Insights into the Fungal Community and Functional Roles of Pepper Rhizosphere Soil under Plastic Shed Cultivation

Round 1

Reviewer 1 Report

This study highlights the importance of soil as a specific niche for some rare fungal members that could be fundamental for the health of soils cultivated under plastic shed .

The results increase information to the body of knowledge about pepper cultivation under plastic shed because it covers interesting information on fungal communities associated to bulk/rhizosphere soil of pepper and their possible role on growth, yield and quality of this economically important crop culture.

I have , however some suggestion to improve the MS:

Line164: Maybe it is more correct to use defense from nematodes instead of defensive of nematodes. If I understand well, here they are plant that need to be defended from.pathogen agents not nematodes itself (e.g with the help of some kind of endobacteria such Wolbachia that contributes to immune defense to nematodes) or I am wrong?

3.3“Shifts in the fungal functional genes in soil” In my opinion this title is not correct; because you have assigned “functional role/ecological guilds to fungal OTUs ” by inference on the base of phylogenetic assignation and you haven't investigated particular  genes and/or their expression.

With FunGuild you can represent main trophic mode assignments but you cannot tigthly related them to particular genes or metabolic pathways.

So please try to soften this statement everywhere in the MS.

You could eventually discuss this last point on the light of the more progress that has been done for prokaryotes for which correlating genome annotations with functional information , such as metabolic pathways, is already possible with FAPROTAX (Functional Annotation of Prokaryotic Taxa), a manually curated functional annotation database (Louca et al. 2016).

line 248: what does you mean here with upregulation? You are not discussing about gene expression. See comment above.

Author Response

Line164: Maybe it is more correct to use defense from nematodes instead of defensive of nematodes. If I understand well, here they are plant that need to be defended from. pathogen agents not nematodes itself (e.g with the help of some kind of endobacteria such Wolbachia that contributes to immune defense to nematodes) or I am wrong?

Answer: Thanks for your comments. Done as suggested. We revised the description as that “This genus was also reported to be beneficial for the defense from nematodes and nutrient uptake by habanero chili pepper (Capsicum Chinese Jacq.), thereby facilitating plant growth”. (See Lines 166)

3.3 “Shifts in the fungal functional genes in soil” In my opinion this title is not correct; because you have assigned “functional role/ecological guilds to fungal OTUs” by inference on the base of phylogenetic assignation and you haven't investigated particular genes and/or their expression.

With FUNGuild you can represent main trophic mode assignments but you cannot tightly related them to particular genes or metabolic pathways.

So please try to soften this statement everywhere in the MS.

You could eventually discuss this last point on the light of the more progress that has been done for prokaryotes for which correlating genome annotations with functional information, such as metabolic pathways, is already possible with FAPROTAX (Functional Annotation of Prokaryotic Taxa), a manually curated functional annotation database (Louca et al. 2016).

Answer: Thanks for your comments. Firstly, according to your suggestion, the title of 3.3 has been revised into that “3.3. Shifts in the fungal functional role in soil”. (See Lines 227)

Secondly, we thoroughly revised the manuscript according to the insightful suggestions from you. For example:

1) The description of “fungal function” was revised as “fungal functional roles” in the whole manuscript. (See Lines 2-3, 19, 31, 55, 66, 72, 74, 77, 81, 139, 142, 227, 228, 230, 232, 239, 265, 266, 269, 271, 272, 279 and 307)

2) We revised the description as that “Based on the trophic mode of fungal OTU, the fungal community functional roles could be divided into nine categories, including saprotroph, pathotroph, symbiotroph, saprotroph-symbiotroph, pathotroph-saprotroph-symbiotroph and other (Fig. 4a)” in the line 232-234.

3) We deleted some of the contents that were related to particular genes in order to make the information expressed in the manuscripts more accurate. (See Lines 258)

Thirdly, it is true that some progress has been made for prokaryotes by connecting genome annotations with functional information (Louca et al., 2016). However, compared to prokaryotes, eukaryotic organisms such as fungi, have large genomes and relatively few fully-sequenced species (Nguyen et al., 2016). The connection between fungal genomes and functional information still needs further investigation. Recently, the FUNGuild analysis method has been developed as a tool used to taxonomically parse fungal OTUs by ecological guild (Nguyen et al., 2016). In many studies, the method of FUNGuild was often applied to explore the changes of the plant-soil fungal community (Kolarikova et al., 2017; Feng et al., 2019; Lian et al., 2019). Therefore, in the manuscript, we mentioned that “Recently, the python-based FUNGuild tool has been used to identify specific ecological categories of fungi through its simple and convenient data processing [19]. Feng et al. reported that some fungi play an important role in restoring the stability of aggregates with FUNGuild [20]. Other studies have used FUNGuild to elucidate the specific roles played by different fungal communities in rhizosphere soils and compost [21, 22]”. (See Lines 67-72)

The FAPROTAX (Functional Annotation of Prokaryotic Taxa), as you mentioned, was generally used for bacteria and archaea (Louca et al., 2016). In this manuscript, as mentioned above, we mainly used FUNGuild to parse fungal OTUs by ecological guild and we have discussed some functional role of some guilds. For example, in lines 249 “Saprotroph is related to the role of fungi in decomposition” and in lines 252-254 “some pathogens attack the root system of crops, while others, such as some fungal parasites, could promote the healthy growth of crops”. Moreover, according to your suggested reference, we also added some discussion about fungal functional roles and plant species, which is that “Some studies have suggested that the changes of fungal functional roles were related to many factors, and plant species were also considered as important influencing factors [42, 43]. Therefore, the difference between rhizosphere and non-rhizosphere fungal functional roles can be attributed to plant species and environmental factors”. (See Lines 261-264)

Line 248: what does you mean here with upregulation? You are not discussing about gene expression. See comment above.

Answer: We revised the description as that “The increase of pathotrophic fungi in rhizosphere soil could be ascribed to the plant roots, although some pathogens attack the root system of crops, while others, such as some fungal parasites, could promote the healthy growth of crops”. (See Lines 251-254)

Again, we really appreciate all the comments and suggestions for this manuscript.

Reference:

Feng, H.J., Wang, S.Y., Gao, Z.D., Wang, Z.K., Ren, X.Q., Hu, S.W., Pan, H. 2019. Effect of land use on the composition of bacterial and fungal communities in saline-sodic soils. Land Degradation & Development, 30, 1851-1860.

Kolarikova, Z., Kohout, P., Kruger, C., Janouskova, M., Mrnka, L., Rydlova, J. 2017. Root-associated fungal communities along a primary succession on a mine spoil: Distinct ecological guilds assemble differently. Soil Biology & Biochemistry, 113, 143-152.

Lian, T.X., Mu, Y.H., Jin, J., Ma, Q.B., Cheng, Y.B., Cai, Z.D., Nian, H. 2019. Impact of intercropping on the coupling between soil microbial community structure, activity, and nutrient-use efficiencies. Peerj, 7.

Louca, S., Parfrey, L.W., Doebeli, M. 2016. Decoupling function and taxonomy in the global ocean microbiome. Science, 353, 1272-1277.

Nguyen, N. H.; Song, Z. W.; Bates, S. T.; Branco, S.; Tedersoo, L.; Menke, J.; Schilling, J. S.; Kennedy, P. G., FUNGuild: An open annotation tool for parsing fungal community datasets by ecological guild. Fungal Ecol. 2016, 20, 241−248.

Author Response File: Author Response.docx

Reviewer 2 Report

This a nice piece of work. I think that the manuscript should be accepted for publication after minor revision. Several minor comments related to some question to clarify some aspects of the methodology, and the way of expression some sentences are included in the attached file.

Comments for author File: Comments.pdf

Author Response

1. Please clarify why the number of replicates used in the study...Based on what???.

Answer: Thanks for your comments. The number of replicates used in the study was selected to ensure the reliability and accuracy of the field experiment data. Therefore, according to our experiment and the reported references together, we revised the description into that “Five replicates were sampled to ensure the reliability of the field experiment data [2]”. (See Lines 91-92)

2. Throughout the manuscript there are some minor typo mistakes and grammatical english aspects that I recommend the authors should carefully read again the manuscript to solve them.

Answer: We thoroughly revised the manuscript according to the rigorous suggestions from you. For example:

“Nanodrop” was revised into “Nano drop” (See Lines 108)

“Lefse” was revised into “LEfSe” (See Lines 158)

“saprotrophy” was revised into “saprotroph” (See Lines 257, 260)

“dietyl phthalate” was revised into “diethyl phthalate” (See Lines 292)

“diisobutyl phthalate” was revised into “di-isobutyl phthalate” (See Lines 292, 294)

“dinbutyl phthalate” was revised into “di-n-butyl phthalate” (See Lines 292, 294)

“dinoctyl phthalate” was revised into “di-n-octyl phthalate”. (See Lines 292, 294)

Once again, we really appreciate all the comments and suggestions for this manuscript.

Author Response File: Author Response.docx