Modeling blackleg severity and incidence in canola and identification of molecular markers linked to a Leptosphaeria maculans avirulence gene

Canola is the most important oilseed crop in Canad,a.Blackleg caused by Leptospheria maculans is the rnost important disease in canola in the prairie provinces.While the sexual spores have been implicated as primary inoculum, there has not much work done on the asexual pycnidiospores and their role in primary infection in Canada.With the frnding of new blackleg pathogenicity groups in the prairies, there has been renewed interest in understanding the host-pathogen interactions, and the role of R-genes and corresponding avirulence genes in the pathogen.Therefore, the two main goals of the epidemiolo gy part of this study were to investigate the role of pycnidiospores of Leptosphaeria maculans ondisease development on plants infected at the cotyledon (i.0), 3-leaf (1, 03) and 6-leaf (1, 06) stages of Brassica napus cvs.Westar and Q2 under field conditions and to develop nonlinear weather-based models to describe blackleg severity and incidence of the infected plants.The results suggested that pycnidiospores were a source of primary inoculum for 2004 and 2005.The results showed that the mean blackleg severity of the plants infected at the cotyledon stage was significantly higher than plants infected at the other two stages.These results also confirmed that the mean blackleg severity of the plants infected at the 3-leaf stage was higher than plants infected at the 6-leaf stage.Blackleg incidence in the plants infected at the cotyledon stage was significantly higher than the other two stages.No signifcant difference in disease incidence was observed between the plants infected al3-leaf and 6-leaf stage.Pearson's correlation coefficient showed that two weather variables, total rainfall per week (R) and average maximum temperature per week (Tmx), werc significantly An atternpt was also made to develop molecular markers for the corresponding avirulence gene in L. mqculans to the LepRI resistance gene in B. napus, and to obtain fuither evidence of a gene-for-gene interaction.The segregation of 94 F1 progen! of Z. maculans from a cross of avirulent isolate (99-56) and virulent isolate (g7-41) on the monogenic line ddm-126s-1 containing LepRI was studied.The progenies were isolated and phenotyped for reactions on the line ddm-1 26s-1.The population segregated in 1 :1 ratio,44 avirulent and 50 virurent d : 0.79, p :0.66), indicating single gene contror of 99-56 avirulence on dm-726s-1.Bulked segregant analysis was used to identiflz two IV Sequence-related amplified polymorphic (SRAP) markers closely linked to the avirulence gene.We have named this avirulence gene AvrLepRI.Two flanking SRAP markers, SA7-108 and SA7-490, were linked to AvrLepRI at 9.6 and 7.8 cM, respectively.This work present further evidence for a gene-for-gene rnodel which has been suggested previuosly in this pathosystem and represents an initial step toward map-based cloning of this gene.V CHAPTER 1 1.2.1.6.3Pathogenicity trait Various cultivars and lines of B. napus were used to differentiate L. maculans and L. biglobosa isolates into dfferent subgroups called pathogenicity groups (PG).An initial B. nqpus differential set comprising of Westar, Glacier and Quinta was used to classify the isolates into three pathogenicity groups; PG2 (avirulent on cvs Quinta and Glacier), PG3 (avirulent on cv Quinta, virulent on cv.Glacier) and PG4 (virulent on all three cultivars, Koch et al, 1991).In another differential set, Westar was replaced by adding winter type B. napus cvs.Liberon and Jet Neuf to the set, leading to the description of six PG that were termed A1 to A6 (Badaw y et al, 1gg1).PG4 isolates were thus further divided into A1 (virulent on Jet Nuef) and A5 (avirulent on Jet Neuf); PG3 into A2 (virulent on Jet Neuf) and A6 (avirulent on Jet Neuf); and PG2 into A4 and A3 (virulent and avirulent on Jet Neuf, respectively).Cotyledon inoculations on eight B. napus differentials (Liberon, Quinta, Glacier, Jet Neuf, Doublol, Karat, R83-14.DH41 and R83-14.DH26) were used to subdivide European PG3 and PG4 isolates into seven subgroups (Kuswinanti et al, 1995).Kerri (1999) subdivided PG2 isolates into 15 subgroups and PG3 and PG4 into eight subgroups based on differential set of Westar, Glacier, Quinta, Quantum , Sentry, Sprint,Val-1 and Dac-1.1.2.1.6.4Molecular traits Different molecular tools (AFLP, Rep-PCR fingerprints, ITS-RFLP) with combination of morphological and biological criteria were used to divide the L. maculans species in two different species L. maculans and Z. biglobosa (Mendes-Pereira 2003 and

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