Carotenoids and Their Biosynthesis in Fungi

Round 1

Reviewer 1 Report

This manuscript is of interesting topic can be accepted in its current form after adding an Abbreviations section after the abstract to ease the reading of its contents.

Author Response

Abbreviations section

This has been added

Reviewer 2 Report

The review manuscript (molecules-1593182) summarized the current advances of carotenoid biosynthesis in fungi and the progress and perspectives of metabolic pathway engineering in yeast. This review is well organized and the literatures are thoroughly documented. Overall, it is well written and easy to understand. I spotted the following minor typos or places to be changed:

1. Line 38, change “were” to “was”;
2. Line 81, change “heterologous” to “heterologously”;
3. Line 115, change “hydroxylated” to “hydroxylation”;
4. Line 116, change “cyclized” to “is cyclized”;
5. Line 120, 121, 122, italicize N. crassa, Fusarium, and F., respectively;
6. Line 133, change “accumulates” to “is accumulated”;
7. Line 182, delete “Haxius 1950”;
8. Line 308, italicize crtI and crtY;
9. Line 350, reword the sentence “Important was the included crtYB genei It favors the”;
10. Line 357, change “lead” to “led”;
11. Line 362, change “the accumulating β-carotene converted” to “the accumulated β-carotene was converted”;
12. Line 367, change “structures” to “structures of”;
13. Line 375, change “feed” to “fed”;
14. Line 398, change “combined” to “combination”;
15. Line 399, delete “of”.

Author Response

Suggested corrections

They were very helpful and have all been included.

Reviewer 3 Report

The topic of the manuscript is interesting. Indeed, to my knowledge information on carotenoid presence in fungi is scattered. On the other hand, I found some important points that should be addressed.

First of all, the levels of carotenoids in fungi have not been reported. I suggest to enter this information in Table 1 and to comment it as related to the amounts found in common food sources (see: Ann. Rev. Food Sci. Tech. 2021, 12, 14.1–14.28 and Crit. Rev. Food Sci. Nutr. 2021, 1–51.)

Secondly, the advantages in considering fungi as sources of carotenoids as an alternative to animal or plant sources have not been explained, while at least one reason should be discussed. One point could be that fungi are easy to be grown, eventually using byproducts of food processes, thus they are suitable sources in the perspective of the circular economy (see: Trends Food Sci. Technol. 2021, 114, 323-332 and Food Funct.  2018, 9, 1353–1372.). Other reasons could be related to bioaccessibility of carotenoid from fungi, or else to their stability of carotenoid within the fungal matrix. Is there any literature information on this?

As a third point, possible application strategies for carotenoids obtained from fungi should be drawn, also discussing if they need to be extracted from the fungal matrix or the fungal biomass can be applied as such.

Author Response

1. Levels of carotenoids

They are not available from most species of Table 1. In general, concentration of fungal carotenoids are low compared to other organisms. The following sentences were included into paragraph 3:

Most of the carotenoids of this table were only structurally analysed and identified without the determination of their concentrations. Only for a few species, the concentrations of their specific carotenoids were quantified. Their values range from 100 to 200 µg/g dw or were even lower [20,31,50,54,57].

 

2. Advantages in considering fungi as sources of carotenoids as an alternative

The following was included

at the end of section 5.1.:

Many yeast are able to grow on agricultural waste materials. For example, enhanced zeaxanthin synthesis was obtained with engineered X. denrorhous growing on wheat straw hemicellulose hydrolysate [99]. Furthermore, Y. lipolytica was improved by metabolic engineering to utilize of a wide range of substrates [103].

at section 5.3:

Transformation with extracellular and cell-bound lipases enabled a β-carotene producing strain to grow on olive oil exclusively as a way to use olive mill waste as a substrate [110].

at the end of Conclusion:

This includes production of carotenoids by growing on agroindustrial waste materias. This recycling process may work with the original strains or with those further engineered for the utilization of a specific substrate.

 

3. Application strategies

This was included in section 5.1:

β-Carotene production with B. trispora is at the moment the only large scale industrial process for fungal carotenoids. This carotenoid is recovered by extraction and supplied in a crystal form as a colorant or as provitamin A [79].

 

Round 2

Reviewer 3 Report

The manuscript has been improved. On the other hand, the levels of carotenoids in fungi are very low. Hence, this reviewer thinks that there should be a better clarification regarding the needs of obtaining carotenoids form fungi, with respect to other abundant sources.

A better introduction of the topic would increase the impact of the study

Author Response

The following clarifications have been made by additions

to Introduction:

and are therefore accumulated in much lower concentrations than by plants and algae. However, fungal carotenoids are different to those found elsewhere offering a structural diversity. In order to increase the levels of commercially interesting carotenoid in different fungal species and to match the concentrations of other organisms, classical and metabolic engineering procedures have been established which will be referred to.

and to Conclusion:

Apart from mass produced β-carotene with B. trispora which already made it into the market, astaxanthin from engineered X. dendrorhous with a content of almost 1% of cell mass in non-optimized laboratory cultures [97] can compete with astaxanthin from Haematococcus species especially as it is accumulating in a non-esterified form [79]. Metabolic engineering of Y. lipolytica [93] and particularly X. dendrorhous [99] resulted in synthesis of the rare carotenoid zeaxanthin matching the concentrations of the high-yield mutant of Dunaliella salina [112].