Analysis of differential metabolites and metabolic pathways of mono- and inter-cropped wheat in response to Blumeria graminis f. sp. tritici infection

Wheat and faba bean intercropping can alleviate the occurrence and severity of wheat powdery mildew. However, the physiological mechanism of intercropping improving wheat disease resistance remains unclear. This study aimed to understand the metabolic differences between mono- and inter-cropped wheat in response to Blumeria graminis f. sp. tritici infection and reveal the physiological mechanism of intercropping for improving wheat resistance to powdery mildew. In this study, the following three nitrogen (N) application levels were established: 75 mg·kg⁻¹ (N1), 150 mg·kg⁻¹ (N2), and 225 mg·kg⁻¹ (N3). Following inoculation with B. graminis f. sp. tritici, the occurrence of powdery mildew in mono- and inter-cropped wheat was investigated, and the metabolomics of mono- and inter-cropped wheat in response to infection with B. graminis f. sp. tritici were analyzed by UPLC-MS/MS, using a widely targeted metabolomic method. The results showed that N levels and N levels × planting patterns significantly affected the incidence and disease indices of powdery mildew in wheat. Under all three N levels, wheat intercropping with faba bean reduced the incidence of wheat powdery mildew by 25.54%–38.81% and decreased the disease index by 20.11%–21.97% relative to mono-cropped wheat (MW), and the intercropping control effect under the N1 level was better than that under N2 and N3 conditions. A total of 822 differential metabolites were detected in the mono- and inter-cropped wheat leaves, of which 69, 52, and 88 were detected at the N1, N2, and N3 levels, respectively. Intercropping of wheat and faba bean regulated flavonoids, alkaloids, amino acids and derivatives, and phenolic acids in wheat leaves compared to MW. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of differential metabolites showed that they were mainly enriched in the biosynthesis of amino acids, metabolic pathways, and secondary metabolites. Among them, metabolites with significant differences were enriched in metabolic pathways at the N1 and N2 levels, and metabolites with significant differences were enriched in amino acid biosynthesis under N stress conditions (N1 and N3). Further analysis of the metabolites from the top 10 up- and down-regulated genes revealed that intercropping upregulated glutathione (G-SH), L-tryptophan, L-asparagine, and L-glutamine in wheat leaves at the N1 level relative to MW, and upregulated L-asparagine, L-homomethionine, and L-tryptophan in intercropped wheat leaves at the N3 level relative to MW. In addition, a few metabolites, including alkaloids, phenolic acids, and organic acids, in wheat leaves were regulated by intercropping compared with MW under the N1 and N3 levels. In conclusion, the response of wheat to powdery mildew infection was regulated by N levels. Metabolites involving amino acids and derivatives, alkaloids, phenolic acids, and organic acids in wheat leaves are regulated by intercropping during B. graminis f. sp. tritici infection and induce different physiological reactions, possibly being one of the mechanisms by which intercropping improves wheat powdery mildew resistance. Intercrop-regulated amino acids and their derivatives under N stress are closely associated with wheat powdery mildew resistance. The present study identified the different responses of mono- and inter-cropped wheat to disease infection via metabolic analysis, facilitating a comprehensive understanding of crop diversity for the management of pests and diseases.