The Molecular Basis of Demethylation Inhibitor Fungicide Resistance in Cercospora beticola and the Role of Seed Inoculum in Cercospora Leaf Spot Disease of Sugar Beet

Cercospora leaf spot (CLS) is the most destructive foliar disease of sugar beet worldwide. CLS is caused by the filamentous fungus Cercospora beticola. Disease management currently relies upon timely application of fungicides, but reliance on certain chemical classes has led to the development of resistance in multiple C. beticola populations. One such class is the demethylation inhibitor (DMI) fungicides of which the genetic basis of resistance has been unclear. Therefore, the first objective of this PhD research was to perform a genome-wide association study on 190 C. beticola isolates to identify mutations associated with tetraconazole (a common DMI) fungicide resistance. Whole genome resequencing identified multiple novel loci associated with sensitivity to tetraconazole including a pleiotropic drug resistance ATP-binding cassette transporter, a regulator of G-protein signaling domain (RGD) protein, a DYRK protein kinase and mutations within and upstream of the gene encoding the DMI target Cytochrome P450 51 (CYP51). This demonstrated the genetic complexity in resistance to DMIs and suggested the involvement of cellular signaling and multidrug resistance as well as target site mutations. The second objective of this research was to investigate the potential of seedborne C. beticola to initiate CLS disease in sugar beet. We showed that viable C. beticola was present in commercial sugar beet seed lots and could function as primary inoculum to cause CLS symptoms in seedlings. All strains identified were resistance to QoI fungicide chemistries and most were also resistant to DMI fungicides. Detection of C. beticola DNA in xylem sap suggested that the fungus may be systemically colonizing the plant via the vascular system. Long-read nanopore sequencing detected other potential pathogenic fungi in seed DNA (e.g. Fusarium and Alternaria spp.) that may also act as primary inoculum sources for important sugar beet diseases. This PhD research has improved our understanding of the development of DMI fungicide resistance in C. beticola, as well as highlighted the importance of seed inoculum in the manifestation of CLS in sugar beet, both of which could improve CLS disease management in the future.