The influence of a soil amendment on the abundance and interaction of arbuscular mycorrhizal fungi with arable soils and host winter wheat
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Arbuscular mycorrhizal (AM) fungi have been shown to be associated with an estimated 70 % of vascular terrestrial plants. Such relationships have been shown to be sensitive to soil disturbance, for example, tillage in the preparation of a seed bed. From the application of arable soil management, AM fungal populations have been shown to be negatively impacted in abundance and diversity, reducing plant growth and development. The present study aims to utilise two sources (multipurpose compost and a commercial inocula) of mycorrhizal fungi for the amendment of arable soils supporting Zulu winter wheat under controlled conditions and quantify plant growth responses. A total of nine fields across three participating farms were sampled, each farm practicing either conventional, reduced, or zero tillage soil management exclusively. Soil textures were assessed for each sampled soil. Via the employment of AM fungal symbiosis quantification methods, AM fungi were compared between soil amendments and their effects on crop growth and development. The present study was able to quantify a mean 6 cm increase to crop height ( P <0.001), 10 cm reduction to root length corresponding with a 2.45-fold increase in AM fungal arbuscular structures ( P <0.001), a 1.15-fold increase in soil glomalin concentration corresponding to a 1.26-fold increase in soil carbon, and a 1.32-fold increase in the relative abundance of molecular identified AM fungal sequences for compost amended soils compared to control samples. Mycorrhizal inocula, however, saw no change to crop height or root length, AM fungal arbuscules were reduced by 1.43-fold, soil glomalin was additionally reduced by 1.55-fold corresponding to a reduction in soil carbon by 1.31-fold, and a reduction to relative AM fungal species abundance by 1.26-fold. The present study can conclude the addition of compost as an arable soil amendment is more beneficial for the restoration of AM fungi beneficial to wheat production and soil carbon compared to the addition of a commercial mycorrhizal inocula.
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I am pleased to tell you that your article has now been accepted for publication in Access Microbiology.
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There are some final minor amendments we would ask you to implement, and please pay particular attention to the suggested scoring system or a robust justification for your method for % root colonisation.
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Comments to Author
The aim of this paper is to compare the performance of compost and a commercial AMF inoculum on the growth of Zulu winter wheat in different soil types under different cultivation systems. Th authors may have been able to draw more conclusions if they had less variables and more information on the composition (ratio of different AMF) of the inoculum. However, the study's aim is to compare a commercial inoculum vs compost and this information is not readily available and the study is a valid one. I have detailed some comments/issues below that I think need to be addressed before publication % root length colonised by AM Fungi The authors measure root colonisation as arbuscular density rather than % colonisation which is the norm. The authors should have included something on this in the methods …
Comments to Author
The aim of this paper is to compare the performance of compost and a commercial AMF inoculum on the growth of Zulu winter wheat in different soil types under different cultivation systems. Th authors may have been able to draw more conclusions if they had less variables and more information on the composition (ratio of different AMF) of the inoculum. However, the study's aim is to compare a commercial inoculum vs compost and this information is not readily available and the study is a valid one. I have detailed some comments/issues below that I think need to be addressed before publication % root length colonised by AM Fungi The authors measure root colonisation as arbuscular density rather than % colonisation which is the norm. The authors should have included something on this in the methods section and also provide some references where this method has been used instead of % colonisation. I am not sure "having discussed this with others.." is sufficient evidence that this approach is sound. I know it can be difficult to determine % colonisation but in those instances a scoring system (cf Giovannetti & Mosse (1980) New Phytologist 84 487) is often used. The authors need to provide a more robust justification for its use Line # 181 Section 2.7 Determination of intracellular arbuscular structure Reference # cited is incorrectly and should be [34]. The method used is one developed by the authors and published in Access Microbiology in 2019. As the method is not widely used the authors should provide the details in the current manuscript rather than just refer the reader to the original reference Line #340 Section 3.3 Mycorrhizal abundance and diversity Universal fungal primers were used so the authors can't assume that the sequences are AMF. Title needs to be edited
Please rate the manuscript for methodological rigour
Satisfactory
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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The reviewer has indicated that not all reviewer suggestions were fully implemented into the revised manuscript. I would like to give you the opportunity to address these points and also encourage particular attention to point 4 from reviewer 2 which argues that the manuscript requires images to support the findings of this work. I would accommodate a prolonged revision period to allow for these images to be captured, so please let us know if you need additional time.
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Comments to Author
The MS has been significantly improved. However, some of the issues raised in the original submission have not yet been resolved in the revised version. 1: Properties of AM fungal inoculum means: i) name of AM fungal species it contains, ii) % of root length colonized by AM fungi, and iii) the number of spores per g of inoculum. AM fungi is one of the main variables of this research, however its details are rather vague and insufficient to support the M&M and result section. 2: "Rhizophagus intraradices" has been misspelt 14 times in the revised MS. 3. Genus "Rhizophagus intraradices" is the new name of "Glomus intraradices", while "Funneliformis mosseae" is the new name of "Glomus mosseae". Please be consistent with the nomenclature. (Authors may visit http://www.amf-phylogeny.com/ …
Comments to Author
The MS has been significantly improved. However, some of the issues raised in the original submission have not yet been resolved in the revised version. 1: Properties of AM fungal inoculum means: i) name of AM fungal species it contains, ii) % of root length colonized by AM fungi, and iii) the number of spores per g of inoculum. AM fungi is one of the main variables of this research, however its details are rather vague and insufficient to support the M&M and result section. 2: "Rhizophagus intraradices" has been misspelt 14 times in the revised MS. 3. Genus "Rhizophagus intraradices" is the new name of "Glomus intraradices", while "Funneliformis mosseae" is the new name of "Glomus mosseae". Please be consistent with the nomenclature. (Authors may visit http://www.amf-phylogeny.com/ or https://invam.ku.edu/nomenclature for more details). 4. Without the visual support of AM fungal colonization in the roots, all the data and effects pertaining to AM fungi are rather arbitrary. I must advise the authors to please concern a researcher with considerable expertise in the field of AM fungi to revise the MS. Thanks.
Please rate the manuscript for methodological rigour
Very poor
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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The reviewers have highlighted major concerns with the work presented. Please ensure that you address their comments. Reviewer 1 has highlighted major concerns with the analysis and interpretation in the results section. Please address these concerns as thoroughly as possible.
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Comments to Author
--- Abstract: * An ideal abstract should be started with background of study followed by problems identified and methodologies adapted. An abstract ends with core findings in the form of results and future perspectives (if any). * Please reduce the abstract to around 200 words. # Line 9-22: Please make it very clear and concise describing the background of study followed by problems identified and then Methodologies adapted to solve the problems. # Line 22-30: These are the core findings of the research, which must be described on a quantitative scale. Introduction: * Please correct the sentence along with punctuation marks. * Before describing aims, please mention the identified problems and the gap between the earlier research studies and the identified problems. Material and Methods: # Line 86-99: …
Comments to Author
--- Abstract: * An ideal abstract should be started with background of study followed by problems identified and methodologies adapted. An abstract ends with core findings in the form of results and future perspectives (if any). * Please reduce the abstract to around 200 words. # Line 9-22: Please make it very clear and concise describing the background of study followed by problems identified and then Methodologies adapted to solve the problems. # Line 22-30: These are the core findings of the research, which must be described on a quantitative scale. Introduction: * Please correct the sentence along with punctuation marks. * Before describing aims, please mention the identified problems and the gap between the earlier research studies and the identified problems. Material and Methods: # Line 86-99: Please take these lines under a different heading viz., "2.1 Field study". # Line 99-111: Please retain these lines under heading viz., "2.2 Experimental design". # Line 103-105: Please take these lines out and put them under a separate heading "2.3 Am fungal inoculum" and describe all possible properties including and the species it contains. * Headings "soil type" and "soil nutrient testing" should be merged under a separate heading "Physicochemical properties of the soil". * Ergosterol HPLC: Please mention all the modifications done in the original methodology. * Glomalin related soil protein (GRSP) extraction: Instead of citing two many methods, please describe the modified method adapted in this research, so that it can be repeated by the other researchers of the area. * Soil carbon via loss on ignition: This heading should be merged in heading "Physicochemical properties of the soil". Result: * Please support the data on AM fungal structure with a photo plate showing AM fungal colonization in the roots of Zulu winter wheat. Discussion: * Some old citations can be omitted to reduce the number of references. ---
Please rate the manuscript for methodological rigour
Poor
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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Comments to Author
The author of the manuscript "The influence of a soil amendment upon the abundance and interaction of arbuscular mycorrhizal fungi with arable soils and host winter wheat" is describing a large set of pot trials to assess the effect of compost and AMF inoculation on wheat growth using several soil types with different management practice backgrounds. The introduction is clearly written and easy enough to follow. While the principal idea of the study is interesting, there are several serious shortcomings in the data analysis and data interpretation that need to be addressed before the manuscript can be published. General comments Many aspects of the methodology are either very brief or are missing entirely. While I appreciate that many standard methods are tedious to be described in detail, I would …
Comments to Author
The author of the manuscript "The influence of a soil amendment upon the abundance and interaction of arbuscular mycorrhizal fungi with arable soils and host winter wheat" is describing a large set of pot trials to assess the effect of compost and AMF inoculation on wheat growth using several soil types with different management practice backgrounds. The introduction is clearly written and easy enough to follow. While the principal idea of the study is interesting, there are several serious shortcomings in the data analysis and data interpretation that need to be addressed before the manuscript can be published. General comments Many aspects of the methodology are either very brief or are missing entirely. While I appreciate that many standard methods are tedious to be described in detail, I would recommend to provide some short basic principals nonetheless. For instance the nutrient analysis could be supplemented with information on what method has been used i.e. spectrophotometry. Methodology of the Illumina sequencing work is completely missing and not reproducible. The authors need to provide a walkthrough of the methods used, especially for the data analysis which is entirely missing. Missing is also a deposit of the sequencing data in NCBI or EBI. Likewise, sequence data analysis seems to be incomplete. No alpha or beta diversity has been shown, only a selection of relative abundance data were shown, mostly only based on presence or absence. The description of the results suggest that the author is not aware that the generated data are of relative abundance and not absolute abundance. From the method description and the result section it appears that data sets were interpreted without posthoc test and only on occasions a posthoc test was applied. Likewise, it appears the author has carried out cherry picking of the data by only conducting t-tests for selected pairs of data. If correct, this is not a sound analysis of the data and I recommend starting over with the data analysis. Consequently, the results and the discussion have to be rewritten from the ground up. Specific comments L1 on instead of upon? L43 Please briefly elaborate how AMF can mobilize plant unavailable nutrients; mechanical forcing? acid exudation? bacterial interaction? enzymatic activity? L49 revise use of monumentally L98 is that not general fungal biomass? L99 200g seems to be quite small for a pot experiment L109-10 Watered not based on pot weight? Specify plant growth chamber details ie.e. make and model L86-111 After sampling the soil, was the soil homogenized for potting? If so, then the history of management i.e. ZT, RT is taken into account only. This needs to be made clearer. L117, 123, 133, 136, Maybe adding a brief overview of the method in a couple of sentences? L120 What does that mean? I am confused by this statement, were pots fertilized and if so, what content? L150-2 Further details needed on the chosen primers, illumina sequencing procedure and especially the data analysis. What pipeline was used etc. Data need to be added into repository and repository IDs provided. L163 data tested for normality? L166 Which posthoc test was selected? L174 total nitrogen? PO4? total P? P2O5? Table 1 - Table could do with improved formatting Are the 6 soils utilized below from 3 farms that have 1-3 soils employing different types of tilling (RT, CT, NT) OR is the sandy loam coming from one farm that employs different tilling? Why measured per volume? Value in kg/ha equivalent? Change to non ambigus SI unit L182 Compost AND AMF or just compost? What about the added mineral fertilizer? L207pp Does that mean the results further above are without posthoc test? What would be the value of reporting them here? I strongly suggest to only report results after posthoc testing. Figure 2 Change label (b) Add letter indicating significant differences? L226 Any significance to report? L233pp As above, please don't report statistical findings without a posthoc test, only report fully analysed data. L267 I am not sure what this means. Does it mean T test were conducted between selected pairs? If so, is this cherry picking? I recommend avoiding this. Please redo the statistical analysis. L290pp Alpha diversity? Beta diversity? Abundance is only relative abundance. This section (3.3) is incomplete and potentially misleading. L299 what is meant with unique? L300 Considering that 25 wt/wt is compost, will these be dominating the AMF community structures? L309 remove "difference" Table 2 All species names in italics, please. Discussion was not reviewed as the text is based on a compromised result section.
Please rate the manuscript for methodological rigour
Very poor
Please rate the quality of the presentation and structure of the manuscript
Satisfactory
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
Yes: Data cherry picking
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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