Gene expression profiling of pathogenicity factors in Puccinia triticina race 162-A

Abstract

Puccinia triticina, the causal agent of brown rust, is a significant pathogen of wheat (Triticum aestivum L.) responsible for substantial yield reductions globally. This study focused on the molecular characterization and pathogenicity analysis of P. triticina Race 162-A, which has emerged as the predominant and rapidly evolving race in northern India. Uredospores of Race 162-A were obtained from the Regional Station of the Indian Institute of Wheat and Barley Research, Flowerdale, Shimla, Himachal Pradesh, for subsequent investigation. A 334 base pair fragment of the cytochrome oxidase subunit I (COX-1) gene was amplified from the genomic DNA of Race 162-A using polymerase chain reaction (PCR). The amplicon was subsequently cloned into the pTZ57R/T vector and subjected to Sanger sequencing. Comparative sequence analysis of the obtained sequence (GenBank Accession No. MN718664) revealed no nucleotide polymorphisms in Race 162-A relative to other P. triticina races. Phylogenetic analysis revealed four distinct clades, designated as Clades A, B, C, and D. Clade A comprised races of Puccinia triticina and exhibited close evolutionary relatedness to Clade B, which contained P. graminis f. sp. tritici races. In contrast, Clades C and D were phylogenetically divergent from Clades A and B, encompassing P. striiformis and P. coronata races, respectively. The temporal expression profiles of four genes—PtMAPK1, PtCYC1, PtCNB, and PtRTP—were analyzed at various time points post-inoculation in both a moderately resistant wheat cultivar (HD 2967) and a susceptible cultivar (Agra Local). The observed expression patterns of these genes suggested their potential role in pathogen establishment within the host. In the susceptible cultivar, the expression levels of all four genes exhibited a significant upregulation at 4 days after inoculation (DAI), corresponding with the pathogen's infection establishment phase. Contrastingly, in the moderately resistant cultivar (HD 2967), elevated expression of these genes persisted until 10 DAI. Protein-protein interaction (PPI) network analysis clarified the functional roles of these genes in wheat resistance. The analysis revealed important interactions involved in stress signalling, protein folding, and calcium signalling pathways.