ABSTRACT
Different peptaibols were observed in both fresh and frozen marine sediments collected from a marine area devoted to bivalve culture (Fier d'Ars, Atlantic coast, France). The identification of the peptaibols was based on a three-step mass-spectrometric analysis: observation of doubly charged ions with a characteristic isotopic profile, cleavage and observation of C- and N-terminal fragments, and partial sequencing of the N-terminal segments. The MS characteristics indicated numerous similarities between the peptaibols detected and those produced by different strains of Trichoderma species isolated from fresh sediments. Peptaibols were also detected in mussel samples collected at the same site. This constitutes the first observation of contamination of the marine human-food chain by fungal metabolites. Since peptaibols were readily observed both in fresh sediments and in samples kept frozen for several years, these compounds can be considered as stable markers of the development of Trichoderma in the marine environment.
Subject(s)
Anti-Bacterial Agents/metabolism , Fungi/growth & development , Marine Biology , Peptides/metabolism , Fungi/metabolism , Protein Conformation , Spectrometry, Mass, Electrospray IonizationABSTRACT
Peptaibols are known membrane-modifying peptides that were recently detected in marine sediments and mussels collected from a shellfish farming area (Fier d'Ars, Atlantic coast, France). In this investigation, embryotoxicity bioassays with oysters (Crassostrea gigas) were performed to assess acute toxicity of alamethicin and different groups of peptaibols produced by a Trichoderma longibrachiatum strain isolated from marine environment. C. gigas embryos appeared very sensitive to all the metabolites examined with higher toxic effects for long-sequence peptides (EC50 ranging from 10 to 64 nM). D-shaped larvae with mantle abnormality were particularly noticed when peptaibol concentrations increased. Disturbances of embryogenesis were also observed following exposure to organic and aqueous extract of sediments from Fier d'Ars (EC50=42.4 and 6.6 g L(-1) dry weight, respectively). Although peptaibol concentrations measured in these sediments could explain only a part of the toxic effects observed, this study suggests that these mycotoxins can induce larval abnormalities in a population of exposed animals at environmentally realistic concentrations. Their detection in coastal areas devoted to bivalve culture should be taken into account.
Subject(s)
Anti-Bacterial Agents/toxicity , Crassostrea/drug effects , Embryo, Nonmammalian/drug effects , Peptides/toxicity , Water Pollutants, Chemical/toxicity , Alamethicin/toxicity , Animals , Biological Assay/methods , Crassostrea/chemistry , Female , France , Geologic Sediments/chemistry , Male , Reproducibility of Results , Trichoderma/chemistryABSTRACT
Extraction followed by reverse phase liquid chromatography (LC)/electrospray ionization-ion trap-mass spectrometry (ESI-IT-MS) analysis has been successfully developed for the determination of peptaibols, fungal toxic metabolites, in marine sediments. Spiking experiments showed that the mean recovery of target compounds exceeded 85% at a spiking level of 10 ng/g of sediment (wet weight). Detection and quantification limits were 250 and 830 pg/g of sediment, respectively. The method developed constituted the first sensitive assay for quantification of peptaibol trace amounts in a natural environment. A concentration of 5 ng/g in sediment samples collected from Fier d'Ars was found.
Subject(s)
Geologic Sediments/chemistry , Peptides/analysis , Spectrometry, Mass, Electrospray Ionization/methods , Alamethicin/isolation & purification , Calibration , Chromatography, Liquid , Complex Mixtures/chemistry , Environment , Peptaibols , Peptides/isolation & purification , Reproducibility of Results , SolventsABSTRACT
The production of peptaibols by a marine-related Trichoderma longibrachiatum strain was studied using electrospray ionisation multiple-stage ion trap mass spectrometry (ESI-MSn-IT) and gas chromatography/electron impact mass spectrometry (GC/EI-MS). Two major groups of peptaibols were identified, those with long sequences (20 amino acids) and others with short sequences (11 amino acids). This paper describes the methodology used to establish the sequences of short peptaibols in a mixture without previous individual separation. Nine peptaibols were identified. Among them, eight are new, namely as trichobrachin A I-IV (Aib9-Pro10 sequence) and as trichobrachin B I-IV (Val9-Pro10 sequence). Original Pro6-Val7 and Val9-Pro10 sequences have to be noted.
