Comparative analysis of distinct phenotyping methods for assessing wheat resistance and pathogen virulence among Fusarium species causing head blight disease

Abstract

Developing disease-resistant crops is a critical strategy for reducing chemical treatments and mitigating plant disease outbreaks, particularly amid global environmental changes. Fusarium head blight (FHB), caused by a complex of Fusarium species, is one of the most devastating cereal diseases, leading to significant economic losses and contamination of grain with harmful mycotoxins that threaten global cereal production and human health. The high variability in virulence within the complex of Fusarium spp and the lack of efficient high-throughput screening methods have impeded the development of resistant cultivars and made large-scale virulence testing labor-intensive and time-consuming. This study evaluates the efficacy of detached leaf, coleoptile, and seedling assays as high-throughput alternatives to the standard head infection assay for assessing the virulence of Fusarium species and differentiating wheat genotypes by resistance or susceptibility. Two near-isogenic wheat lines, one carrying FHB resistance loci and the other without, were used to assess the virulence of four Fusarium species. The seedling and coleoptile assays showed strong concordance with the traditional head infection assay, accurately reflecting differences in disease severity across Fusarium species and between wheat lines. Conversely, the detached leaf assay provided some differentiation among species but was inconsistent in identifying differences between plant genotypes. Across all assays, F. graminearum consistently exhibited the highest virulence, causing severe disease in leaves, stems, seedlings, and heads, while F. poae was the least virulent. Interestingly, F. culmorum and F. avenaceum displayed tissue-specific variability. These findings establish the coleoptile and seedling assays as rapid, high-throughput alternatives for breeding programs, accelerating the identification of FHB-resistant genotypes and reducing the reliance on the labor-intensive head assay.

Keywords: Cereals; Fusarium head blight; Head infection; High-throughput phenotyping.

Fig. 1

Lesions on the stems and leaves in coleoptile assay were measured 7 days after inoculation. a) A representative image from different replicates of the susceptible wheat genotype (bbaa) infected by F. graminearum. b) Effect of Fusarium species treatment on coleoptile stem lesion size (mm) on resistant (3B5A) and susceptible (bbaa) wheat for the coleoptile assay. Different letters above each boxplot indicate significant (α = 0.05) differences in group means as determined by the Tukey HSD Post-hoc test (n = 12). c) Scatterplot comparing stem lesion size in coleoptile assay to head infection assay. The regression line indicates a significant correlation (P = 1.76-05), with the shaded area representing the 95% confidence interval. d) Effect of Fusarium species treatment on coleoptile leaf lesion size (mm) on resistant (3B5A) and susceptible (bbaa) wheat for the coleoptile assay. e) Scatterplot comparing leaf lesion size in coleoptile assay to head infection assay. The regression line indicates a significant correlation (P = 4.58E-03), with the shaded area representing the 95% confidence interval

Fig. 2

a) Effect of Fusarium species treatment on disease index (0–4) on resistant (3B5A) and susceptible (bbaa) wheat for the seedling assay 14 days post inoculation. Different letters above each boxplot indicate statistically significant (α = 0.05) differences in group means as determined by the Tukey HSD Post-hoc test (n = 15). b) Scatterplot comparing seedling disease index in seedling assay to head infection assay. The regression line indicates a significant correlation (P = 1.10E-05), with the shaded area representing the 95% confidence interval

Fig. 3

Lesions on the leaves in the detached leaf infection assay were measured three days post-inoculation. Wheat leaves inoculated with sterile water (control, left) are compared to leaves inoculated with F. graminearum (right) in the susceptible wheat cultivar. a) A representative image of the setup used for assessing fungal infection. Wheat leaves were placed on water-agar media, followed by inoculation with spore suspensions of different Fusarium species. b) Effect of Fusarium species treatment on lesion area (%) on leaves of resistant (3B5A) and susceptible (bbaa) wheat. Different letters above each boxplot indicate statistically significant (α = 0.05) differences in group means as determined by the Tukey HSD Post-hoc test (n = 18). c) Scatterplot comparing leaf lesion in leaf assay to head infection assay. The regression line indicates a significant correlation (P = 6.37E-04), with the shaded area representing the 95% confidence interval

Fig. 4

Infection assessment on spikes were measured at 10 days post inoculation with a head assay. Spikes of (a) resistant (3B5A) and (b) susceptible (bbaa) wheat cultivars were inoculated with spore suspensions of various Fusarium species to evaluate pathogen impact and cultivar response. (c) Disease symptoms were evaluated by assessing the extent of bleaching and necrosis on the spikelet. (d) Effect of Fusarium species treatment on head infection rate (%) on resistant and susceptible wheat. Different letters above each boxplot indicate statistically significant (α = 0.05) differences in group means as determined by the Tukey HSD Post-hoc test (n = 8)