Podium Presentation in Room 4 on Thursday at 9:20 (Chair: Chris Cox)
Authors: Madison McMinn (1,2), Sankha S. Basu (3), Begona Gimenez-Cassina Lopez (1), Michael S. Regan (1), Elizabeth C. Randall (4), Amanda R. Clark (1), Christopher Cox (5), Gary R. Kinsel (6), Mary Kinsel (6), Jeffrey N. Agar (2) and Nathalie Y.R. Agar (1,4,7).
Rapid detection of pathogens in clinical tissue specimens remains a critical diagnostic need. Culture-based approaches cannot be performed rapidly, and histological-based approaches lack the specificity needed for confident pathogen identification. Although nucleic acid-based approaches provide increased sensitivity and specificity, technical complexity and high costs coupled with time-consuming steps limit their application. Over the past decade, Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI TOF MS) has emerged as a powerful diagnostic tool for identification of bacterial isolates. A recently described variation on MALDI, termed metal oxide laser ionization (MOLI), involves using a metal oxide as the matrix. Unlike other metal oxides, cerium(IV) oxide (CeO2) demonstrates a unique property of laser induced catalytic side chain cleavage of fatty acids. MOLI MS has also shown promise as a tool for bacterial identification by leveraging lipidomic differences between bacterial strains, but has been limited to target plate analysis. In this study, we further characterize the catalytic mechanism for CeO2 mediated fatty acid catalysis and demonstrate its potential as a tool for pathogen detection in tissue specimens using MOLI MSI.
The catalytic properties of CeO2-mediated MOLI MS were investigated using target plate analysis using lipid standards to determine how variations in lipid structure affect CeO2-mediated cleavage. These methods were further applied to reference bacterial strains to build on previously demonstrated MOLI MS techniques. We developed and optimized a method to perform MOLI MSI using multiple mass spectrometers on normal and intracranial tumor mouse brains. To reduce turnaround time and increase clinical applicability, we tested and optimized instrument parameters and multiple sample preparation techniques. To assess the potential of MOLI MSI to detect pathogens directly from tissue, we utilized a pseudo-infection model by spotting bacterial lipid extracts on mouse brain tissue sections and imaged by MOLI MSI.
CeO2 induced catalysis was applied to bacterial strains to generate fatty acid signatures. MOLI MSI applied to normal mouse brain revealed differentiable fatty acyl pools in myelinated and non-myelinated regions and distinct fatty acyl signatures in tumor regions of a glioblastoma PDX mouse model. In the pseudo-infection model, the spotted regions were molecularly resolved from the supporting mouse brain tissue by the diagnostic odd-chained fatty acids and reflected control bacterial MOLI MS signatures. Both spotting of CeO2 and slide-precoating allowed for rapid preparation.
We demonstrated the application of CeO2 as a catalyst for MSI and the potential for MOLI MSI as a tool for direct identification of pathogens from infected tissues.
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