Development of a diagnostic assay for differentiation between genetic groups in clades I, II, III and IV of Puccinia graminis f. sp. tritici

Authors: Szabo, Les; OLIVERA, PABLO - University Of Minnesota; WANYERA, RUTH - Kenya Agricultural And Livestock Research Organization; VISSER, BOTMA - University Of The Free State; Jin, Yue

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/18/2022
Publication Date: 8/1/2022
Citation: Szabo, L.J., Olivera, P.D., Wanyera, R., Visser, B., Jin, Y. 2022. Development of a diagnostic assay for differentiation between genetic groups in clades I, II, III and IV of Puccinia graminis f. sp. tritici. Plant Disease. 106(8):2211-2220. https://doi.org/10.1094/PDIS-10-21-2161-RE.
DOI: https://doi.org/10.1094/PDIS-10-21-2161-RE

Interpretive Summary: Wheat stem rust, caused by Puccinia graminis f. sp. tritici, has re-emerged as a serious disease. New variants (races) of the wheat stem fungal pathogen have appeared in Africa, Europe and the Asia that overcome host resistance genes that are critical to protecting U.S. wheat crop. Monitoring the global spread of these new variants is an important component of the management of this disease. The standard method of identifying race variants of the wheat stem pathogen uses a bioassay with 20 wheat lines with different stem rust resistance genes The bioassay takes several weeks to over a month to complete and requires living samples. We analyzed a diverse set of pathogen isolates and developed a rapid molecular assay based on 17 DNA markers to place the isolates into 12 genetic groups. The new assay was 100% accurate in identifying strains belonging to these 12 genetic groups and had no false positives in a test set of 94 isolates. The assay has built in redundancy so that minor genetic changes or errors will not affect the accuracy of the results. Genetic groups of wheat stem rust pathogen found in the U.S. are distinct from these new variants and are easily differentiated by this assay. A test set of samples from Africa demonstrated the functionality of this assay with non-living samples. This research will be used by scientists involved in global surveillance and pathogen population studies and for diagnosticians involved in monitoring plant pathogens.

Technical Abstract: Wheat stem rust has re-emerged as a serious disease caused by new variants of Puccinia graminis f. sp. tritici. Variants with significant virulences and broad geographical distribution include Ug99 race group, race TTRTF, and TKTTF race group. Genetic analysis has placed isolates representing these critical new virulent races into 12 genetic groups that make up clades I to IV. Development of molecular diagnostic assay for these 12 genetic groups will be an important component of global surveillance efforts. A single-nucleotide polymorphism database was mined for candidate markers that would differentiate between these 12 genetic groups. Thirty-five candidate markers were screened and a core set of 17 markers were tested against a set of 94 isolates representing a broad range of genotypic and race phenotypic range. This core set of markers were 100% accurate in identifying the 12 genetic groups for 52 clade I to IV isolates and no false positives were observed with no-target isolates. The assay has built in redundancy so that minor genetic changes, or errors in genotyping calling will not affect the accuracy of the results. This assay is also affective in identifying the genetic groups in clade V from Germany and Georgia, the three main subgroups in North American clade VI, and the clade VII consisting of TTTTF race found in North and South America. This assay provides a rapid diagnostic tool for both living and non-living samples for detection the critical new race or race groups of P. graminis f. sp. tritici.