Genetically Modified Material Research Articles

Landscape gene flow, coexistence and threshold effect: The case of genetically modified herbicide tolerant oilseed rape ( Brassica napus)

Globally there have been a number of concerns about the development of genetically modified crops many of which relate to the implications of gene flow at various levels. In Europe these concerns have led the European Union (EU) to promote the concept of ‘coexistence’ to allow the freedom to plant conventional and genetically modified (GM) varieties but to minimise the presence of transgenic material within conventional crops. Should a premium for non-GM varieties emerge on the market, the presence of transgenes would generate a ‘negative externality’ to conventional growers. The establishment of maximum tolerance level for the adventitious presence of GM material in conventional crops produces a threshold effect in the external costs. The existing literature suggests that apart from the biological characteristics of the plant under consideration (e.g. self-pollination rates, entomophilous species, anemophilous species, etc.), gene flow at the landscape level is affected by the relative size of the source and sink populations and the spatial arrangement of the fields in the landscape. In this paper, we take genetically modified herbicide tolerant oilseed rape (GM HT OSR) as a model crop. Starting from an individual pollen dispersal function, we develop a spatially explicit numerical model in order to assess the effect of the size of the source/sink populations and the degree of spatial aggregation on the extent of gene flow into conventional OSR varieties under two alternative settings. We find that when the transgene presence in conventional produce is detected at the field level, the external cost will increase with the size of the source area and with the level of spatial disaggregation. On the other hand when the transgene presence is averaged among all conventional fields in the landscape (e.g. because of grain mixing before detection), the external cost will only depend on the relative size of the source area. The model could readily be incorporated into an economic evaluation of policies to regulate adoption of GM HT OSR.

Modelling the persistence of volunteer oilseed rape ( Brassica napus)

The mixing of volunteer oilseed rape ( Brassica napus) within the crop has been tolerated by farmers. However, the coexistence of volunteer and crop populations needs to be tightly controlled where the introduction of unique varieties is accompanied by concern over the associated economic and environmental risks, as with the introduction of genetically modified (GM) varieties. A means is required to explore volunteer population dynamics before GM varieties are widely grown commercially. Yet, few measurements of the physiological status in situ, life-cycle biology or even their decline in the seed bank are available for any type of volunteer population. Accordingly, a model of within-field oilseed rape population dynamics was developed to predict the density of volunteers and the associated impurity (proportion of volunteer plants or seeds in the total number of oilseed rape plants or seeds) under a range of conditions, and to define a framework for exploring persistence of GM and other volunteers in agricultural systems. Simulations were based on a parameterisation of a typical winter oilseed rape cultivar grown in rotation with winter wheat. For all combinations of parameter values considered, the volunteer cohorts declined over time. The results were consistent with observations on the persistence of oilseed rape. Persistence remained high across the range of life-history parameter values estimated for oilseed rape. In the absence of control the simulated volunteer cohort did not fall below 1% of the total oilseed rape density until 16 years after it was sown. The stringent application of a combination of control measures (represented by 100% herbicide efficacy and reduction of harvest seed loss to 1%) reduced impurity to 0.12% within 3 years. National and international legislation has imposed thresholds for labelling of adventitious presence of GM material in foodstuffs, including harvested crops, ranging from 0.5% and 0.9% in the European Union to 5% in Japan. The results of this study indicate that the lower thresholds could be met within a reasonable time-frame (3 years) but only with the rigorous application of volunteer control measures. This might affect the adoption rates of GM and other unique varieties of oilseed rape by reducing the net benefit of these to individual farmers and have a limiting effect on the supposed reduction herbicide use associated with GM herbicide tolerance. Consequently, dedicated production of single varieties, e.g. either GM or non-GM, may be the only way that the coexistence of these crop types can be achieved.

Immunoassay as an Analytical Tool in Agricultural Biotechnology

Immunoassays for biotechnology engineered proteins are used by AgBiotech companies at numerous points in product development and by feed and food suppliers for compliance and contractual purposes. Although AgBiotech companies use the technology during product development and seed production, other stakeholders from the food and feed supply chains, such as commodity, food, and feed companies, as well as third-party diagnostic testing companies, also rely on immunoassays for a number of purposes. The primary use of immunoassays is to verify the presence or absence of genetically modified (GM) material in a product or to quantify the amount of GM material present in a product. This article describes the fundamental elements of GM analysis using immunoassays and especially its application to the testing of grains. The 2 most commonly used formats are lateral flow devices (LFD) and plate-based enzyme-linked immunosorbent assays (ELISA). The main applications of both formats are discussed in general, and the benefits and drawbacks are discussed in detail. The document highlights the many areas to which attention must be paid in order to produce reliable test results. These include sample preparation, method validation, choice of appropriate reference materials, and biological and instrumental sources of error. The article also discusses issues related to the analysis of different matrixes and the effects they may have on the accuracy of the immunoassays.

Kernel lot distribution assessment (KeLDA): a study on the distribution of GMO in large soybean shipments

The reliability of analytical testing is strongly affected by sampling uncertainty. Sampling is always a source of error and the aim of “good” sampling practice is to minimize this error. Generally the distribution of genetically modified (GM) material within lots is assumed to be random in order to use binomial distribution to make inferences. This assumption was never verified in practice and no experimental data investigating the distribution of genetically modified organisms (GMOs) exist. The objectives of the KeLDA project were: (1) to assess the distribution of GM material in soybean lots (2) to estimate the amount of variability of distribution patterns among lots. The GM content of 15 soybean lots imported into the EU was estimated (using real-time PCR methodology) analyzing 100 increment samples systematically sampled from each lot at predetermined time intervals during the whole period of off-loading. The distribution of GM material was inferred by the one-dimensional (temporal) distribution of contaminated increments. All the lots display significant spatial structuring, indicating that randomness cannot be assumed a priori. The evidence that the distribution of GM material is heterogeneous highlights the need to develop sampling protocols based on statistical models free of distribution requirements.

Coherence between Legal Requirements and Approaches for Detection of Genetically Modified Organisms (GMOs) and Their Derived Products

Analytical methods for the qualitative and quantitative detection of genetically modified (GM) products may serve multiple purposes. Legal requirements differ among jurisdictions, ranging from no requirements to mandatory use of event-specific quantitation and implementation of production chain traceability. Although efforts have been taken to harmonize the analytical methodology at national, regional, and international levels, no normative international standards have yet been established. Lack of coherence between analytical methodologies and their applicabilities, on the one hand, and legislation, on the other hand, is a major problem. Here, key points where coherence is lacking are discussed. These include the definition of units of measurements, expression of GM material quantities, terminology, and inconsistent legal status of products derived from related but slightly different transformation routes. Finally, recommendations to improve the coherence are brought forward, including guidance to stakeholders for prediction of product-specific GM material quantities from gene ratios in the originating seed.

“Words of mass destruction”: British newspaper coverage of the genetically modified food debate, expert and non-expert reactions

This article reports the findings of a one-year project examining British press coverage of the genetically modified (GM) food debate during the first half of 2003, and both expert and non-expert reactions to that coverage. Two pro-GM newspapers and two anti-GM newspapers were selected for analysis, and all articles mentioning GM during the period in question were stored in a machine readable database. This was then analyzed using corpus linguistic and discourse analytic techniques to reveal recurrent wording, themes and content. This text analysis was complemented by 12 interviews with experts involved in the communication of GM issues, and 12 focus-group sessions in which members of the public reacted to selected newspaper texts and other GM material. Both in the press and in public reaction, the issue of GM was found to be intimately associated with other political events of the time, notably the invasion of Iraq. Except among experts, there was little awareness of the official national debate and issues were approached in more general terms. Pro-GM characterization of the issues as primarily scientific, both by newspapers and experts, was rejected by the anti-GM press and campaigners, and by the focus-group participants. They assessed the issues in a more global frame, rejecting scientists and companies as unreliable. In addition, they linked both US and British GM policy to the invasion of Iraq, and, by analogy, rejected pro-GM arguments as untrustworthy.

Adventitious presence of GMOs: Scientific overview for Canadian grains

The global expansion in the development and cultivation of genetically modified (GM) crops has increased international concern about adventitious presence of GM materials in non-GM seeds and grains. GM events in canola, corn, soybean, cotton, flax, papaya, potato, squash, sugar beet, and tomato have received regulatory approval in Canada. However, GM cultivars are only in commercial production for canola, corn and soybean. More than 30 GM events have been approved in these three crops. Cases of unapproved adventitious presence of GM materials that have impacted grain trading and handling in Canada and other countries include StarLink™ corn, GT200 canola, GM canola in mustard and recently Bt10 corn. Some countries have established tolerance and traceability requirements for adventitious presence of GMOs, while others are in the process of developing or adopting legislation. The threshold for labeling of adventitious presence of approved GM material in non-GM grain varies from 0.9% (e.g., EU) to 5% (e.g., Japan). Progress has been made in the development of DNA- and protein-based GMO detection methods. However, only a limited number of these detection methods have been internationally validated. The challenges for detection methods include sampling, a lack of certified reference material, a lack of DNA sequence information for the design of event-specific primers, and the sheer number of individual events that may be present and tested for. Current efforts by ISO and CEN will be valuable for establishing harmonized and standardized GMO detection methods. Key words: List of GM events, cases of AP, tolerance, traceability, detection methods, challenges

Honey produced from genetically modified canola (Brassica napus) nectar will not need to be labelled as a GM food under current Australian guidelines

Food or ingredients labelled as genetically modified (GM) contain either new genetic material or protein as a result of genetic modification. In Australia, a 1% threshold, below which labelling is not required, exists for the unintended presence of GM material in non-GM foods. The canola pollen content by dry weight in a range of canola honey samples from diverse geographical areas in Australia was determined to be 0.2 ± 0.12%, well below the 1% threshold. Two GM canola honey samples sourced from Canada contained 0.19 and 0.24% of canola pollen. This work indicates honey derived from GM canola crops will not need to be labelled as a GM food.

Quantitative Detection System for Maize Sample Containing Combined-Trait Genetically Modified Maize

Various countries have established regulations that stipulate the labeling of agricultural commodities, feed, and food products that contain or are made from genetically modified (GM) material or that contain adventitious GM material in amounts that exceed certain threshold levels. While regulations in some countries refer to GM material on a weight per weight (w/w) percentage, the currently applied detection methods do not directly measure the w/w percentage of the GM material. Depending on the particular method and the sample matrix it is applied to, the conversion of analytical results to a w/w percentage is challenging or not possible. The first rapid PCR system for GM maize detection on a single kernel basis has been developed. The equipment for the grinding of individual kernels and a silica membrane-based 96-well DNA extraction kit were both significantly revised and optimized for this particular purpose, respectively. We developed a multiplex real-time PCR method for the rapid quantification of GM DNA sequences in the obtained DNA solutions. In addition, a multiplex qualitative PCR detection method allows for the simultaneous detection of different GM maize traits in each kernel and thereby for identification of individual kernels that contain a combination of two or more GM traits. Especially for grain samples that potentially contain combined-trait GM maize kernels, the proposed methods can deliver informative results in a rapid, precise, and reliable manner.

Lack of detectable allergenicity of transgenic maize and soya samples

The safety issues regarding foods derived from genetically modified (GM) plants are central to their acceptance into the food supply. The potential allergenicity of proteins newly introduced in GM foods is a major safety concern. We sought to monitor, in potentially sensitive human populations, the allergenicity effects of 5 GM materials obtained from sources with no allergenic potential and already under commercialization in the European Union. We have performed skin prick tests with protein extracts prepared from transgenic maize (MON810, Bt11, T25, Bt176) and soya (Roundup Ready) samples and from nontransgenic control samples in 2 sensitive groups: children with food and inhalant allergy and individuals with asthma-rhinitis. We have also tested IgE immunoblot reactivity of sera from patients with food allergy to soya (Roundup Ready) and maize (MON810, Bt11, Bt176) samples, as well as to the pure transgenic proteins (CryIA[b] and CP4 5-enolpyruvylshikimate-3-phosphate synthase). None of the individuals undergoing tests reacted differentially to the transgenic and nontransgenic samples under study. None of the volunteers tested presented detectable IgE antibodies against pure transgenic proteins. The transgenic products under testing seem to be safe in terms of allergenic potential. We propose postmarket testing as an important screening strategy for putative allergic sensitization to proteins introduced in transgenic plants.

The co-existence between transgenic and non-transgenic maize in the European Union: a focus on pollen flow and cross-fertilization

The ongoing discussion on the co-existence between genetically modified (GM) and non-GM crops becomes more important in the European Union (EU). With the recent inscription of 17 GM maize varieties in the common EU catalogue of varieties of agricultural plant species, the acreage of transgenic maize for market purposes is expected to increase in some European countries. In the EU, specific tolerance thresholds have been established for the adventitious and technically unavoidable presence of GM material in non-GM produce, and member states are elaborating legal frames to cope with co-existence. As maize is a cross-pollinated crop relying on wind for the dispersal of its pollen, technical management measures will be imposed to reduce cross-fertilization between transgenic and non-transgenic maize. Various biological, physical and analytical parameters have been identified to play a role in the study of cross-fertilization in maize. This variability may hamper the comparison between research results and may complicate the definition of appropriate isolation distances and/or pollen barriers in order to limit out-crossing. The present review addresses these parameters and proposes containment measures in order to not exceed the legal labeling thresholds in maize.

Polymerase Chain Reaction Technology as Analytical Tool in Agricultural Biotechnology

The agricultural biotechnology industry applies polymerase chain reaction (PCR) technology at numerous points in product development. Commodity and food companies as well as third-party diagnostic testing companies also rely on PCR technology for a number of purposes. The primary use of the technology is to verify the presence or absence of genetically modified (GM) material in a product or to quantify the amount of GM material present in a product. This article describes the fundamental elements of PCR analysis and its application to the testing of grains. The document highlights the many areas to which attention must be paid in order to produce reliable test results. These include sample preparation, method validation, choice of appropriate reference materials, and biological and instrumental sources of error. The article also discusses issues related to the analysis of different matrixes and the effect they may have on the accuracy of the PCR analytical results.

The fate of transgenic sequences present in genetically modified plant products in fish feed, investigating the survival of GM soybean DNA fragments during feeding trials in Atlantic salmon, Salmo salar L

Vegetable protein sources like soybeans, canola and maize gluten are good alternatives to fish meal. However, a large proportion of such products available on the international market may possess genetically modified (GM) components. This report concerns a study to investigate the fate and survival of ingested GM soy DNA fragments (120 and 195 bp) and a 180-bp fragment of the lectin gene of soybean ( Glycine max) during feeding trials with Atlantic salmon post-smolt. Specifically, the study focused on the fate of selected GM soy DNA fragments from feed to fish to investigate their survival through the fish gastrointestinal (GI) tract and whether the DNA could be traced in a variety of fish tissues. Fish were fed three experimental diets for 6 weeks, which were formulated from defined components and represented either GM or non-GM materials (17.2% of the fish meal was replaced with either GM or non-GM soy). A control diet composed of fish meal as the only protein source was used for comparison purposes. The transgenic sequences (120 and 195 bp) and the lectin gene (180 bp) could be detected in the GM soy feed. In the fish GI tract, however, only the smaller DNA fragment (120 bp) could be amplified from the content of the stomach, pyloric region, mid intestine and distal intestine. No transgenic or conventional soy DNA fragments could be detected in liver, muscle or brain tissues resected from sacrificed fish. The sensitivity limit of the method was evaluated to be 20 copies. These data indicate that GM soy transgenic sequences may survive passage through the GI tract but that they cannot be traced in fish tissues.

Detection of genetically modified soya in a range of organic and health food products

A selection of organic/health foods containing soya beans was tested for the presence of genetically modified (GM) material. Out of 25 samples of food products containing unrefined soya ingredients, ten tested positively for the presence of GM material. This was surprising because eight out of the ten GM‐positive samples were either labelled as “GM free” and/or were labelled as “organic”, both of which imply the absence of GM ingredients. In no case did any of the foods tested reach the mandatory 1 per cent threshold required for positive labelling as GM products under current European Union legislation, although one product was close to this limit. However, there was considerable batch‐to‐batch variation in the GM soya content of some of the food products, depending on the purchase date and retailer. The paper discusses the implications of these results regarding international regulations on food labelling and use of “GM free” labels or their equivalent.

Simulating kernel lot sampling: the effect of heterogeneity on the detection of GMO contaminations

Guidelines defining kernel sampling strategies for quality analyses have been provisionally adopted for the detection of genetically modified (GM) contamination in kernel lots. However, these guidelines are not specific for GM material detection and are not intended for the sampling of non-uniform distributions, a probable situation with respect to the presence of GM material in kernel lots. An analysis of the problem of non-random distribution, through the investigation of the effectiveness of different sampling techniques in producing representative bulk samples, is presented. The analysis is based on a two-step modelling procedure: 1) the kernel lot is created and 2) the lot is sampled to produce a bulk sample. This allows the identification of optimal sampling techniques as a function of specific combinations of population characteristics. For each of 5 levels of GM impurity, varying between 0.1% and 2%, we investigated the effect of 5 levels of stratification (lot size=10 7 kernels). Our results indicate: I) For every GM level, the higher the heterogeneity level, the more unstable the GM estimate becomes; even modest levels of stratification affect the stability of GM estimates. 2) As the number of increment samples increases, the coefficient of variation (CV) of the estimate decreases. Although the pattern of decrease remains similar across stratification levels, the estimated CV changes: with low levels of stratification. 50 samples are enough to obtain estimates with CV<10%. In the case of modest levels of stratification even 100 samples are not sufficient to maintain CV<10%. In case of strongly heterogeneous lots estimates based on 100 units have CVs around 50%. At the same time, the likelihood of false negative results increases significantly.

Reliability of food measurements: the application of proficiency testing to GMO analysis

This paper reviews the experience of the Food Analysis Performance Assessment Scheme (FAPAS®) in operating a proficiency testing scheme for the analysis of genetically modified (GM) food. Initial rounds of proficiency testing have shown a tendency for laboratories to over-estimate GM levels, results obtained by polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) detection methods to be significantly different and that data are skewed and not normally distributed until log-transformed. During the initial rounds, it was found that for analysis and quantification of GM material, it was not possible to assign a target value for standard deviation external to the round data, from which performance could be assessed. However, when working in a log scale, the internally derived, robust standard deviation (^σ) was found to be constant and could be used directly to predict a target value (σ) for performance assessment. Results from the first four rounds have provided valuable information and a general overview of laboratory ability. Choosing a target value for standard deviation which reflects current best practice has enabled laboratory performance to be assessed. Issues surrounding the assessment of performance are discussed which highlight some of the implications raised as a result of this initial assessment, regarding the enforcement of European labelling legislation.

The environmental impact of genetically modified (GM) crops: a review

Evaluating environmental impact is more complex than statistical analysis; it requires value judgements about which kinds of environmental impacts are acceptable and which are not. The new EU Directive on the release and commercialization of genetically modified (GM) crops (2001-18-EC) includes a requirement for an assessment of indirect effects on the environment of farming practices associated with the introduction of a GM crop. There is also a requirement for post-commercialization monitoring to address impacts of scale and time. Both of these additions to the regulations present challenges in deciding how environmental impacts should be measured, and in judging their acceptability. Impacts can also be socio-economic. These frequently require scientific tools to help to resolve them. The demands by some food sectors for zero GM content in crop produce is likely to prove difficult or impossible to verify by analytical procedures. It will probably be necessary to agree on levels of tolerance of the presence of GM material in “non-GM” crop produce. It is important that the future use of biotechnology in agriculture is considered in the context of a broader discussion about our vision for agriculture and the environment over the coming decades.

New feeds from genetically modified plants: substantial equivalence, nutritional equivalence, digestibility, and safety for animals and the food chain

The introduction by recombinant DNA technology of new genes into major crops used by animals has raised important questions about the safety of novel feeds. The recently amended European Council Directive 2001/18/EC requires an assessment of risks for human, animals and the environment before viable seeds can be imported or the plant itself can be cultivated in Europe. In addition, the Novel Food and the Novel Food Ingredient Regulation ( EC Council Directive, 1997) covers the use of non-viable products of any genetically modified (GM) plant intended for food purposes. The conceptual basis for the analysis of risks remains that of substantial equivalence. In practice, compositional analysis of key nutrients and key toxicants used to compare a GM plant with its conventional counterpart is the major source of data used to establish substantial equivalence. Although laboratory animals are used in toxicological studies of the products of introduced genes, animals are rarely fed the entire transformed plants or their by-products. Despite not being required or recommended by the existing legislation, many new products have been intensively tested with farm animals to measure effects on performance and animal health, digestibility of key nutrients, wholesomeness and feeding value. In this paper, we consider the value of such studies for establishing the safety of GM feedstuffs for the target animals and whether animal feeding studies are always necessary to establish the safety for the consumer of products of animals fed GM material. Compositional analysis has always shown the genetically modified plants to fall within the range of established values. The equivalence in digestible energy and crude protein between isogenic and transformed plants expressing a wide range of modifications (insect resistance, herbicide tolerance, or the barnase/barstar system of sterility/fertility restoration genes) also has been clearly demonstrated in different species. In none of these experiments was animal performance, whether measured as growth rate, feed efficiency and carcass merit in beef cattle, egg mass in laying hens, milk production, composition and quality in dairy cows or digestibility in rabbits, affected by feeding transformed plants compared to animals fed control or isogenic plants. Detection of chloroplasts-specific gene fragments by polymerase chain reaction showed the presence of plant DNA fragments (199 base pairs) in lymphocytes and duodenal juice of the dairy cow, and in muscle, liver, kidney and spleen of broilers. However, tDNA expressing Bt, usually found as a single copy gene, could not be detected in milk, tissues sample or eggs of livestock fed Bt maize and is probably not cause for concern.

Performance assessment under field conditions of a rapid immunological test for transgenic soybeans

SummaryCurrent market conditions and food regulations make it necessary for international and domestic participants in the agrifood industry to structure supply chains that control the content of genetically modified (GM) material in their products. Tests to detect and/or quantify GM components represent an important tool in maintaining such supply systems. This study assesses the field performance of kits that employ lateral flow immuno‐technology to detect soybeans GM to be resistant to the herbicide glyphosate. Operators at 23 grain‐handling facilities were paid to conduct analyses on a series of blinded samples containing defined proportions of conventional and transgenic soybeans. The observed rate of false positives was 6.7% in an experiment in which the highest level of GM material was 1% and 22.3% in a second experiment in which the highest level of GM material was 10%. This difference may be attributed to increased risk of cross‐contamination with the higher level of transgenic material used in the second study. Samples containing 0.01% GM material were reported as genetically modified 6.70% of the time, while samples containing 0.1, 0.5 and 1% GM material were classified as genetically modified 29.5, 67.7 and 68.2% of the time, respectively. Thus, the frequencies of false negatives were 93.3, 70.5, 33.3 and 31.8% for samples containing 0.01, 0.1, 0.5 and 1.0% GM material. Samples containing 10% GM material were correctly reported as genetically modified in all cases. These results lead to the conclusion that the kit under study is useful in screening for lots of soybeans that contain high levels of GM material, but that, as a field tool, it is not effective in monitoring for GM material at the level of 1.0% or lower. Statistical and immunochemical analyses were carried out in order to assess the relative contributions of various factors to the error observed in these studies. These analyses indicated that limitations in operator performance, not defects in test kit materials, were the primary contributors, while sample size may play a secondary role. As both operator performance and sample size are independent of the specific characteristics of the test kit used in this study, it appears justifiable to generalize conclusions obtained here to other similar test systems.

Antibiotic-resistance-free GM plants

Many fears and concerns are expressed in connection with the use of genetically modified (GM) crops. One is the possibility that antibiotic resistance might be transferred to microorganisms in the wider environment or to those within the gut flora of animals, including humans. Antibiotic resistance is widely used as a marker for selection of GM material. A report by Siriluck Iamtham and Anil Day [Nat. Biotechnol. (2000) 18, 1172–1176], details a procedure for the removal of resistance to the antibiotics streptomycin and spectinomycin from tobacco plants that have been transformed using GM plastids. Although such work might not allay all the problems associated with the use of GM crops, it should help to reduce them.