4.3 italic whereas 4.4 not.

 

Line 518, Duckett et al. reported that…….

 

 

Line 541, reported by Johnson et al.

 

 

 

 

Author Response

Reviewer 3

R3 - Line 37 lower in the control group than in the TMR group? Need improve.

AU: Thank you for your comment. The abstract has a very limited maximum number of words, so we limit its use. Actually, it is the opposite, so we changed the sentence to make it clearer: “Dry matter intake and average daily gain were 13% and 15%, respectively, lower in the TMR treatment than in the Control treatment.”

R3 - Table 1: added the full name of the abbreviate.

AU: Thank you, it has been corrected.

R3 - Table 2: metabolizable energy measured or calculate value.

AU: The value is calculated. We added this information to the footnotes of the table.

R3 - Line 343 CH₄, subscript. Double checked.

AU: Thank you, it has been corrected.

R3 - Line 396 to 397, add the p values.

AU: Thank you, it has been added.

R3 - Line 413, add “,”

AU: Thank you, it has been corrected.

R3 - Line 415, space.

AU: Thank you, it has been corrected.

R3 - 4.1, 4.2, and 4.3 italic whereas 4.4 not.

AU: Thank you, it has been corrected.

R3 - Line 518, Duckett et al. reported that…….

AU: Thank you, it has been corrected.

R3 - Line 541, reported by Johnson et al.

AU: Thank you, it has been corrected.

Reviewer 4 Report

Comments and Suggestions for Authors

The present manuscript aimed to evaluate the effect of replacing a conventional concentrated finishing diet with a diet based on corn silage and spent brewers' grains on meat quality, carcass composition, and the carbon footprint of finishing beef cattle. Although the replacement finishing diet is a viable alternative due to its production cost, the authors should consider the following points to improve the quality of the manuscript

Abstract
L.31. Include the meaning of TMR.

Introduction 
L.52 and 72. CH4. subscript
L.57. Delete "in"

Materials and methods
L.92. Twenty yearling crossbred males
L.102. Delete "LW"
L.109. The composition of the diets used is shown in Table 1.
L.113. change "are" to "were"
L.132. Please be more clear since in table 2 it is difficult to locate what is mentioned
L.136. Delete "in"
L.176-187. Please review this information as it appears to be a results section rather than a materials and methods section
L.191. Consider adjustments to the wording of this methodology
L.193. It would be good to include the author's name
L.193-194. were determined according to the method described by...
L.204-205. Please modify this sentence so that the pH determination is better understood
L.206. in duplicate is very little to do statistical tests (justify)
L.255-257. Give coherence to these sentences
L.277. Be more specific in the statistical analysis used to compare the two experimental groups

Results
L.298. Check this data
L.338. subscripts in CH4 y CO2. Check throughout the manuscript
L.351. 8.91 or 8.94?
L.352. Check if the data is correct (7.07)
L.366. Include the meaning of ScF and ImF
L.382-384. Considering this variability in the results, was the statistical test performed considering equal or unequal variances? It would be good if it were described in the statistical analysis section
L.432. This is table 8. As a recommendation, tables should be found immediately after they are referred to in the text
L.453. Check the data as it does not correspond to the data in the table
L.455-456. Check the data as it does not correspond to the data in the table

Discussion
L.471. Include the authors' names or change the writing style
L.531-532. Check the wording of this sentence
L.541-544. Check the wording of this sentence
L.553. Delete "and"
L.581. , and rumenic acid (c9,t11-18:2).
L.582. Delete "compared a"
L.642. was

 

Comments for author File: Comments.pdf

Author Response

Reviewer 4

 

R4 - L.31. Include the meaning of TMR.

AU: Thank you for your comment. The abstract has a very limited maximum number of words, so we limit its use, but we now added the meaning of TMR.

R4 - L.52 and 72. CH4. Subscript

AU: Thank you, it has been corrected.
R4 - L.57. Delete "in"

AU: Thank you, it has been corrected.

R$ - L.92. Twenty yearling crossbred males

AU: It was changed to ‘Twenty young crossbred males’ at the request of another reviewer.

R4 - L.102. Delete "LW"

AU: Thank you, it has been corrected.

R4 - L.109. The composition of the diets used is shown in Table 1.

AU: Thank you, it has been corrected.

R$ - L.113. change "are" to "were"

AU: Thank you, it has been corrected.

R4 - L.132. Please be more clear since in table 2 it is difficult to locate what is mentioned

AU: The reviewer is right, because it was wrong – it is Table 1 (not Table 2), it has been corrected.

R4 - L.136. Delete "in"

AU: Thank you, it has been corrected.

R4 - L.176-187. Please review this information as it appears to be a results section rather than a materials and methods section

AU: We understand the comment, because these linear regressions were made using the data from results, so the equations were moved to the Results section, in sub-chapter 3.2.

 

R4 - L.191. Consider adjustments to the wording of this methodology

AU: We have taken into account the comments in this section of the methodology.

R4 - L.193. It would be good to include the author's name

AU: Thank you, it has been corrected.

R4 - L.193-194. were determined according to the method described by...

AU: Thank you, it has been corrected.

R4 - L.204-205. Please modify this sentence so that the pH determination is better understood

AU: Thank you, it has been corrected.

R4 - L.206. in duplicate is very little to do statistical tests (justify)

This repetition is just a laboratory replicate, not an experimental replication. Two replicate are enough when they do not differ more than what is allowed in the analytical protocols (the variation allowed differs among the methods). When that happen, the analysis is repeated.  

 

 

R4 - L.255-257. Give coherence to these sentences.

AU: These sentences were not correctly formatted in the manuscript. Thanks for noting, it is now corrected

 

R4 - L.277. Be more specific in the statistical analysis used to compare the two experimental groups

AU: We rephrased the statistical section and added the general type of analysis that was only implicit. For most of the data we fit a linear mixed model where the Diet was the fixed effect and the experimental unit were the pens, and the animal within the pens were treated as subsamples.

 

R4 -L.298. Check this data

AU: Sorry, we are not sure if understand the issue here. Comparing DM intake in the Control (10.6 kg/d) to that of  TMR diet  (9.2 kg/d) we verify that the latter is  13.2% lower than the Control .

(Control – TMR) / Control = ((10.6-9.2)/10.6) x 100 = 13.2%

R4 -L.338. subscripts in CH4 y CO2. Check throughout the manuscript

AU: Thank you, It was formatted incorrectly, but we have corrected it throughout the manuscript.

R4 -L.351. 8.91 or 8.94?

AU: The value 8.91 (which is the average of the two treatments) is correct.

 

L.352. Check if the data is correct (7.07)

AU: Thank you for noting. The correct value is 7.07. Corrected

L.366. Include the meaning of ScF and ImF

AU: ScF (Subcutaneous Fat) and ImF (Intermuscular Fat) had already been defined in the text previously (lines 209 and 217).

R4 - L.382-384. Considering this variability in the results, was the statistical test performed considering equal or unequal variances? It would be good if it were described in the statistical analysis section

AU: We checked the homogeneity of variance and the variances here high and similar among treatments.

R4-.432. This is table 8. As a recommendation, tables should be found immediately after they are referred to in the text

AU: It has been corrected and moved up, but we have moved most of the table to Supplementary Material, as suggested by another reviewer.

R4 -.453. Check the data as it does not correspond to the data in the table

AU: Thank you, it has been corrected.

R4 -.455-456. Check the data as it does not correspond to the data in the table

AU:Thank you, it has been corrected.

Discussion
R4 -L.471. Include the authors' names or change the writing style

AU: Thank you, it has been corrected.

R4 - L.531-532. Check the wording of this sentence

AU: Thank you, it has been corrected.

R4 - L.541-544. Check the wording of this sentence

AU: Thank you, it has been corrected.

R4 - L.553. Delete "and"

AU: Thank you, it has been corrected.

R4 - L.581. , and rumenic acid (c9,t11-18:2).

AU: Thank you, it has been corrected.

R4 -L.582. Delete "compared a"

AU: Thank you, it has been corrected.

R4 -L.642. was

AU: Thank you, it has been corrected.

 

 

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

I find the use of the term "ether extract" more appropriate than "crude fat" because it indicates the method by which the fat, waxes, dyes, etc., were extracted. I believe that mycotoxin testing of the feed is necessary in addition to blood tests. However, I appreciate the effort you have put into it, so I will accept publication of the paper in its current form.

Author Response

R1: I find the use of the term "ether extract" more appropriate than "crude fat" because it indicates the method by which the fat, waxes, dyes, etc., were extracted. I believe that mycotoxin testing of the feed is necessary in addition to blood tests. However, I appreciate the effort you have put into it, so I will accept publication of the paper in its current form.

 

AU: We replaced the designation Crude fat with Ether extract throughout the manuscript.

We agree that mycotoxin evaluation of feed and blood tests on animals would be very important information to gather and will take that into consideration for future experiments. We thank the reviewer for understanding the absence of that data.

Reviewer 4 Report

Comments and Suggestions for Authors

The manuscript improved significantly after the first round of revisions. However, the authors should consider the following points:

L.98. 50 m^2 (superscript).
L.136. Generally tables referred to in the text are placed immediately after the text, not before.
L.189. described by...
L.272. When comparing two experimental groups, a t-Student test is performed and equal or unequal variances are considered, this must be described.
L.356. In the table it appears as 8.94, not 8.91.
L.592-593. Check the wording of this sentence.

Comments for author File: Comments.pdf

Author Response

R2: L.98. 50 m^2 (superscript).

AU: Thank you, it has been corrected.

R2: L.136. Generally, tables referred to in the text are placed immediately after the text, not before.
AU: We agree and appreciate the comment. In fact, Table 1 is first mentioned in the text on line 110 before it appeared. The mention to Table 1 that appears in Line 136 is already the second reference to that table.

 

R2: L.189. described by...

AU: Thank you, it has been corrected.

R2:L.272. When comparing two experimental groups, a t-Student test is performed and equal or unequal variances are considered, this must be described.

AU: The reviewer is correct. We can compare two treatments just using a t-student test, but we can also compare them with more advanced linear models. The more advanced models also solve the simpler cases that are just particular cases of a wide range scope of possible models. In the current mixed model software, like Proc MIXED of SAS, we can also test the variance of each treatment and conduct the comparison considering the variance of each treatment. That was not needed because the variances did not differ significantly between treatments.

We provide here an example, to the reviewer evaluate our analysis, as sometimes the succinct description that appears in the paper maybe be not so clear, particularly for those that do not use SAS.

As example, for the case of C18:1c9 in muscle the SAS script used was

To test the homogenity of variances we run this model

Proc mixed data = fam;

class diet pen;

model c181c9 = diet;

repeated /group = diet;

Lsmeans diet;

run;

 

And for the output we got the information that the variance of Control is 194158 and the variance of TMR is 61376. The difference seems large but the Likelihood Ratio Test indicates us that the variances do not differ ( P = 0.0924).  

Covariance Parameter Estimates
Cov Parm	Group	Estimate
Residual	Diet CONT	194158
Residual	Diet TMR	61376

 

 

Null Model Likelihood Ratio Test
DF	Chi-Square	Pr > ChiSq
1	2.83	0.0924

 

Confirming that the variances do not differ, we can proceed to the final model with a common variance among the treatments and considering animal as subsampling within the pen.

 

Proc mixed data = FAm;

class dieta parque;

model c181c9 = diet;

repeated /sub=pen(dieta) type=cs;

Lsmeans dieta;

run;

And the output looks like this

 

Type 3 Tests of Fixed Effects
Effect	Num DF	Den DF	F Value	Pr > F
Diet	1	2	3.14	0.2185

 

Least Squares Means
Effect	Diet	Estimate	Standard Error	DF	t Value	Pr > |t|
Diet	CONT	559.10	64.5810	2	8.66	0.0131
Diet	TMR	397.29	64.5810	2	6.15	0.0254

 

We rephrased the statistical analysis section and now clearly state that we checked the homogeneity of variance.

 

R2: L.356. In the table it appears as 8.94, not 8.91.

AU: Thank you for the comment. The sentence is “The CO₂ production values were similar for the two treatments when represented as kg/day, averaging 8.91 kg/d.”
Maybe it's not clear, but the value shown is the average of the values for the two treatments (8.94 and 8.88). Since there are no significant differences between the two treatments, we put the average, which is 8.91.

R2: L.592-593. Check the wording of this sentence.

AU: Thank you for alerting us. We have changed the sentence to make it more clear.