Heme Dependent Catalase Conditionally Contributes to Oxygen Tolerance of Tetragenococcus halophilus Strains Isolated from Soy Sauce Moromi

Round 1

Reviewer 1 Report

In this study the authors have worked on elucidating the oxygen tolerance diversity of T. halophilus strains isolated from soy sauce moromi, identification of antioxidant enzymes in T. halophilus, and elucidating the possible mechanism of these enzymes in protection of T. halophilus against oxidative stress. While the scientific merit of the study is of high quality. I would recommend english language check as use of past tense in some places and past participle in other is a little confusing. 

Author Response

Thanks for the reviewer's suggestions. English language editing of this manuscript has been done.

Reviewer 2 Report

As authors mentioned, the understanding on the oxygen tolerance of T. halophilus will help us to optimize its cultivation conditions for fermentation. They presented some physiological data to say that heme-dependent catalase can be a player for oxygen tolerance in T. halophilus, however, the evidence is a little bit indirect. In Fig. 4B, the growth was increased under aerobic condition in the absence of catalase activity. Then, can the growth of R21 and 23 be increased by other mechanisms rather than catalase?  I am reluctant to agree with author’s interpretation on Fig.2 data (line 160-161) when consider incorporating Fig.1.     

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 3 Report

The manuscript adressed three major questions:

what are the differences in oxidative stress tolerance in T. halophilus strains?

which enzymes are involved in oxygen tolerance?

what are the possible mechanisms in protection of T. halophilus against oxidative stress?

The authors showed defferences in oxidation resistance between strains and defined the antioxidant enzymes activities. Their results elucidated the antioxidant mechanism which is based on the heme dependent catalase and contributes to oxygen tolerance in the presence of heme under aerobic conditions. This finding indicated possible optimization of T. halophilus growth through the respiration metabolism.

However there are some important issues that need to be corrected:

The study lacks statistical analysis of the results.

All species names of the LAB strains should be corrected according to the current taxonomy. In April 2020, the previous genus Lactobacillus was divided into 25 genera including the emended genus Lactobacillus, which includes host-adapted organisms, Paralactobacillus and 23 novel genera for which the following names were established: Holzapfelia, Amylolactobacillus, Bombilactobacillus, Companilactobacillus, Lapidilactobacillus, Agrilactobacillus, Schleiferilactobacillus, Loigolactobacilus, Lacticaseibacillus, Latilactobacillus, Dellaglioa, Liquorilactobacillus, Ligilactobacillus, Lactiplantibacillus, Furfurilactobacillus, Paucilactobacillus, Limosilactobacillus, Fructilactobacillus, Acetilactobacillus, Apilactobacillus, Levilactobacillus, Secundilactobacillus and Lentilactobacillus ((Zheng et al. 2020; https://doi.org/10.1099/ijsem.0.004107)).

The generic term ‘lactobacilli’ could be useful to designate all organisms that were classified as Lactobacillaceae until 2020.

The reference [40] should be axchange to one of many publications of prof. Alexandra Gruss or Martin Bastian Pedersen such as the one listed below or their other cited publication within this reference.

Pedersen MB, Garrigues C, Tuphile K, Brun C, Vido K, Bennedsen M, Møllgaard H, Gaudu P, Gruss A. Impact of aeration and heme-activated respiration on Lactococcus lactis gene expression: identification of a heme-responsive operon. J Bacteriol. 2008 Jul;190(14):4903-11. doi: 10.1128/JB.00447-08.

Specific comments below:

Line 18: Correct as follows: "Tetragenococcus halophilus strains are the halophilic lactic acid bacteria (LAB) that are present in ..."

Line 23: Correct as follows: "In this work, the growth of T. halophilus strains isolated from..."

Lines 24-25: Correct as follows: "...sauce moromi under both aerobic and anaerobic conditions was compared and the function of their antioxidant enzymes was investigated."

Line 36: Correct as follows: "...bacteria that are involved..."

Line 37: Correct as follows: "They play important roles in formation of ..."

Lines 38-39: Correct as follows: "...is a species of..." or "...is a group of..."

Line 39: Correct as follows: "...that are present in..."

Line 40: Correct as follows: "...food products..."

Line 47: Correct as follows: "...thus improves.."

Lines 48-49: Correct as follows: "...T. halophilus improves the quality of fermented foods and shortens the fermentation period..."

Line 53: Correct as follows: "...has a great impact.."

Line 57: Correct as follows: "...promotes its adaptation to.."

Line 59: Correct as follows: "...upon osmotic stress.."

Line 80: Correct as follows: "...LOX), help to enhance.."

Line 83: Correct as follows: "...are present in lactobacilli strains...

Lines 140, 142, : Change "tolerant" to: "tolerance"

Line 144: Correct as follows: "...highest growth ratio, indicating the best capability to tolerate oxidative stress."

 

Line 183: Correct as follows: "...catalyzes the conversion of H₂O₂ to water and oxygen."

Lines 184-185: Correct as follows: "...catalyzes the conversion of H₂O₂ to water and oxygen."

Lines 184-186: Correct as follows: "Lactobacilli are generally catalase negative, while T. halophilus showed capability to resist H₂O_2..."

Lines 215-216: Correct as follows: "...that contributes to increasing the content of volatiles, and thus to improvement of flavor in fermented foods..."

Line 224: Correct as follows: "...differences in resistance to oxidative stress and elaboration of the possible mechanism..."

Line 236: Correct as follows: "...is an attribute to be associated..."

 

Line 238: Correct as follows: "...indicating the function..."

Line 240: Correct as follows: "Catalase is the enzyme catalyzing..."

Lines 243-244: Correct as follows: "Catalase from LAB generally groups..."

Line 245: Correct as follows: "...by some lactobacilli,..."

Line 247: Correct as follows: "...and Lactiplantibacillus plantarum..."

Line 248: Correct as follows: "...(Table 1)."

 

 

 

 

 

 

 

 

 

Author Response

Please see the attachment.

Author Response File: Author Response.docx