Histopathological assessment of the crown rot pathogen Fusarium pseudograminearum was performed using fluorescence microscopy of culm tissues of six cereal genotypes grown in inoculated field conditions. Tissue samples were collected at 10, 16, and 22 weeks after planting (WAP). Colonization of culm tissues was initiated through epidermal penetration, most distinctly through stomatal apertures, and progressed into the parenchymatous hypoderm, which exhibited the discoloration used as the basis for visual assessment of disease. Hyphae spread from the culm base vertically through the tissues, initially via the hypoderm and pith cavity. Colonization of sclerified cells occurred later in the disease process. Both xylem and phloem tissues became colonized by 16 WAP in all host genotypes, with colonization being less extensive in the more resistant genotypes. Culms displaying dead head symptoms revealed dense colonization in at least the first three internodes, with frequent xylem vessel and phloem cell occlusions. Paired living culms from the same plants exhibited less extensive colonization. These observations have revealed the ability of F. pseudograminearum to colonize all cell types of nodal and internodal sections, including vascular tissues, across all host genotypes. This study is the first detailed examination of the pattern of F. pseudograminearum colonization in adult hosts and indicates a potential vascular mechanism by which the effects of crown rot are produced.

Assessment among cereal genotypes of relative seedling resistance to the crown rot pathogen Fusarium pseudograminearum has been primarily based on visual discoloration of the leaf sheaths. This study is the first to investigate the relationship between the widely used visual rating of seedling leaf sheath discoloration and the degree of colonization of these tissues by the pathogen, based on quantitative polymerase chain reaction (qPCR) of fungal DNA using primers specific for the translation elongation factor α sequence. Fourteen-day-old seedlings of four hard white spring wheat genotypes which differ in their degree of resistance to the pathogen, based on the expression of visible symptoms, were inoculated using a droplet method and assessed weekly from 7 to 35 days after inoculation (dai) for both discoloration and fungal DNA content per unit of tissue weight. Both visual assessment of disease symptoms and qPCR of fungal biomass indicated significant differences between the partially resistant and susceptible wheat genotypes from 14 dai. Visual discoloration of leaf sheath tissues was strongly correlated with fungal biomass estimated by qPCR in all four genotypes; however, this correlation became weaker with increasing time after inoculation. Significant correlations between these parameters were indicated at 14, 21, and 28 dai whereas, by 35 dai, the correlation was not significant. Evaluation of plants at 14 dai provided a rapid test which gave clear discrimination between lines for both parameters and was the time point of closest correlation between fungal colonization and disease symptoms. Symptom expression at all times following inoculation was accompanied by tissue infection, and at no time was symptomless infection observed under this screening environment. These qPCR results confirm that visual assessments of disease symptoms reflect the extent of tissue colonization by the pathogen in recently colonized tissues and confirm the validity of visual assessments for disease rating in high-throughput screening of breeding materials.

A novel fluorescence‐based differential staining procedure has been developed for the rapid visualization of Fusarium pseudograminearum hyphae in cereal tissues. Infected tissues are stained with safranin, which binds to host cell walls, while hyphal tissues are stained with the fluorescent dye solophenyl flavine 7GFE. The method is rapid, does not require clearing of culm cross‐sections, and provides high contrast informative images of fungal and plant structures during pathogenesis.

Fusarium crown rot is a significant disease of durum wheat (Triticum turgidum L. var. durum), which exhibits high levels of disease susceptibility. The most extreme symptom of crown rot is a prematurely senescing culm that typically fails to set grain. Individual crown rot-affected durum wheat plants displaying both nonsenescent and prematurely senescent culms were harvested to compare visual discoloration, Fusarium pseudograminearum biomass, and vascular colonization in culm sections sampled at three different heights above the crown. Field samples of EGA Bellaroi were collected at Wellcamp, QLD, in 2011, 2012, 2013, and 2014, and of Hyperno at Narrabri, NSW, in 2014. Prematurely senescent culms exhibited greater visual discoloration, F. pseudograminearum biomass, and vascular colonization than nonsenescent culms in each year they were examined. The extent of these differences varied between environments and timing of collection in each year. Vascular colonization initially occurred in xylem vessels and spread into phloem tissues as disease severity increased. The increased presence of hyphae in vascular bundles of prematurely senescing culms provides strong evidence for the hypothesis that restriction of water and nutrient movement in a diseased culm is a key factor in crown rot severity.

Histopathological assessment of infection by the crown rot pathogen Fusarium pseudograminearum in wheat seedling tissues was performed using fluorescence microscopy. The coleoptiles and leaf sheaths of four host cultivars of differing susceptibility were examined. Leaf sheaths were most frequently penetrated via stomata, indicated by initial lesions forming at the guard cells. Internally, cell wall penetration was facilitated by penetration structures which appeared as hyphal swellings or septate foot‐shaped appressoria. Colonization of leaf sheaths resulted in the re‐emergence of hyphae from stomata on both surfaces of the sheath. These hyphae are hypothesized to have two major roles; first as exploratory hyphae for colonization of new tissues, and secondly as sites of profuse conidial production. The formation of conidia on the leaf sheath surface was only recorded on the most susceptible bread wheat genotype. No other major differences in host–pathogen interactions were observed among these cultivars. Almost all cell types in the leaf sheath tissues were extensively colonized, except for the vascular bundles and silica cells. This investigation provides the first comprehensive assessment of F. pseudograminearum infection structures and growth patterns during the infection of wheat seedlings.

Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), the causal agent of common rootrot (CRR) and foliar spot blotch (SB) diseases in