Comparative genomics identifies genetic markers linked to structural variations that differentiate Puccinia graminis tritici and secalis formae speciales

Stem rust is a serious disease of many gramineous plants including small grain staples such as wheat, barley, rye and triticale. Separate formae speciales (ff. spp.) of the causal fungus, Puccinia graminis, predominantly infect certain host plant genera. However, despite these taxonomic subdivisions, many P. graminis ff. spp. are genetically too similar to distinguish using existing genetic markers. For those infecting cereals, this is particularly challenging for P. graminis f. sp. tritici (Pgt) and P. graminis f. sp. secalis (Pgs). Herein we harnessed newly available genomic data for 39 Pgt and Pgs isolates and identified four regions of structural variation that were used to design four simple PCR‐based assays to distinguish the aforementioned formae speciales. These genomic regions display large presence/absence variation between Pgt and Pgs isolates, and yet a high degree of sequence conservation within shared neighbouring regions, facilitating primer design. We also confirmed lack of amplification in host plant genera analysed, which included assessment of the shared alternate host of Pgt and Pgs, Berberis vulgaris. Accurate classification of P. graminis ff. spp. infections on B. vulgaris is exceptionally valuable to rapidly define plants harbouring P. graminis inoculum when adjacent to cereal crops. Finally, we demonstrated utility of these four genetic markers to correctly distinguish a genetically diverse array of Pgt and Pgs isolates. This strategy could now be readily applied to other pathogens of interest, which will be of increasing value as genomic resources continue to rapidly expand for many key biotic threats to agricultural productivity.