Molecular Systematics of the Genus Hypocrea in Europe

The majority of ascomycetes (`sac fungi`) is characterized by life cycles containing a sexual (teleomorph) and an asexual form (anamorph). Within the ascomycetes the order Hypocreales contains many species, which have economic and ecological importance, as pathogens of economically important crops (potato, tomato, bean, peanut, cacao, apple and pear trees), as pathogens of cultivated mushrooms, as sources of pharmaceuticals and mycotoxins, as biological control agents of plant pathogenic fungi, as sources of industrial enzymes, and even as a source of food. Particularly important is the genus Trichoderma, which is one of the principal components of fungi in soil. Studies of this genus are therefore mainly carried out with material isolated from soil. Teleomorphs of this genus belong to the genus Hypocrea and occur commonly on wood, other plant material and bracket fungi. The morphological variation within Hypocrea is limited in such a degree, that the identification of species relying solely on morphological examination is impossible and leads to misidentifications. Therefore, descriptions of species in the popular mycological literature refer to species complexes. For accurate identification of the species of Hypocrea it is essential to study their morphology, culture them, study the morphology of their Trichoderma anamorphs and to sequence certain genes. It is unknown, how many and which species of Hypocrea occur in Europe. The project is therefore designed to (1) assess the biodiversity of Hypocrea species occurring in Europe after collecting Hypocrea teleomorphs in (Central) European forests, (2) compare the identity, number and abundance of species with that of other regions like East Asia, South America and Indonesia, (3) compare the phylogenetic classification (obtained by evaluation and comparison of gene sequences) with that based on morphological characters of the species found, (4) detect new species of Hypocrea, (5) identify as yet unknown connections between Hypocrea teleomorphs and Trichoderma anamorphs, and (6) analyze ecological data obtained for the Hypocrea species in Central Europe.

Species of the mould genus Trichoderma are "biochemical factories" which produce a large number of substances, such as antibiotics (e.g. peptaibols) and enzymes like cellulases (to degrade plant material) and chitinases (to degrade other fungi). Thus, they are applied in several spheres of life such as detergents, textile and paper industry, but also in the biological control of harmful or pathogenic fungi. Most species of Trichoderma have been isolated from soil. The sexual stage of this genus, Hypocrea, forms often brightly coloured pillows, pustules or crusts on wood and bark, other fungi (mainly polypores) and more rarely on grasses. Ca. 450 names have been established but it is unknown how many true species exist which form the sexual stage. The objective of this project therefore was to investigate the biodiversity of Hypocrea, i.e. how many and which species exist in Europe. To achieve this fungal specimens were collected each year. Several methods were applied to identify and to characterise the species: morphology of the sexual state (Hypocrea), preparation of pure cultures and the Trichoderma states (anamorphs) from ascospores, morphology of the anamorphs, growth experiments to characterise cultures and determine growth rates, and extraction of DNA, determination of sequences of 3-4 genes and of the positions on the phylogenetic tree of the genus Hypocrea/ Trichoderma based on gene sequences. More than 500 fresh specimens were collected in Europe within 3.5 years and more than 400 cultures were prepared from them. 60 species were identified using gene sequences in combination with morphological and ecological data. With recently published species at least 70 species occur in Europe. This result exceeds initial estimations of 15-20 species by far. More than 25 are new, of these 3 have been described. All relevant data of 30 species have been recorded, and all species are planned to be described in a European monograph, with some in few additional single papers, within a follow-up project. The cultures are being deposited in an official strain collection (CBS, Utrecht), where they are freely accessible after their publication. The cultures represent an enormous potential for future research on the nature and applicability of secondary metabolites and other aspects of Trichoderma.