The Hypocrea lixii/Trichoderma harzianum species aggregate contains a group of taxa (H. lixii/T.harzianum , T. aggressivum , T. tomentosum , T. cerinum , T. velutinum , H. tawa ) of which some (e. g. T. harzianum) are important for biocontrol of plant pathogenic fungi in agriculture, whereas others are aggressive pathogens of Agaricus spp. and Pleurotus spp. in mushroom farms (T. aggressivum), or opportunistic pathogens of immunocompromised mammals including humans (T. harzianum). We characterized the evolutionary properties of three genomic regions in Hypocrea/Trichoderma: the internal transcribed spacer regions ITS1 and 2 of rDNA, the large intron of translation elongation factor 1-alpha (tef1a), and a portion of the large exon of the endochitinase 42 gene (ech42 ), selected the best model which describes the evolution of every fragment, tested the molecular clock hypothesis and made an estimation of the usability of the combined three fragments data matrix for the phylogenetic analysis of the genus as a whole as well as on the level of the holomorphic H. liaxii/T. harzianum species clade and separate clonal lineages. To this end, we applied Bayesian phylogenetic inferences to 124 sequences of ITS1 and 2 and of the large tef1a intron, and to 64 ech42 gene sequences to resolve the evolution of H. lixii/T. harzianum with respect to the position of other taxa with closely related phenotypes. The resulting phylogram clearly identified T.aggressivum, T. velutinum, H. tawa, T. cerinum and T. tomentosum as phylogenetic species, and in addition identified three new unknown phylogenetic species as members of this clacle. The clear distinction between T. tomentosum and T. cerinum was not recognized in all trees, but was supported by multivariate analysis of phenotype micro arrays. In contrast, H. lixii/T. harzianum did not form a single phylogenetic species in this study, as its monophyly was not supported in any analysis. Strains morphologically identified as H. lixii or T. harzianum formed either a proliferating megaclade (three loci combined tree) or appeared as several independent branches on the star-like phylograrm (ITS1 and 2 and large tef1a intron) . However most terminal clacles had a high posterior probability and thus fulfilled the criterion to consider their member isolates as a phylogenetic species. The clade containing the ex-neotype strain of T. harzianum included exclusively clonally propagating isolates from temperate climates, and was clearly distinguished from all other "H. lixii/T. harzianum" lineages by multivariate analysis of phenotype microarray data. We propose that only this lineage should be regarded as T. harzianum sensu stricto, and that the other lineages be regarded as T.harzianum sensu lato until phenetic characters have been defined which allow their description as separate taxa.