Leaf Endophytes Relationship with Host Metabolome Expression in Tropical Gymnosperms

Plant-microbe interactions play a pivotal role in shaping host fitness, especially concerning chemical defense mechanisms. In cycads, establishing direct correlations between specific endophytic microbes and the synthesis of highly toxic defensive phytochemicals has been challenging. Our research delves into the intricate relationship between plant-microbe associations and the variation of secondary metabolite production in two closely related Zamia species that grow in distinct habitats; terrestrial and epiphytic. Employing an integrated approach, we combined microbial metabarcoding, which characterize the leaf endophytic bacterial and fungal communities, with untargeted metabolomics to test if the relative abundances of specific microbial taxa in these two Zamia species were associated with different metabolome profiles. The two species studied shared approximately 90% of the metabolites spanning diverse biosynthetic pathways: alkaloids, amino acids, carbohydrates, fatty acids, polyketides, shikimates, phenylpropanoids, and terpenoids. Co-occurrence networks revealed positive associations among metabolites from different pathways, underscoring the complexity of their interactions. Our integrated analysis demonstrated to some degree that the intraspecific variation in metabolome profiles of the two host species was associated with the abundance of bacterial orders Acidobacteriales and Frankiales, as well as the fungal endophytes belonging to the orders Chaetothyriales, Glomerellales, Heliotiales, Hypocreales, and Sordariales. We further associate individual metabolic similarity with four specific fungal endophyte members of the core microbiota, but no specific bacterial taxa associations were identified. This study represents a pioneering investigation to characterize leaf endophytes and their association with metabolomes in tropical gymnosperms, laying the groundwork for deeper inquiries into this complex domain.