3.4 | Metabolomic comparison of fenced and unfencedL. benzoin plants
We identified 1,225 metabolite features in all L. benzoin samples by conducting untargeted metabolomics analysis. PCA did not indicate clear treatment- or species-based subgrouping of the samples (Figure 5A ). Hence, we followed a supervised approach (PLS-DA and HCA) to probe for metabolites that varied among the samples based on treatment (fence vs no fence) (Figure 5B , D ). Both methods (PLS-DA and HCA) identified similar significant metabolite features with similar patterns of accumulation between the fenced and unfenced plots (Figure 5C, D ).
We used the metabolite features that we identified to obtain putative prediction of the chemical identities of the 1,225 metabolite features and the metabolic pathways that they are assigned to using the GSEA functional analysis tool. Our analysis identified glyoxylate and dicarboxylate metabolism (P = 0.01) and pentose phosphate pathway (P = 0.02) as significantly overrepresented pathways (Appendix Table 4 ). The glyoxylate and dicarboxylate pathway is not involved in plant defense directly; however, the possible role of the pentose phosphate pathway in pathogen defense has been shown in Arabidopsis thaliana . .