Occurrence and characterization of peptaibols from Trichoderma citrinoviride, an endophytic fungus of cork oak, using electrospray ionization quadrupole time-of-flight mass spectrometry

Abstract

A cork oak endophytic strain of Trichoderma citrinoviride, previously selected for its antagonistic potential against various fungal pathogens involved in oak decline, was screened for the production of bioactive secondary metabolites. From liquid culture a mixture of polypeptide antibiotics (peptaibols) belonging to the paracelsin family was isolated and characterized. This peptide mixture was purified by column chromatography and preparative TLC on silica gel, and separated by analytical HPLC. It was analysed by MALDI-TOF MS and nano-ESI-QTOF MS. Tandem mass experiments were performed to determine the amino acid sequences based on the fragmentation pattern of selected parent ions. The mixture comprised 20-residue peptides with Cterminal phenylalaninol and N-terminal acetylation. Twenty-eight amino acid sequences were identified, and amino acid exchanges were located in positions 6, 9, 12 and 17. Among them, seven sequences are new as compared to those reported in the database specifically for peptaibols and in the literature. In addition, we obtained experimental evidence suggesting the existence of non-covalent dimeric forms (homo-and hetero-) of the various peptaibol species. The peptide mixture showed strong antifungal activity toward seven important forest tree pathogens, and it was highly toxic in an Artemia salina (brine shrimp) bioassay. These results emphasize the cryptic role of endophytic fungi as a source of novel bioactive natural products and biocontrol agents.

Endemic and Emerging Pathogens Threatening Cork Oak Trees: Management Options for Conserving a Unique Forest Ecosystem

“…Promising results for the control of oak pathogens have been achieved in in vivo and in vitro bioassays using mutualistic endophytic fungi of the Trichoderma genus (Linaldeddu et al 2007b). In particular, one endophytic strain of Trichoderma citrinoviride from cork oak was found to produce in liquid culture a mix of polypeptide antibiotics (peptaibols) that were very effective against seven forest tree pathogens, including D. corticola (Maddau et al 2009). …”

Two Classes of New Peptaibols Are Synthesized by a Single Non-ribosomal Peptide Synthetase of Trichoderma virens

“…The role of peptaibols in sporulation and mycoparasitism has been hypothesized (24), and their involvement in induced disease resistance in plants has been demonstrated (18,(25)(26)(27). Recently, a microheterogenous mixture of 20-residue peptaibols isolated from an endophytic biocontrol strain of Trichoderma citrinoviride has been demonstrated to have antifungal activities against several important forest tree pathogens, thus implicating their role in biocontrol (28). The 18-residue peptaibols from T. harzianum have been shown to exhibit mammalian cytotoxicity (29).…”

Specific SCAR markers and multiplex real-time PCR for quantification of two Trichoderma biocontrol strains in environmental samples

“…; Ascomycetes, Hypocreales) are used as biocontrol agents (BCA) against soilborne plant pathogenic fungi (Turoczi et al 1996;Howell 2003;Benítez et al 2004). They produce extracellular enzymes (Haran et al 1996) and/or antifungal antibiotics (Ghisalberti and Rowland 1993;Maddau et al 2009), act as competitors or direct predators (mycoparasites) to fungal pathogens (Haran et al 1996;Tseng et al 2008), promote plant growth (Vinale et al 2009), or induce resistance in plants (Vargas et al 2008;Uchiyama et al 2009). …”