Application of raman spectroscopy and mass spectrometry to study growth and interaction processes of the white-rot fungus schizophyllum commune

Menezes, Riya Christina

S. commune is a wood-decay fungus that causes white-rot and is ubiquitously found. Its abundance can not only be attributed to its prolific outbreeding (>23,000 mating types), but also to its superior competition ability. This thesis, therefore, focuses mainly on studying secondary metabolites produced by S. commune upon interaction with various white-rot fungi, and nuclear migration-a life cycle step. It also showcases various ways in which, established techniques like Raman microscopy and mass spectrometry can be effectively employed in microbiology. Firstly, aspects of nuclear migration, and chromophores present in S. commune were investigated using live-cell imaging with Raman microspectroscopy and coherent anti-Stokes Raman scattering (CARS), and fluorescent microscopy. Nuclear migration is required for dikaryotization, which is necessary for mushroom formation. Nuclei could not be located in this study, as Raman microscopy has limited use in live specimens, and CARS microscopy lacked the contrast to image molecules beyond lipids. The second study comprised of the identification of secondary metabolites produced by S. commune during interactions with wood-decay fungi and investigation of their role. S. commune being a wood-rotting fungus encounters other competitor microorganisms competing for space and nutrient sources in the forest ecosystem. We developed a fast, less-invasive way to identify, quantify and image metabolites of S. commune interacting with competitor species, using the pigment indigo as an example. This combination of Raman microscopy and Liquid Extraction Surface Analysis coupled with high-resolution mass spectrometry opens new avenues to investigate fungal and bacterial metabolites directly from the sample plates without extensive preparation. Finally, the role of an auxin in the fungus, its biosynthesis and significance was investigated. It was shown that wild type S. commune strains constitutively produced indole-3-acetic acid.

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Menezes, Riya Christina: Application of raman spectroscopy and mass spectrometry to study growth and interaction processes of the white-rot fungus schizophyllum commune. 2015.

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