Review Reports - Effect of Nitrogen Fertilization and Inoculation of Durum Wheat with <i>Fusarium pseudograminearum</i> on Yield, Technological Quality and Gluten Protein Composition

Round 1

Reviewer 1 Report

The manuscript Agronomy-2442331 „Effect of Nitrogen Fertilization and Inoculation of Durum Wheat with Fusarium pseudograminearum on Yield, Technological Quality and Gluten Protein Composition”, presented for peer review is interesting, but it has some shortcomings, which I will mention below:

Fusarium crown rot (FCR), caused by the fungal pathogen Fusarium pseudograminearum, is a chronic disease of wheat and barley in many arid and semi-arid cropping regions of the world, including Australia. Among cool-season cereals, bread wheat and, particularly, durum wheat is susceptible to F. pseudograminearum, although there is a distinct range of reactions to FCR in bread wheat. As a result of an effect on grain development, FCR incidence and severity are negatively correlated with grain yield, kernel number per head, kernel weight, tiller height and straw weight. In Australia, a lot of research results clearly show that FCR is an economically important cereal disease.

1. Section 3 result under Fig 7: „Dough strength (gluten index, GI) showed significant
effects by genotype and disease.”- incomprehensible

2. Section Discussion, I don't quite understand the sentence: "This would help to explain why there was more impact on grain yield and some quality measures from FCR inoculation in 2020 and not in 2021. In 2020, high residual N levels reduced N responsive soil with limited impacts on durum quality.”
Consider: .... some quality measures from FCR - i.e. what features, I would break this sentence into two which will be more readable. Please consider the offer.

3. Conclusions: I don't understand the sentence: Consider: " However, cooler and wetter conditions in 2021 prevented the expression of FCR despite inoculation of plants and due to soil management in the summer of 2021 prior to sowing, good nitrogen responses in terms of grain yield and grain protein content were obtained with consequent effects on grain and semolina traits.” - this is incomprehensible, it needs to
be corrected.

4. My comment, what is the purpose of the authors writing about data that they do not show in the manuscript?- (data not shown)

5. So, the nutrient status of the soil has also been reported to affect FCR disease development. For instance, studies have suggested that nitrogen fertilizers in the field can increase the disease incidence and severity of root diseases caused by Fusarium pathogens in wheat. I am missing in conclusion and whether FCR develops more in wet or dry weather conditions. Did the large different doses of nitrogen have a significant effect on the development of FCR in dry or wet years? How the moisture and nutritional status of the soil, as well as the presence of non-pathogenic or beneficial microbes, affect FCR inoculum levels and disease development. The role of nitrogen :

Filip, Ewa et al. “An Overview of Factors Affecting the Functional Quality of Common Wheat (Triticum aestivum L.).” International Journal of Molecular Sciences 24 (2023): n. pag.

6. Self-citations of two authors were detected ( Sissons, M.; Simpfendorfer, S.) and large errors in the citation of literature, many items were not quoted, e.g.: 17, 19-24, 26, 29, 36, 39.

Author Response

We thank the reviewer for their helpful comments which we have addressed in point form below.

Section 3 result under Fig 7: „Dough strength (gluten index, GI) showed significant
effects by genotype and disease.”- incomprehensible

We have revised the text to make clearer as “The ANOVA on the 2020 data showed significant effects on dough strength (gluten index, GI) from genotype and FCR application (Table S1).”

Section Discussion, I don't quite understand the sentence: "This would help to explain why there was more impact on grain yield and some quality measures from FCR inoculation in 2020 and not in 2021. In 2020, high residual N levels reduced N responsive soil with limited impacts on durum quality.”
Consider: .... some quality measures from FCR - i.e. what features, I would break this sentence into two which will be more readable. Please consider the offer.

We have revised the text to make clearer as “These overall differences in rainfall and temperature between the seasons, with 2020 being hotter and drier, are known to favour expression of FCR infection resulting in more impact on grain yield [21] and some quality measures (TW, MPT, SCR and especially GI) in 2020.”

Conclusions: I don't understand the sentence: Consider: " However, cooler and wetter conditions in 2021 prevented the expression of FCR despite inoculation of plants and due to soil management in the summer of 2021 prior to sowing, good nitrogen responses in terms of grain yield and grain protein content were obtained with consequent effects on grain and semolina traits.” - this is incomprehensible, it needs to
be corrected.

We have revised the text to make clearer as “The cool and wet conditions during the growing season in 2021 limited disease expression and FCR impact on yield and quality despite infection of plants. Residual soil N levels at sowing where much higher in 2020 than in 2021 which limited the effects of N treatments on yield and quality in the first year of experimentation. The lowering of residual soil N levels prior to sowing in 2021 resulted in good nitrogen responses in terms of grain yield and grain protein content being obtained with consequent effects on other technological properties such as TW, TGW, SY, L*, b*, MPT, RBD and GI.”

My comment, what is the purpose of the authors writing about data that they do not show in the manuscript?- (data not shown)

We have removed all reference to “data not shown” and replaced many of the suppl. Tables with two; Means and ANOVA results, Tables S1 & S2.

So, the nutrient status of the soil has also been reported to affect FCR disease development. For instance, studies have suggested that nitrogen fertilizers in the field can increase the disease incidence and severity of root diseases caused by Fusarium pathogens in wheat. I am missing in conclusion and whether FCR develops more in wet or dry weather conditions. Did the large different doses of nitrogen have a significant effect on the development of FCR in dry or wet years? How the moisture and nutritional status of the soil, as well as the presence of non-pathogenic or beneficial microbes, affect FCR inoculum levels and disease development. The role of nitrogen :

Filip, Ewa et al. “An Overview of Factors Affecting the Functional Quality of Common Wheat (Triticum aestivum L.).” International Journal of Molecular Sciences 24 (2023): n. pag.

We suggest that clarification of reviewers points 2 and 3 should have fixed this.

Presence of non-pathogenic or beneficial microbes was not specifically explored within this current study. However, these factors are unlikely to have impacted on the outcomes of this study as natural suppression of FCR has never been reported to the authors knowledge. There have been previous studies on the biological control of FCR but this involves culturing and introduction of elevated populations of specific microbes (e.g. Trichoderma spp.) rather than a natural process that may have been occurring in the fields where this experimentation was conducted

Self-citations of two authors were detected ( Sissons, M.; Simpfendorfer, S.) and large errors in the citation of literature, many items were not quoted, e.g.: 17, 19-24, 26, 29, 36, 39.

The use of our citations was either for reference to a method or because the work was relevant to the Australian situation where there are no other suitable references that could have been used. As for no listing in the main text of the references numbered, only #26 (now 33) was not listed, now corrected. All others are mentioned where appropriate. An explanation might be that if the reviewer is searching for example, for ref 39 using "[39]" as a search term, it will not be found because it is represented in the text as [38,39]. We checked a recent publication from this journal and have adjusted the reference list to match the required format.

Author Response File: Author Response.pdf

Reviewer 2 Report

The research entitled (Effect of Nitrogen Fertilization and Inoculation of Durum Wheat with Fusarium pseudograminearum on Yield, Technological Quality and Gluten Protein Composition) investigate the effects of FCR infection in the presence of different N fertiliser application rates in two commercial durum wheat varieties on grain yield, grain and semolina quality and relate the changes obtained to the mature grain gluten composition.

1. First the fungal name in the title and abstract must be in italics.

2. The abstract part missing numerical results.

3. In all the manuscript the fungal scientific names and the plants scientific names must be in italics.

4. The introduction part has very little information about Fusarium disease, the authors need to add more informations and relevant references.

5. What are the plant seed sources?

6. What are the moisture contents of the soil, humidity, and the speed of the wind during the experiment?

7. What is the source of the fungal isolate used and on what basis it was identified?

8. During these treatments what are the pathogenicity levels and the symptoms degrees?

9. The research needs extensive English editing and grammar corrections.

10. The reference section needs to revise one by one.

The research needs extensive English editing and grammar corrections

Author Response

We thank the reviewer for their helpful comments which we have addressed point form below.

First the fungal name in the title and abstract must be in italics.

All mention of the fungal name in the manuscript (excluding references) have been italicised

The abstract part missing numerical results.

We have revised the abstract and added some numerical data as suggested

In all the manuscript the fungal scientific names and the plants scientific names must be in italics.

As for 1.

The introduction part has very little information about Fusariumdisease, the authors need to add more informations and relevant references.

The introduction has been revised to include more background information about FCR with 5 additional references included

What are the plant seed sources?

We have inserted this text in section 2.2 “Each year seed of the two durum varieties was sourced from the same seed increase site grown under irrigation in the preceding season to ensure high levels of germination and vigor of the planting seed.”

What are the moisture contents of the soil, humidity, and the speed of the wind during the experiment?

We inserted this into section 2.2 “In both seasons the trial sites had full soil moisture profiles (~220 mm plant available water content 0-120 cm) due to high rainfall (471.2 mm in 2020 and 466.0 mm in 2021) in the 5-6 months prior to sowing (Table 1). Sowing dates were 9^th June 2020 and 6^th July 2021.”

FCR is not a leaf disease, so humidity and wind speed are not relevant to infection. FCR infection and expression is related to soil moisture (i.e. rainfall) and stress (temperature and moisture) during grain fill which is why we provided data for these parameters in Table 1.

What is the source of the fungal isolate used and on what basis it was identified?

We have inserted this into section 2.2 “Inoculum was an equal mixture of five separate isolates of F. pseudograminearum collected postharvest from commercially infected wheat crops around Moree and Narrabri in NNSW in December 2019. Single spore isolates were established to ensure purity and identification as F. pseudograminearum confirmed through quantitative PCR [18]. The same five isolates were used to prepare inoculum for the field experiments in 2020 and 2021.”

During these treatments what are the pathogenicity levels and the symptoms degrees?

We have included data in Suppl. Tables S3&S4 that show fusarium infection levels and these tables are referred to in section 3.2.

The research needs extensive English editing and grammar corrections.

We have checked the English and grammar carefully and hope these changes improves the document suitably

The reference section needs to revise one by one.

We have revised the reference list in line with this journals style

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

The authors enhanced the manuscript and gave sufficient answers to the questions. Only Figure 13 (A-D) has an insufficient resolution and needs to be improved.