1. Functional Analysis of Essential Genes in Aspergillus Fumigatus

Aflatoxins are toxic and carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and A. parasiticus. Media with simple sugars are more favorable for aflatoxin biosynthesis, media containing 0.1 M, or higher, of a suitable carbohydrate source will support biosynthesis of aflatoxin. This research is aimed at identifying genes differentially expressed and regulated in response to changes of different carbon sources. RNA was purified from the fungal mycelia harvested at six time points post shifting and aflatoxin contents were measured. Aflatoxin production is detected as early as 16 hours after shifting from a low carbon medium, Yeast Extract (YE) to a high carbon medium, Yeast Extract Sucrose (YES). Aflatoxin levels were reduced from 3 hour up to 12 hours time points post-shift. Aflatoxin biosynthesis recovered to the initial baseline level after 18 hours post shifting to YES. At the 24 hours time point, the aflatoxin level was increased to as high as 10 fold of the initial levels. Gene expression profiling using microarray identified a total of 1958 expressed genes across all experiments as analyzed by TIGR MeV software program. These included most of the aflatoxin pathway genes. Analysis of the microarray data using oneway ANOVA determined 62 expressed genes were significant. Targeted investigation into the functions of these highly expressed genes could help to identify potential genes for signal transduction between carbon assimilation and secondary metabolite production and for regulation of aflatoxin formation. 44. ASPERGILLUS FLAVUS GENOME SEQUENCE AND COMPARATIVE ANALYSIS WITH ASPERGILLUS ORYZAE Gary A. Payne, Jiujiang Yu, Beth L. Pritchard, Jennifer Wortman, Natalie Fedorova, Masayuki Machida, Katsuya Gomi, Keietsu Abe, Doug E. Brown, Ralph A. Dean, Deepak Bhatnagar, Thomas E. Cleveland, William Nierman; 1 North Carolina State University, 840 Main Campus Dr., Raleigh, NC, 27695-7567, USA; 2 USDA/ARS/SRRC, 1100 Robert E. Lee Blvd. New Orleans, LA, 70124, USA. Phone: 301-795-7570 Fax: 301-838-0208, [email protected]. 3 The Institute for Genomic Research, 9712 Medical Center Dr., Rockville, MD, 20850, USA 4 Applied Gene Technology Research Group, National Institute of Advanced Industrial Science and Technology (AIST), Central 6, 1-1, Higashi, Tsukuba, Ibaraki, 305-8566, Japan 5 The, Graduate School of Agricultural Science, Tohoku University, 1-1, Tsutsumidori-Amamiyamachi Aoba-ku, Sendai 981-8555, Japan Aspergillus flavus attacks several crop species and produces the potent carcinogen aflatoxin in nuts and agricultural grains. A 5X genome sequence has been assembled into 79 scaffolds ranging in size from 4.5 Mb to 1.0 kb. The 36.3 Mb genome contains 13,071 genes, 99.6% of which reside in the 16 largest scaffolds. The sequence data have been deposited at NCBI and is also available at (http://www.aspergillusflavus.org) through a web browser that allows visualization of Blast matches to genes, proteins and genomic sequence of other Aspergillus species, alignments of A. flavus ESTs, and GO annotations. Fortunately, a whole genome sequence is also available for A. oryzae, a closely related fungus that does not produce aflatoxins and has lost the ability to infect plants, animals and humans. Comparative genomic analysis between the two species indicates that the genome of A. flavus is very similar to that of A. oryzae, sharing greater than 96% sequence identity. Further, these two fungi have a similar number of genes necessary for secondary metabolism. The predicted number of genes for polyketide synthases (34), non-ribosomal peptide synthases (22), and cytochrome P450s (122) is similar to that predicted for A. oryzae, 31, 24, and 151, respectively. Author Abstract Abbas, Hamed K. 43 Abe, Keietsu 23,30,44 Allen, Gethin 1 Archer, David B. 9 Arentshorst, Mark 32 Azuma, Nobuhiro 13 Baker, S. E. 41 Baraaksma, Machtelt 31 Bayram, Özgür 2,10,34 Bhatnagar, Deepak 30,43,44 Biesemann, Christoph 2 Birch, Mike 1 Bland, John M. 42 Braaksma, Machtelt 3 Braus, Gerhard H. 2,10,25,34,35 Brown, Doug E. 30,44 Busch, Silke 10,25,35 Calderone, Richard 15 Calvo, Ana 4 Campbell, Bruce C. 14,43 Christmann, Martin 35 Cleveland, Thomas E. 30,42,43,44 Cobeño, Laura 4 Davis, Meryl A. 33 de Vries, R. P. 5 Dean, Ralph A. 30,44 Debets, Fons 29,41 Dou, Xiaowei 28 Dyer, P 41 Dzikowska, Agniesza 6 Eikmeyer, Felix G. 3 Endo, Yoshikazu 6 Espeso, Eduardo 4 Eyres, G. 41 Fedorova, Natalie 30,44 Flitter, Simon 21 Frisvad, Jens C. 7 Garnet, Oumaïma 15 Gomi, Katsuya 8,23,30,36,44 Goosen, T. 9 Grundmann, Alexander 37 Hata, Yoji 8 Heerikhuisen, Margreet 31 Helmstaedt, Kerstin 10 Hill, Terry W. 11 Hoekstra, Rolf 29 Horio, Tetsuya 28 Hynes, Michael J. 33 Irniger, Stefan 34 Iversen, J. J. L. 9 Jacobus, Carrie 12 Jørgensen, T. R. 9 Kanamaru, Kyoko 13 Kato, Masashi 6,13 Kim, H. Stanley 42,43 Kim, Jong 14,43 Kobayashi, Tetsuo 6,13 Kocsubé, S. 41 Kosaric, Olja 27 Krappman, Sven 2 Lamarre, Claude 15 Lane, L. 41 Langeveld, Sandra 40 Lanthaler, Karin 19 Latgé, Jean-Paul 15,24 Leeder, Abigail C. 16,17 Li, Shu-Ming 37 Liu, Hui-Lin 18 Lockhart, Lesley 19 Lokman, B. Christien 31 Loprete, Darlene M. 11 Machida, Masayuki 23,30,44 Maiya, Shubha 37 Matsushika, Akinori 13 May, Gregory S. 14 Meijer, Susan 20 Meyer, Vera 21,22 Mizuno, Takeshi 13 Mizutani, O. 23 Mortensen, Uffe 26,27 Mouyna, Isabelle 24 Nahlik, Krystyna 25,35 Nakajima, T. 23 Nielsen, Jakob 26 Nielsen, Jens 20 Nielsen, Michael Lynge 26,27 Nierman, William C. 30,42,43,44 Oakley, Berl R 28 Obata, Hiroshi 8 Ogasawara, Hironobu 8 Ogbuadika, C. 9 Olsson, Lisbeth 20 Osmani, Stephen 18,28,38,39 Pál, Károly 29,41 Panagiotou, Gianni 20 Payne, Gary A. 12,30,43,44 Peij, Noël N. M. E. 40 Pel, Herman 40 Pritchard, Beth L. 30,44 Punt, Peter 3,31 Ram, Arthur 21,22,32,40 Reiser, Kathrin 33 Robertson, Dominique 12 Samson, Robert A. 7 Sano, M 23 Sari, Faith 34 Scheffler, Brian E. 42,43 Schwier, Elke U. 25,35 Shiina, M. 23 Shintani, T. 36 Stinnett, Suzanne M. 4 Szafron, Łukasz 6 Takagi, S. 36 Takahashi, Saori 8 Teepe, Annette G. 11 Tokuoka, M. 36 Turner, Geoffrey 1,16,17,19,37 Ukil, Leena 38,39 van de Vondervoort, Peter J. I. 40 van den Hondel, Cees 3,9,21,22,31,32 van der Werf, Mariët J. 3 van Grieken, C. 5 van Zeijl, Cora 31 vanKuyk, P. A. 5 Varga, János 29,41 Visser, Jaap 40 Voßkuhl, Katja 10 Walczuk, Bogusław 6 Watanabe, T. 23 Wilkinson, Jeffery R. 42,43 Woloshuk, Charles P. 12 Wortman, Jennifer 30,44 Wösten, H. A. B. 5 Yamagata, Y. 23 Yamashino, Takashi 13 Yu, Jiujiang 14,30,42,43,44 Zarrin, Majid 16