Homologs of ToxB, a host-selective toxin gene from Pyrenophora tritici-repentis, are present in the genome of sister-species Pyrenophora bromi and other members of the Ascomycota Rachael M. Andrie 1 , Conrad L. Schoch, Rebecca Hedges 2 , Joseph W. Spatafora, Lynda M. Ciuffetti * Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331, USA Received 30 April 2007; accepted 23 October 2007 Available online 30 October 2007 Abstract Pyrenophora tritici-repentis requires the production of host-selective toxins (HSTs) to cause the disease tan spot of wheat, including Ptr ToxA, Ptr ToxB, and Ptr ToxC. Pyrenophora bromi, the species most closely related to P. tritici-repentis, is the causal agent of brown leaf spot of bromegrass. Because of the relatedness of P. bromi and P. tritici-repentis, we investigated the possibility that P. bromi con- tains sequences homologous to ToxA and/or ToxB, the products of which may be involved in its interaction with bromegrass. Multiplex polymerase chain reaction (PCR) revealed the presence of ToxB-like sequences in P. bromi and high-fidelity PCR was used to clone sev- eral of these loci, which were subsequently confirmed to be homologous to ToxB. Additionally, Southern analysis revealed ToxB from P. bromi to have a multicopy nature similar to ToxB from P. tritici-repentis. A combination of phylogenetic and Southern analyses revealed that the distribution of ToxB extends further into the Pleosporaceae, and a search of available fungal genomes identified a dis- tant putative homolog in Magnaporthe grisea, causal agent of rice blast. Thus, unlike most described HSTs, ToxB homologs are present across a broad range of plant pathogenic ascomycetes, suggesting that it may have arose in an early ancestor of the Ascomycota. Ó 2007 Elsevier Inc. All rights reserved. Keywords: Bromegrass; Brown leaf spot; gpd; Host-selective toxins; ITS; MAT-2 HMG box; Multi-gene phylogeny; Ptr ToxB; Tan spot; Wheat; Pyrenophora tritici-repentis; Pyrenophora bromi; Ascomycota; Plant pathogen 1. Introduction Host-selective toxins (HSTs), a diverse set of molecules known only to be produced by plant pathogenic fungi, function as essential disease determinants in a number of plant host–pathogen interactions (reviewed in Walton, 2000; Markham and Hille, 2001; Wolpert et al., 2002). They are implicated in pathogenesis/virulence because they are toxic only to hosts susceptible to the fungus, and in turn, toxin production by the fungal pathogen is strictly correlated with disease elicitation on a susceptible host. For a number of HSTs, causality in disease has been con- firmed molecularly (Panaccione et al., 1992; Ciuffetti et al., 1997; Yoder et al., 1997; Tanaka et al., 1999; Chur- chill et al., 2001; Johnson et al., 2001). HSTs range from low-molecular weight products of multifunctional enzymes or complex enzymatic pathways encoded for by complex gene clusters to protein products of single genes. In general, the genetic determinants of their production are discontin- uously distributed in that they are present only in HST- producing, pathogenic isolates and are completely absent in nonpathogenic counterparts (Ahn and Walton, 1996; 1087-1845/$ - see front matter Ó 2007 Elsevier Inc. All rights reserved. doi:10.1016/j.fgb.2007.10.014 * Corresponding author. Fax: +1 541 737 3573. E-mail address: ciuffetL@science.oregonstate.edu (L.M. Ciuffetti). 1 Present address: Division of Infectious Diseases, Department of Medicine, Oregon Health and Science University, Portland, OR 97239, USA. 2 Present address: Monoclonal Antibody Facility, Institute of Neuro- science, University of Oregon, Eugene, OR 97403, USA. www.elsevier.com/locate/yfgbi Available online at www.sciencedirect.com Fungal Genetics and Biology 45 (2008) 363–377