670 Angelo MAZZAGLIA 1 , Naldo ANSELMI 1 , Saverio VICARIO 2 and Andrea VANNINI 1 *  Department of Plant Protection, University of Studies of Tuscia, via S. Camillo de Lellis, 01100 Viterbo, Italy.  Department of Biology, University Tor Vergata, via Orazio Raimondo, 00173 Roma, Italy. E-mail : vanniniunitus.it Received 10 July 2000 ; accepted 8 December 2000. Species belonging to Hypoxylon and related genera are bark pathogens and wood decay agents of angiosperms, sometimes able to live asymptomatically in host tissues. In the present work the ITS regions of the ribosomal DNA from 21 isolates belonging to five species were sequenced and then compared with each other. A phylogenetic parsimony analysis among isolates was carried out. ITS2 appears to be the most informative among the sequenced portions of rDNA. Results are strictly consistent with the latest taxonomic arrangements obtained with classical morphological approaches. The ITS sequence analysis is an efficient and accurate method for discriminating among taxa of Hypoxylon and related genera. INTRODUCTION Hypoxylon is a conspicuous and widespread genus of the family Xylariaceae, especially in subtropical and tropical regions. Some of them are saprobes. However, most are pathogens of angiosperms, causing bark necrosis, or wood decay (Manion & Griffin 1986, Rayner & Boddy 1986, Vannini & Scarascia Mugnozza 1991, Whalley 1996). Some are able to spend long periods asymptomatically in the host tissues as vegetative mycelium, suggesting a true endophytic habit of these fungi, which is extremely important in their ecological adaptation (Petrini & Petrini 1985, Petrini 1996). The taxonomy of Hypoxylon has been subject to several rearrangements during this century depending on the morphological traits considered. Miller (1961) did an extensive and complete study of the genus Hypoxylon, modifying his previous scheme (Miller 1928) and proposing four Sections for Hypoxylon. In this monograph Miller gave a relatively broad definition of the genus, including also species from other genera of the family such as Nummularia, now Biscogniauxia, which was later reinstated as genus by Pouzar (1979). Recently Ju & Rogers (1996) established a stricter Hypoxylon. According to their concept, a species belongs to this genus only if satisfies four main features : stromata unipartite ; ascus apical apparatus broader than tall ; stromatal tissue solid and homogeneous ; and an anamorph broadly referable to Nodulisporium. * Corresponding author. It is evident that taxonomy of Hypoxylon and related genera based on analysis of morphological traits requires time and specific experience in order to came to satisfactory classi- fication. Furthermore, classification of the species is mainly based on the teleomorph and stromata as commonly found in nature, as well as anamorphs in culture. Teleomorph are not formed in culture. A complementary approach to fungal taxonomy or phylogeny is represented by DNA analysis of specific genome regions. Methods that can be used include random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphisms (RFLPs) that permit evaluation of phylogenetic relationships without knowing the nucleotide sequence. However, DNA sequence-based methods are more accurate and allow more estimation of precise differences and storage of data for further investigations. Sequence data of rDNA subunits provide a powerful tool for evaluating phylogenetic relationship among different taxa, being highly correlated within species but relatively variable at higher taxonomic levels (among species, genera, etc.). Among rDNA subunits, the sequence of the Internal Transcribed Spacers (ITS) flanking the 58S gene, are commonly used to study phylogeny of several organisms including Oomycota (Cooke & Duncan 1997) and fungi (Saenz & Taylor 1999, Harrington & Potter 1997, Liu, Rogers & Ammirati 1997, Chillali et al. 1998, Aanen et al. 2000). The aim of this work is to estimate phylogenetic relationships among five species from Hypoxylon and related genera of the Xylariaceae with sequence data of ITS1 58SITS2 region to distinguish among the species and genera. Mycol. Res. 105 (6) : 670–675 (June 2001). Printed in the United Kingdom. Sequence analysis of the 58S rDNA and ITS regions in evaluating genetic relationships among some species of Hypoxylon and related genera