Drs. George Lacy and John Cairns of Virginia Polytechnic Institute and State University request funds to determine (1) if recombinant DNA may be dispersed into or from other bacteria in plants as well as in selected microcosm habitats, (2) if genetically engineered microorganisms (GEMs) survive and/or colonize these habitats, and (3) if the presence of genetically engineered microorganisms (GEMs) have an effect upon bacterial community structure. The GEMs to be used are constructed from the pathogenic bacterium Erwinia carotovora, which causes soft rot or many plants, has distinct pthogenic and resident phases on or in living plants, and survives transiently in, but does not colonize, soil or aquatic habitats. These objectives will test the hypotheses that (1) recombinant DNA can be dispersed from GEMs to native bacteria, (2) that GEMs survive the habitats into which they are placed, even if at low concentrations, and (3) that GEMs will compete with and displace native species, thus producing alterations in normal community structure. The frequency for dispersal of recombinant DNA from the GEMs and its integration in the genomes of other microbes will be examined from three points of view: (1) in vitro studies will determine the frequency for mobilization of recombinant DNA conjugation and transformation under optimal laboratory conditions, (2) in situ microcosm studies will determine detectable frequencies for genetic mobilization by any genetic mechanism, and (3) in plants studies will determine if transfer may occur within diseased plants. In these experiments, the effect of genetic similarity of DNA recipients will be followed using taxonomically less related organisms (Bacillus and Pseudomonas spp.) isolated from the microcosms, related organisms (Erwinia and Klebsiella spp.), and closely related organisms (strains of E. carotovora EC14 from which the GEMs are derived. Effects upon bacterial community structure will be measured in microcosms representing soil and aquatic habitats into which the pathogen is released naturally in high to low populations by decomposition of diseased plants. Estimates of survivorship and/or colonization by the introduced GEMs will be determined by comparing the populations reached by the GEMs with those reached by the wildtype pathogen and total bacterial and actinomycetal populations. Populations of specific genera will be monitored, including Bacillus, Clostridium, Streptomyces, and fluorescent Pseudomonas spp. by viable count procedures. This area of research in microbial ecology is highly topical and needed at the national level. There is a great paucity of data in these areas and both industry and regulatory agencies are being inhibited in progress with GEMs because of the lack of fundamental information. The research will be funded jointly with the EPA. The investigators and their institution are highly regarded and suited for this research. The Program recommends funding.
