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Chapman, Peter J. as First Author
Chapman, P.J., M. Shelton, Magda Grifoll and S. Selifonov. 1995. Fossil Fuel Biodegradation: Laboratory Studies. EPA/600/J-95/434. Environ. Health Perspect. 103(Suppl 5):79-83. (ERL,GB 871).
Biodegradation of the polycyclic aromatic hydrocarbons of creosote by undefined
bacterial cultures was shown to be accompanied by the accumulation of neutral
and acidic oxidation products. Formation of a number of identified neutral
products is accounted for by demonstration of anomalous actions of an arene
dioxygenase on the benzylic methylene and methylene carbons of napthenoaromatic
hydrocarbons. Both neutral and acidic water-soluble fractions are also formed
when various mixed bacterial cultures degrade weathered crude oil. While
constituents of these fractions are not yet identified, the neutral materials
have been shown to be toxic to developing embryos of invertebrates. These
observations are discussed in relation to chemical and toxicological
assessments of biodegradation of the complex chemical mixtures of fossil fuels.
Chapman, P.J. 1988. Constructing Microbial Strains for Degradation of Halogenated Aromatic Hydrocarbons. In: Environmental Biotechnology: Reducing Risks from Environmental Chemicals Through Biotechnology. Gilbert S. Omenn et al., Editor. Plenum Press, New York, NY. Pp. 81-95. (ERL,GB X568).
This paper examines methods that have been used to isolate and to construct
bacteria capable of growing aerobically with chlorinated aromatic compounds,
including chlorinated hydrocarbons. It also describes some recent work in this
area of research.
Chapman, Peter J. as Contributing Author
Selifonov, Sergey A., Peter J. Chapman, Simon B. Akkerman, Jerome E. Gurst, Jacqueline M. Bortiatynski, Mark A. Nanny and Patrick G. Hatcher. 1998. Use of 13C Nuclear Magnetic Resonance to Assess Fossil Fuel Biodegradation: Fate of [1-13C]Acenaphthene in Creosote Polycyclic Aromatic Compound Mixtures Degraded by Bacteria. Appl. Environ. Microbiol. 64(4):1447-1453. (ERL,GB 1021).
[1-13C]acenaphthene, a tracer compound with a nuclear magnetic resonance
(NMR)-active nucleus at the C-1 position, has been employed in conjunction with
a standard broad-band-decoupled 13C-NMR spectroscopy technique to study the
biodegradation of acenaphthene by various bacterial cultures degrading aromatic
hydrocarbons of creosote. Site-specific labeling at the benzylic position of
acenaphthene allows 13C-NMR detection of chemical changes due to initial
oxidations catalyzed by bacterial enzymes of aromatic hydrocarbon catabolism.
Biodegradation of [1 -13 C]acenaphthene in the presence of naphthalene or
creosote polycyclic aromatic compounds (PACs) was examined with an undefined
mixed bacterial culture (established by enrichment on creosote PACs), and with
isolates of individual naphthalene- and phenanthrene-degrading strains from
this culture. From 13C-NMR spectra of extractable materials obtained in time
course biodegradation experiments under optimized conditions, a number of
signals were assigned to accumulated products such as 1-acenaphthenol,
1-acenaphthenone, acenaphthene-1,2-diol and naphthalene 1,8-dicarboxylic acid,
formed by benzylic oxidation of acenaphthene and subsequent reactions. Limited
degradation of acenaphthene could be attributed to its oxidation by napthalene
1,2-dioxygenase or related dioxygenases, indicative of certain limitations of
the undefined mixed culture with respect to acenaphthene catabolism.
Coinoculation of the mixed culture with cells of acenaphthene-grown strain
Pseudomonas sp. A2279 mitigated accumulation of partial transformation products
and resulted in more complete degradation of acenaphthene. This study
demonstrates the value of the stable isotope labeling approach and its ability
to reveal incomplete mineralization even when as little as 2 to 3% of substrate
is incompletely oxidized, yielding products of partial transformation. The
approach outlined may prove useful in assessing bioremediation performance.
Pinkart, Holly C., Richard Devereux and Peter J. Chapman. 1998. Rapid Separation of Microbial Lipids Using Solid Phase Extraction Columns. J. Microbiol. Methods. 34(1):9-15. (ERL,GB 1038).
A method was developed to rapidly separate lipid classes commonly found in
microbial communities. The method is based on the use of aminopropyl solid
phase extraction columns to separate polyhydroxyalkoanates (PHA),
phospholipids, sterols, triglycerides, diglycerides, monoglycerides, and steryl
esters. Recoveries of all lipid classes, with the exception of PHA and sterols,
ranged from 91% to greater than 99%. PHA were recovered at 69% of the standard,
and sterols from 82-84% of the standard. When applied to the analysis of lipids
extracted from the cyanobacterium Spirulina platensis, the method afforded
excellent recovery and separation of phospholipids and diglycerides including
saturated, monounsaturated and polyunsaturated fatty acids. The S. platensis
lipids also contained hydrocarbons and phytol recovered in the steryl ester and
diglyceride fractions, respectively. This method provided a high yield,
specific and rapid separation of microbial lipids with little contamination
from other lipid groups, and will be useful for the characterization of
microbial communities in environmental samples.
Leblond, Jeffrey D. and Peter J. Chapman. 2000. Lipid Class Distribution of Highly Unsaturated Long Chain Fatty Acids in Marine Dinoflagellates. J. Phycol. 26(6):1103-1108. (ERL,GB 1094).
The very long chain highly unsaturated C28 fatty acids, octacosaheptaenoic
[28:7(n-6)] and octacosaoctaenoic acid [28:8(n-3)], were found to be associated
with phospholipids, obtained by fractionation of total lipid extracts into
distinct lipid classes, in 4 and 6, respectively, of 16 examined
dinoflagellates. An interfraction comparison of fatty acids associated with
phospholipids and glycolipids has also shown that the phospholipid fractions
contained the majority (over 75% in 12 of 16 strains) of docosahexaenoic acid
[22:6(n-3)] and traces of tetracosanoic acid (24:0). By contrast, the highly
unsaturated C18 fatty acids octadecatetraenoic [18:4(n-3)] and
octadecapentaenoic acid [18:5(n-3)] were primarily recovered from a
chloroplast-associated glycolipid fraction comprised of
monogalactosyldiacylglycerol, digalactosyldiacylglycerol, and
sulfoquinovosyldiacylglycerol. In 12 of 16 strains, an interfraction comparison
showed that over 90% of 18:5(n-3) was found to be associated with
glycolipids.These findings indicate that the C28 fatty acids are located and
probably synthesized in the cytoplasm or in an organelle other than the
chloroplast, possibly with 22:6(n-3) and 24:0 as precursors, whereas the C18
fatty acids 18:4(n-3) and 18:5(n-3) are glycolipid constituents apparently
synthesized within the chloroplast. The function(s) of these C28 fatty acids as
components of phospholipids in cellular membranes is currently unknown.
Leblond, Jeffrey D. and Peter J. Chapman. 2002. Survey of the Sterol Composition of the Marine Dinoflagellates Karenia brevis, Karenia mikimotoi, and Karlodinium micrum: Distribution of Sterols Within Other Members of the Class Dinophyceae. EPA/600/J-01/420. J. Phycol. 38(8):670-682. (ERL,GB 1149).
The sterol composition of different marine microalgae was examined to determine
the utility of sterols as biomarkers to distinguish members of various algal
classes. For example, members of the class Dinophyceae possess certain 4-methyl
sterols, such as dinosterol, which are rarely found in other classes of algae.
The ability to use sterol biomarkers to distinguish certain dinoflagellates
such as the toxic species Karenia brevisHansen and Moestrup, responsible for
red tide events in the Gulf of Mexico, from other species within the same class
would be of considerable scientific and economic value. Karenia brevis has been
shown by others to possess two majors sterols, (24S)-4a-methyl-5a
-ergosta-8(14),22-dien-3b-ol (ED) and its 27-nor derivative (NED), having novel
structures not previously known to be present in other dinoflagellates. This
prompted the present study of the sterol signatures of more than 40
dinoflagellates. In this survey, sterols with the properties of ED and NED were
found in cultures of K. brevis and shown also to be the principal sterols of
Karenia mikimotoi Hansen and Moestrup and Karlodinium micrum Larsen,two
dinoflagellates closely related to K. brevis. They are also found as minor
components of the more complex sterol profiles of other members of the
Gymnodinium/Peridinium/Prorocentrum (GPP) taxonomic group. The distribution of
these sterols is consistent with the known close relationship between K.
brevis, K. mikimotoi,and K. micrum,and and serves to limit the use of these
sterols as lipid biomarkers to a few related species of dinoflagellates.
Leblond, Jeffrey D., Terence J. Evens and Peter J. Chapman. 2003. Biochemistry of Dinoflagellate Lipids, with Particular Reference to the Fatty Acid and Sterol Composition of a Karenia brevis Bloom. EPA/600/J-03/491. Phycologia. 42(4):324-331. (ERL,GB 1160).
The harmful marine dinoflagellate, Karenia brevis (Dinophyceae), frequently
forms large toxic blooms in the waters off of the west coast of Florida (USA)
and is responsible for massive fish kills and public health concerns. Despite
decades of field studies on this organism, no investigation has yet
characterized the lipid composition of a K. brevis bloom. To address this lack
of information, samples from a 1999 K. brevis bloom from the northwest Florida
coast were analyzed for their fatty acid and sterol composition. Fatty acids
found in lipid fractions containing membrane phospholipids,
chloroplast-associated glycolipids, and storage triglycerides differed
significantly. The glycolipid fraction was found to contain octadecapentaenoic
acid [18:5(n-3)], a fatty acid commonly associated with dinoflagellates. The
phospholipid fraction was found to contain small amounts of two recently
described, highly unsaturated fatty acids, octacosaoctaenoic acid [28:8(n-3)]
and octacosaheptaenoic acid [28:7(n-6)]. Fatty acids from the triglyceride
fraction were more abundant than those associated with glycolipid or
phospholipids. Sterols were found mainly as free sterols and were dominated by
two compounds (24S)-4a-methyl-5a-ergosta-8(14),22-dien-3b-ol (ED) and its
27-nor derivative (NED). The lipid composition of these samples very closely
resembles laboratory-grown cultures of K. brevis and serves to provide an in
situ field validation of past laboratory examinations of this organism. The
implications of our data are discussed in the context of the physiological
autecology of K. brevis, in the form of a minireview on the biochemistry of
dinoflagellate lipids, as studied in both the laboratory and the environment.
Leblond, Jeffrey D. and Peter J. Chapman. Unpublished. Polyunsaturated C27 Hydrocarbons in the Marine Dinoflagellate, Pyrocystis lunula (Dinophyceae): Preliminary Characterization. J. Phycol. 33 p. (ERL,GB 1179).
The lipids of different algal species have revealed a diversity of fatty acids,
sterols, and hydrocarbons, several of which are considered of chemotaxonomic
value and candidates for useful chemical biomarkers with the potential for
characterizing phytoplankton community composition. Neutral fractions obtained
from the lipids of two strains of Pyrocystis lunula, in the course of
characterizing over forty dinoflagellates, were found to contain an abundant
quantity of long-chain polyunsaturated hydrocarbons, along with previously
reported keto-steranes. Gas chromatographic examination showed a retention time
of the predominant hydrocarbon, relative to standards, which together with its
molecular weight (m/z = 364) and mass spectrum suggested a multiply unsaturated
C27 compound (C27H40) with eight double bonds. Reduction (Adams catalyst) gave
a product identical to authentic n-heptacosane indicating a straight chain
hydrocarbon. While the positions and stereochemistry of double bonds have not
been established, the carbon number of this hydrocarbon and the number of
double bonds strongly suggest a relationship to, and formation by
decarboxylation of, the recently described, long-chain polyunsaturated C28
fatty acid, [28:8(n-3)], shown to be a constituent of dinoflagellate
phospholipids. This hydrocarbon was not found in any other genus of the over
forty examined dinoflagellates, nor was it found in isolates of two other
species of Pyrocystis, P. fusiformis and P. noctiluca. The function(s) of this
compound in P. lunula is currently unclear.
Leblond, Jeffrey D. and Peter J. Chapman. 2004. Sterols of the Heterotrophic Dinoflagellate, Pfiesteria piscicida (Dinophyceae): Is There a Lipid Biomarker?. J. Phycol. 40(1):104-111. (ERL,GB 1191).
Within U.S. waters, blooms of the dinoflagellate, Pfiesteria piscicida, have
been recorded on an almost regular basis in the Chesapeake Bay and surrounding
mid-Atlantic regions for the last two decades. Despite the apparent
significance of such blooms to the environment and human health, and the
attendant economic consequences, little work has addressed the physiology and
biochemistry, particularly that of sterol composition, of P. piscicida. GC-MS
characterization of trimethylsilyl ether derivatives of sterols from free
sterol and sterol ester fractions was performed in an effort to determine
whether P. piscicida produces unique sterols that may serve as potential
biomarkers. This characterization revealed that, like most dinoflagellates, the
majority of sterols was present as free sterols. Furthermore, the profile of
free sterols was found to resemble those of photosynthetic dinoflagellates,
with the dominant compound being the previously reported dinoflagellate sterol,
dinosterol. A number of other 4a-methyl-substituted sterols and steroidal
ketones common to other dinoflagellates were also identified. No strong
candidate(s) for a unique sterol biomarker was present.
Leblond, Jeffrey D., Jeremy L. Dahmen, Rebecca L. Seipelt, Matthew J. Elrod-Erickson, A. Bruce Cahoon, Rodney Kincaid, James C. Howard, Terence J. Evens and Peter J. Chapman. 2005. Lipid Composition of Chlorarachniophytes (Chlorarachniophyceae) from the Genera Bigelowiella, Gymnochlora, and Lotharella. J. Phycol. 41:311-321. (ERL,GB 1218).
The Chlorarachniophyceae are unicellular eukaryotic algae characterized by an
amoeboid morphology that may be the result of secondary endosymbiosis of a
green alga by a nonphotosynthetic amoeba or amoeboflagellate. Whereas much is
known about the phylogeny of chlorarachniophytes, little is known about their
physiology, particularly that of their lipids. In an initial effort to
characterize the lipids of this algal class, four organisms from three genera
were examined for their fatty acid and sterol composition. Fatty acids from
lipid fractions containing chloroplast-associated glycolipids, storage
triglycerides, and cytoplasmic membrane-associated polar lipids were
characterized. Glycolipid-associated fatty acids were of limited composition,
principally eicosapentaenoic acid [20:5(n-3)] and hexadecanoic acid (16:0).
Triglyceride-associated fatty acids, although minor, were found to be similar
in composition. The polar lipid fraction was dominated by lipids that did not
contain phosphorus, and had a more variable fatty acid composition with 16:0
and docosapentaenoic acid [22:5(n-3)] dominant along with a number of minor C18
and C20 fatty acids. Crinosterol and one of the epimeric pair
poriferasterol/stigmasterol were the sole sterols. Several genes required for
synthesis of these sterols were computationally identified in Bigelowiella
natans. One sterol biosynthesis gene showed the greatest similarity to SMT1 of
the green alga, Chlamydomonas reinhardtii. However, homologues to other
species, mostly green plant species, were also found. Further, the method used
for identification suggested that the sequences have been transferred to a
genetic compartment other than the likely original location, the nucleomorph
nucleus.
Shields, Malcolm S., Stacy O. Montgomery, Peter J. Chapman, Stephen M. Cuskey and P.H. Pritchard. 1989. Novel Pathway of Toluene Catabolism in the Trichloroethylene-Degrading Bacterium G4. EPA/600/J-89/168. Appl. Environ. Microbiol. 55(6):1624-1629. (ERL,GB 668). (Avail. from NTIS, Springfield, VA: PB90-129537)
o-Cresol and 3-methylcatechol were identified as successive transitory
intermediates of toluene catabolism by the trichloroethylene-degrading
bacterium G4. The absence of a toluene dihydrodiol intermediate or toluene
dioxygenase and toluene dihydrodiol dehydrogenase activities suggested that G4
catabolizes toluene by a unique pathway. Formation of a hybrid species of 18O-
and 16O- labeled 3-methylcatechol from toluene in an atmosphere of 18O2 and 16O
2 established that G4 catabolizes toluene by successive monooxygenations at the
ortho and meta positions. Detection of trace amounts of 4-methylcatechol from
toluene catabolism suggested that the initial hydroxylation of toluene was not
exclusively at the ortho position. Further catabolism of 3-methylcatechol was
found to proceed via catechol-2,3-dioxygenase and hydroxymuconic semialdehyde
hydrolase activities.
Mueller, James G., Peter J. Chapman and P. Hap Pritchard. 1989. Creosote-Contaminated Sites: Their Potential for Bioremediation. EPA/600/J-89/170. Environ. Sci. Technol. 23(10):1197-1201. (ERL,GB 671). (Avail. from NTIS, Springfield, VA: PB90-129552)
The demonstrated ability to biologically transform toxic organic chemicals into innocuous compounds under laboratory conditions has led to the view that microbial systems may be employed advantageously to remediate hazardous waste sites polluted with similar materials. Hence, bioremediation is a rapidly developing technology with the potential to provide efficient and economic means of removing organic pollutants from contaminated materials. However, bioremediation is currently restricted by a variety of factors. One of the most significant of these is the difficulty of effectively degrading complex mixtures of chemicals as often found at hazardous waste sites. Although preliminary evidence shows that such treatments are potentially effective in situ, additional data on the performance of bioremediation efforts in large scale field trials is also needed. Bioremediation of creosote-contaminated materials is reviewed here by characterizing coal-tar creosote, identifying techniques for assessing the biodegradability of its many chemical constitutents, examining known routes of microbial transformation of these chemicals, and reviewing the performance of previous bioremediation efforts. This approach is developed as a model system to project the potential application of bioremediation to ameliorate environments contaminated by complex mixtures of structurally diverse hazardous chemicals.
Mueller, James G., Peter J. Chapman and P. Hap Pritchard. 1989. Action of a Fluoranthene-Utilizing Bacterial Community on Polycyclic Aromatic Hydrocarbon Components of Creosote. EPA/600/J-89/425. Appl. Environ. Microbiol. 55(12):3085-3090. (ERL,GB 674). (Avail. from NTIS, Springfield, VA: PB90-245721)
Cultures enriched by serial transfer through a mineral salts medium containing
fluoranthene were used to establish a stable, 7-membered bacterial community
from a sandy soil highly contaminated with coal-tar creosote. This community
exhibited an ability to utilize fluoranthene as sole carbon source for growth
as demonstrated by increases in protein concentration and changes in absorption
spectra when grown on fluoranthene in liquid culture. Biotransformation of
other polycyclic aromatic hydrocarbons (PAHs) was verified by demonstrating
their disappearance from an artificial PAH mixture using capillary gas
chromatography. When grown on fluoranthene as sole carbon source and
subsequently exposed to fluoranthene plus 16 additional PAHs typical of those
found in creosote, this community transformed all PAHs present in this defined
mixture. After 3 days of incubation, 13 of the original 17 PAH components were
degraded to levels below the limit of detection (10 ng/L). Continued incubation
resulted in extensive degradation of the remaining 4 compounds. The ability of
this community to utilize a high molecular weight PAH as sole carbon source, in
conjunction with its ability to transform a diverse array of PAHs, suggests
that it may be of value in the bioremediation of environments contaminated with
PAHs such as those impacted by creosote.
Mueller, James G., Peter J. Chapman, Beat O. Blattmann and P. Hap Pritchard. 1990. Utilization of Fluoranthene by Pseudomonas paucimobilis Strain EPA505. In: Gas, Oil, Coal, and Environmental Biotechnology II. EPA/600/D-89/254. Cavit Akin and Jared Smith, Editors. Institute of Gas Technology, Chicago, IL. Pp. 243-253. (ERL,GB 690).
Pseudomonas paucimobilis strain EPA505 was previously purified from a
7-membered bacterial community originally isolated from a creosote-contaminated
soil for its ability to degrade polycyclic aromatic hydrocarbon (PAH)
components of creosote. The unique ability of this organism to utilized
fluoranthene as sole source of carbon and energy for growth in pure culture was
demonstrated by increase in bacterial biomass, changes in UV-absorption,
decrease in aqueous fluoranthene concentration, and the production of
metabolites when fluoranthene was supplied as sole carbon source in liquid
culture. Compounds accumulating in fluoranthene culture medium during growth of
EPA505 have been distinguished by HPLC and UV-absorption properties. Based on
precedents established for bacterial degradation of similar compounds,
speculative pathways are proposed to illustrate the novel biochemistry employed
by strain EPA505 in the utilization of fluoranthene. Whereas utilization of
fluoranthene appears to involve previously undefined variations on established
oxygenation and ring cleavage processes, these findings suggest the potential
of this and other organisms for accelerating the biotransformation of other
environmental pollutants currently considered recalcitrant to microbiological
attack. Hence, strain EPA505 and other organisms similarly isolated for their
ability to degrade fluoranthene and related compounds may prove useful to
remediation efforts employing biological processes.
Mueller, James G., Peter J. Chapman, Beat O. Blattmann and P. Hap Pritchard. 1990. Isolation and Characterization of a Fluoranthene-Utilizing Strain of Pseudomonas paucimobilis. EPA/600/J-90/109. Appl. Environ. Microbiol. 56(4):1079-1086. (ERL,GB 691). (Avail. from NTIS, Springfield, VA: PB90-264698)
A soil bacterium capable of utilizing fluoranthene as the sole source of carbon
and energy for growth was purified from a seven-member bacterial community
previously isolated from a creosote waste site for its ability to degrade
polycyclic aromatic hydrocarbons. By standard bacteriological methods, this
bacterium was characterized taxonomically as a strain of Pseudomonas
paucimobilis and was designated strain EPA505. Utilization of fluoranthene by
strain EPA505 was demonstrated by increase in bacterial biomass, decrease in
aqueous fluoranthene concentration, and transient formation of transformation
products in liquid cultures where fluoranthene was supplied as the sole carbon
source. Resting cells grown in complex medium showed activity toward
anthraquinone, benzo[b]fluorene, biphenyl, chrysene, and pyrene as demonstrated
by the disappearance of parent compounds or changes in their UV absorption
spectra. Fluoranthene-grown resting cells were active against these compounds
as well as 2,3-dimethylnaphthalene, anthracene, fluoranthene, fluorene,
naphthalene, and phenanthrene. These studies demonstrate that organic compounds
not previously reported to serve as growth substrates can be utilized by axenic
cultures of microorganisms. Such organisms may possess novel degradative
systems that are active toward other compounds whose biological degradation has
been limited because of inherent structural considerations or because of low
aqueous solubility.
Mueller, James G., Suzanne E. Lantz, Beat O. Blattmann and Peter J. Chapman. 1991. Bench-Scale Evaluation of Alternative Biological Treatment Processes for the Remediation of Pentachlorophenol- and Creosote-Contaminated Materials: Slurry-Phase Bioremediation. EPA/600/J-91/331. Environ. Sci. Technol. 25(6):1055-1061. (ERL,GB 721). (Avail. from NTIS, Springfield, VA: PB92-129683)
Performance data on slurry-phase bioremediation of pentachlorophenol (PCP)- and
creosote-contaminated sediment and surface soil were generated at the
bench-scale level. Aqueous slurries, containing 0.05% Triton X-100 to
facilitate the soil washing process and to help stabilize the suspensions, were
prepared from sediment and surface soil freshly obtained from the American
Creosote Works Superfund site at Pensacola, Florida. Slurries (1.1 L) were
incubated for 30 days in separate, 1.5 L bioreactors operated in the batch mode
at 28.5°C with continuous mixing (300 rpm). Dissolved oxygen and pH were
maintained at 90% and pH=7.0. Samples removed with time from each reactor were
extracted and analyzed by gas chromatography for PCP and 42 monitored creosote
constitutents to delineate the activity of indigenous microorganisms. Changes
in microbial biomass were also recorded. Excluding PCP, benzo[b]fluoranthene,
benzo[k]fluoranthene and indeno[123-cd]pyrene, slurry-phase bioremediation of
highly contaminated sediment (pH adjusted) resulted in rapid and extensive
biodegradation (3-5 days to biodegrade >50% of targeted compounds) of monitored
constituents. Conversely, microbial activity in surface soil slurries was
slower (14-21 days required to biodegrade greater than 50% of targeted
compounds) and generally confined to the more readily biodegradable,
lower-molecular-weight compounds. These data suggest that slurry-phase
bioremediation strategies can be effectively employed to remediate
creosote-contaminated materials.
Mueller, James G., Suzanne E. Lantz, Beat O. Blattmann and Peter J. Chapman. 1991. Bench-Scale Evaluation of Alternative Biological Treatment Processes for the Remediation of Pentachlorophenol- and Creosote-Contaminated Materials: Solid-Phase Bioremediation. EPA/600/J-91/335. Environ. Sci. Technol. 25(6):1045-1055. (ERL,GB 722). (Avail. from NTIS, Springfield, VA: PB92-129725)
Bench-scale biotreatability studies were performed to evaluate the potential
for using a solid-phase bioremediation process to ameliorate pentachlorophenol
(PCP)- and creosote-contaminated sediment and surface soil present at the
American Creosote Works Superfund site, Pensacola, Florida. Surface soil and
sediment were contaminated with approximately 1 and 7 % (weight basis) organic
pollutants, respectively, but the more recalcitrant creosote constitutents
(i.e., high-molecular-weight polycyclic aromatic hydrocarbons) were
proportionately higher in the surface soil indicative of creosote weathering.
The effects of tilling and fertilization on the rate and extent of
biodegradation of PCP and 42 targeted creosote constituents by indigenous
microflora were monitored by gas chromatographic analysis of organic extracts
of soil and sediment. Changes in microbial populations (total heterotrophs and
the number of phenanthrene-degraders) were also recorded. Specially designed
"landfarming chambers" allowed for the quantitative analysis of targeted
pollutants lost through abiotic processes (i.e., leaching, adsorption,
volatilization). Data corrected for these losses represented an accurate
assessment of biological activity towards creosote and PCP. In general,
solid-phase bioremediation resulted in slow and predictable losses of targeted
pollutants (i.e., low-molecular-weight creosote constituents were more readily
biodegraded than higher-molecular-weight contaminants), and the more
recalcitrant pollutants (e.g., PCP) tended to persist. Biodegradation in
surface soils was stimulated by the addition of inorganic soluble nutrients,
but this treatment had little effect with the more highly contaminated,
alkaline sediments. Performance data from these studies suggest that full-scale
site remediation employing solid-phase bioremediation strategies may not
effectively meet acceptable treatment standards in the time defining these
studies.
Mueller, James G., Douglas P. Middaugh, Suzanne E. Lantz and Peter J. Chapman. 1991. Biodegradation of Creosote and Pentachlorophenol in Contaminated Groundwater: Chemical and Biological Assessment. EPA/600/J-91/328. Appl. Environ. Microbiol. 57(5):1277-1285. (ERL,GB 728). (Avail. from NTIS, Springfield, VA: PB92-129659)
Shake flask studies examined the rate and extent of biodegradation of
pentachlorophenol (PCP) and 42 components of coal-tar creosote present in
contaminated groundwater recovered from the American Creosote Works (ACW)
Superfund site, Pensacola, Florida. The ability of indigenous soil
microorganisms to remove these contaminants from aqueous solutions was
determined by gas chromatographic analysis of organic extracts of biotreated
groundwater. Changes in potential environmental and human health hazards
associated with the biodegradation of this material were determined at
intervals by Microtox® assays and fish toxicity and teratogenicity tests. After
14 days incubation at 30°C, indigenous microorganisms effectively removed 100,
99, 94, 88 and 87% of measured phenolic, lower-molecular-weight polycyclic
aromatic hydrocarbons (PAH), S-heterocycle, N-heterocycle and O-heterocycle
constituents of creosote, respectively. However, only 53% of the
higher-molecular-weight PAHs were degraded; PCP was not removed. Despite the
removal of a majority of the organic contaminants through biotreatment, only a
slight decrease in toxicity and teratogenicity of biotreated groundwater was
observed. These data suggest that toxicity and teratogenicity are associated
with compounds difficult to treat biologically, and that one may not
necessarily rely on indigenous microorganisms to effectively remove these
compounds in a reasonable time span; to this end, alternative or supplemental
approaches may be necessary. Similar measures of toxicity and teratogenicity of
treated material may offer a simple, yet important, guide to bioremediation
effectiveness.
Shields, Malcolm S., Stacy O. Montgomery, Stephen M. Cuskey, Peter J. Chapman and P.H. Pritchard. 1991. Mutants of Pseudomonas cepacia G4 Defective in Catabolism of Aromatic Compounds and Trichloroethylene. EPA/600/J-91/337. Appl. Environ. Microbiol. 57(7):1935-1941. (ERL,GB 730). (Avail. from NTIS, Springfield, VA: PB92-129741)
Pseudomonas cepacia strain G4 possesses a novel pathway of toluene catabolism
that is shown to be responsible for the degradation of trichloroethylene (TCE).
This pathway involves conversion of toluene via o-cresol to 3-methylcatechol.
In order to determine the enzyme of toluene degradation that is responsible for
TCE degradation, chemically induced mutants, blocked in the toluene ortho
-monooxygenase (TOM) pathway of G4, were examined. Mutants of the phenotypic
class designated TOM A- were all defective in their ability to oxidize toluene,
o-cresol, m-cresol, and phenol, suggesting that a single enzyme is responsible
for conversion of these compounds to their hydroxylated products
(3-methylcatechol from toluene, o-cresol, and m-cresol and catechol from
phenol) in the wild type. Mutants of this class did not degrade TCE. Two other
mutant classes blocked in toluene catabolism, TOM B-, which lacked
catechol-2,3-dioxygenase or TOM C-, which lacked 2-hydroxy-6-oxoheptadienoic
acid hydrolase activity, were fully capable of TCE degradation. Therefore, TCE
degradation is directly associated with the monooxygenation capability
responsible for toluene, cresol and phenol hydroxylation.
Resnick, Sol M. and Peter J. Chapman. 1994. Physiological Properties and Substrate Specificity of a Pentachlorophenol-Degrading Pseudomonas Species. EPA/600/J-94/444. Biodegradation. 5(1):47-54. (ERL,GB 750). (Avail. from NTIS, Springfield, VA: PB95-112157)
A bacterial strain capable of utilizing pentachlorophenol (PCP) as sole source
of carbon and energy for growth was isolated from enrichment cultures
containing 100 mg/1 PCP in a mineral salts medium inoculated with contaminated
soil from a lumber treatment waste site. The isolate, designated strain SR3,
was identified as a species of Pseudomonas by virtue of its physiological and
biochemical characteristics. Mineralization of PCP by Pseudomonas sp. strain
SR3 was demonstrated by loss of detectable PCP from growth medium,
stoichiometry of chloride release (5 equivalents of chloride per mole of PCP),
and formation of biomass consistent with the concentration of PCP mineralized.
PCP-induced cells of strain SR3 showed elevated rates of oxygen consumption in
the presence of PCP, and with different chlorinated phenols, with complete
degradation of 2,3,5,6-, 2,3,6-, 2,4,6-, 2,4, and 2,6-chloro-substituted
phenols. Concentration of PCP up to 175 mg/liter supported growth of this
organism, but maximal rates of PCP removal were observed at a PCP concentration
of100 mg/liter. Based on its degradative properties, Pseudomonas sp. strain SR3
appears to have utility in bioremediation of soil and water contaminated with
PCP.
Jeffrey, Wade H., Stephen M. Cuskey, Peter J. Chapman, Sol Resnick and Ronald H. Olsen. 1992. Characterization of Pseudomonas putida Mutants Unable to Catabolize Benzoate: Cloning and Characterization of Pseudomonas Genes Involved in Benzoate Catabolism and Isolation of a Chromosomal DNA Fragment Able to Substitute for xylS in Activation of the TOL Lower-Pathway Promoter. EPA/600/J-92/381. J. Bacteriol. 174(15):4986-4996. (ERL,GB 766). (Avail. from NTIS, Springfield, VA: PB93-121135)
Mutants of Pseudomonas putida mt-2 that are unable to convert benzoate to
catechol were isolated and grouped into two classes: those that did not
initiate attack on benzoate and those that accumulated
3,5-cyclohexadiene-1,2-diol-1-carboxylic acid (benzoate diol). The latter
mutants, represented by strain PP0201, were shown to lack benzoate diol
dehydrogenase (benD) activity. Mutants from the former class were presumed
either to carry lesions in one or more subunit structural genes of benzoate
dioxygenase (benABC) or the regulatory gene (benR) or to contain multiple
mutations. Previous work in this laboratory suggested that benR can substitute
for the TOL plasmid-encoded xylS regulatory gene, which promotes gene
expression from the OP2 region of the lower or meta pathway operon.
Accordingly, structural and regulatory gene mutations were distinguished by the
ability of benzoate-grown mutant strains to induce expression frm OP2 without
xylS by using the TOL plasmid xylE gene (encoding catechol 2,3-dioxygenase) as
a reporter. A cloned 12-kb BamHI chromosomal DNA fragment from the P.
aeruginosa PAO1 chromosome complemented all of the mutations, as shown by
restoration of growth on benzoate minimal medium. Subcloning and deletion
analyses allowed identification of DNA fragments carrying benD, benABC, and the
region possessing xylS substitution activity, benR. Expression of these genes
was examined in a strain devoid of benzoate-utilizing ability, Pseudomonas
fluorescens PFO15. The disappearance of benzoate and the production of catechol
were determined by chromatographic analysis of supernatants from cultures grown
with casamino acids. When P. fluorescens PFO15 was transformed with plasmids
containing only benABCD, no loss of benzoate was observed. When either benR or
xylS was cloned into plasmids compatible with those plasmids containing only
the benABCD regions, benzoate was removed from the medium and catechol was
produced. Regulation of expression of the chromosomal structural genes by benR
and xylS was quantified by benzoate diol dehydrogenase enzyme assays. The
results obtained when xylS was substituted for benR strongly suggest an
isofunctional regulatory mechanism between the TOL plasmid lower-pathway genes
(via the OP2 promoter) and chromosomal benABC. Southern hybridizations
demonstrated that DNA encoding the benzoate dioxygenase structural genes showed
homology to DNA encoding toluate dioxygenase from the TOL plasmid pWW0, but
benR did not show homology to xylS. Evolutionary relationships between the
regulatory systems of chromosomal and plasmid-encoded genes for the catabolism
of benzoate and related compounds are suggested.
Eaton, Richard W. and Peter J. Chapman. 1992. Bacterial Metabolism of Naphthalene: Construction and Use of Recombinant Bacteria to Study the Ring Cleavage of 1,2-Dihydroxynaphthalene and Subsequent Reactions. EPA/600/J-93/059. J. Bacteriol. 174(23):7542-7554. (ERL,GB 795). (Avail. from NTIS, Springfield, VA: PB93-168938)
The reactions involved in the bacterial metabolism of naphthalene to salicylate
have been reinvestigated using recombinant bacteria carrying genes cloned from
the NAH7 plasmid. When intact cells of Pseudomonas aeruginosa PAO1 carrying DNA
fragments encoding the first three enzymes of the pathway were incubated with
naphthalene, they formed products of the dioxygenase-catalyzed ring cleavage of
1,2-dihydroxynaphthalene. These products were separated by chromatography on
Sephadex G-25 and identified by 1H and 13C nuclear magnetic resonance
spectroscopy and gas chromatography-mass spectrometry as
2-hydroxychromene-2-carboxylate (HCCA) and
trans-o-hydroxybenzylidenepyruvate (tHBPA). HCCA was detected as the first
reaction product in these incubation mixtures by its characteristic UV
spectrum, which slowly changed to a spectrum indicative of an equilibrium
mixture of HCCA and tHBPA. Isomerization of either purified product occurred
slowly and spontaneously to give an equilibrium mixture of essentially the same
composition. tHBPA is also formed from HCCA by the action of an isomerase
enzyme encoded by the NAH7 plasmid. The gene encoding this enzyme, nahD, was
cloned on a 1.95 kb KpnI-BglII fragment. Extracts of Escherichia coli JM109
carrying this fragment catalyzed the rapid equilibrium of HCCA and tHBPA.
Metabolism of tHBPA to salicylaldehyde by hydration and aldol cleavage is
catalyzed by a single enzyme encoded by a 1 kb MluI-StuI restriction fragment.
Mechanisms for both the isomerase- and hydratase/aldolase- catalyzed reactions
are proposed. The salicylaldehyde dehydrogenase gene, nahF, was cloned on a
2.75 kb BamHI fragment which also carries the naphthalene dihydrodiol
dehydrogenase gene, nahB. By identifying the enzymes encoded by various clones,
the gene order for the nah operon was shown to be p, A, B, F, C, E, D.
Grifoll, Magdalena, Sergey A. Selifonov and Peter J. Chapman. 1995. Transformation of Substituted Fluorenes and Fluorene Analogs by Pseudomonas sp. Strain F274. EPA/600/J-02/250. Appl. Environ. Microbiol. 61(9):3490-3493. (ERL,GB 908).
Pseudomonas sp. F274, previously shown to catabolize fluorene via fluorenone
and its angular dioxygenation, 2',3'-dihydroxy-2-carboxybiphenyl, phthalate and
protocatechuate, was examined for its abililty to transform substituted
fluorenes and S- and N-heterocyclic analogues. Halogen- and methyl-substituted
fluorenes were metabolized to the correspondingly substituted phthalates via
attack on the unsubstituted ring. In the case of 1-methylfluorene, initial
oxidation of the methyl group to carboxyl prevented all other transformations
but 9-monooxygenation. The strain also oxidized S-heteroatoms and benzylic
methylenic groups of fluorene analogues. No angular dioxygenation of S- and
N-heterocycles was observed.
Grifoll, Magdalena, Sergey A. Selifonov, Charylene V. Gatlin and Peter J. Chapman. 1995. Actions of a Versatile Fluorene-Degrading Bacterial Isolate on Polycyclic Aromatic Compounds. Appl. Environ. Microbiol. 61(10):3711-3723. (ERL,GB 909).
Pseudomonas cepacia F297 showed the ability to grow with fluorene as a sole
source of carbon and energy; its growth yield corresponded to assimulation of
about 40% of fluorene carbon. Accumulation of a ring meta-cleavage product
during growth and identification of 1-indanone in growth media and washed cell
suspensions suggest that strain F297 metabolizes fluorene by mechanisms
analogous to those of naphthalene degradation. In addition to fluorene, strain
F297 utilized for growth a wide variety of polycyclic aromatic compounds
(PACs), including naphthalene, 2,3-dimethylnaphthalene, phenanthrene,
anthracene, and dibenzothiophene. Fluorene-induced cells of the strain also
transformed 2,6-dimethylnaphthalene, biphenyl, dibenzofuran, acenaphthene and
acenaphthylene. Identification of products formed from those substrates (GC-MS)
in washed cell suspensions, indicates that Pseudomonas cepacia F297 carries out
the following reactions: (i) aromatic ring oxidation and cleavage, apparently
using the pyruvate released for growth, (ii) methyl group oxidations, (iii)
methylenic oxidations, and (iv) S-oxidations of aromatic sulfur heterocycles.
Strain F297 grew with a creosote-PAC mixture, producing almost complete removal
of all aromatic compounds containing 2-3 rings in 14 days, as demonstrated by
GC analysis of the remaining PACs recovered from cultures. Identification of
key chemicals confirmed that not only are certain compounds depleted, but also
the anticipated reaction products are found.
Eaton, Richard W. and Peter J. Chapman. 1995. Formation of Indigo and Related Compounds from Indolecarboxylic Acids by Aromatic Acid-Degrading Bacteria: Chromogenic Reactions for Cloning Genes Encoding Dioxygenases That Act on Aromatic Acids. EPA/600/J-96/165. J. Bacteriol. 177(23):6983-6988. (ERL,GB 920).
The p-cumate-degrading strain Pseudomonas putida F1 and the m- and p
-toluate-degrading strain P. putida mt-2 transform indole-2-carboxylate and
indole-3-carboxylate to colored products identified here as indigo, indirubin,
and isatin. A mechanism by which these products could be formed spontaneously
following dioxy-genase-catalyzed dihydroxylation of the indolecarboxylates is
proposed. Indolecarboxylates were employed as chromogenic substrates for
identifying recombinant bacteria carrying genes encoding p-cumate dioxygenase
and toluate dioxygenase. Dioxygenase gene carrying bacteria could be readily
distinguished as dark green-blue colonies among other colorless recombinant
Escherichia coli colonies on selective agar plates containing either
indole-2-carboxylate or indole-3-carboxylate.
Selifonov, Sergey A., Magdalena Grifoll, Richard W. Eaton and Peter J. Chapman. 1996. Oxidation of Naphthenoaromatic and Methyl-Substituted Aromatic Compounds by Naphthalene 1,2-Dioxygenase. EPA/600/J-96/161. Appl. Environ. Microbiol. 62(2):507-514. (ERL,GB 930).
Oxidation of acenaphthene, acenaphthylene and fluorene was examined with
recombinant strain Pseudomonas aeruginosa PA01 (pRE695), expressing naphthalene
dioxygenase genes cloned from plasmid NAH7. Acenaphthene underwent
monooxygenation to 1-acenaphthenol with subsequent conversion to
1-acenaphthenone, cis- and trans- acenaphthene-1,2-diols, while acenaphthylene
was dioxygenated to give cis-acenaphthene-1,2-diol. Non-specific dehydrogenase
activities present in the host strain led to the conversion of both
acenaphthene-1,2-diols to 1,2-acenaphthoquinone. The latter was spontaneously
oxidized to naphthalene-1,8-dicarboxylic acid. No aromatic ring dioxygenation
products were detected from acenaphthene and acenaphthylene. Mixed
monooxygenase and dioxygenase actions of naphthalene dioxygenase on fluorene
yielded products of benzylic 9-monooxygenation, aromatic ring dioxygenation, or
both. Action of naphthalene dioxygenase on a variety of methyl-substituted
aromatic compounds, including 1,2,4-trimethylbenzene and isomers of
dimethylnaphthalene, resulted in formation of benzylic alcohols, methyl group
monooxygenation products, which were subsequently converted to the
corresponding carboxylic acids by dehydrogenase(s) in the host strain. Benzylic
monooxygenation of methyl groups was strongly predominant over aromatic ring
dioxygenation and essentially non-specific with respect to the substitution
pattern of the aromatic substrates. In addition to monooxygenating benzylic
methyl and methylene groups, naphthalene dioxygenase behaved as a
sulfoxygenase, catalyzing monooxygenation of the sulfur heteroatom of
3-methylbenzothiophene.
Mueller, James G., Suzanne E. Lantz, Beat O. Blattmann, Douglas P. Middaugh and Peter J. Chapman. 1990. Alternative Biological Treatment Processes for Remediation of Creosote-Contaminated Materials: Bench-Scale Treatability Studies. EPA/600/9-90/049. U.S. Environmental Protection Agency, Environmental Research Laboratory, Gulf Breeze, FL. 74 p. (Avail. from NTIS, Springfield, VA: PB91-179085)
Bench-scale biotreatability studies were performed to determine the most
effective of two bioremediation application strategies to ameliorate creosote-
and pentachlorophenol (PCP)-contaminated soils present at the American Creosote
Works Superfund site, Pensacola, Florida: solid-phase bioremediation or
slurry-phase bioremediation. When indigenous microorganisms were employed as
biocatalysts, solid-phase bioremediation was slow and ineffective (8-12 weeks
required to biodegrade greater than 50% of resident organics). Biodegradation
was limited to lower-molecular-weight constituents rather than the more
hazardous, higher-molecular-weight (HMW) compounds); PCP and HWM polycyclic
aromatic hydrocarbons (PAHs) containing 4 or more fused rings resisted
biological attach. Moreover, supplementation with aqueous solution of inorganic
nutrients had little effect on the overall effectiveness of the treatment
strategy. Alternatively, slurry-phase bioremediation was much more effective:
greater that 50% of targeted organics were biodegraded in 14 days. Again,
however, more persistent contaminants, such as PCP and HMW PAHs, were not
extensively degraded when subjected to the action of indigenous microorganisms.
Shields, Malcolm S., Stacy O. Montgomery, Peter J. Chapman, Stephen M. Cuskey and Parmely H. Pritchard. 1990. Involvement of a Toluene Degradative Pathway in the Biodegradation of Trichloroethylene by Pseudomonas cepacia Strain G4. In: Bioremediation of Hazardous Wastes. EPA/600/9-90/041. U.S. Environmental Protection Agency, Office of Research and Development, Biosystems Technology Development Program, Washington, DC. Pp. 49-52. (ERL,GB X721).
The oxidation of trichloroethylene (TCE) by bacteria possessing enzymes that act on toluene is now established for three species: P. cepacia, P. putida, and P. mendocina. In all cases an oxygenase acting directly on the aromatic ring of toluene is responsible for an attack on the alternate substrate TCE. P. cepacia, strain G4, was shown to catabolize toluene by a novel pathway. HPLC and GC/MC analysis indicated toluene hydroxylations at first the ortho and then meta positions to sequentially form o-cresol and 3-methylcatechol. Analysis of pathway intermediates (by GC/MS) formed in a defined atmosphere of 18O2 and 16O2 confirmed the sequential nature of this monooxygenation activity and the source of the oxygen as O2. The generation of several mutants unable to metabolize a variety of related aromatic compounds--toluene, phenol, cresol, catechol and hydroxymuconic semialdehyde (HMS)--revealed that the only mutants that also suffered loss of TCE-metabolizing ability were those that coincidentally lost the ability to hydroxylate toluene, phenol, o- or m-cresol. The principle of toluene ring oxygenation has led to the identification of strains of Nocardia, Alcaligenes, and Acinetobacter as capable of TCE degradation.
Eaton, Richard W., Peter J. Chapman and P. Hap Pritchard. 1990. Degradation of Heterocyclics. In: Bioremediation of Hazardous Wastes. EPA/600/9-90/041. U.S. Environmental Protection Agency, Office of Research and Development, Biosystems Technology Development Program, Washington, DC. Pp. 46-48. (ERL,GB X722).
Many contaminants of ground water are heterocyclic compounds that are potentially hazardous to human health and to the environment. These classes of compounds along with many others are formed as combustion products in the generation of synthetic fuels from fossil fuels. Groundwater contamination by these chemicals has been shown in the vicinity of coal gasification plants, oil shale-retorting facilities, and at wood treatment facilities where coal-tar derived creosote has been used. To develop biological approaches to treat sites contaminated with these and other chemicals, the microbiology and biochemistry of their biodegradation must be understood. This research is designed to provide such insights.
Mueller, James G., Peter J. Chapman, Parmely H. Pritchard, Ron Thomas, Ellis L. Kline and Suzanne E. Lantz. 1990. Development of a Sequential Treatment System for Creosote-Contaminated Soil and Water: Bench Studies. In: Bioremediation of Hazardous Wastes. EPA/600/9-90/041. U.S. Environmental Protection Agency, Office of Research and Development, Biosystems Technology Development Program, Washington, DC. Pp. 42-43. (ERL,GB X723).
A triphasic, sequential treatment system for the potential remediation of environments contaminated by mixtures of hazardous wastes has been co-developed by Southern BioProducts (SBP), Incorporated, Atlanta, GA and the Gulf Breeze Environmental Research Laboratory, Gulf Breeze, FL. This approach integrates established soil-washing technology (phase I) with dewatering and chemical fractionation through a patented filtration process (phase II). Together, these steps reduce the volume of material to be subsequently treated from 100 to 100,000 fold. Volume reduction of this magnitude facilitates phase III of this system: biodegradation of susceptible pollutants employing EPA/SBP-patented bacteria capable of utilizing high-molecular-weight creosote constituents as growth substrates. The efficacy of this treatment system on creosote-contaminated soil and water is currently being evaluated at the bench-scale level.
Mueller, James G., Peter J. Chapman and Parmely H. Pritchard. 1990. Aerobic Biodegradation of PAHs. In: Bioremediation of Hazardous Wastes. EPA/600/9-90/041. U.S. Environmental Protection Agency, Office of Research and Development, Biosystems Technology Development Program, Washington, DC. Pp. 23-24. (ERL,GB X725).
Biological degradation represents the major route through which polycyclic aromatic hydrocarbons (PAHs) and other organic chemicals are removed from contaminated environments. In most cases, lower molecular weight PAHs containing two or three rings are readily degraded biologically. Conversely, higher-molecular-weight PAHs are considered resistant to biological action and tend to persist in contaminated environments. Unfortunately, higher molecular weight PAHs represent the greatest risk to public and environmental health. For bioremediation to be considered as an acceptable remedial action alternative for PAH-contaminated sites (i.e., creosote waste sites, coal gasification sites, petroleum refineries), biotreatment processes must be capable of destroying these chemicals. The isolation of microbial systems capable of utilizing high-molecular-weight PAHs (e.g., fluoranthene) as sole sources of carbon and energy for growth partially addresses this challenge.
Folsom, Brian R., Peter J. Chapman and Parmely H. Pritchard. 1990. Performance of a Recirculating Bioreactor for the Degradation of TCE. In: Bioremediation of Hazardous Wastes. EPA/600/9-90/041. U.S. Environmental Protection Agency, Office of Research and Development, Biosystems Technology Development Program, Washington, DC. Pp. 6-8. (ERL,GB X726).
Of the volatile organic chemicals found as common ground-water contaminants, trichloroethylene (TCE) has received significant attention; and as a result, a number of bacterial systems with the ability to degrade TCE by cometabolism are now recognized. In these systems, TCE is degraded by bacterial enzymes that are typically expressed following induction with other chemicals. Previously, one of these TCE-degrading organisms was isolated by investigators at the Gulf Breeze Environmental Research Laboratory. This organism has subsequently been indentified as Pseudomonas cepacia strain G4 and requires either toluene, o-cresol, m-cresol, or phenol for induction of TCE degradative enzymes. A novel toluene degradative pathway involving sequential hydroxylation of toluene at ortho and meta positions to form 3-methylcatechol has been characterized. TCE is completely degraded to CO2, Cl, and unidentified, nonvolatile products by this organism.
Morris, Pamela J., Michael E. Shelton and Peter J. Chapman. 1995. Co-Contaminated Sites: Biodegradation of Fossil Fuels in the Presence of PCBs. In: Bioremediation of Recalcitrant Organics. Robert E. Hinchee, Daniel B. Anderson and Ronald E. Hoeppel, Editors. Battelle Press, Columbus, OH. Pp. 123-130. (ERL,GB X823).
Polychlorinated biphenyl (PCB)-contaminated sites are often co-contaminated with fossil fuels making biodegration more difficult. Our current studies examine biodegradation of the fossil fuel components of two PCB-contaminated sites: (1) a former racing Drag Strip soil contaminated with Aroclor 1242 and (2) a sediment from Silver Lake contaminated with Aroclor 1260. The sandy surface soil at the Drag Strip site contains 1.9% organic carbon and 1.5% fossil fuel component. Analysis of the solvent-extractable organic fraction, by alumina column chromatography, shows the distribution of organics to be 91.2% hydrocarbons, 7.8% polars, and 1.1% asphaltenes. This oil is extremely weathered and contains few readily biodegradable components. Enrichments have yielded undefined mixed cultures of bacteria capable of extensive degradation of components of both the Drag Strip and Silver Lake site materials. One culture, enriched from a creosote-contaminated soil adjacent to a utility pole, transformed approximately 28% and 37% (by weight) of the Drag Strip and Silver Lake oils, respectively. While the presence of fossil fuels has been shown to inhibit aerobic PCB degradation, our studies show that the presence of PCBs negatively impacts fossil fuel biodegradation. continuing studies will examine the nature of PCB inhibition of fossil fuel biodegradation.
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