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ABSTRACT
Based on morphological and immunological studies, we hypothesize that there are two lineages within the Sclerotiniaceae, a family of plant-infecting ascomycetes in the order Helotiales: 1) genera producing sclerotia, which are tuberlike, melanized masses of hyphae, and 2) genera producing substratal stromata, which are mats of compact hyphae that incorporate plant tissues. We sequenced the Internal Transcribed Spacer (ITS 1), defined by primers ITS 1 and 2, in 43 isolates: 29 sclerotial isolates (19 species in 9 genera), 11 substratal isolates (8 species in 4 genera), and 3 outgroup isolates in the Leotiaceae (3 species in 3 genera). Direct, double-stranded sequencing yielded ca 170 bases for sclerotial isolates and ca 200 bases for substratal and outgroup isolates. MACVECTOR and MULT ALIN were used for global alignment, and multiple alignment with hierarchical clustering, respectively. The Internal Transcribed Spacer showed close similarity among most of the sclerotial taxa (76 to 100% similarity to Sclerotinia sclerotiorum). This supports our hypothesis that a sclerotial lineage exists and suggests that this lineage has evolved relatively recently. Isolates of the asexual (mitotic) species Sclerotium cepivorum showed 98% similarity to those of the genus Sclerotinia. Sequence divergence was greater (45 to 65% similarity to S. sclerotiorum) amongst the substratal taxa and our outgroups. Parsimony analysis produced one statistically strongly supported tree for a group of species in the genus Rutstroemia, including Sclerotinia homoeocarpa. Although such subclusters of species can be distinguished using parsimony analysis, we conclude that a substratal Hneage cannot be discerned based on sequence data from the ITS. Among these more distantly related taxa, including some substratal ingroup taxa and the outgroup taxa, ITS 1 is saturated with changes and shows relatively equal dissimilarity. The variation observed in the ITS does not resolve among more distantly related taxa.