Preliminary Exploration of Molecular Methods for the Specific Detection and Differentiation of the Bdellovibrios

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Abstract: 9615515, PI-Williams: Bdellovibrios are procaryotic, parasitic predators which prey on susceptible gram-negative bacilli. Based on the observation that bdellovibrios reduce the population of its prey in laboratory vessels, a logical question is, does the predatory bacterium play some role in the control of microbial populations in nature? Adequate experimentation to resolve this question has not been reported. This is largely due to the limitation in technology to detect and monitor the predator in the environment. The only method currently available for the specific detection of bdellovibrios is the culture technique. As with nearly all cultural methods this technique has substantial limitations. Serological and molecular probes and polymerase chain reaction (PCR) primers which allow for the specific detection of a number of bacterial species have been constructed. However such probes have not been developed for the bdellovibrios. A probe for the bdellovibrios willould allow studies which monitor the numbers of these predators in the environment. The ability to detect those which are parasitic within host cells would allow an estimation of the number of infected cells in the environment and hence define the impact of bdellovibrios on populations of susceptible bacteria. The aim of this researchproposal is to develop a genetic probe and PCR primers which can be used for the specific detection of bdellovibrios in aquatic environments. Initial efforts will be made to identify unique sequences among known nucleic acid sequences for bdellovibrios and to test these as probes and as PCR primers. The objectives and experimental approaches to developing a probe specific for the bdellovibrio group or subgroups of these organisms are described below. 1. Identification and synthesis of specific DNA oligonucleotide sequences for use as detection probes and PCR primers; identification of candidate genomic restriction fragment sequences for use as DNA probes and their production by cloning and/or PCR synthesis; identification of candidate rRNA sequences and construction of rRNA probes. 2. To examine the use and sensitivity of PCR to detect bdellovibrios in laboratory microcosms. 3. To examine the use of genus and species specific probes to confirm the identification of plaques (colonies) of bdellovibrios recovered on culture medium.