Visualizing bacterial cells and describing their responses to the environment are difficult tasks. Their small size is the chief reason for the difficulty, which means that we must often use many millions of cells in a sample in order to determine what the average response of the bacteria is. However, an average response can sometimes mask important events in bacterial physiology, which means that our understanding of these organisms will suffer. We have used a variety of instruments to visualize bacterial cells, all of which tell us something different about the sample. We use a fluorescence activated cell sorter to sort cells based on the fluorescence provided by bioreporter genes, and these can be used to select for particular genetic mutations. Cells can be visualized by epifluorescent microscopy, and sensitive photodetectors can be added that allow us to find a single bacterial cell that is fluorescent or bioluminescent. We have also used standard photomultipliers to examine cell aggregates as field bioreporter microorganisms. Examples of each of these instruments show how our understanding of bacterial physiology has changed with the technology.
Oak Ridge National Laboratory and Quantum Magnetics, Inc. are exploring novel landmine detection technologies. Technologies considered here include bioreporter bacteria, swept acoustic resonance, nuclear quadrupole resonance (NQR), and semiotic data fusion. Bioreporter bacteria look promising for third-world humanitarian applications; they are inexpensive, and deployment does not require high-tech methods. Swept acoustic resonance may be a useful adjunct to magnetometers in humanitarian demining. For military demining, NQR is a promising method for detecting explosive substances; of 50,000 substances that have been tested, one has an NQR signature that can be mistaken for RDX or TNT. For both military and commercial demining, sensor fusion entails two daunting tasks, identifying fusible features in both present-day and emerging technologies, and devising a fusion algorithm that runs in real-time on cheap hardware. Preliminary research in these areas is encouraging. A bioreporter bacterium for TNT detection is under development. Investigation has just started in swept acoustic resonance as an approach to a cheap mine detector for humanitarian use. Real-time wavelet processing appears to be a key to extending NQR bomb detection into mine detection, including TNT-based mines. Recent discoveries in semiotics may be the breakthrough that will lead to a robust fused detection scheme.
Bioremediation is the use of living systems, usually microorganisms, to treat a quantity of soil or water for the presence of hazardous wastes. Bioremediation has many advantages over other remediation approaches, including cost savings, versatility, and the ability to treat the wastes in situ. In order to study the processes of microbial bioremediation, we have constructed bacterial strains that incorporate genetically engineered bioreporter genes. These bioreporter genes allow the bacteria to be detected during in situ processes, as manifested by their ability to bioluminesce or to fluoresce. This bioreporter microorganisms are described, along with the technology for detecting them and the projects which are benefiting from their application.
This work involves the design and development of species specific gene (DNA) probes designed to detect Pseudomonas species. A non radioactive detection system for DNA was designed to detect the presence of these microbes in environmental samples for biomonitoring applications. The details of the methodology and their applicability for various environmental analyses is discussed below.
A remote detection system for in-situ monitoring of the bioluminescence produced by a microbial "reporter" strain grown in rotary film bioreactors has been developed. Two systems were developed: one using a low-temperature photomultiplier (PMT), and one using an avalanche photodiode. The photodiode system was preferred for its sensitivity and portability. Testing was performed on a recombinant constitutive light producing strain. The results indicate that the optical monitor is sensitive enough for direct microbial detection for hazardous waste application.
Conference Committee Involvement (1)
Advanced Environmental, Chemical and Biological Sensing Technologies