INVESTIGATION INTO THE MATING SYSTEM AND POPULATION STRUCTURE OF COLLETOTRICHUM TRUNCATUM FROM LENTIL

Colletotrichum truncatum is the pathogenic agent of lentil anthracnose, a disease responsible for severe yield losses in Western Canada. This ascomycetous fungus is believed to reproduce asexually under field conditions, but the sexual stage, Glomerella truncata, has been obtained under laboratory conditions. Preliminary studies on a limited number of isolates suggested that C. truncatum exhibits the typical bipolar mating system of heterothallic ascomycetous fungi; however, this result had to be confirmed on a larger number of isolates, especially considering that all other Colletotrichum species studied to date do not seem to conform to this system. The aim of this study was to increase understanding of the mating system of C. truncatum. Based on preliminary information, it was hypothesized: 1) that C. truncatum is a heterothallic fungus with a unilocus, biallelic mating system; and 2) that sexual reproduction occurs in the field. These hypotheses were tested by i) performing classical mating studies to confirm heterothallism and bipolarity of C. truncatum; ii) analyzing the genetic basis for cross fertility with the use of molecular markers; iii) probing for the presence of genes responsible for mating types; iv) determining if different mating types coexist on a small geographical scale; and v) assessing the genetic diversity of field isolates of C. truncatum by conducting a molecular population study, and determine the extent of linkage disequilibrium. Twenty-one isolates were crossed in all possible combinations on lentil material. When each isolate was used alone, no sexual structures were detected. However, when two different isolates were put in contact on lentil stems, perithecia were produced in 22% of the cases. The 21 field isolates fell into two mating incompatibility groups (IG), supporting the bipolar nature of the mating system of C. truncatum. Molecular markers differing between two parental isolates were found in their progeny, confirming heterothallism. The results are consistent with the typical mating system of ascomycete, in which mating types are controlled by the mating type (MAT) locus with two alleles, MAT1-1 and MAT1-2, each controlling a mating type. However, in C. truncatum, degenerate primers targeting conserved regions of MAT1-2 revealed that all isolates tested carried a copy of MAT1-2, independently of their mating incompatibility group. This is not consistent with the typical mating system of heterothallic ascomycetes, suggesting that, as proposed for other Colletotrichum species, the mating system of C. truncatum is likely controlled at least partially by another system. A molecular population study undertaken on C. truncatum isolates from different Saskatchewan locations and crop years showed that genetic diversity was low and suggested a high level of clonality. However, a moderate level of linkage disequilibrium existed in the population, indicating the possibility of at least some level of sexual reproduction. This study also showed that isolates of different mating types were spatially coexisting in the same field, sometimes on the same plant, suggesting that geographical isolation is probably not a factor restricting sexual reproduction