Root rot of citrus trees caused by Phytophthora nicotianae is responsible for severe economic losses in citriculture. Use of
resistant rootstocks is an effective method of managing this problem, however, breeding and selection of new citrus rootstocks is
a time-consuming undertaking. The objective was to develop a method for the rapid assessment of rootstocks for P. nicotianae
tolerance, using a metabolomics approach to identify metabolic markers for the phenotypic trait of tolerance. Sixteen citrus
rootstocks were inoculated with P. nicotianae in the greenhouse for determination of relative tolerance/susceptibility. Healthy
citrus roots from four tolerant and four susceptible rootstocks were used for metabolite analysis with the objective of identifying
potential biomarkers. Organic solvent extractions of the roots were prepared and analysed by mass-spectrometry based liquid
chromatography, which produced 367 ion features (retention time and m/z). Orthogonal partial least squares discriminant
analysis of peak abundance using MarkerLynx software allowed for the identification of ion features that differentiate tolerant
and susceptible rootstocks. Using descriptive and inferential statistics based on the ion features of uninoculated tolerant vs.
susceptible rootstocks, applying logistic regression, 14 top markers were identified and two of them (22.03_259.0975 and
22.21_313.1445: retention time (rt) and mass to charge ratio (m/z) were accepted as potential metabolic markers. A model that
can potentially predict tolerance in citrus rootstocks with >98% accuracy is presented.