The development of transgenic crops as production platforms for biogenic agents will largely depend on the success of efforts to confine the genes and their expressed proteins in field environments. We have used quantitative exposure assessment to evaluate how management practices affect materials escape due to outcrossing by pollen flow or grain loss during harvest operations. Specifically, we study the use of maize to produce biogenic agents within field-confined systems. Decision trees representing simplified schemes of fully conforming (designed to comply with current regulatory standards for field confined trials), partially conforming, and non-conforming management practices were developed. Exemplifying assumptions and published data for conformance and material fate probabilities were used in Monte Carlo simulations to forecast materials escape by pollen outcrossing and harvest operations from a 1 ha source field. Deterministic analyses showed fully conforming confinement management restricted materials loss to low levels (for this example, outcrossing produced <1 in 106 kernels in receptor fields). The corresponding high-end (90th percentile) probabilistic result was 16- and 4333-fold higher (relative to deterministic outcrossing = 1) for outcrossing and harvest loss, respectively. For partially conforming practice, high-end outcrossing ranged from 100- to >15 000-fold over the base result in receptor fields, and harvest loss was >10 000-fold over the base result. For non-conforming practice, high-end outcrossing produced >15 000-fold greater kernels in receptor fields and high-end harvest loss was at least 19 000-fold greater. Deterministic estimates of off-field loss by machine transfer are as much as 30 000-fold higher for non-conforming operations relative to the base case of pollen outcrossing. Better knowledge of failure frequencies for confinement management practices, improved physical models of materials flows, refined analysis of confinement loss probabilities using quantitative tools, and decision analysis to improve and audit management system performance are all needed to extend understanding of confinement integrity beyond the exemplifying case used here.

The safety and impact on the environment of transgenic crops are important issues, and studies have shown that pollen from transgenic Bt (Bacillus thuringiensis) corn (Zea mays L.) may kill nontarget insects. To develop an algorithm for assessing the environmental effect of transgenic crops, we arranged a field experiment in Tsukuba, Japan. Pollen dispersal and deposition were measured inside and outside a cornfield throughout the flowering period. Weather conditions such as wind speed and direction were measured at the same time. Pollen dispersal peaked 1 week after the start of flowering and continued for 12 days thereafter. The variation in daily pollen dispersal was similar at all observation points. Both pollen dispersal and deposition decreased exponentially with distance from the cornfield on all days. We estimated potential pollen deposition with a quasi-mechanistic model that incorporates the effects of wind direction, wind speed, and flowering intensity. The daily potential deposition was summed over the flowering period, and then the relationship between distance from the cornfield and the integrated potential deposition was estimated. It was possible to show the effective area of the environmental risk zone posed by genetically modified pollen by combining the distance/deposition relationship with the dose/response relationship derived from a laboratory assay. The algorithm described here can be applied to various wind-pollinated plants to estimate both potential and integrated pollen deposition.

Hybridization of Brassica napus L. (oilseed rape) and Raphanus raphanistrum L. (wild radish) has been demonstrated, and may be the first step towards introgression of transgenes in this wild relative. If wild radish were to display a new adaptive advantage by expressing the transgene, this could modify the ecological balance of species within the agro-ecosystem. To determine if transgenes remained stable in the hybrid, the frequency of herbicide resistance was studied over four advanced generations of hybrid progeny (G8 to G11) that were subjected to herbicide selection pressure. It is expected that hemizygous resistant plants containing an herbicide resistance transgene back-crossed to wild radish would have 50% resistant progeny. In each of the G8 to G11 generations, only 18% of the progeny from resistant plants were resistant. The chromosome complement of herbicide-susceptible progenies, analyzed at G9, was not different from that of wild populations of wild radish. Herbicide-resistant G9 progeny showed higher chromosome instability, and one third of the progeny contained a supernumerary chromosome. These results suggest that in the presence of herbicide selection pressure, the transgene for herbicide resistance would be maintained despite a lack of stabilized introgression. In the absence of selection, the frequency of resistance in the population is expected to decline.

The effect of food components on degradation of DNA by DNase I (EC 3.1.21.1) was monitored by electrotransformation of Escherichia coli, making it possible to determine the number of plasmid molecules capable of giving rise to transformed cells. The transformation frequency increased linearly with the plasmid number within the range of 2 × 106 to 2 × 1010. DNA degradation was reduced by one order of magnitude in the presence of 0.05% (w.v–1) maltol or 1 mM putrescine. Complete inhibition of degradation was observed with ≥0.2% (w.v–1) maltol, ≥0.01% (w.v–1) octyl gallate or ≥0.5 mM of spermine. To monitor degradation of plant DNA during food processing, a real-time PCR system was established. The ratio of copy numbers of a potato gbss DNA fragment of 325 bp and a nested 96 bp fragment was determined. The latter served as internal reference for normalization. The system made it possible to exclude process-dependent changes of DNA concentration in the food matrix. Processing of genetically modified potatoes to dried potato sticks, crisps or flakes was studied and drying steps were shown to exert the strongest effect on DNA degradation, resulting in a drop of the ratio from 0.73 to 0.16.

Impact assessment studies rely on relevant sampling methods. Correct evaluation of methods can be done by their concurrent use in the same sampling site and period. Collecting webs of Theridion impressum L. Koch (Araneae: Theridiidae) may serve as an arthropod sampling method: empty cuticles of prey items remain attached to the back side of the leaves with adhesive silk. Our study aimed to compare the applicability of the two methods concurrently (spider web survey and whole plant visual sampling) in a risk assessment study. The spider web survey recorded more predatory insect families than the whole plant visual sampling. Both methods were able to detect significant differences in the quantity of predatory insects in Bt vs. isogenic plots, but not in the same taxa (Nabidae: spider-web, 2001, Bt > Iso; Coccinellidae: plant sampling, 2001, Iso > Bt; Welsh-test, P < 0.05); thus, they could not confirm each other. The lack of confirmation can be explained by differences in the sensitivity and selectivity of the two methods. A web survey of T. impressum has the practical advantage that although we concentrate only on the one species during field sampling, we gain additional information on a wide range of foliage-dwelling arthropods. Due to several biological uncertainties, interpretation and explanation of the results remain problematic. Thus, additional research – based on in-situ observation and recording of T. impressum-prey interactions – is necessary before we could propose web survey method as a complementary tool in ecological impact assessment.