Identifying genetic diversity of avirulence genes in Leptosphaeria maculans using whole genome sequencing.

ABSTRACT

Next generation sequencing technology allows rapid re-sequencing of individuals, as well as the discovery of single nucleotide polymorphisms (SNPs), for genomic diversity and evolutionary analyses. By sequencing two isolates of the fungal plant pathogen Leptosphaeria maculans, the causal agent of blackleg disease in Brassica crops, we have generated a resource of over 76 million sequence reads aligned to the reference genome. We identified over 21,000 SNPs with an overall SNP frequency of one SNP every 2,065 bp. Sequence validation of a selection of these SNPs in additional isolates collected throughout Australia indicates a high degree of polymorphism in the Australian population. In preliminary phylogenetic analysis, isolates from Western Australia clustered together and those collected from Brassica juncea stubble were identical. These SNPs provide a novel marker resource to study the genetic diversity of this pathogen. We demonstrate that re-sequencing provides a method of validating previously characterised SNPs and analysing differences in important genes, such as the disease related avirulence genes of L. maculans. Understanding the genetic characteristics of this devastating pathogen is vital in developing long-term solutions to managing blackleg disease in Brassica crops.