Septoria tritici blotch (STB; Zymoseptoria tritici) is the most important leaf disease of wheat in Northern and Western Europe. The problem of fungicide resistance in Z. tritici populations is challenging future control options. In order to investigate differences in azole performances against STB, 55 field trials were carried out during four seasons (2015-2018). These trials were undertaken in ten different countries across Europe covering a diversity of climatic zones and agricultural practices. During all four seasons, four single azoles (epoxiconazole, prothioconazole, tebuconazole and metconazole) were tested. Increasing variability in the performances of these azoles against STB was observed across Europe. The efficacy of the tested azoles varied considerably across the continent and between countries. The shifts in disease control from these commonly used azoles were confirmed by increasing EC 50 values for epoxiconazole, prothioconazole-desthio and metconazole. The sensitivity towards tebuconazole remained relatively low across the four years. The frequencies of CYP51 mutations varied substantially across Europe, with a clear pattern of significantly decreasing frequencies of D134G, V136A and S524T in the local Z. tritici populations from west to east. In contrast, no major differences were seen for CYP51 mutations V136C, A379 and I381V. The four azoles showed different levels of cross-resistance, which again depended on specific CYP51 mutations. Across the four seasons, the single azoles increased the yields between 9 and 11% on average.

Demethylation inhibitor (DMI) and succinate dehydrogenase inhibitor (SDHI) fungicides are currently relied upon for the control of septoria tritici blotch (STB) in European wheat fields. However, multiple mutations have occurred over time in the genes encoding the targeted proteins that have led to a practical loss of fungicide efficacies. Among the different amino acid substitutions in Zymoseptoria tritici associated with resistance to these fungicides, S524T in CYP51 (DMI target) and H152R in SdhC (SDHI target) are regarded as conferring the highest resistance factors to DMI and SDHI, respectively. To facilitate further studies on the monitoring and selection of these substitutions in Z. tritici populations, a multiplex allele‐specific quantitative PCR (qPCR) assay allowing for estimation of both allele frequencies in bulk DNA matrices was developed. The assay was then used on complex DNA samples originating from a spore trap network set up in Belgium, Denmark, Sweden, and Ireland in 2017 and 2018, as well as on leaf samples with symptoms. The S524T allele was present in all field samples and its proportion was significantly higher in Ireland than in Belgium, whereas the proportion of H152R was only sporadically present in both countries. The frequency of S524T varied greatly in the airborne inoculum of all four countries; however, the H152R allele was never detected in the airborne inoculum. The method developed in this study can be readily adopted by other laboratories and used for multiple applications including resistance monitoring in field populations of Z. tritici.

BACKGROUND: Over the past decade, demethylation inhibitor (DMI) and succinate dehydrogenase inhibitor (SDHI) fungicides have been extensively used to control to septoria tritici blotch, caused by Zymoseptoria tritici on wheat. This has led to the development and selection of alterations in the target-site enzymes (CYP51 and SDH, respectively). RESULTS: Taking advantage of newly and previously developed qPCR assays, the frequency of key alterations associated with DMI (CYP51-S524T) and SDHI (SDHC-T79N/I, C-N86S and C-H152R) resistance was assessed in Z. tritici-infected wheat leaf samples collected from commercial crops (n = 140) across 14 European countries prior to fungicide application in the spring of 2019. This revealed the presence of a West to East gradient in the frequencies of the most common key alterations conferring azole (S524T) and SDHI resistance (T79N and N86S), with the highest frequencies measured in Ireland and Great Britain. These observations were corroborated by sequencing (CYP51 and SDH subunits) and sensitivity phenotyping (prothioconazole-desthio and fluxapyroxad) of Z. tritici isolates collected from a selection of field samples. Additional sampling made at the end of the 2019 season confirmed the continued increase in frequency of the targeted alterations. Investigations on historical leaf DNA samples originating from different European countries revealed that the frequency of all key alterations (except C-T79I) has been gradually increasing over the past decade. CONCLUSION: Whilst these alterations are quickly becoming dominant in Ireland and Great Britain, scope still exists to delay their selection throughout the wider European population, emphasizing the need for the implementation of fungicide antiresistance measures.

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