Bioremediation Potential of White-rot Fungi: Treatment of PAH-contaminated Soil

Kari Steffen*, Elke Lang**, Carsten i. d. Wiesche**, Frantisek Zadrazil**, Annele Hatakka*

* Department of Applied Chemistry and Microbiology, PO-Box 56, Viikki Biocenter,
FIN-00014 University of Helsinki, Finland

** Department of Soil-Biology, Federal Agricultural Research Center of, Bundesallee 50,
D-38116 Braunschweig, Germany

White-rot fungi are wood decomposting basidiomycetes which are capable of degrading not only lignin, but also various recalcitrant environmental pollutants. Among these contaminants are polycyclic aromatic hydrocarbons (PAHs). PAHs are common environmental pollutants formed during incomplete incineration of organic material and found in coal and tar producing sites. Many of the higher molecular PAHs (five or more rings) are considered to be mutagenic and carcinogenic. Therefore especially PAH-contaminated soil is a health hazard for humans if the contaminated dust is formed an inhaled. Many PAHs, for example benzo(a)pyrene (BaP), undergo a change in the liver catalyzed by cytochrome P₄₅₀ of mammals which results in the formation of highly carcinogenic epoxides. Due to the health risk it is important to map contaminated sites and try to clean them up. Conventional methods like incineration and soil washing are expensive, and cheaper biological treatment is favored. White-rot fungi have been found to posses a good potential for PAH-contaminated soil bioremediation due to their ligninolytic exoenzymes, e. g. lignin peroxidases, manganese peroxidases (MnPs) and laccases. The research of PAH-contaminated soil treatment is mainly focused on the search for an ideal fungus, capable of growing, excreting its enzymes and degrading or inactivating PAH-compounds in the soil.

Two different methods were used in this study to test four white-rot fungi: Bjerkandera adusta, Trametes hirsuta, Phlebia radiata and Pleurotus sp. P1 Florida. Erlenmeyer flasks were used for the first experiment. The contaminated soil and straw, the latter to be used as a substrate for the fungus, were mixed, autoclaved, and the fungus was added as a liquid culture. For the second experiment, glass tubes (23,5cm long; diameter 2,7cm) were used. Straw with pregrown mycelia of the fungus and the contaminated native soil (not autoclaved) were not mixed but placed in contact with each other. The mineralisation of BaP was followed with ¹⁴C-labeled BaP and the formed ¹⁴CO₂ was measured with a liquid scintillation counter. Another glass tube experiment with unlabeled BaP was performed to follow the enzyme activities of MnP and laccase during the BaP breakdown. The enzymes were extracted from the soil with buffers.

Only 1% of the total added ¹⁴C-BaP was recovered as ¹⁴CO₂ in the flask experiment for B. adusta, T. hirsuta and P. radiata. Pleurotus was not tested. No mineralisation of BaP was observed by these fungi in the tube experiment. They were either killed or did not grew into the soil. This was mainly because they could not compete against the natural microflora of the soil. Pleurotus grew actively into the soil and mineralized 6% of the applicated ¹⁴C-BaP in 20 weeks. Pleurotus also produced reasonable amounts of active MnP and laccase in the soil. The MnP activity correlated for the first 8 weeks with the mineralisation of BaP.

The results show that Pleurotus sp. P1 Florida is capable of mineralizing BaP in soil. The main reason for this is the competitiveness of the fungus against other soil microorganisms and its growth into the soil. An other reason is that Pleurotus excretes high amounts of MnP and laccase, possibly also other enzymes into the soil, which are capable of mineralizing high molecular weight PAHs. It is also obvious that most of the white-rot fungi are not able to grow into soil and therefore they are not useful for the bioremediation of contaminated soil.