The necrotrophic fungal pathogen Pyrenophora tritici‐repentis (Ptr) causes the major wheat disease tan spot, and produces multiple necrotrophic effectors that contribute to virulence. The proteinaceous effector ToxA induces necrosis in wheat genotypes possessing the Tsn1 gene, although the importance of the ToxA–Tsn1 interaction itself in varietal disease development has not been well studied. Here, 40 Australian spring wheat varieties were assessed for ToxA sensitivity and disease response to a race 1 wildtype Ptr isolate and ToxA‐deleted strain at both seedling and tillering growth stages. ToxA sensitivity was generally associated with disease susceptibility, but did not always predict spreading necrotic symptoms. Whilst the majority of Tsn1 varieties exhibited lower disease scores following toxa mutant infection, several exhibited no distinct differences between wildtype and toxa symptoms. This implies that ToxA is not the major determinant in tan spot disease development in some host backgrounds and indicates the presence of additional effectors. Unexpectedly, several tsn1 varieties exhibited a reduction in disease severity following toxa mutant inoculation, which may suggest an indirect role for ToxA in pathogen fitness. Additionally, increased chlorosis was observed following toxa mutant infection in three varieties, and further work is required to determine whether this is likely to be due to ToxA epistasis of ToxC symptoms. Taken together, these observations demonstrate that Ptr interacts with the host in a complex and intricate manner, leading to a variety of disease reactions that are dependent or independent of the ToxA–Tsn1 interaction.

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The virulence of 57 Australian isolates of Pyrenophora tritici‐repentis (Ptr), a necrotrophic fungal pathogen responsible for the major wheat disease tan spot, was assessed through plant infection assays. Isolates collected from the northern, southern, and western wheat‐cropping regions of Australia were evaluated against 16 Australian bread wheat cultivars under controlled growth conditions. Following infection, the wheat panel displayed varying disease symptoms ranging from tiny necrotic specks to spreading chlorotic and necrotic lesions. Analysis of variance indicated that the wheat cultivar exhibited a greater effect on the disease response, explaining 62.7% of the variation, in comparison to the isolate (10.4%). The interaction between the cultivar and the isolate was statistically significant and was attributed to 9.8% of the total variation. All Ptr isolates examined were able to cause disease, but did not display a clear distinction in virulence on the wheat panel investigated, instead showing subtle differences in aggressiveness. Based on the disease responses, there was no obvious pattern between isolate aggressiveness and cropping region. Some cultivars, such as Hydra, exhibited an effective level of resistance in relation to the panel of isolates tested. All 57 Ptr isolates were found to possess the ToxA effector gene and lack the ToxB effector gene. The gene expression level of ToxA was up‐regulated at 3 days postinfection in both ToxA‐sensitive and ‐insensitive cultivars, independent of ToxA–Tsn1 recognition.

Tan spot disease is caused by Pyrenophora tritici-repentis (Ptr), one of the major necrotrophic fungal pathogens that affects wheat crops globally. Extensive research has shown that the necrotrophic fungal effectors ToxA, ToxB and ToxC underlie the genetic interactions of Ptr race classification. ToxA and ToxB are both small proteins secreted during infection, however the structure of ToxC remains unknown. In line with the recent discovery of ToxC1 gene that is involved in ToxC production, a subset of 68 isolates collected from the Australian wheat cropping regions were assessed for presence of all three effectors by pathotyping against four tan spot wheat differential lines and PCR amplification of ToxA, ToxB and ToxC1. Based on the disease phenotypes, the 68 isolates were grouped into two races with 63 classified as race 1 and five as race 2. A representative selection of each race was tested against eight Australian commercial wheat cultivars and showed no distinction between the virulence levels. Sequencing of ToxA showed that both races had identical gene sequences of haplotype PtrA1. All the race 1 isolates possessed ToxC1 but three race 2 isolates also contained ToxC1 despite being unable to induce a spreading chlorotic symptom on the ToxC differential line. QTL mapping confirmed the absence of the ToxC-Tsc1 association in disease response caused by the ToxC1-containing race 2 isolate, however ToxC1 expression was detected during plant infection. Altogether, these results suggest that there is a complex regulatory process involved in the production of ToxC within the Australian race 2 isolates.