Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSproblem is that, although the effectiveness of P. gigantea as a bio-control agent has empirically been shown, the long term biological effect of this funguson conifer trees as well as on other soil micro-flora has not been empirically proven. We investigated the impact of P. gigantea treatment on stumpmycobiota using metagenomic pyrosequencing approach as this has not been done before. Samples from forest sites pre-treated with P. gigantea for 1, 6and 13 years ago were collected, DNA was isolated and pyrosequenced. Similarly samples were also collected from untreated stumps within the sameforest sites. Sequences were quality trimmed using Mothur software. After trimming we had 26 398 sequences from 53 117. For the extraction of the fullITS1 of the nuclear ITS region FungalITSextractor was used and these sequences were clustered at 97% similarity using cdhit-454 with the most abundantsequence types serving as cluster seeds. The most frequent sequence type in each cluster was used for the BLAST searches against NCBI BLASTN and ³ 97%similarity across the entire length of the pairwise alignment was taken to indicate conspecificity. Differences between control and treated stumps weretested statistically with Paired-Sample T-test (SPSS 19). Also diversity indexes and similarity indexes between controls and treated were calculated usingEstimates 8.2.0. After one year of the clear-cut we found from Phlebiopsis gigantea-treated stumps 107 different fungal OTUs and from non-treatedstumps 119 fungal OTUs, from which they shared 102 OTUs. After 6 years we observed from treated stumps 118 fungal OTUs and from non-treated 134fungal OTUs and they shared 99 OTUs. After 13 years we found from treated stumps 131 OTUs and from non-treated 139 OTUs and shared OTU numberwere 109. However there were no statistical differences between control and treatment. Based on our results our primary conclusion is that stumptreatment should continue as there is no obvious adverse effect on the other stump mycobiota.703. Diversity of yeast and mold species in different cheeses. Nabaraj Banjara 1 , Kenneth Nickerson 2 , Heather Hallen-Adams 1 . 1) Food Science andTechnology, University of Nebraska-Lincoln, Lincoln, NE; 2) Biological Sciences, University of Nebraska-Lincoln, Lincoln, NE.The yeast and mold diversity from different commercial cheeses collected from local markets (Lincoln, NE, USA) was studied using microbial counting andmolecular biology approaches. Twenty one distinct types of cheese samples were investigated. Briefly, 10 grams of each cheese sample was homogenizedin distilled water, serially diluted from 10 to 10 -6 , grown in Yeast Extract Glucose Chloramphenicol Agar (YGC) and the population was counted fromdilution plates. Yeasts and molds were identified by amplification and sequence analysis of the nuclear ribosomal RNA genes, using ITSIF and TW13primers. Debaryomyces hansenii was the predominant fungal species in most of the cheeses (59% of samples at up to 5.7 x 10 6 CFU/ gram). Other fungiisolated included D. fabryi, D. prosopidis, D. subglobosus, Penicillium roqueforti and Candida sake. In our samples, farmstead cheese had the highest (2.1 x10 8 CFU/gram) and Swiss cheese the lowest (8x10 3 CFU/gram) fungal population.704. Heterologous expression and characterization of soil organic matter-specific proteases secreted by the ectomycorrhizal fungus Paxillus involutus.Morten N. Grell 1 , Linas Pupelis 1 , Tomas Johansson 2 , Firoz Shah 2 , Lene Lange 1 , Anders Tunlid 2 . 1) Section for Sustainable Biotechnology, Department ofBiotechnology, Chemistry and Environmental Engineering, Aalborg University Copenhagen, Denmark; 2) Microbial Ecology Group, Department of Biology,Lund University, Sweden.Paxillus involutus (Batsch) Fr. (Basidiomycetes; Boletales) is widely distributed in the Northern hemisphere, and is one of the best-studiedectomycorrhizae fungi, especially with respect to its ecology and physiology. In a study on the mechanisms by which Paxillus involutus degrade complexorganic matter extracted from plant litter material, transcriptomes were sequenced using the 454 technology and NimbleGen microarrays produced. Anumber of genes encoding extracellular enzymes showed an increased transcript level during degradation of soil organic matter (SOM), as compared withgrowth on a defined medium (MNM). They were suggested to constitute a Fenton-like, radical-based biodegradation system that disrupts the organicmatter-protein complexes thereby mobilizing embedded nitrogen (Rineau et al. 2012, Environm. Microbiol. 14, 1477-1487). Supporting this, a number ofprotease genes were found to have a significantly increased SOM/MNM expression value and to be upregulated during growth on protein-rich substrates.Most highly expressed were aspartic, metallo, and serine proteases. This is in agreement with biochemical analysis of SOM degradation. To study substratespecificity and regulation of selected proteases in detail these are expressed in the yeast Pichia pastoris.705. Effector proteins in fungal defense against fungivorous nematodes: Targets and functional significance. Therese Wohlschlager 1 , StefanieSchmieder 1 , Alex Butschi 2 , Paola Grassi 3 , Alexander Titz 4 , Stuart Haslam 3 , Michael Hengartner 2 , Markus Aebi 1 , Markus Künzler 1 . 1) Institute of Microbiology,ETH Zürich, Switzerland; 2) Institute of Molecular Life Sciences, University of Zürich, Switzerland; 3) Division of Molecular Biosciences, Imperial College,London, United Kingdom; 4) Department of Chemistry, University of Konstanz, Germany.The defense of fungi against fungivores is largely based on the production of intracellular toxins. A significant proportion of these toxins are peptides andproteins that are synthesized by the ribosome and stored in the cytoplasm. Protein toxins include lectins that target specific glycoepitopes in the intestineof the fungivore upon ingestion and kill the fungivore by a yet unknown mechanism. In our laboratory, we focus on the functional characterization offungal protein toxins that are directed against nematodes. We use the model nematode Caenorhabditis elegans to identify the targets and to study thetoxicity mechanism of these fungal defense effector proteins in the nematode. In addition, we employ the fungivorous nematodes Aphelenchus avenaeand Bursaphelenchus willibaldi to study the diversity, the functional significance and the transcriptional regulation of these proteins in the fungus.Recently, we identified a nematotoxic lectin from the mushroom Laccaria bicolor that is homologous to animal lectins involved in innate immunity againstbacteria. We found that the nematotoxicity of the lectin is based on its specific binding to methylated fucose residues on nematode N-glycans. Amonganimals, this epitope is only present in worms and molluscs but not in insects or vertebrates. We performed affinity chromatography of C. elegans wholeworm protein extracts using the L. bicolor lectin and other nematotoxic fungal lectins recognizing protein-bound glycans. The results of this analysissuggest that these lectins target the same set of glycoproteins in the nematode intestine and may confer toxicity by a common mechanism. In order toaddress the functional significance of these proteins for fungal defense against fungivorous nematodes, we expressed some of the fungal proteinsdisplaying toxicity towards C. elegans, in the filamentous ascomycete Ashbya gossypii. These transformants were fed to A. avenae and the propagation ofthe fungivorous nematode on the various transformants was determined. Expression of some effector proteins significantly inhibited propagation of thenematode suggesting that these proteins have a role in fungal defense against these organisms. Experiments addressing the relative fitness of the variousA. gossypii transformants upon selective pressure of feeding by A. avenae are under way.706. Microfluidic platforms for monitoring interactions between fungi and bacteria. Martina A. Stöckli 1 , Claire E. Stanley 2 , Pauli T. Kallio 1 , MarkusKünzler 1 , Andrew J. deMello 2 , Markus Aebi 1 . 1) Microbiology and Immunology, ETH Zurich, Zurich, Switzerland; 2) Chemistry and Applied Biosciences, ETHZurich, Zurich, Switzerland.Bacteria and fungi share many microhabitats where they interact with each other. These interactions are diverse and play important roles in certainhuman infections, as well as in the biological control of plant diseases. The basis of these interactions, however, is not well understood, as theirinvestigation is technically challenging. Conventional microscopic approaches are limited, in that they can only monitor interactions at a particular time294