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Title: Genetic Analysis of Stress Responses in Soil Bacteria for Enhanced Bioremediation of Mixed Contaminants

Abstract

In order to realize the full potential of bioremediation, an understanding of microbial community and individual bacterial responses to the stresses encountered at contaminated sites is needed. Knowledge about genetic responses of soil and subsurface bacteria to environmental stresses, which include low nutrients, low oxygen, and mixed pollutants, will allow extrapolation of basic principles to field applications, either using indigenous bacteria or genetically engineered microorganisms. Defining bacterial responses to those stresses presents an opportunity for improving bioremediation strategies, both with indigenous populations and genetically-engineered microbes, and should contribute to environmental management and restoration actions that would reduce the cost and time required to achieve OEM's clean up goals. Stress-inducible genes identified in this project can be used as molecular probes for monitoring performance of indigenous bacteria as well as the effectiveness of bioremediation strategies being employed. Knowledge of survival and catabolic plasmid stability of indigenous bacteria will be needed for devising the most effective bioremediation strategy. In addition, stress-inducible regulatory elements identified in this project will be useful for creating genetically-engineered microorganisms which are able to degrade hazardous wastes under stress conditions at contaminated sites. One of the model organisms, Deinococcus radiodurans, is a stress-resistant bacterium. Thus, in addition tomore » serving as a model for gene regulation in Gram-positive organisms, it may have specific application at aerobic DOE sites where combinations of contaminants produce a particularly stressful environment. Similarly, the use of Sphingomonas F199, isolated from a depth of 407 m at the Savannah River site (Fredrickson et al., 1991), may have relevance to deep subsurface bioremediation applications, where indigenous or engineered microorganisms adapted to the that environment are needed. In addition, F199 contains aromatic oxygenases that are relevant to degradation of contaminants at that site and is representative of a large class of similar organisms from Savannah River Identification of the genes responsive to different stresses encountered at contaminated sites will provide a basic understanding of stress responses in soil bacteria and can lead to improved strategies for bioremediation. Enhanced in situ removal of hazardous wastes by stimulating growth of indigenous bacteria with nutrients or electron acceptors such as oxygen has been demonstrated. However, how much and how often to apply these supplements has largely been determined empirically. As a result, a controlled, reproducible, and properly managed degradation of pollutants in the environment is difficult to achieve. Genes inducible by low nutrient and low oxygen conditions can serve as markers for determining the minimal amount of supplements needed. The disappearance and reappearance of such stress responses will determine how much and when nutrients and oxygen are needed to be applied or reapplied. Similar applications of stress-inducible markers are already being applied in bacterial cultures in solution (Selifonova and Eaton, 1996). Stress responses induced by pollutants also have potential use as a biological index for the performance of indigenous bacteria during bioremediation as well as a microbiological risk assessment index for environmental pollutants. For instance, measurement of the stress responses of contaminant-degrading microorganisms would provide information complementary to measurement of enzymatic activity. This more complete picture of the physiological state of the desired organisms can be used to predict their performance. Finally, prior knowledge of the stress responses of competing bacteria could be used to predict their environmental competitiveness. Promoters from stress inducible genes will facilitate the construction of genetically engineered microorganisms in which the expression of the catabolic genes is uncoupled from both microbial growth and the utilization of the pollutant as the carbon source. The application of genetically engineered organisms in bioremediation requires the design of gene expression systems that function under environmental conditions and are cost effective. The promoter, the genetic regulatory element that directs the use of the gene, plays the central role in gene expression systems. The ideal promoter for environmental applications should possess two qualities: (1) it does not require the addition of exogenous compounds for activation, and (2) it is active under nutrient-limited conditions and not dependent on cell growth for activity. Promoters that are expressed constitutively meet the first quality. However, such promoters usually require active cell growth for expression and thus incur the increased cost of constant nutrient addition.« less

Authors:
Publication Date:
Research Org.:
Pacific Northwest National Lab., Richland, WA (US)
Sponsoring Org.:
USDOE Office of Environmental Management (EM) (US)
OSTI Identifier:
827357
Report Number(s):
EMSP-55031
R&D Project: EMSP 55031; TRN: US200425%%617
Resource Type:
Technical Report
Resource Relation:
Other Information: PBD: 31 Dec 2000
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 54 ENVIRONMENTAL SCIENCES; 63 RADIATION, THERMAL, AND OTHER ENVIRONMENTAL POLLUTANT EFFECTS ON LIVING ORGANISMS AND BIOLOGICAL MATERIALS; BACTERIA; BINDING ENERGY; BIOREMEDIATION; CARBON SOURCES; GENE REGULATION; GENETICS; MICROORGANISMS; OXYGENASES; RISK ASSESSMENT; SAVANNAH RIVER; SOILS; VALENCE

Citation Formats

Wong, Kwong-Kwok. Genetic Analysis of Stress Responses in Soil Bacteria for Enhanced Bioremediation of Mixed Contaminants. United States: N. p., 2000. Web. doi:10.2172/827357.
Wong, Kwong-Kwok. Genetic Analysis of Stress Responses in Soil Bacteria for Enhanced Bioremediation of Mixed Contaminants. United States. doi:10.2172/827357.
Wong, Kwong-Kwok. Sun . "Genetic Analysis of Stress Responses in Soil Bacteria for Enhanced Bioremediation of Mixed Contaminants". United States. doi:10.2172/827357. https://www.osti.gov/servlets/purl/827357.
@article{osti_827357,
title = {Genetic Analysis of Stress Responses in Soil Bacteria for Enhanced Bioremediation of Mixed Contaminants},
author = {Wong, Kwong-Kwok},
abstractNote = {In order to realize the full potential of bioremediation, an understanding of microbial community and individual bacterial responses to the stresses encountered at contaminated sites is needed. Knowledge about genetic responses of soil and subsurface bacteria to environmental stresses, which include low nutrients, low oxygen, and mixed pollutants, will allow extrapolation of basic principles to field applications, either using indigenous bacteria or genetically engineered microorganisms. Defining bacterial responses to those stresses presents an opportunity for improving bioremediation strategies, both with indigenous populations and genetically-engineered microbes, and should contribute to environmental management and restoration actions that would reduce the cost and time required to achieve OEM's clean up goals. Stress-inducible genes identified in this project can be used as molecular probes for monitoring performance of indigenous bacteria as well as the effectiveness of bioremediation strategies being employed. Knowledge of survival and catabolic plasmid stability of indigenous bacteria will be needed for devising the most effective bioremediation strategy. In addition, stress-inducible regulatory elements identified in this project will be useful for creating genetically-engineered microorganisms which are able to degrade hazardous wastes under stress conditions at contaminated sites. One of the model organisms, Deinococcus radiodurans, is a stress-resistant bacterium. Thus, in addition to serving as a model for gene regulation in Gram-positive organisms, it may have specific application at aerobic DOE sites where combinations of contaminants produce a particularly stressful environment. Similarly, the use of Sphingomonas F199, isolated from a depth of 407 m at the Savannah River site (Fredrickson et al., 1991), may have relevance to deep subsurface bioremediation applications, where indigenous or engineered microorganisms adapted to the that environment are needed. In addition, F199 contains aromatic oxygenases that are relevant to degradation of contaminants at that site and is representative of a large class of similar organisms from Savannah River Identification of the genes responsive to different stresses encountered at contaminated sites will provide a basic understanding of stress responses in soil bacteria and can lead to improved strategies for bioremediation. Enhanced in situ removal of hazardous wastes by stimulating growth of indigenous bacteria with nutrients or electron acceptors such as oxygen has been demonstrated. However, how much and how often to apply these supplements has largely been determined empirically. As a result, a controlled, reproducible, and properly managed degradation of pollutants in the environment is difficult to achieve. Genes inducible by low nutrient and low oxygen conditions can serve as markers for determining the minimal amount of supplements needed. The disappearance and reappearance of such stress responses will determine how much and when nutrients and oxygen are needed to be applied or reapplied. Similar applications of stress-inducible markers are already being applied in bacterial cultures in solution (Selifonova and Eaton, 1996). Stress responses induced by pollutants also have potential use as a biological index for the performance of indigenous bacteria during bioremediation as well as a microbiological risk assessment index for environmental pollutants. For instance, measurement of the stress responses of contaminant-degrading microorganisms would provide information complementary to measurement of enzymatic activity. This more complete picture of the physiological state of the desired organisms can be used to predict their performance. Finally, prior knowledge of the stress responses of competing bacteria could be used to predict their environmental competitiveness. Promoters from stress inducible genes will facilitate the construction of genetically engineered microorganisms in which the expression of the catabolic genes is uncoupled from both microbial growth and the utilization of the pollutant as the carbon source. The application of genetically engineered organisms in bioremediation requires the design of gene expression systems that function under environmental conditions and are cost effective. The promoter, the genetic regulatory element that directs the use of the gene, plays the central role in gene expression systems. The ideal promoter for environmental applications should possess two qualities: (1) it does not require the addition of exogenous compounds for activation, and (2) it is active under nutrient-limited conditions and not dependent on cell growth for activity. Promoters that are expressed constitutively meet the first quality. However, such promoters usually require active cell growth for expression and thus incur the increased cost of constant nutrient addition.},
doi = {10.2172/827357},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2000},
month = {12}
}