Rapid characterisation of aquatic hyphomycetes by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Molecular methods in fungal taxonomy are well established; they are, however, relatively labor-intensive, requiring large investments in time and resources and considerable expertise. Protein fingerprinting using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) provides a rapid, reliable and economical alternative method of characterisation of fungal isolates. The objective of our study was to evaluate the suitability of MALDI-TOF MS for the characterisation of aquatic hyphomycetes, a polyphyletic group of fungi that play a crucial role in organic matter decomposition in lotic ecosystems and are sensitive to environmental change. We first evaluated the efficiency of 13 protein extraction methods combining chemical and mechanical treatments on three aquatic hyphomycete species that are commonly associated with decomposing leaves in streams. This identified mycelium age as a key parameter affecting extraction efficiency. We then built a spectral database of 34 isolates in 23 aquatic hyphomycete species. Characterisation of all isolates was confirmed by morphological criteria and internal transcribed spacer (ITS) rDNA sequencing. The dendrogram resulting from MALDI-TOF MS based on mass spectrum similarity indicated a good and relevant taxonomical discrimination; the tree structure was comparable to that of the phylogram based on ITS sequences; in addition, MALDI-TOF MS provided a greater taxonomic precision than classical morphological methods. MALDI-TOF MS can be used for a rapid characterisation and identification of aquatic hyphomycete species. It provides a valuable alternative approach to established methods routinely used in fungal taxonomy and offers tremendous potential for ecological and ecotoxicological studies.