A sensitive method for examining whole-cell biochemical composition in single cells of filamentous fungi using synchrotron FTIR spectromicroscopy Konstantin Jilkine a , Kathleen M. Gough a , Robert Julian b , Susan G.W. Kaminskyj c, * a Department of Chemistry, University of Manitoba, Winnipeg, MB, Canada R3T 2N2 b Synchrotron Radiation Centre (SRC), University of Wisconsin at Madison, Stoughton, WI 53589, USA c Department of Biology, University of Saskatchewan, Saskatoon, SK, Canada S7N 5E2 Received 9 July 2007; received in revised form 7 October 2007; accepted 19 October 2007 Abstract Cell function is related to cell composition. The asexual state of filamentous fungi (molds and mildews) has two main life cycle stages: vegetative hyphae for substrate colonization and nutrient acquisition, and asexual spores for survival and dispersal. Hyphal composition changes over a few tens of microns during growth and maturation; spores are different from hyphae. Most biochemical analyses are restricted to studying a few components at high spatial resolution (e.g. histochemistry) or many compounds at low spatial resolution (e.g. GC–MS). Synchrotron FTIR spectromicroscopy can be used to study fungal cell biology by fingerprinting varieties of carbohydrates, proteins, and lipids at about 6 lm spatial resolution. FTIR can distinguish fungal species and changes during hyphal growth, and reveals that even fungi grown under optimal vs mildly stressed conditions exhibit dramatic biochemical changes without obvious morphological effects. Here we compare hypha and spore composition of two fungi, Neurospora and Rhizopus. There are clear biochemical changes when Neurospora hyphae commit to spore development, during spore maturation and following germination, many of which are consistent with results from molecular genetics, but have not been shown before at high spatial resolution. Rhizopus spores develop within a fluid-containing sporangium that becomes dry at maturity. Rhi- zopus spores had similar protein content and significantly more carbohydrate than the sporangial fluid, both of which are novel findings. Ó 2007 Elsevier Inc. All rights reserved. Keywords: Filamentous fungi; Neurospora; Rhizopus; Spores; Synchrotron FTIR spectromicroscopy 1. Introduction Fungi play key roles in the environment, having major positive impacts on terrestrial plant survival through mycor- rhizal symbioses, being essential for herbivore nutrition through assisting cellulose digestion, and being critical to recycling dead plant materials. Some species are used in ancient and modern biotechnology, and as model experi- mental systems. Other species, both as plant and human pathogens, are established or emerging threats. Fungi inter- act with their surroundings through their cell walls and by means of the compounds they secrete (particularly for nutri- ent acquisition) or store (particularly for defense or repro- duction). The fungal wall supports and protects the cytoplasm, maintains cell shape, and permits force genera- tion required for substrate penetration [1–3]. Cell function is intrinsically related to cell composition. The asexual state of filamentous fungi (molds and mildews) has two main life cycle stages: vegetative hyphae for sub- strate colonization and nutrient acquisition [1,4] and spores for survival and dispersal (e.g. [5]). The sexual state is tran- sient at best. The vegetative life cycle of a typical mold begins with spore germination followed by development 0162-0134/$ - see front matter Ó 2007 Elsevier Inc. All rights reserved. doi:10.1016/j.jinorgbio.2007.10.023 * Corresponding author. Tel.: +1 306 966 4422; fax: +1 306 966 4461. E-mail address: Susan.Kaminskyj@usask.ca (S.G.W. Kaminskyj). www.elsevier.com/locate/jinorgbio Available online at www.sciencedirect.com Journal of Inorganic Biochemistry xxx (2007) xxx–xxx JOURNAL OF Inorganic Biochemistry ARTICLE IN PRESS Please cite this article in press as: K. Jilkine et al., J. Inorg. Biochem. (2007), doi:10.1016/j.jinorgbio.2007.10.023