Effects of Italian Ryegrass (IRG) Supplementation on Animal Performance, Gut Microbial Compositions and Odor Emission from Manure in Growing Pigs

Round 1

Reviewer 1 Report

 

In this manuscript, firstly, Sungkwon Park et al. analyzed the chemical composition of Italian ryegrass (IRG), and then evaluated their animal performance, gut microbial compositions, odor emission from manure in growing pigs. It is meaningful work since IRG is a supplementary feedstuff for pigs. At the same time, the results of the study were well discussed. But there also have many places, especially grammatical structure or common grammatical errors, need to be corrected/added in the article.

• Title of manuscript: I think it should be corrected: Effects of Italian ryegrass supplementation on animal performance, gut microbial compositions and odor emission from manure in growing pigs
• Line 9,12: add -relative to before “abundances”, e.g., increased/decreased relative abundances
• Line 19: delete Volatile fatty acids
• Line 25: add Korea’s national regulation
• Line 42, 45: add –s to effects
• Line 43: add –s to compositions
• Line 51, 52: Complete the number? and year? for guidelines set
• Line 57: delete –s to the end of a diet
• Line 69: were is replaced by was
• Recheck from line 79 to 94: e.g., “BOD concentrations were” are replaced by BOD concentration was
• Line 141: discussion is singular noun, not plural noun
• Line 147: control diet/ no control diets
• Line 157: delete “phrase” for the feeding trail
• Line 162: both … treatments increased significantly ….
• Line 170: should be corrected: Chemical characterization of manure from pigs fed diets increasing levels of IRG
• Line 176: Decreasing effect in VFA was…
• Line 195: ..effected the phenols and indoles levels in …
• Line 197 and 198: should be corrected: Manure …. had lower 10 and 32% phenols than those in control.
• Line 203-204: delete “the” between lower and degradability; between greater and excretion
• Line 219: should be corrected: The chemical parameters of manure are used ….
• Line 245-246: … influenced by changes in bacterial diversity and composition from diet formulation
• Line 250-252: should be presented in materials and methods
• Line 261: add –s to the end of figure: figures
• Line 269-271: delete numbers as 262, ….225
• Line 271: correct: gram-positive/negative bacteria
• Line 273: “included in the families” is replaced by belong to
• Line 281: at phylum level
• Line 284: at genus level
• Line 294: a diet, not a diets
• Line 298: in the colon
• Line 300: …that an increase in relative abundances….
• Line 307: correct: carbohydrate fermentation
• Line 324: delete P < 0.05
• Line 332: rewrite the conclusions
• Line 339-341: rewrite this phrase, it should be corrected: IRG supplementation in pig diets modulate relative abundances of detailed bacteria????

Author Response

Response to Reviewer 1 Comments

Point 1:
Line 9,12: add -relative to before “abundances”, e.g., increased/decreased relative abundances
→ Revised
Line 19: delete Volatile fatty acids
→ Revised
Line 25: add Korea’s national regulation
→ Revised
Line 42, 45: add -s to effects
→ Revised
Line 43: add -s to compositions
→ Revised

Response 1:
All comments of point 1 are revised in manuscript according to reviewer’s opinion.

Point 2:
Line 51, 52: Complete the number? and year? for guidelines set

Response 2:
Sentence is revised according to reviewer’s comment as follow; All procedures were conducted in accordance with guidelines No. 16 set by the National Institute of Animal Science Animal Care and Use Committee at 2008.

Point 3:
Line 57: delete -s to the end of a diet
→ Revised
Line 69: were is replaced by was
→ Revised
Recheck from line 79 to 94: e.g., “BOD concentrations were” are replaced by BOD concentration was
→ Revised
Line 141: discussion is singular noun, not plural noun
→ Revised
Line 147: control diet/ no control diets
→ Revised
Line 157: delete “phrase” for the feeding trail
→ Revised
Line 162: both … treatments increased significantly ….
→ Revised
Line 170: should be corrected: Chemical characterization of manure from pigs fed diets increasing levels of IRG
→ Revised
Line 176: Decreasing effect in VFA was…
→ Revised
Line 195: ..effected the phenols and indoles levels in …
→ Revised
Line 197 and 198: should be corrected: Manure …. had lower 10 and 32% phenols than those in control.
→ Revised
Line 203-204: delete “the” between lower and degradability; between greater and excretion
→ Revised
Line 219: should be corrected: The chemical parameters of manure are used ….
→ Revised
Line 245-246: … influenced by changes in bacterial diversity and composition from diet formulation
→ Revised

Response 3:
All comments of point 3 are revised in manuscript according to reviewer’s opinion.

Point 4:
Line 250-252: should be presented in materials and methods

Response 4:
Sentence is revised according to reviewer’s opinion as follow; The IRG 1.0% group was chosen for analysis of the changes in bacterial community based on overall odor reducing efficiency of manure in the current study. Concomitant to previous result, control (IRG 0%) and IRG 1.0% treatment group showed different correlation in the level of odorous compounds with the structure of bacterial community.

Point 5:
Line 261: add -s to the end of figure: figures
→ Revised
Line 269-271: delete numbers as 262, ….225
→ Revised
Line 271: correct: gram-positive/negative bacteria
→ Revised
Line 273: “included in the families” is replaced by belong to
→ Revised
Line 281: at phylum level
→ Revised
Line 284: at genus level
→ Revised
Line 294: a diet, not a diets
→ Revised
Line 298: in the colon
→ Revised
Line 300: …that an increase in relative abundances….
→ Revised
Line 307: correct: carbohydrate fermentation
→ Revised
Line 324: delete P < 0.05
→ Revised
Line 332: rewrite the conclusions
→ Revised
Line 339-341: rewrite this phrase, it should be corrected: IRG supplementation in pig diets modulate relative abundances of detailed bacteria????
→ Revised

Response 5:
All comments of point 5 are revised in manuscript according to reviewer’s opinion.

Author Response File: Author Response.docx

Reviewer 2 Report

The study assessed the inclusion of Italian ryegrass in the pig diet in different percentages (0.5%, 1% and 1.5%). The aim of the study was to investigate the impact of fermentable carbohydrates present in the Italian ryegrass on the animal performance, odorous chemical, and bacterial composition of manure. This study presents several interesting point in order to reduce the environmental impact of the pig farm facilities, varying microbiota fermentation.

Some suggestions are reported below:

Line 22-24 change in: Odor emitting from swine manure accounted for 34.6 % from that of total livestock facilities and contributed to civil complaints, affecting environment, animal health and production [2].

Line 32 change in: Control of odor generation and attenuation of nutrient excretion depends on the essential formulation and the balance of nutrients in the animal diet.

Line 35-38 change in: In particular, fermentable carbohydrate (FC) containing dietary fiber has been known to relieve constipation, increase nutrient digestibility and improve gut  health, resulting in reducing odor emission via incorporating excessive nitrogen into the microbial biomass nitrogen [7-9, add  DOI: https://doi.org/10.1080/1828051X.2020.1749899].

Line 38-41 change in: Italian ryegrass (IRG) can be cultivated abundantly in Korea and used as a feed ingredient which levels of acid detergent fiber, neutral detergent fiber, acid detergent lignin, and soluble and insoluble dietary fiber and non-structural carbohydrates are similar to those of sugar beet pulp, thus can provide abundant FC [10, add: DOI: https://doi.org/10.1007/BF02194097]

Line 42-43: However, there are limited data on dietary IRG fiber content and the effect of supplementation on odor generation and gut microbial composition in pigs

Line 51-52: Please report the number of ethical permissions

Line 62: Please add more information on drying process of ryegrass

Line 68-70 change in: From each metabolism crates manure was collected at twice weekly, added in enclosed manure storage container, and stored in the freezer (-20°C) until analysis

Line 85-86: delete “The values were determined by the difference in weight between before and after of dry”

Line 93-96: Please some description of know methods (e.g. Kjeldahl) could be resumed

Line 114: Please specify which kind of standard (internal or external) was used for chromatographic determination

Table 4 change the title as follow: “Concentrations of volatile fatty acids and odorous compounds in manure from pigs fed diets with increasing levels of IRG”.

Table 4: Please insert the complete name of volatile fatty acids

Line 203-204 change in: The lower the degradability of fiber is in the hindgut, the greater the excretion of amino acids is.

Table 5: Please improve table editing

Line 271-272: Bacteria of gram positive by 344 (60%) and gram negative by 225 (40%) were classified in these genera

Figure 3: in order to improve the comprehension of the figure could be better to differentiate largely the colours used for each phyla

Line 300: change 5 in five

Line 304, 322 and 338:  considering that microbiota data regard only the comparison between 0 and 1% is better to specify every time IRG (1%)

Author Response

Response to Reviewer 2 Comments

Point 1:
Line 22-24 change in: Odor emitting from swine manure accounted for 34.6 % from that of total livestock facilities and contributed to civil complaints, affecting environment, animal health and production [2].
→ Revised

Line 32 change in: Control of odor generation and attenuation of nutrient excretion depends on the essential formulation and the balance of nutrients in the animal diet.
→ Revised

Line 35-38 change in: In particular, fermentable carbohydrate (FC) containing dietary fiber has been known to relieve constipation, increase nutrient digestibility and improve gut health, resulting in reducing odor emission via incorporating excessive nitrogen into the microbial biomass nitrogen [7-9, add  DOI: https://doi.org/10.1080/1828051X.2020.1749899].
→ Revised

Line 38-41 change in: Italian ryegrass (IRG) can be cultivated abundantly in Korea and used as a feed ingredient which levels of acid detergent fiber, neutral detergent fiber, acid detergent lignin, and soluble and insoluble dietary fiber and non-structural carbohydrates are similar to those of sugar beet pulp, thus can provide abundant FC [10, add: DOI: https://doi.org/10.1007/BF02194097]
→ Revised

Line 42-43: However, there are limited data on dietary IRG fiber content and the effect of supplementation on odor generation and gut microbial composition in pigs
→ Revised

Response 1:
All comments of point 1 are revised in manuscript according to reviewer’s comments.

Point 2:
Line 51-52: Please report the number of ethical permissions

Response 2:
Sentence is revised according to reviewer’s comment as follow; All procedures were conducted in accordance with guidelines No. 16 set by the National Institute of Animal Science Animal Care and Use Committee at 2008.

Point 3:
Line 62: Please add more information on drying process of ryegrass

Response 3:
Sentence is revised according to reviewer’s comment as follow; IRG was seeded in October and harvested at the heading stage in April of the next year. After naturally dry under the sun, IRG hay was ground into particles with less than 2 mm size.

Point 4:
Line 68-70 change in: From each metabolism crates manure was collected at twice weekly, added in enclosed manure storage container, and stored in the freezer (-20°C) until analysis
→ Revised

Line 85-86: delete “The values were determined by the difference in weight between before and after of dry”
→ Revised

Response 4:
All comments of point 4 are revised in manuscript according to reviewer’s comments.

Point 5:
Line 93-96: Please some description of know methods (e.g. Kjeldahl) could be resumed

Response 5:
Sentence is revised according to reviewer’s comment as follow; The Kjeldahl procedure for total nitrogen determination is widely used for animal feed, fertilizer quality, environmental samples, and biological analysis [13]. Nitrogen was quantified to three steps of digestion, distillation, and titration. Organic nitrogen containing samples are digested with sulfuric acid to ammonium sulfate; the ammonium is then liberated by raising the pH and measured by titration [14].

Point 6:
Line 114: Please specify which kind of standard (internal or external) was used for chromatographic determination

Response 6:
Sentence is revised according to reviewer’s comment as follow; Concentrations of VFA, phenols, and indoles were determined by the external standard quantitation method with standard reagents of each compounds from Sigma-Aldrich (MO, USA).

Point 7:
Table 4: Please insert the complete name of volatile fatty acids

Response 7:
Complete names of volatile fatty acids are added in the table legend under the Table 4.

Point 8:
Table 4 change the title as follow: “Concentrations of volatile fatty acids and odorous compounds in manure from pigs fed diets with increasing levels of IRG”.
→ Revised

Line 203-204 change in: The lower the degradability of fiber is in the hindgut, the greater the excretion of amino acids is.
→ Revised

Table 5: Please improve table editing
→ Revised

Response 8:
All comments of point 8 are revised in manuscript according to reviewer’s comments.

Point 9:
Line 271-272: Bacteria of gram positive by 344 (60%) and gram negative by 225 (40%) were classified in these genera

Response 9:
Sentence is revised according to comments of reviewer 1 (Line 269-271: delete numbers as 262, ….225; Line 271: correct: gram-positive/negative bacteria) and reviewer 2 as follow; Bacteria of gram-positive by 60% and gram-negative by 40% were classified in these genera.

Point 10:
Figure 3: in order to improve the comprehension of the figure could be better to differentiate largely the colours used for each phyla
→ Revised

Line 300: change 5 in five
→ Revised

Line 304, 322 and 338:  considering that microbiota data regard only the comparison between 0 and 1% is better to specify every time IRG (1%)
→ Revised

Response 10:
All comments of point 10 are revised in manuscript according to reviewer’s comments.

Author Response File: Author Response.docx