Need for Mitigation and Termination Strategies
Mitigation and termination are topics of relevance to introduction of organisms to the
environment. There has been much discussion of the potential adverse effect of such
introductions in general. The Risk Assessment will have identified any specific concerns, and in
most cases will have noted that all possible events cannot be identified. Hence, especially for
field trials, permission to conduct the release includes requirements for developing protocols for
identifying the onset of an adverse effect and for terminating the trial
and mitigating the effect. It must be pointed out that field trials involving microbes and most
plants cannot generally be said to be contained. The proper term is confined.
Environmental Issues
While there has been much public concern and speculation about the possibility of
adverse environmental effects from the environmental application of runaway engineered
organisms, there is little evidence of adverse effect. The possibility of effects on nontarget
organisms, on biological cycles, and on human health has been discussed. Yet, to date, after
almost 5000 field tests of engineered microbes or plants, there is no record of such problems
arising. In one instance (Short et al. 1991) researchers evaluating the efficacy of Pseudomonas
strains that had been engineered to degrade 2-4 Dichlorophenoxyacetate found that 2-4
Dichlorophenol (a toxic intermediate metabolite) accumulated in the soil. The accumulation of
2-4 Dichlorophenol resulted in a loss of 90% of the fungal population in the soil, soil
decontaminated resulted in replenishment of normal flora. Nevertheless, the possibility of an
adverse effect requires that environmental applications be reviewed for safety considerations.
Cavalieri (1991) has proposed that microcosms be used to predict the environmental
consequences of the application of engineered microorganisms. Microcosms can provide
information about persistence, survival, and specific effects of the modified microbe in question
relative to the unmodified host. While the information from microcosms may not be entirely
representative of results under field conditions, it will provide a basis for deciding whether or
how field testing should proceed. Similarly, based on microcosm data, the process could be
modified, safety precautions instituted or devised, provisions for confinement or mitigation
devised, and effective monitoring protocols designed.
Total eradication of unwanted microorganisms is rare, but reduction to acceptable levels
(i.e., below the level of unacceptable economic or health impact) is possible. Absolute
containment of microorganisms is not possible and, based on experience with both beneficial and
detrimental microorganisms, not essential (Vidaver and Stotzky 1992). Vidaver and Stotzky
propose the use of the more realistic term "confinement" in lieu of containment. Confinement
does not imply that the microbe will not spread beyond the point of application, but rather that it
can be effectively managed and adverse effects minimized. Most microorganisms are confined
biologically by their individual requirements for nutrients, moisture, and sensitivity to
environmental conditions (i.e., their niche).
Additional strategies involve the use of debilitated organisms or the construction and use
of safe cloning vectors with limited ability to transfer or survive outside the original host and the
use of replicons sensitive to temperature or other environmental factors. The use of debilitated
microbes is not practical for environmental applications. However, several conditionally lethal
systems for the control of released bacteria have been designed and tested. These include a
temperature sensitive system ( e.g. where DNA repair does not occur at cold temperatures), a
conditionally lethal construct wherein the organism has an inducible metabolic pathway that can
be activated only by the presence of an innocuous chemical not normally present in the
environment of the microbe and a "suicide" gene which will destroy a key feature without which
the cell cannot survive. The gene is controlled by the presence (induced) or absence
(derepressed) of the waste in question. If the waste concentration falls below a critical level, the
gene is activated level; Alternatively the gene is always active and a second gene provides
protection. Activity of the second gene is controlled by the concentration of the waste being
treated.
Decontamination (or mitigation) of the environment of microbes has been studied and is
discussed by Vidaver and Stotzky (1992). It is important to keep in mind that each situation is
different and that procedures for decontamination will differ. A case by case approach is
essential. The type of organism, the physical environment, the nature of the modification and the
season must all be considered. Knowledge about the organism, whether it is a wild type or has
been modified, is critical to designing a decontamination protocol. Vidaver and Stotzky list
specific methods of decontaminating soils, plants, and animals, if they are contaminated with a
hazardous microbe, and provide an indication of the time required to achieve effectiveness.
Details of soil sterilization to decrease the bacterial levels at the site are also specified. Specific
soil fumigants in common use in the USA are identified.
Plants and animals must be considered because of the possibility of escape during a field
application. Thus, in the event of contamination of animals (straying on the site) incineration,
quarantine or slaughter could be employed immediately to minimize spread of the
microorganism. Birds, rodents and runoff water must be considered as alternate sources of
microbial dispersal. Plants which are growing on the site may be contaminated with the microbe.
If the microbe is considered a hazard the plants should be immediately destroyed (i.e. burning,
tillage) or quarantined if a future use is being considered. Long term solutions are presented for
use in the event the project has along life span and the problem is recurrent. The issue of physical
security, especially with animals, cannot be overstressed. Strong, tall fences will eliminate the
presence of most unwanted mammals and insure against trespassers.
Use of a fumigant will significantly lower the densities of all microorganisms present in
the soil. Sterilization is not achieved. As a result, over time, the remnants of the microbial flora
will reproduce and cell densities will increase. The new population may be similar to the
previous one, in terms of types and relative numbers of individual types or it may radically differ,
depending on which portion of the population survived the fumigation and at what level. There
is a possibility that the introduced microbe could be the dominant type. For this reason, it is
recommended that the treated site be reinoculated with uncontaminated soil from the surrounding
area. This will most likely result in replacement of the original indigenous microbial flora and
significantly decrease the probability that the introduced microbe would flourish.
