Nitrogen availability might play an essential role in plant diseases by enhancing fungal cell growth and influencing the expression of genes required for successful pathogenesis. Nitrogen availability could modulate secondary metabolic pathways as evidenced by the significant differential expression of several core genes involved in mycotoxin biosynthesis and genes encoding polyketide synthase/nonribosomal peptide synthetases, cytochrome P450 and carbohydrate-active enzymes inFusarium sacchari, grown on different nitrogen sources. A combined analysis was carried out on the transcript and metabolite profiles of regulatory metabolic processes and the virulence ofFusarium saccharigrown on various nitrogen sources. The nitrogen regulation of the gibberellin gene cluster included the metabolic flux and multiple steps of gibberellin synthesis. UHPLC-MS/MS-based metabolome analysis revealed the coordination of these related transcripts and the accumulation of gibberellin metabolites. This integrated analysis allowed us to uncover additional information for a more comprehensive understanding of biological events relevant to fungal secondary metabolic regulation in response to nitrogen availability.