Review Reports

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The study investigated the diversity of Fusarium spp. of FGSC causing Fusarium head blight (FHB) on winter wheat in Poland in 2014-2017 and 2019 and also examined the potential impact of weather or environmental factors such as rainfalls and temperature on the fungal pathogen diversity. The samples collected during 5-year period at 26 sampling sites provided a valuable source to investigate the temporal and geographical dynamics of the fungal pathogen causing FHB and to examine the correlation between fungal incidence and the environmental factors. Authors stated in the methods/materials section that the number of samples used for data collection/analysis varied from year to year, suggesting that the number of samples collected at each sampling sites varied year to year. However, the data presented in the manuscript were a cumulative data of 5-year period (except the supplemental data) that was based on pg of Fusarium DNA in mcg of wheat DNA. Authors used these data sets to investigate the geographical variation in the fungal pathogen occurrence. The manuscript didn’t show if all wheat samples were tested positive for Fusarium by qPCR or if there are any wheat samples tested negative. Since the amount of the fungal DNA relative to the wheat DNA will be affected by the number of fungal positive samples and the number of wheat samples, to better understand the presented data in the manuscript, it is recommended to include the detail of the number of samples collected from each sampling site in each of 5-year period together with summarized qPCR data. In addition, authors may need to thoroughly examine any possible effect of the varying sample numbers at each sampling site/year on the data interpretation. Overall the manuscript provide a merit for plant pathologists to understand the FHB incidence and the environmental factors contributing the temporal dynamics of fungal population structure in FGSC, and it is recommended for publication with minor, but necessary, revision.

 

Line 40: F. graminearum

Line 68-69: italicize ‘Microdochium nivale’

Line 94: Does this mean that since 2017, the two winter wheat cultivars in all 26 experimental stations have been replaced to Hondia, a moderately resistance cultivar? If so, is there any possibility that the occurrence of Fusarium spp. during the test period was affected by this change?

Line 100: The number of samples collected from each location in each year need to be included in the manuscript. This information can be critical to understand better the variation in the occurrence of Fusarium spp. in different regions and also in different years.

Line 196-201: This paragraph described the rainfall variations in May-July of 2014-2019. However, it is not clear if the description in the text matched to the data in the figure. Was the rainfall variation based on IQR or IQR+whiskers (or IQR+whiskers+outliers)? Re-confirmation is needed for clarification. Also, it is recommended to double-check the temperature variation in the text and the temperature data in the figure (in Line 178-184).

Line 216-217: It is not clear what ‘trace amount of Fusarium DNA’ means. Clarification needed. In addition, this section described about the percentage of samples that have Fusarium DNA. However, there is no data in the manuscript to confirm this statement.

Line 210-230: The data in Figure 3 and 4 are pg of Fusarium DNA in mcg of wheat DNA. However, in line 100, authors said that the number of samples collected at 26 sampling sites were varied in different years. As commented above, it may be necessary to include this variation in the data presentation and interpretation.

Line 275: Table 2 was not cited in the text.

Line 281-283: There is no data to support what was stated in the text. It is recommended to include the supporting data.

Line 315-322: In this section, authors mentioned about flowering, ripening, stem elongation and milk-dough development to explain the correlation between Fusarium DNA content and the two weather variables. If Figure 6 include those wheat developmental stage, it will be of help for readers to understand the data better.

 

 

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files

General comments: Authors stated in the methods/materials section that the number of samples used for data collection/analysis varied from year to year, suggesting that the number of samples collected at each sampling sites varied year to year. However, the data presented in the manuscript were a cumulative data of 5-year period (except the supplemental data) that was based on pg of Fusarium DNA in mcg of wheat DNA. Authors used these data sets to investigate the geographical variation in the fungal pathogen occurrence. The manuscript didn’t show if all wheat samples were tested positive for Fusarium by qPCR or if there are any wheat samples tested negative. Since the amount of the fungal DNA relative to the wheat DNA will be affected by the number of fungal positive samples and the number of wheat samples, to better understand the presented data in the manuscript, it is recommended to include the detail of the number of samples collected from each sampling site in each of 5-year period together with summarized qPCR data. In addition, authors may need to thoroughly examine any possible effect of the varying sample numbers at each sampling site/year on the data interpretation. Overall the manuscript provide a merit for plant pathologists to understand the FHB incidence and the environmental factors contributing the temporal dynamics of fungal population structure in FGSC, and it is recommended for publication with minor, but necessary, revision.

Response: The number of samples from a specific location remained the same each year. However, the number of sampling locations varied over the years, although we tried to include at least one location from each province. Unfortunately, in some years, samples from several regions were missing, as shown in Figure S1. We added Table 2 showing the percentage of samples with Fusarium DNA detected, as well as the percentage of samples in which Fusarium DNA was at a trace amount. 

Comment 1: Line 40: F. graminearum; Line 68-69: italicize ‘Microdochium nivale’

Response: All text was corrected and reviewed to ensure proper italicization.

Comment 2: Line 94: Does this mean that since 2017, the two winter wheat cultivars in all 26 experimental stations have been replaced to Hondia, a moderately resistance cultivar? If so, is there any possibility that the occurrence of Fusarium spp. during the test period was affected by this change?

Response: Cultivar Bamberka was replaced by Hondia in 2017. Cultivar Arkadia was sampled every year. Comparing results from 2014-2016 and 2107, 2019, there was no effect from the cultivar change. In 2017, the level of DAN was high, and in 2019, levels were similar to 2016.

Comment 3: Line 100: The number of samples collected from each location in each year need to be included in the manuscript. This information can be critical to understand better the variation in the occurrence of Fusarium spp. in different regions and also in different years.

Response: We added Table 1 showing the number of samples and some explanation of this in Material an Methods

Comment 4: Line 196-201: This paragraph described the rainfall variations in May-July of 2014-2019. However, it is not clear if the description in the text matched to the data in the figure. Was the rainfall variation based on IQR or IQR+whiskers (or IQR+whiskers+outliers)? Re-confirmation is needed for clarification. Also, it is recommended to double-check the temperature variation in the text and the temperature data in the figure (in Line 178-184).

Response: Indeed, it was very unclear. So, we rewrote these lines and improved Figure 2. 

Comment 5: Line 216-217: It is not clear what ‘trace amount of Fusarium DNA’ means. Clarification needed. In addition, this section described about the percentage of samples that have Fusarium DNA. However, there is no data in the manuscript to confirm this statement.

Response: An explanation of 'trace amount' was added to the Materials and Methods section. We also included Table 2, which shows the percentage of samples with detected Fusarium DNA as well as the percentage of samples with quantified Fusarium DNA.

Comment 6: Line 210-230: The data in Figure 3 and 4 are pg of Fusarium DNA in mcg of wheat DNA. However, in line 100, authors said that the number of samples collected at 26 sampling sites were varied in different years. As commented above, it may be necessary to include this variation in the data presentation and interpretation.

Response: Numbers of samples collected and analyzed in different years were shown and explained. 

Comment 7: Line 275: Table 2 was not cited in the text.

Response: corrected

Comment 8: Line 281-283: There is no data to support what was stated in the text. It is recommended to include the supporting data.

Response: Lines were rewritten and additional supporting data were added (Figure S3).  

Comment 9: Line 315-322: In this section, authors mentioned about flowering, ripening, stem elongation and milk-dough development to explain the correlation between Fusarium DNA content and the two weather variables. If Figure 6 include those wheat developmental stage, it will be of help for readers to understand the data better.

Response: Figure 6 was improved.

Reviewer 2 Report

Comments and Suggestions for Authors

Please see the attachment. 

Comments for author File: Comments.pdf

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files.

General comments: However, the narrative occasionally lacks depth in discussing the implications of mycotoxin risks, despite briefly mentioning them. Some sections feel descriptive without sufficient integration of prior Polish studies for context. The study skips 2018 without explanation, potentially biasing temporal trends.

Response: We improved the discussion section, also mentioning the mycotoxin issue. Polish as well as German studies were more deeply discussed. Lack of 2018 data was explained.  

Comments 1:  Lines 1–3: The title is concise and accurately reflects the study's scope; however, consider adding "DNA-based quantification" to emphasize the novel methodological angle, as this is central to the work.

Response: We changed the title accordingly to: DNA-based quantification of Fusarium species in winter wheat grain in Poland from 2014 to 2017 and 2019.

Comments 2: Lines 25–27: Add "mycotoxins" or "F. graminearum" for specificity, given their prominence in the abstract and introduction.

Response: Added F. graminearum. Results for mycotoxins were not presented in this paper.

Comments 3: Line 15-16 claims "F. graminearum s.s. being the most dominant," but results (lines 220–227) show year-to-year variation (e.g., F. poae dominant in 2015), which could mislead; revise to "predominantly" or specify the overall trend.  

 Response: These lines were rephrased. 

Comments 4: Line 20–23: Positive precipitation correlation and negative temperature link are clear but could quantify (e.g., "r = 0.XX").  

Response: We added r values for Spearman rank correlations with precipitation and temperature. 

Comment 5: Line 65–66 reference to maize area (1,814,974 ha in 2024) is forward-looking but verify source accuracy (https://kukurydza.info.pl/—ensure it's peer-reviewed).  

Response: We added references from FAO and Eurostat. Acreage for 1990 was corrected as the value of 59,000 regarded only grain maize.

Comment 6: Line 52–56 on Northeast vs. Southwest Europe could cite more recent data (post-2020) for currency

Response: Added some references, including one review. However, there are only a few articles on Fusarium species on wheat published after 2020. Some of them were cited in the Discussion.

Comment 7: Line 100: note varying sample numbers/year, but do not specify them; add a table for transparency.  

Response: Table was added (Table 1)

Comment 8: Line 145–146: Standard curves are described, but efficiency values (e.g., R²) should be reported.  

Response: Efficacy values were reported.

Comment 9: Line 172: Weather data windows are mentioned but not defined. Clarify as "92 phenological periods across sites/years."

Response: Changed accordingly. 

Comment 10: Line 185–191 repeats "lowest in 2014/highest variation in 2019" phrasing—minor redundancy with lines 178–180; consolidate.  

Response: We rephrased these sentences.

Comment 11: Line 219: "significantly higher" claims need p-values or stats.  

Response: A description of the statistical analysis was added to the Materials and Methods section, and a corresponding description was included in the title of Figure 3.

Comment 12: Expect integration of PCA results (mentioned in the abstract) and correlations in the discussion section.

Responses:  The Results of PCA and correlation analysis were discussed