Molecular manipulation of the mating-type system and development of a new approach for characterizing pathogen virulence in Pyrenophora tritici-repentis

Authors: AMEEN, GAZALA - North Dakota State University; KARIYAWASAM, GAYAN - North Dakota State University; SHI, GONGJUN - North Dakota State University; Friesen, Timothy; Faris, Justin; ALI, SHAUKAT - South Dakota State University; RASMUSSEN, JACK - North Dakota State University; LIU, ZHAOHUI - North Dakota State University

Submitted to: Fungal Genetics and Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/14/2017
Publication Date: 10/16/2017
Citation: Ameen, G., Kariyawasam, G., Shi, G., Friesen, T.L., Faris, J.D., Ali, S., Rasmussen, J.B., Liu, Z. 2017. Molecular manipulation of the mating-type system and development of a new approach for characterizing pathogen virulence in Pyrenophora tritici-repentis. Fungal Genetics and Biology. 109:16-25. https://doi.org/10.1016/j.fgb.2017.10.004.

Interpretive Summary: Tan spot of wheat is a major problem for growers throughout the world with regular losses of 5 to 15% when environmental conditions are favorable to the causal fungal pathogen Pyrenophora tritici-repentis (Ptr). Ptr is a self fertilizing pathogen, meaning that the generation of sexual populations that can be used for characterizing virulence factors have been difficult. In the current work, we have manipulated the mating type genes of Ptr in order to force the pathogen to outcross, making it possible to harvest progeny that can be used in genetically mapping simple and complex traits including but not limited to virulence. This accomplishment lays the foundation for future work in characterizing this pathogen and how it is able to obtain nutrients from wheat, ultimately resulting in economic losses to growers worldwide.

Technical Abstract: The ascomycete Pyrenophora tritici-repentis (Ptr) is an important fungal pathogen worldwide that causes tan spot of wheat. The fungus is self-fertile because each isolate contains both mating type (MAT) idiomorphs. In this work, we developed knockouts of the MAT genes in Ptr and tested fertility of the knockout strains and outcrossing between the knockout strains carrying the opposite mating type. The fungal strains with deletions of either MAT1-1-1 or MAT1-2-1 did not form mature pseudothecia making them functionally heterothallic. The cross between the heterothallic strains of the same isolate (86-124) was fully fertile with the only difference compared to the homothallic wild type strain being the slightly lower percentage of pseudothecium formation. However, the cross between 86-124 (race 2, ToxA-containing isolate) and DW5 (race 5, ToxB-containing isolate) was partially fertile and had fewer mature pseudothecia. Furthermore, most mature asci produced only two or four instead of eight functional ascospores. A collection of ascospores from this cross was obtained and genotyped for the presence of the ToxA, ToxB and MAT genes as well as simple sequence repeat markers. The segregation of these genes and markers and recombination of different allele types at these loci was observed. This work clearly demonstrates that the fungus requires both MAT genes for sexual production and can undergo outcrossing and sexual recombination. It also establishes a new and practical way for further characterizing fungal virulence in Ptr through the development of segregating fungal populations and subsequent genetic analysis.