Use of Field pathogenomics approach for Puccinia striiformis f. sp. tritici race identification and phylogenomic delineation in North India.

Stripe rust of wheat caused by Puccinia striiformis f. sp. tritici (Pst) is among the top 10 most important fungal phytopathogens in the world, threatening the global wheat production. Continuous pathogen evolution and air borne nature of the pathogen, increased the importance of pathogen population structure studies in case of wheat-Puccinia pathosystem. The current study aimed to characterize the Pst pathotype distribution in North India. A total of 61 stripe rust infected samples were collected from sub-mountainous areas of Punjab (50), Himachal Pradesh (HP) (8) and Jammu & Kashmir (J&K) (3) during 2021-24. Virulence profiling results of 58 samples using Indian stripe rust differentials revealed the prevalence of five pathotypes (238S119, 110S119, 46S119, 47S103 and 6S0) in North India. Among which 238S119 was the most predominant (51.72%) one followed by 110S119 (27.59%) and 46S119 (17.24%). No virulence was found against stripe rust resistant Yr5, Yr10, Yr15, Yr24 and YrSP genes indicating these genes are the most effective resistant genes in North India providing complete resistance till date. A total of 23 field infected samples were used for transcriptome analysis (RNA-seq) generating an average of 31.77 million raw reads. The clustered assembly generated mean assembly size of ∼ 45.10 Mb containing 7,587-2,49,571 contigs with 47.99% GC content, N50 value of 749.87 with 7.48-94.90% BUSCO score. Phylogenomic analysis of farmer's field samples revealed the existence of three different Pst lineages in North India i.e., Punjab lineage, Himachal lineage and Kashmir lineage, where Himachal lineage exhibited highest genetic diversity. A total of three (3) SSR and 14 KASP markers developed in previous studies were tested on farmer's field isolates for pathotype identification. Eight KASP markers showed reproducible polymorphism for pathotype specific detection. Correlating the KASP assay with results of virulence profiling revealed that these markers have the ability to detect the virulence change from five established pathotypes under field conditions. The findings of present study provided better understanding in Pst pathotype distribution in North India and pathotype detection using consistent polymorphic markers along with deciphering the behaviour of Pst under field conditions and deducing their evolutionary relationship in North India.