Fungal decomposition dynamics using Fourier transform infrared spectroscopy and atomic force microscopy

I studied the decomposition of leaf material at the hyphal scale of resolution and the chemical and physical changes occurring at the fungal/substrate interface using Fourier transform infrared (FTIR) spectroscopy and atomic force microscopy (AFM). The chemical composition of 20 fungal isolates was determined using FTIR and FTIR-attenuated total reflectance (FTIR-ATR) microscopy, with the intention of subtracting fungal FTIR spectra from decomposing leaf spectra in a separate experiment. I found fungi are difficult to differentiate using FTIR spectra. I performed a twelve month decomposition study placing leaves of white oak, black huckleberry, and pitch pine in leaf litter of the New Jersey pinelands. FTIR-ATR spectra of the leaves at 10x10 micron areas were taken at 0, 6, and 12 months. I found leaf chemistry of oak and huckleberry was similar, but different from pine, prior to decomposition. As decomposition progressed, the chemistry of the leaves appeared to become more similar. Subtracting fungal spectra from the leaf spectra was not possible and complicated analysis of leaf spectra. AFM was combined with FTIR-ATR to study the fungal hypha/leaf interface at the level of an individual hypha. It was not possible to scan a leaf on the AFM, so a flatter substrate was required. I attempted to cast starch:lignin biofilms as a simple leaf model using published methods, however it was found that during the film making process the lignin may have been degraded. Instead, manufactured cellophane was used as a model for cellulose. Three fungal species were grown on cellophane squares, and AFM and FTIR-ATR imaging utilized to determine the chemical and physical properties of the cellophane adjacent to the hyphae. Fungal species capable of producing cellulase caused a change in the physical characteristics and chemistry of cellophane adjacent to fungal hyphae. These experiments demonstrate the chemistry of fungal hyphae and decomposing leaves using FTIR-ATR and chemical and physical changes occurring in a substrate during fungal decomposition using FTIR-ATR and AFM. The novel use of FTIR-ATR and AFM to investigate the fungus/substrate interface at the scale of an individual fungal hypha introduces new methods for studying fungi at this scale of resolution.