Unlocking the Compositional and Isotopic Complexity of the Hydrothermal Lipidome: High-Resolution Insights from the Guaymas Basin

Hydrothermal environments are extreme ecosystems characterized by steep thermal gradients, dynamic redox conditions, and energy-rich fluid flow that foster unique microbial communities and biogeochemical processes. Despite decades of research, hydrothermal sediments, such as those found within the Guaymas Basin, continue to yield novel discoveries and serve as model systems for studying carbon cycling and microbial lipid adaptation. My project explores the hydrothermal lipidome of the Guaymas Basin by integrating three complementary techniques. First, we introduce an isotopologue-based approach for compound-specific isotope analysis (CSIA) for the detection of methanotrophic isotopic signatures that go beyond the limitations of conventional gas-chromatography - isotope ratio mass spectrometry (GC-IRMS). To unlock its full potential, we combine this approach with feature-based molecular networking (FBMN), which provides comprehensive compositional insight into the hydrothermal lipidome, allowing the identification of distinct molecular signatures and their environmental controls and discerning a potential methanotrophic origin. To expand on the limited spatial resolution of traditional lipidomics, we employed matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) for micrometer-scale mapping of biomolecules on the undisturbed sediment core, revealing millimeter-scale shifts in lipid distributions tied to the intense redox stratification. Overall, this multidimensional approach uncovers the molecular and isotopic complexity of the Guaymas Basin sediments, governed by steep thermal and geochemical gradients.