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Three avirulence genes, AvrLm1, AvrLm6, and AvrLm4-7, were recently identified in Leptosphaeria maculans and found to be localized as solo genes within large noncoding, heterochromatin-like regions mainly composed of retrotransposons, truncated and degenerated by repeat-induced point mutation (RIP). The Rlm6 resistance gene has been overcome within 3 years in outdoor experiments in France and, here, we investigate the molecular basis of evolution toward virulence at the AvrLm6 locus. A region of 235 kb was sequenced in a virulent isolate and showed the deletion of AvrLm6 and three divergent mosaics of retrotransposons. AvrLm6 was found to be absent from 66% of 70 virulent isolates, with multiple events of deletion. The sequencing of virulent alleles in 24 isolates revealed a few cases of point mutations that had created stop codons in the sequence. The most frequent mutation events, however, were RIP, leading to the modification of 4 to 9% of the bases compared with the avirulent allele and generating 2 to 4 stop codons. Thus, RIP is described for the first time as an efficient mechanism leading to virulence and the multiple patterns of mutation observed suggest that multiple RIP events could occur independently in a single field population during 1 year.

Leptosphaeria maculans, the cause of stem canker of oilseed rape (OSR), exhibits gene-for-gene interactions with its host plant. The race structure of L. maculans was assessed on the basis of the analysis of 1011 isolates collected in France between 1990 and 2000, with regards to three AVR genes, AvrLm1, AvrLm2 and AvrLm4. The effect of selection pressure, due to large-scale cropping of Rlm1 cultivars, on the evolution of races of the fungus was also evaluated. The results revealed a scarcity or complete absence of isolates harbouring AvrLm2, whereas isolates harbouring AvrLm4 were present at a variable level, that was as high as 17.2–31.2% depending on the sample year and location. When obtained from rlm1 cultivars, isolates harbouring AvrLm1 always represented more than 83% of the populations until the 1997–1998 growing season. As a consequence, the Rlm1 cultivars had been highly efficient at controlling the disease and were grown on an estimated 43.7% of the total French acreage in OSR in 1998–1999. However, the increased commercial success of Rlm1 cultivars was paralleled by a decrease in the proportion of isolates harbouring AvrLm1 in 1997–1998 and 1998–1999. This resulted in less than 13% of isolates harbouring AvrLm1 in populations being collected from rlm1 cultivars in 1999 and 2000, and contributed to the loss of efficiency of the Rlm1 resistance in the field. The present study is an illustration of one round of a `boom and bust' cycle that occurred for a pathosystem where it has never been reported before. These data and the high evolutionary potential of L. maculans are fully supportive of one pathogen species with a high risk of breaking down resistance genes in OSR and suggest that the development of integrated strategies aiming at maximising the durability of novel resistance is now a priority for this pathosystem. [ABSTRACT FROM AUTHOR]

Leptosphaeria maculans (anamorph Phoma lingam), the ascomycete causing stem canker of crucifers, is a species complex that can be separated into at least seven distinct subgroups using a combination of biochemical and molecular criteria. In the present study sequences of the entire ITS region, including the 5.8S rDNA, of 38 isolates representing the seven subgroups, along with specimens from culture collections, were analysed, compared to those of closely related Leptosphaeria species, and the phylogeny inferred using parsimony and distance analyses. A well-supported clade encompassed all isolates of the seven subgroups along with L. conferta, a known saprobe of dried crucifer stems. The L. maculans isolates were further separated into two well-supported clades corresponding to L. maculans s. str. and the recently named L. biglobosa. Parsimony and distance analyses further separated groups within both species, usually corresponding to specific host plants or geographic origin, e.g. L. maculans ‘brassicae’ from cultivated Brassica, L. maculans ‘lepidii’, from Lepidium sp., L. biglobosa ‘brassicae’, from various Brassica species, L. biglobosa ‘thlaspii’ from Thlaspi arvense, L. biglobosa ‘erysimii’ from Erysimum sp., and L. biglobosa ‘canadensis’ mostly found in central Canada. The oldest L. maculans specimens maintained in international collections clustered with either L. maculans ‘brassicae’, L. biglobosa ‘brassicae’, or a still different group closely related to L. biglobosa ‘thlaspii’. The evolutionary relationships between the seven infraspecific groups are discussed in terms of phytopathological relevance and species isolation linked with specific life cycle, geographic isolation or host specificity. [ABSTRACT FROM AUTHOR]