Bioremediation of waste gas and soil by black extremotolerant fungi

Recalcitrant environmental pollutants originating from recent and past industry as well as from sinks used for deposit of contaminated sludge are leaking into soil and water and seriously threaten not only the environment but also our food safety and our drinking water. Chemical and physical cleaning methods are by far too expensive and too intricate to be performed on a large scale. Bioremediation thus far has been concentrated on bacteria, however with several obstacles. A special group of fungi , namely the "black yeasts", is very tolerant to adverse conditions (acid, heavy metals, temporal desiccation, low N and P supply, extreme temperatures, UV and radioactive irradiation). The potential of these fungi to degrade hydrocarbons and xenobiotics has been demonstrated for several species. Therefore these fungi are highly promising candidates for bioremediation. Two hundred fungi from 4 different phylogenetic groups will be screened for their ability to degrade benzene-derivates, alkanes and PCBs. These strains will be provided by the CBS-KNAW Fungal Biodiversity Center which hosts the largest collection of black yeasts. Further, highly contaminated sites will be screened for new fungal species by using selective media. The transcriptome of promising candidates will be sequenced (Ion Proton), and compared with protein expression profiles in order to identify the responsible enzymes and biodegradation pathways. The project will be the basis for biotechnological applications in industrial filters and for the set-up of ready to use kits (inocula) for environmental remediation.

For several years it has been known that black micro-fungi or black yeasts belong to the most stress resistant organisms on Earth. These fungi inhabit rock in all region of the Earth including hot and cold deserts and the top of the highest mountains. Some species growth in highly polluted water and others in abandoned nuclear reactors. Also the human environment was conquered by these fungi: They can be found in bathrooms, in sauna facilities and in dishwashers. In rare cases black fungi can cause serious or even fatal infections. Because the black fungi are used to live in environments with poor nutrient availability they are adapted to use any chemical substance including toxins and other compounds that cannot be used as a nutrient by other organisms. This latter quality lead the Extremophile Center to launch a FWF-funded project in which 163 black yeasts were screened for their potential to filter exhaust fumes and clean contaminated soils. Nineteen strains were able to degrade at least one of the three tested hydrocarbons, i.e. toluene, hexadecane and biphenyl. Only two fungi were able to use all three aliphatic compounds as sole carbon source: Exophiala mesophila and Cladophialophora immunda. In order to get a deeper insight into the cellular processes underlying the toluene degradation, both genomes were sequenced, assembled and annotated. The transcriptome of C. Immunda growing on toluene as sole carbon source was sequenced and subsequently analyzed by bioinformatic. The molecular methods have been optimized for this special group of fungi and the bioinformatic analysis was streamlined with robust and reproducible software-pipelines that can be used to analysis the genome and transcriptome of any other fungi A selective isolation method that was developed during this project allowed us to identify new hydrocarbon-degrading strains in addition to the 163 tested strains.. Also here fungi with dark pigmentation exhibited a higher hydrocarbon-degrading potential than less pigmented species. Because both fungi - Exophiala and Cladophialophora are closely related to human pathogen strains, plans for using them as biofilters have been shelved for the time being. For future projects we will focus on those non-pathogens that have been freshly isolated from the environment.