Among plant pathogens, the necrotrophic fungus Botrytis cinerea is one of the most prevalent, leading to severe crop damage. Studies related to its colonization of different plant species have reported variable host metabolic responses to infection. In tomato, high N availability leads to decreased susceptibility. Metabolic flux analysis can be used as an integrated method to better understand which metabolic adaptations lead to effective host defence and resistance. Here, we investigated the metabolic response of tomato infected by B. cinerea in symptomless stem tissues proximal to the lesions, with a reconstructed metabolic model constrained with a large and consistent metabolic dataset acquired under four different N supplies, throughout 7 days post inoculation (dpi). An overall comparison of 48 flux solution vectors of Botrytis- and mock-inoculated plants showed that fluxes were higher in Botrytis-inoculated plants, and the difference increased with a reduction in available N, accompanying an unexpected increase in radial growth. Despite higher fluxes, such as those involved in cell wall synthesis and other pathways, fluxes related to glycolysis, the TCA cycle, amino acids and protein synthesis were limited under very low N, which might explain the enhanced susceptibility. Limiting starch synthesis and enhancing fluxes towards redox and specialized metabolism also contributed to defence independent of N supply.Read less <