survival and spread of genetically manipulated organisms (GMOs) in river
sediments, soils and agricultural environments
Genetically engineered micro-organisms (GEMs) are likely to be accidentally
released to, or deliberately used in, aquatic or terrestrial habitats.
Potential environmental uses of GEMs include waste treatment, detoxification,
control of agricultural pests and plant growth stimulation. Little is
currently known about gene flow or inoculant survival in natural populations
of bacteria so the fate of GEMs or their DNA is unknown. Studies on plasmid
transfer and inoculant survival in soil and water are therefore necessary.
This research aimed to study the transfer of natural plasmids, isolated
from soil and water, and of recombinant plasmid vectors between species
of Escherichia coli, Pseudomonas and Alcaligenes.
Heavy metal resistance genes and lactose-utilisation genes were used as
Approach and methodology
Plasmids are self-replicating units of DNA. We investigated the roles
of the key genetical and environmental factors that might limit their
transfer. Laboratory microcosms were designed to investigate their transfer
and survival in water and soil. The survival and mobility of genetically
marked plant beneficial pseudomonads was studied in soil microcosms and
lysimeters, to simulate an agricultural soil environment. To ensure the
validity of the results recently isolated, natural strains of bacteria
and plasmids were used. Laboratory and field results were compared whenever
Main findings and outcome
Plasmids were found to transfer actively in the conditions pertaining
in soil and river epilithon. Transfer between a wide variety of bacteria,
including heterotrophs, sulphate reducing bacteria and cyanobacteria,
was shown to be possible. Retrotransfer allowed movement of small recombinant
plasmids directly into natural bacterial communities. Plasmids from aquatic
bacteria were clearly shown to be able to capture recombinant DNA by retrotransfer.
Similarity was demonstrated between plasmid transfer on stones in a stream
microcosm and in situ in the river studies. Survival and mobility
studies of plant growth beneficial pseudomonads demonstrated that bacteria
could survive well in agricultural soils, especially when a dry inoculum
was used. Pseudomonad mutants with impaired survival were isolated and
shown to have the potential for DNA containment.
Important methodological advances were made in this study. These included
a gene escape model which used a transfer proficient E. coli containing
a heavy-metal resistance gene cassette, and an efficient marker system
for soil pseudomonads with a novel lactose utilisation DNA insert.
The potential for retrotransfer in aquatic bacteria was shown. Likewise,
a stream microcosm for simulating plasmid transfer between GEMs on river
stones was developed.
January 1989 December 1990
of the project:
This project was continued in EC project: BIOT-CT91-0284
Laboratory of Genetics & Biotechnology (SCK/VITO)
Instituto de Biologia Experimental e Tecnologica (IBET)
University of Wales
College of Cardiff (UK)
University of Ghent (BE)