The survival of genetically engineered Erwinia carotovora L-864, with a kanamycin resistance gene inserted in its chromosome, was monitored in the water and sediment of aquatic microcosms. The density of genetically engineered and wild-type E. carotovora strains declined at the same rate, falling in 32 days below the level of detection by viable counts. We examined the impact of the addition of genetically engineered and wild-type strains on indigenous bacteria belonging to specific functional groups important in nutrient cycling. For up to 16 days, the densities of total and proteolytic bacteria were significantly higher (P < 0.05) in microcosms inoculated with genetically engineered or wild-type E. carotovora, but by 32 days after inoculation, they had decreased to densities similar to those in control microcosms. Inoculation of genetically engineered or wild-type E. carotovora had no apparent effect on the density of amylolytic and pectolytic bacteria in water and sediment. Genetically engineered and wild-type E. carotovora did not have significantly different effects on the densities of specific functional groups of indigenous bacteria (P > 0.05).

An inexpensive, quantitative, and sensitive technique was developed for detection of genetically engineered Erwinia carotovora in soil samples. Enrichment media, antibiotic resistance, and most probable number (MPN) analysis were used to enumerate as few as 1 to 10 target cells/10 g soil. The MPN technique recovered significantly higher cell densities than plating; however, densities estimated by the two techniques were strongly correlated. After inoculation of soil microcosms with genetically engineered E. carotovora, a decline rate of 1.2 log units/g soil/10 days and then subsequent disappearance was observed using the MPN technique.

Genetically engineeredErwinia carotovora persisted significantly longer in thermally perturbed microcosms (35 days) than in nonstressed microcosms (5 days). Decreased pressure of competitors and predators and increased nutrient availability were examined as the most probable reasons for greater vulnerability of perturbed microcosms to colonization by genetically engineered microorganisms (GEMs). Indigenous bacteria that competed with GEMs for the same nutrient sources (protein, cellulose, pectate) were present immediately after perturbation in densities one to two orders of magnitude lower than in unperturbed microcosms, but their populations increased to densities significantly higher than in unperturbed microscosms 10 to 15 days after inoculation. Predators of bacteria (protozoans, cladocerans, nematodes, and rotifers) were present during the experiment in unperturbed microcosms, while dense populations of bacteriovorous nanoflagellates developed in perturbed microcosms. Preemptive inoculation of perturbed microcosms with GEMs did not have a longlasting effect on the recovery of total, proteolytic, cellulolytic, and pectolytic bacteria in perturbed microscosms, indicating the absence of competitive exclusion.

No abstract

Abstract-The release of genetically engineered organisms into the environment has raised concerns about their potential ecological impact. In this study, genetically engineered Erwinia carotovora strains expressing varying levels of reduced phytopathogenicity and wildtype E. carotovora strains were used in aquatic and soil microcosms to assess the survival, intraspecific competition, and effects upon specific groups of indigenous bacteria. In aquatic microcosms, the densities of Erwinia genetically engineered organisms (GEMs) and wildtype strains declined and fell below the detectable limit of plate counts 15 d after the microcosms were inoculated. In aquatic microcosms, engineered E. carotovora did not exhibit a competitive advantage over the wildtype. The effect of engineered and wildtype E. carotovora on densities of total and selected bacteria was not significantly different. Treatment with engineered E. carotovora did not change biomass values of the receiving community but did cause a transitory increase in metabolic activity. In aquatic microcosms, the inability of genetically engineered E. carotovora to persist, displace resident species, or affect the metabolic activity of aquatic communities indicates the low risk of adverse ecological consequences in aquatic ecosystems. Unlike previous investigations involving soil microcosms, densities of both the genetically engineered and wildtype E. carotovora remained at detectable levels over 60 d in both agricultural clay and forest loam soils. The type of soil significantly affected the survival of the GEM and the wildtype. The sorptive properties of clay particles, as well as low concentrations of soil nutrients and organic matter, may have contributed to the unexpected patterns of GEM and wildtype survival.