Mechanisms of induced susceptibility to Diplodia tip blight in drought-stressed Austrian pine

Plants experiencing drought stress are frequently more susceptible to pathogens, likely via alterations in physiology that create favorable conditions for pathogens. Common plant responses to drought include the production of reactive oxygen species (ROS) and the accumulation of free amino acids (AAs), particularly proline. These same phenomena also frequently occur during pathogenic attack. Therefore, drought-induced perturbations in AA and ROS metabolism could potentially contribute to the observed enhanced susceptibility. Furthermore, nitrogen (N) availability can influence AA accumulation and affect plant resistance, but its contributions to drought-induced susceptibility are largely unexplored. Here we show that drought induces accumulation of hydrogen peroxide (H2O2) in Austrian pine (Pinus nigra Arnold) shoots, but that shoot infection by the blight and canker pathogen Diplodia sapinea (Fr.) Fuckel leads to large reductions in H2O2 levels in droughted plants. In in vitro assays, H2O2 was toxic to D. sapinea, and the fungus responded to this oxidative stress by increasing catalase and peroxidase activities, resulting in substantial H2O2 degradation. Proline increased in response to drought and infection when examined independently, but unlike all other AAs, proline further increased in infected shoots of droughted trees. In the same tissues, the proline precursor, glutamate, decreased significantly. Proline was found to protect D. sapinea from H2O2 damage, while also serving as a preferred N source in vitro. Fertilization increased constitutive and drought-induced levels of some AAs, but did not affect plant resistance. A new model integrating interactions of proline and H2O2 metabolism with drought and fungal infection of plants is proposed.