The Transition Period Updated: A Review of the New Insights into the Adaptation of Dairy Cows to the New Lactation

Round 1

Reviewer 1 Report

The manuscript by Mezzetti et al, is a review on the pathophysiological metabolic and immune system adaptations during the transition period.
In my opinion, the manuscript does not need major revisions. 

As a first suggestion, I would ask the authors to limit the number of references to articles from the last 20 years. I understand that many historical citations must be included, but I would still limit the number of references to the last two decades. Please at least replace citations older than the 2000.

Line 233: What do you mean with metaflammation"? or meta-inflammation?

Lines 430-434 .. : Monensin has effect on many biochemical parameters (blood NEFA, BHB, glucose, urea, AST, ALT, GGT, TG ,CHL, bilirubin, total protein, etc) as described in the review by Mammi et al., 2021, Animals.

In any case, the effect of CRC monensin on BHB, NEFA and glucose levels is robust. As also confirmed by a recent article , lower levels of BHB and NEFA, correspond to higher levels of glucose in cows treated with monensin ( Fiore et al., 2021, Journal of Dairy Research).

Lines 441-494: Regarding the chapter  "Predictive markers to identify risky animals":

- the reader's expectations may be different from what is written. Typically when talking about predictive markers or biomarkers, someone might start thinking about blood biochemical analyses or the newer metabolomics analyses (1H-NMR, GC, GC-TLC, HPLC, etc.). I ask you to focus the title on rumination times or something else inherent to what you have written.

- Suggestions: Saha et al., (2019 - Animals) have found that the decrease in rumen pH has been associated with a linear increase in the content of all VFAs in the rumen fluid and with changes in the relative proportions of acetate and propionate, so a latent variable was extracted to represent the overall rumen acidity condition. Cows with different ruminal acidity spent a similar amount of time ruminating during the day, but their RT differed after the morning feeding.

Author Response

R1

The manuscript by Mezzetti et al, is a review on the pathophysiological metabolic and immune system adaptations during the transition period.
In my opinion, the manuscript does not need major revisions. 

As a first suggestion, I would ask the authors to limit the number of references to articles from the last 20 years. I understand that many historical citations must be included, but I would still limit the number of references to the last two decades. Please at least replace citations older than the 2000.

AU: We thanks this reviewer for his positive feedback and for his precious suggestions. We tried to avoid using citations that were older than 2000 (except for Drackley 1999 and Kherli 1989, that revested a particular importance for the topic of this paper).

Line 233: What do you mean with metaflammation"? or meta-inflammation?

AU: metaflammation is a chronic low-grade inflammation (see 10.1016/j.arr.2017.10.003)

Lines 430-434 .. : Monensin has effect on many biochemical parameters (blood NEFA, BHB, glucose, urea, AST, ALT, GGT, TG ,CHL, bilirubin, total protein, etc) as described in the review by Mammi et al., 2021, Animals. In any case, the effect of CRC monensin on BHB, NEFA and glucose levels is robust. As also confirmed by a recent article , lower levels of BHB and NEFA, correspond to higher levels of glucose in cows treated with monensin ( Fiore et al., 2021, Journal of Dairy Research).

AU: We modified the section including the following statement: “These positive effect of monensin on rumen fermentation is reflected by trends of several plasma analytes (i.e reduced BHB and NEFA concentrations and higher plasma glucose) in cows treated with monensin (Fiore et al., 2021; Mammi et al., 2021).” Thank you

Lines 441-494: Regarding the chapter "Predictive markers to identify risky animals":

- the reader's expectations may be different from what is written. Typically, when talking about predictive markers or biomarkers, someone might start thinking about blood biochemical analyses or the newer metabolomics analyses (1H-NMR, GC, GC-TLC, HPLC, etc.). I ask you to focus the title on rumination times or something else inherent to what you have written.

AU: We changed the title to “Early detection of risky animals through plasma analytes trends and behavioral patterns”. Thank you

- Suggestions: Saha et al., (2019 - Animals) have found that the decrease in rumen pH has been associated with a linear increase in the content of all VFAs in the rumen fluid and with changes in the relative proportions of acetate and propionate, so a latent variable was extracted to represent the overall rumen acidity condition. Cows with different ruminal acidity spent a similar amount of time ruminating during the day, but their RT differed after the morning feeding.

AU: We added this at L 496: “Saha et al., 2019 have found that cows undergoing a subclinical rumen acidosis condition (as reflected by the increased VFAs content, altered acetate to propionate ratio and decreased pH of the rumen fluid) spent a similar amount of time ruminating during the day as compared to healthy animals, but their RT differed after the morning feeding”. Thank you

Reviewer 2 Report

Very informative review of an "extended" transition period. The narrative "flowed" well. I think the title should indicate that a review had being carried out. The approach to your review was relevant and comprehensive.  It highlighted the importance of "optimal welfare" as well as a "balanced diet". It also highlighted how  prognostic behaviour and physiology could be used as a means to prompt an earlier intervention.  

Comments and Suggestion:

Line 241-243: ".. dairy cows raised in commercial TMR systems typically undergo 3 or 4 important changes in diet composition from dry-off to early lactation, to fit with relevant changes in their requirements,," In an appendix, give examples of these 3 to 4 changes in diet composition. 

Line 258-260:  Please elaborate further the following statement: -

" ..dietary-driven changes in the rumen bacterial community have been demonstrated to affect the infiltration of several immune mediators in the rumen through saliva.."

Line 295-298: Rewrite this sentence as it does not make sense.

Line 313: Give examples from the literature of "a soft and comfortable lying surface and adequate ventilation.."

Line 318-320: Quote from the literature what the "ideal stocking rate" would be. What is the ideal "resting space".  What is meant when you say that ".. each pen should be sized using 120-150% average calving rate (depending on average stay)..". According to the literature, what is the recommended pen size?

Line 324: Mention the "detrimental effects".

Line 325-328: Wathes et al., 2007 is not a suitable reference 

Line 335-336: give options from the literature for hiding

Line 338: what is meant by the phrase ".. and a blind may benefit periparturient cows.." 

;

Author Response

Very informative review of an "extended" transition period. The narrative "flowed" well. I think the title should indicate that a review had being carried out. The approach to your review was relevant and comprehensive.  It highlighted the importance of "optimal welfare" as well as a "balanced diet". It also highlighted how prognostic behaviour and physiology could be used as a means to prompt an earlier intervention.

AU: We greatly appreciate such a positive feedback by the reviewer. We modified the title as follows: “The transition period updated: a review of the new insights into the adaptation of dairy cows to the new lactation”. Thank you

Comments and Suggestion:

Line 241-243: ".. dairy cows raised in commercial TMR systems typically undergo 3 or 4 important changes in diet composition from dry-off to early lactation, to fit with relevant changes in their requirements," In an appendix, give examples of these 3 to 4 changes in diet composition.

AU: We included Table 1 to satisfy this request. Thank you.

Line 258-260:  Please elaborate further the following statement: -" ..dietary-driven changes in the rumen bacterial community have been demonstrated to affect the infiltration of several immune mediators in the rumen through saliva.."

AU: We changed the sentence as follows: “Furthermore, dietary-driven changes in the rumen pH and bacterial community interplay with rumen epithelial cells, that regulate the infiltration of several immune mediators in the rumen through saliva” Thank you

Line 295-298: Rewrite this sentence as it does not make sense.

AU: We are afraid that line numbering in the reviewed document does not matches with the one used by reviewers anymore. We can’t find the sentence this reviewer is referring to.

Line 313: Give examples from the literature of "a soft and comfortable lying surface and adequate ventilation.."

AU: We included them within brackets: “In all transition pens, a soft and comfortable lying surface and adequate ventilation should be provided (i.e. dry and deep layer of shavings, straw, or sand as bedding material and a minimum airspeed of 1 m s-1)  , as well as feed bunks with headlocks (76 cm wide) — these are preferred to post-and-rail design (Fregonesi et al., 2007b; Tucker et al., 2009; van Gastelen et al., 2011; Campler et al., 2014; Mondaca, 2019)”. Thank you

Line 318-320: Quote from the literature what the "ideal stocking rate" would be. What is the ideal "resting space".  What is meant when you say that ".. each pen should be sized using 120-150% average calving rate (depending on average stay)..". According to the literature, what is the recommended pen size?

AU: We added the reference. We defined ideal resting space within brackets (>13 m2 per cow). Thank you.

Line 324: Mention the "detrimental effects".

AU: We included them within brackets: “(i.e. reducing feed and water intake, increasing standing time and decreasing lying time)”. Thank you

Line 325-328: Wathes et al., 2007 is not a suitable reference

AU: We added 10.3168/jds.2016-10987. Thank you

Line 335-336: give options from the literature for hiding

AU: We included them within brackets, as follows: “(i.e. a plywood barrier covering half of the pen)”

Line 338: what is meant by the phrase ".. and a blind may benefit periparturient cows.."

AU: We rephrased the sentence as follows: “A series of papers (Creutzinger et al., 2021a; b; c) demonstrated that providing the maternity pen with additional space and a physical blind (created using plastic road barriers and plywood, a steel gate, and shade cloth) may benefit periparturient cows and heifers”. Thank you.

Reviewer 3 Report

General comments:

Overall, the review manuscript by Mezzetti et al is very well written. Based on some new perspectives of impacts of metabolic changes and immune dysfunctions on transition dairy cows, the authors proposed a wider time frame for transition period (TP) as compared to the ‘conventional’ TP definition. The authors also reviewed recent publications in management/nutritional strategies for a ‘good’ transition period. In conclusion, the roots of health problems occurring at the onset of lactation could originate before the ‘conventional’ transition period (even preceding the dry-off period).

Major points:

1. Lines (L) 178-202: A potential genetic contribution on the likelihood of developing systemic inflammation is not deeply reviewed/discussed. Some examples for different breeds need to be discussed. How genetic factors are linked to systemic inflammation and metabolic disorders during the transition period?
2. L441-499: I would suggest the authors to review more studies which focused on predictive biomarkers (in blood or urine, during the dry-off period) for periparturient diseases of dairy cows. To make it simple, a summary Table could be added for this section. There are some new metabolomics studies for identification of biomarkers for different metabolic/infectious diseases (e.g., ketosis, mastitis, metritis, lameness, retained placenta, and milk fever) in dairy cows. Since several studies reported predictive biomarkers for diseases at 8 weeks before parturition. These inputs could serve as strong evidence to support the overall idea of current manuscript that TP could be extended from beginning of dry-off (or even before the dry-off).

Minor points:

1. Lines (L) 104-111. How about BHBA level in pre-ketotic cows (e.g., during the dry off period, or before calving)? Even BHBA level might be under normal range, increased level of BHBA in pre-ketotic cows (during the dry-off, no NEB) could be a risk factor for ketosis during the early lactation. This is important to link metabolic disorders and immune dysfunction/inflammation, which is the driving cause?
2. L147-148: Please clarify here, what does ‘altered immune function during the dry-off period in SCK model’ mean? Immune suppression or immune response activation?

Author Response

Overall, the review manuscript by Mezzetti et al is very well written. Based on some new perspectives of impacts of metabolic changes and immune dysfunctions on transition dairy cows, the authors proposed a wider time frame for transition period (TP) as compared to the ‘conventional’ TP definition. The authors also reviewed recent publications in management/nutritional strategies for a ‘good’ transition period. In conclusion, the roots of health problems occurring at the onset of lactation could originate before the ‘conventional’ transition period (even preceding the dry-off period).

Major points:

1. Lines (L) 178-202: A potential genetic contribution on the likelihood of developing systemic inflammation is not deeply reviewed/discussed. Some examples for different breeds need to be discussed. How genetic factors are linked to systemic inflammation and metabolic disorders during the transition period?

To the best of our knowledge, the precise genetic mechanism underlying a successful adaptation to the new lactation has not been identified yet. We tried to rephrase the paragraph as follows: “A clear explanation behind the mechanism responsible for the onset of such an early and persistent inflammatory condition in some animals still lacking. Recently, Milanesi et al. (2021) found the expression of two inflammatory biomarkers related to the acute phase response (paraoxonase and ceruloplasmin) to be primarily under cis regulation in dairy cattle. This significant genetic association was identified in two dairy breeds (Italian Holstein and Italian Simmental), characterized by highly different selection histories, immune-metabolic status and production performances (Bomba et al., 2015; Marras et al., 2015; Lopreiato et al., 2019). Thus, a different expression of inflammatory biomarkers driven by genotype could account for the different inflammatory conditions found by Cattaneo et al. (2021) before the dry-off. Despite that, the relationship between the candidate causative variants identified by Milanesi et al. (2021) and the animals’ performances during the TP has not yet been investigated, and the contribution of genotype on the successful adaptation of dairy cows to the new lactation still to be elucidated.”

We also added some extra information regarding genomic information on health issues in the last paragraph of the review (Genomic information to prevent metabolic dysfunctions).

1. L441-499: I would suggest the authors to review more studies which focused on predictive biomarkers (in blood or urine, during the dry-off period) for periparturient diseases of dairy cows. To make it simple, a summary Table could be added for this section. There are some new metabolomics studies for identification of biomarkers for different metabolic/infectious diseases (e.g., ketosis, mastitis, metritis, lameness, retained placenta, and milk fever) in dairy cows. Since several studies reported predictive biomarkers for diseases at 8 weeks before parturition. These inputs could serve as strong evidence to support the overall idea of current manuscript that TP could be extended from beginning of dry-off (or even before the dry-off).

AU: We included the following statement at the onset of the chapter on predictive markers: “Several studies have been performed on biological fluids of transitioning dairy cows (i.e., urine, blood, saliva, rumen fluid) using different analytical procedures. Research efforts have focused on identifying candidate biomarkers that might reflect ‘at risk’ condition for developing early lactation diseases (i.e., ketosis, mastitis, metritis, lameness, retained placenta, and milk fever). Considering the extent of this topic, this review will be only focused on predictive markers allowing a close monitoring of the immune status of dairy cows that are approaching to calving.”

Minor points:

1. Lines (L) 104-111. How about BHBA level in pre-ketotic cows (e.g., during the dry off period, or before calving)? Even BHBA level might be under normal range, increased level of BHBA in pre-ketotic cows (during the dry-off, no NEB) could be a risk factor for ketosis during the early lactation. This is important to link metabolic disorders and immune dysfunction/inflammation, which is the driving cause?

AU: We are grateful with this reviewer for raising this point. We proposed a linkage between immune activation during the dry period and BHB release before calving, hypothesizing this could serve as a driving factor for developing ketosis in early lactation. Such a relationship is explained at L 127-135, as follows : “Greater IFN-γ production from leukocytes consequential to their activation may induce insulin resistance during the dry period (Choi et al., 2018), thus accounting for the simultaneous increase of plasma concentrations of glucose, nonesterified fatty acids (NEFA), and BHB before calving. The high circulating glucose levels and the anorexic power exerted by high circulating NEFA, BHB, and PICs concentrations could reduce feed intake around calving (Kuhla, 2020). Such a feed intake depression, together with the energy requirement related to the activation of the immune system in the dry period, worsen the NEB in early lactation (Ingvartsen and Moyes, 2012; Bertoni and Trevisi, 2013). Severe NEB in early lactation induces milk yield losses and boosts the mobilization of lipid sources, increasing the NEFA load on liver metabolism.”

1. L147-148: Please clarify here, what does ‘altered immune function during the dry-off period in SCK model’ mean? Immune suppression or immune response activation?

AU: We clarified the statement as follows: “the role exerted by altered immune function during the dry period in the SCK model (i.e. an immune response activation followed by a transient immune suppressive status)”. Thank you

Round 2

Reviewer 3 Report

The authors have done a great job in this revised version. Just a minor point needs to be addressed. 

Minor point 1: Please cite some representative references for lines 452-455 'Research efforts have focused on identifying candidate biomarkers that might reflect ‘at risk’ condition for developing early lactation diseases (i.e., ketosis, mastitis, metritis, lameness, retained placenta, and milk fever). 

Author Response

We addressed the comment by including three references supporting our statement. Despite that, if this reviewer could provide any suggestion for a suitable reference that could improve the quality of our manuscript we will be glad to include it in the text. Thank you