Biodiscovery of Potential Antibacterial Diagnostic Metabolites from the Endolichenic Fungus Xylaria venustula Using LC–MS-Based Metabolomics

SIMPLE SUMMARY: In this study, we determined the bioactivities and chemical natures of three species of lichen Usnea and their associated endolichenic fungi (ELF) through metabolomics. We found significant differences in the antibacterial activities and the metabolites produced by the host lichen and its ELF, with the latter targeting a wider scope of organisms. We also discovered potential key metabolites produced by ELF that are yet to be reported. This study shows the application of metabolomics in rapidly identifying bioactive metabolites that are of significance in the discovery of new drugs. ABSTRACT: Three species of the lichen Usnea (U. baileyi (Stirt.) Zahlbr., U. bismolliuscula Zahlbr. and U. pectinata Stirt.) and nine associated endolichenic fungi (ELF) were evaluated using a metabolomics approach. All investigated lichen crude extracts afforded antibacterial activity against Staphylococcus aureus (minimum inhibitory concentration (MIC): 0.0625 mg/mL), but none was observed against Escherichia coli, while the ELF extract Xylaria venustula was found to be the most active against S. aureus (MIC: 2.5 mg/mL) and E. coli (MIC: 5 mg/mL). X. venustula was fractionated and tested for to determine its antibacterial activity. Fractions XvFr1 to 5 displayed bioactivities against both test bacteria. Selected crude extracts and fractions were subjected to metabolomics analyses using high-resolution LC–MS. Multivariate analyses showed the presence of five secondary metabolites unique to bioactive fractions XvFr1 to 3, which were identified as responsible for the antibacterial activity of X. venustula. The p-values of these metabolites were at the margin of significance level, with methyl xylariate C (P_60) being the most significant. However, their high variable importance of projection (VIP) scores (>5) suggest these metabolites are potential diagnostic metabolites for X. venustula for “dual” bioactivity against S. aureus and E. coli. The statistical models also showed the distinctiveness of metabolites produced by lichens and ELF, thus supporting our hypotheses of ELF functionality similar to plant endophytes.