ABSTRACT

THOMAS, ANNA. Biology, Epidemiology and Population Genomics of Pseudoperonospora cubensis, the Causal Agent of Cucurbit Downy Mildew. (Under the direction of Dr. Peter S. Ojiambo and Dr. Ignazio Carbone).

Cucurbit Downy Mildew (CDM) caused by the obligate ,

Pseudoperonospora cubensis, is economically the most important disease of cucurbits worldwide. During the past two decades, a resurgence of CDM has occurred around the world resulting in severe disease epidemics. In the United States, resurgence of the disease occurred in 2004 and resulted in significant crop losses. Introduction of a new lineage, a new pathotype or new genetic recombinants of the pathogen have been suggested as potential reasons for this resurgence. The potential for sexual reproduction, the current pathotype structure of P. cubensis in the United States, and the role different cucurbit host types and geography play in the shaping the genetic structure of P. cubensis were investigated.

Pairing assays involving known mating type tester strains and unknown isolates revealed, for the first time, the presence of both A1 and A2 mating types in the United States.

Mating type was found to be significantly (P < 0.0001) associated with host type with isolates collected from cucumber being primarily of the A1 mating type, while those from squash and watermelon being of the A2 mating type. Similarly, a significant (P = 0.0287) association of mating type with geography was also observed. Isolates collected from northern tier states of Michigan, New Jersey, New York and Ohio were all A1, while isolates belonging to either the A1 or A2 mating type were present in equal proportions in southern- tier states of Alabama, Florida, Georgia, North Carolina, South Carolina and Texas.

Plasmolysis test showed that about 40% of the oospores produced were viable and this suggests that oospores may play a role in the epidemiology of CDM in the United States. Compatibility assays were conducted using 22 isolates and fifteen host differentials to classify the isolates into pathotypes. Based on the compatibility with the differential host set, five pathotypes (1, 3, 4, 5 and 6) were identified among the 22 isolates. Pathotypes 1 and 3 had not been previously described in the United States and isolates of these two new pathotypes were also compatible with Poinsett 76, a cultivar of C. sativus known to be resistant to CDM prior to 2004. Virulence within the pathogen population was expressed based on virulence factors, virulence phenotypes and virulence complexity. The number of virulence factors ranged from 2 to 8 indicating a complex virulence structure with 77% of the isolates having 5 to 8 virulence factors. Thirteen virulence phenotypes were identified and the mean number of virulence factors per isolate and mean number of virulence factors per virulence phenotype was 5.05 and 5.23, respectively, indicating that complex isolates and phenotypes contributed equally to the complex virulence structure of P. cubensis. Gleason and Shannon indices of diversity were 3.88 and 2.32, respectively and indicated a diverse virulence structure of P. cubensis population within the United States.

Comparative whole genomic analysis of nine isolates collected from diverse cucurbit hosts revealed the presence of two distinct evolutionary lineages. In addition, phylogenetic analysis showed that P. cubensis and its sister species, P. humuli, shared a recent common ancestor. A reconstruction of ancestral recombination suggested a hybrid origin of lineage II of P. cubensis, with P. humuli and lineage I of P. cubensis as putative parents. Subsequently, the role of different cucurbit host types in shaping the structure of P. cubensis population was investigated using 93 isolates with the double digest Restriction Associated DNA Sequencing technology. Many fixed polymorphisms throughout the genome separated lineage II that included isolates mainly from Cucumis spp. from lineage I that comprised i