Three endochitinase-encoding genes, cr-ech58, cr-ech42 and cr-ech37 were identified and characterised from the mycoparasitic C. rosea strain IK726. The endochitinase activity was specifically induced in media containing chitin or Fusarium culmorum cell walls as sole carbon sources. RT-PCR analysis showed that the three genes were differentially expressed. The expression of the cr-ech42 and cr-ech37 genes was triggered by F. culmorum cell walls and chitin whereas glucose repressed their expression. In contrast, the expression of cr-ech58 was not triggered by F. culmorum cell walls and chitin, suggesting a different role for this endochitinase. Phylogenetically, the cr-ech42 and cr-ech37 genes showed to be orthologous to endochitinase 42 and 37 kDa encoding genes from other mycoparasitic fungi, while no orthologous gene for the cr-ech58 gene was found. Three genetically modified mutants of C. rosea were made by disruption of the endochitinase genes via Agrobacterium-mediated transformation and their biocontrol activity was evaluated. While in planta bioassays showed no significant difference in biocontrol efficacy between the disruptants and the wildtype, the real time RT-PCR analysis showed that disruption of each endochitinase gene affected the activity of C. rosea during interaction with F. culmorum in liquid cultures. Electronic supplementary material  The online version of this article (doi:10.1007/s00294-008-0199-5) contains supplementary material, which is available to authorized users.