?,EPA
United States
Environmental Protection
Agency
EPA/540/MR-93/514
June 1993
SUPERFUND INNOVATIVE
TECHNOLOGY EVALUATION
Demonstration Bulletin
Fungal Treatment Bulletin
USEPA-RREL/USDA-FPL
Technology Description: Fungal Treatment is a bioremedia-
tion process utilizing white-rot fungi as a means of destroying
organic contaminants in soils. The Fungal Treatment technology
was developed jointly by the Forest Products Laboratory (FPL) of
the U.S. Department of Agriculture's Forest Services in Madison,
Wl, and the U.S. EPA Risk Reduction Engineering Laboratory's
Biosystems Branch. The Fungal Treatment demonstration was
conducted on the site of a former wood treating facility, the
Brookhaven Wood Preserving (BWP) in Brookhaven, MS. The
goal of this demonstration was to determine the use of Fungal
Treatment to destroy pentachlorophenol (POP) and select poly-
nuclear aromatic hydrocarbons (PAHs: creosote constituents) in
contaminated soil.
White-rot fungi are the major degraders of lignin, a polymeric
component of wood which is resistant to biodegradation. Interest
in these fungi for biotechnological applications has increased
during the past decade, spurred by the ability of these organisms
to degrade a wide variety of hazardous compounds (including
POP and PAHs). The fungal treatment process involves inocula-
tion of the contaminated soil with selected fungal strain(s) fol-
lowed by addition of nutrients (if needed), irrigation, and aeration
of the soil through tilling/cultivating to provide optimal fungal
growth conditions. Inoculation is accomplished by physically mix-
ing the soil and the inoculum. Mixing can be performed in solids
mixing equipment, or in-situ by placing the inoculum on the
contaminated soil and tilling until the two are thoroughly mixed. In
the case of ex-situ mixing, the soil-inoculurn mix must then be
spread over the ground. Land farming procedures such as irriga-
tion, aeration and nutrient addition are then implemented periodi-
cally to sustain the fungal activity within the soil matrix As a
result of the fungal activity, the hazardous compounds are trans-
formed and become irreversibly bound to soil organic matter, in
which state they are not biologically active and thus do not
present toxicity problems. The fungal treatment can take several
weeks to several months to achieve the desired level of contami-
nant reductions.
Two species of white-rot fungi, Phanerochaete chrysosporium
and Phanerochaete sordida, have exhibited the best growth po-
tential in soil contaminated with wood preserving wastes. Studies
conducted by the developer have indicated that lignin-degrading
enzymes generated by these fungal species are capable of
oxidizing and detoxifying some normally recalcitrant PAHs. In
addition, these fungal species have been known to detoxify
phenolic compounds (PCP is one of them) by methylation of the
phenolic group. The methylated product is susceptible to further
transformation by the fungi.
Prior to treatment, a treatability study either in the field or in a
laboratory is performed using representative contaminated soil.
Based on the treatability data, the developer will determine: (1)
the applicable fungal species, (2) the inoculum loading level, (3)
length of treatment time, and (4) the need for nutrients or other
soil supplements.
Waste Applicability: This Fungal Treatment has been tested
for treatment of soils contaminated with organic wood preserving
compounds such as pentachlorophenol (PCP) and select poly-
nuclear aromatic hydrocarbons (PAHs) found in creosote. Warm
temperatures (greater than 80°F) and sufficient moisture (greater
than 30%) in the target matrix are desirable for the optimal
growth of the fungus and, thus, for the degradation of the con-
taminants.
Demonstration Results: Over a 20-wk period, from June
1992 through November 1992, the Fungal Treatment technology
was tested under actual field conditions. The test matrix was
produced from excavated wood preserving wastewater treatment
sludges from unlined onsite surface impoundments. This soil-like
material was screened to remove solid debris larger than three
inches in size, and further mixed/diluted with clean soil to bring
the PCP concentration down to an acceptable level for fungal
growth and viability. This test soil was then used in the test and
the two control plots as described below.
On the BWP site a Test Plot and two Control Plots, A and B,
were constructed. In Control Plot B, the test soil mix only was
homogenized and placed as a 10-in. bed on top of a sand layer.
In Control Plot A, the test soil mix was homogenized with the
sterile (i.e., non-inoculated or non-fungal) spawn material at a
10:1 w/w ratio and placed as a 10-in. bed on a sand layer. In the
Test Plot, the test soil mix was homogenized with the fungal
inoculum at a 10:1 w/w ratio and placed as a 10-in. bed on a
sand layer. The Test and the two Control Plots were rototilled/
cultivated about once a week over the duration of the study. Soil
temperatures were recorded on a daily basis. Moisture content of
the soil from each of the plots was determined on a weekly basis.
If the moisture content in a plot dropped below the target level, it
was irrigated with leachate collected from the same plot and/or
with the municipal tap water. Soil samples from each of the plots
were collected at the start of the study, and then at the end of the
5th, 9th, and 20th weeks of the study.
Data from the Demonstration are undergoing final review. Some
key initial findings are as follows:
/-v-y
U%) Printed on Recycled Paper
-------
• Levels of POP and the target PAHs found in the underlying sand For Further Information:
layer and the leachate from each of the plots were insignificant,
indicating low leachability and loss of these contaminants due to EPA Project Manager:
periodic irrigation of the soil and heavy rainfall. Kim Lisa Kreiton
U.S. EPA
• Levels of POP, the target PAHs, and dioxins in the active air Risk Reduction Engineering Laboratory
samples collected during the soil tilling events were insignifi- 26 West Martin Luther King Drive
cant, indicating a very low potential of air-borne contaminant Cincinnati, OH 45268
transport as a result of Fungal Treatment activities. (513) 569-7328
An Applications Analysis Report and a Technology Evaluation Developer:
Report describing the complete Fungal Treatment SITE demon- EPA/Biosystems- John Glaser
stration will be available in Summer of 1993. (513) 569-7568
_. . , „ USDA/FPL: Richard Lamar
The developers collected data independently and will produce (608) 231 -9200
results via other publications.
•U.S. Government Printing Office: 1983 — 750-071/80011
United States
Environmental Protection Agency
Center for Environmental Research Information
Cincinnati, OH 45268
Official Business
Penalty for Private Use
$300
EPA/540/MR-93/514
BULK RATE
POSTAGES FEES PAID
EPA
PERMIT No. G-35
------- |