Program Book - 27th Fungal Genetics Conference

FULL POSTER SESSION ABSTRACTSPia Laine 2 , Lars Paulin 2 , Taina Lundell 1 . 1) Department of Food and Environmental Sciences, Division of Microbiology, Fungal Biotechnology Laboratory; 2)Institute of Biotechnology, DNA Sequencing and Genomics Laboratory, Viikki Campus, University of Helsinki, FINLAND.The efficient wood-degrading white-rot basidiomycete Phlebia radiata Fr. is able to degrade all the main components of lignocellulose and secretes arepertoire of lignin-converting peroxidases and oxidases as well as carbohydrate-acting enzymes. P. radiata is the second species of the genus Phlebia tobe genome sequenced, thus giving more insight in to the phlebioid clade of the order Polyporales, class Agaricomycetes, and comparative data forfunctional analyses on white-rot fungal mechanisms of wood decay and decomposition of lignin. We carried out transcriptome sequencing and secretomeanalyses on the Finnish isolate 79. Molecular systematics of the genus Phlebia was inferred by perfoming a four-gene study on North-European Phlebiaspp. isolates including 10 species. Ribosomal RNA-encoding (SSU 18S rDNA; ITS1-5.8S-ITS2; ITS2-28S) regions and two protein-coding genes (gapdh, rpb2)were partially PCR-amplified with fungal or basidiomycete-specific primer pairs. Phylogenetic sequence analyses resulted with no single taxonomic clusterof Phlebia. The genus is obviously polyphyletic, and the various species were scattered together with other genera of Polyporales, like Phanerochaete andRhizochaete, in the families Corticiaceae and Meruliaceae. P. radiata was the most related with Phlebia acerina and P. rufa along with P. tremellosa and P.brevispora, whereas P. subserialis and Phlebiopsis gigantea are more distant. P. radiata transcriptome analysis resulted with 6 590 unique gene transcripts,and similarity searches with blastx (E-value cut-off 1e-6) matched 77% of the unique transcripts to known or predicted protein-coding gene sequences.Functional annotation assigned Gene Ontology term for 54% of the gene transcripts. Most of the genes were annotated with term nucleotide bindingwhereas 7% were oxidoreductases. Among these were several lignin-modifying enzymes (class II heme-including peroxidases and laccases). From thedataset, 16% of the gene transcripts were unknown thus representing proteins potentially unique to P. radiata. Also, the mitochondrial genome of over150 kb in size shows unique features and high degree of genetic flexibility. These results provide an insight into gene content of P. radiata and genomeleveltranscriptional information on fungal genetic machinery for growth on complex liquid media.329. Characterization of molecular mechanisms underlying the multi-drug-resistant phenotypes of Mycosphaerella graminicola field isolates. SelimOmrane 1 , Anne-Sophie Walker 1 , Hind Sghyer 1 , Catherine Lanen 1 , Lamia Aouini 2 , Gert Keema 2 , Sabine Fillinger 1 . 1) BIOGER, INRA, Thiverval-Grignon, France;2) Plant Research International, Wageningen University, Wageningen, The Netherlands.Multidrug resistance (MDR) is a common trait developed by many organisms to counteract chemicals and/or drugs used against them. The basic MDRmechanism is relying on an overexpressed efflux transport system that actively expulses the toxic agent outside the cell. In fungi, MDR (or PDR) has beenextensively studied in Saccharomyces cerevisiae and Candida albicans, but also plant pathogenic fungi, e.g., Botrytis cinerea, Oculimacula yallundae andMycosphaerella graminicola are concerned by this phenomenon. In agriculture, it is currently under investigation if MDR strains may threaten the efficacyof current fungicide treatments. MDR strains were detected in septoria leaf blotch (M. graminicola) field populations since 2008. These strains are slightlymore resistant to DMI (inhibitor of the sterol 14 a-demethylase) fungicides than comparable cyp51 genotypes and cross-resistant to fungicides withdifferent modes of action. The identification of the molecular mechanism explaining the MDR phenotype in two isolated strains (MDR6 and MDR7) wasthe main goal of this study. By the use of C14-prochloraz, a DMI, we demonstrated increased fungicide efflux in both MDR strains in comparison tosensitive strains. RNA-sequencing led to the identification of several overexpressed transporter genes, out of which one MFS (major facilitator family)transporter had particularly abundant mRNA in both MDR strains. Crosses between both MDR strains showed that mdr6 and mdr7 loci are closely linked.We applied bulk-progeny sequencing to progeny of the crosses MDR6 x sensitive and MDR7 x sensitive in order to map the genomic regions co-segregatingwith the MDR phenotypes. SNP frequency analysis in sensitive and resistant bulks showed a clear co-segregation between phenotypes and the left arm ofchromosome 7. This region harbors a gene cluster including the MFS transporter gene mentioned above. After sequencing the mfs promoter, we identifieda 514 bp insertion in both MDR strains. Further studies are needed to validate the role of this insertion leading putatively to mfs overexpression and toclarify its relation to the MDR phenotype in the two studied strains. Financial support: Arvalis Institut du Vegetal, BASF Agro SAS, Bayer SAS, DuPont deNemours SAS, Syngenta Crop Protection AG.330. Candidate pathogenesis gene identification via Ustilago maydis `first gen` genomic analyses. M.E. Donaldson 1 , S. Meng 2 , B.J. Saville 1,3 . 1)Environmental & Life Sciences, Trent University, Peterborough, Ontario, Canada; 2) Lineberge Comprehensive Cancer Center, School of Medicine,University of North Carolina at Chapel Hill, Chapel Hill, N.C., USA; 3) Forensic Science Program, Trent University, Peterborough, ON, Canada.Three different approaches were used to identify candidate pathogenesis genes for the model plant pathogen Ustilago maydis (DC) Corda: 1) suppressivesubtractive hybridization (SSH) cDNA library analysis, 2) a bioinformatics approach, selecting genes unique among pathogenic basidiomycete fungi, and 3)microarray hybridization analyses. The SSH cDNA library was constructed to capture U. maydis genes expressed in planta, as well as Zea mays genes upregulatedduring the infection process. The resulting ESTs represented 23 U. maydis, and 159 Zea mays transcripts, respectively. Analysis of the U. maydistranscripts revealed 14 genes which have been previously characterized, 8 genes of unknown function, and 1 putative non-coding RNA. The fact that notall of the transcripts code secreted proteins is consistent with the observation by others that not all U. maydis effectors have recognizable secretionsignals. RT-PCR results supported in planta transcript levels for a subset these genes. To evaluate the three strategies, one gene from each series ofexperiments was chosen for selective gene deletion experiments. Deletion strains were created for: 1) a hypothetical gene (um03046) that had highrepresentation in the SSH cDNA library, 2) a conserved hypothetical gene (um01632) unique among basidiomycetes, and 3) the calcineurin B regulatorysubunit (cnb, um10226), identified through microarray hybridization as two-fold more highly expressed in the dikaryotic filamentous growth formcompared to the diploid filamentous form. All U. maydis deletion strains were capable of mating on DCM medium containing charcoal. Mutant U. maydisclones disrupted for um03046 and um01632 did not show aberrant growth phenotypes; and the morphology of these cells was indistinguishable fromwild-type strains. In contrast, the cnb mutants did not appear to separate after budding. In pathogenesis assays, the um03046 and um01632 mutants wereslightly more, or less pathogenic than wild-type infections, respectively. Results for the cnb mutants were more striking, with a marked 77% decrease inthe disease index, compared to wild-type infections. Together, these results support SSH cDNA library and microarray hybridization analyses as useful toolsin identifying genes expressed during specific stages of in planta development and those genes involved in pathogenesis.331. A biocontrol agent among pathogens : How Pseudozyma flocculosa genome relates to singular lifestyle. F. Lefebvre 1 , D.L. Joly 2 , G. Bakkeren 2 , F.Belzile 3 , R.R. Bélanger 1 . 1) Centre de recherche en horticulture, Département de phytologie, Université Laval, Quebec, QC, Canada; 2) Pacific Agri-FoodResearch Centre, Agriculture and Agri-Food Canada, Summerland, BC, Canada; 3) Institut de biologie intégrative et des systèmes, Département dephytologie, Université Laval, Québec, QC, Canada.Most fungal species belonging to the Ustilaginales are well known for their pathogenic activity towards a variety of plant species. One interestingexception is Pseudozyma flocculosa that rather acts as a biocontrol agent against powdery mildews. In order to better understand the factors underlyingthese opposed lifestyles among closely related organisms, the genome of P. flocculosa was first sequenced and annotated on the basis of homology to27th Fungal Genetics Conference | 201