Genetic engineering of crops as potential source of genetic hazard in the human diet.

The benefits of genetic engineering of crop plants to improve the reliability and quality of the world food supply have been contrasted with public concerns raised about the food safety of the resulting products. Debates have concentrated on the possible unforeseen risks associated with the accumulation of new metabolites in crop plants that may contribute to toxins, allergens and genetic hazards in the human diet. This review examines the various molecular and biochemical mechanisms by which new hazards may appear in foods as a direct consequence of genetic engineering in crop plants. Such hazards may arise from the expression products of the inserted genes, secondary or pleiotropic effects of transgene expression, and random insertional mutagenic effects resulting from transgene integration into plant genomes. However, when traditional plant breeding is evaluated in the same context, these mechanisms are no different from those that have been widely accepted from the past use of new cultivars in agriculture. The risks associated with the introduction of new genes via genetic engineering must be considered alongside the common breeding practice of introgressing large fragments of chromatin from related wild species into crop cultivars. The large proportion of such introgressed DNA involves genes of unknown function linked to the trait of interest such as pest or disease resistance. In this context, the potential risks of introducing new food hazards from the applications of genetic engineering are no different from the risks that might be anticipated from genetic manipulation of crops via traditional breeding. In many respects, the precise manner in which genetic engineering can control the nature and expression of the transferred DNA offers greater confidence for producing the desired outcome compared with traditional breeding.     

The impact of genetic modification of human foods in the 21st century: a review

Genetic engineering of food is the science which involves deliberate modification of the genetic material of plants or animals. It is an old agricultural practice carried on by farmers since early historical times, but recently it has been improved by technology. Many foods consumed today are either genetically modified (GM) whole foods, or contain ingredients derived from gene modification technology. Billions of dollars in U.S. food exports are realized from sales of GM seeds and crops. Despite the potential benefits of genetic engineering of foods, the technology is surrounded by controversy. Critics of GM technology include consumer and health groups, grain importers from European Union (EU) countries, organic farmers, environmentalists, concerned scientists, ethicists, religious rights groups, food advocacy groups, some politicians and trade protectionists. Some of the specific fears expressed by opponents of GM technology include alteration in nutritional quality of foods, potential toxicity, possible antibiotic resistance from GM crops, potential allergenicity and carcinogenicity from consuming GM foods. In addition, some more general concerns include environmental pollution, unintentional gene transfer to wild plants, possible creation of new viruses and toxins, limited access to seeds due to patenting of GM food plants, threat to crop genetic diversity, religious, cultural and ethical concerns, as well as fear of the unknown. Supporters of GM technology include private industries, research scientists, some consumers, U.S. farmers and regulatory agencies. Benefits presented by proponents of GM technology include improvement in fruit and vegetable shelf-life and organoleptic quality, improved nutritional quality and health benefits in foods, improved protein and carbohydrate content of foods, improved fat quality, improved quality and quantity of meat, milk and livestock. Other potential benefits are: the use of GM livestock to grow organs for transplant into humans, increased crop yield, improvement in agriculture through breeding insect, pest, disease, and weather resistant crops and herbicide tolerant crops, use of GM plants as bio-factories to yield raw materials for industrial uses, use of GM organisms in drug manufacture, in recycling and/or removal of toxic industrial wastes. The potential risks and benefits of the new technology to man and the environment are reviewed. Ways of minimizing potential risks and maximizing the benefits of GM foods are suggested. Because the benefits of GM foods apparently far outweigh the risks, regulatory agencies and industries involved in GM food business should increase public awareness in this technology to enhance worldwide acceptability of GM foods. This can be achieved through openness, education, and research.     

Opportunities and surprises in crops modified by transgenic technology: metabolic engineering of benzylisoquinoline alkaloid, gossypol and lysine biosynthetic pathways

The development of new or improved traits in plants, whether that is through traditional genetic modification and selection or through transgenic technologies, is associated with the potential risk of unintended changes with harmful or unacceptable consequences. The greater definition and precision of transgenic modification and the regulatory oversight of such technology may, however, confer advantages in safety and efficacy. This bears considerable relevance to the use of transgenic-based metabolic engineering in agricultural trait development. Metabolic engineering seeks to modify the amounts or chemical structures within selected biosynthetic routes without introducing inadvertent effects on other metabolic pathways. Examples discussed here include attempts to; (i) modify benzylisoquinoline alkaloid biosynthesis in poppy, (ii) improve the nutritional value of maize by increasing levels of free lysine, and (iii) increase the nutritional value of cottonseed by eliminating gossypol production. Clearly, evaluation of the efficacy (and unintended consequences) of such approaches is vital. A role for metabolomics in the compositional and metabolite analyses of new plant varieties derived from transgenic-based metabolic engineering is discussed. Major themes discussed in this review include; (i) the heightened level of scrutiny associated with genetically modified (GM) crop evaluations has markedly contributed to the safety in the adoption of transgenic technology, and (ii) the nature of any introduced trait may prove more relevant to safety assessments than the means by which the trait is introduced.     

Genetic modification and plant food allergens: risks and benefits

Plant genetic engineering has the potential to both introduce new allergenic proteins into foods and remove established allergens. A number of allergenic plant proteins have been characterized, showing that many are related to proteins which have potentially valuable properties for use in nutritional enhancement, food processing and crop protection. It is therefore important to monitor the allergenic potential of proteins used for plant genetic engineering and major biotechnology companies have established systems for this. Current technology allows gene expression to be down-regulated using antisense or co-suppression and future developments may allow targeted gene mutation or gene replacement. However, the application of this technology may be limited at least in the short term by the presence of multiple allergens and their contribution to food processing or other properties. Furthermore, the long-term stability of these systems needs to be established as reversion could have serious consequences.     

Transgenic plants and biosafety: science, misconceptions and public perceptions

One usually thinks of plant biology as a non-controversial topic, but the concerns raised over the biosafety of genetically modified (GM) plants have reached disproportionate levels relative to the actual risks. While the technology of changing the genome of plants has been gradually refined and increasingly implemented, the commercialization of GM crops has exploded. Today's commercialized transgenic plants have been produced using Agrobacterium tumefaciens-mediated transformation or gene gun-mediated transformation. Recently, incremental improvements of biotechnologies, such as the use of green fluorescent protein (GFP) as a selectable marker, have been developed. Non-transformation genetic modification technologies such as chimeraplasty will be increasingly used to more precisely modify germplasm. In spite of the increasing knowledge about genetic modification of plants, concerns over ecological and food biosafety have escalated beyond scientific rationality. While several risks associated with GM crops and foods have been identified, the popular press, spurred by colorful protest groups, has left the general public with a sense of imminent danger. Reviewed here are the risks that are currently under research. Ecological biosafety research has identified potential risks associated with certain crop/transgene combinations, such as intra- and interspecific transgene flow, persistence and the consequences of transgenes in unintended hosts. Resistance management strategies for insect resistance transgenes and non-target effects of these genes have also been studied. Food biosafety research has focused on transgenic product toxicity and allergenicity. However, an estimated 3.5 x 10(12) transgenic plants have been grown in the U.S. in the past 12 years, with over two trillion being grown in 1999 and 2000 alone. These large numbers and the absence of any negative reports of compromised biosafety indicate that genetic modification by biotechnology poses no immediate or significant risks and that resulting food products from GM crops are as safe as foods from conventional varieties. We are increasingly convinced that scientists have a duty to conduct objective research and to effectively communicate the results--especially those pertaining to the relative risks and potential benefits--to scientists first and then to the public. All stakeholders in the technology need more effective dialogues to better understand risks and benefits of adopting or not adopting agricultural biotechnologies.     

Molecular characterization for food safety assessment of a genetically modified late blight resistant potato: an unusual case

Standard food safety assessments of genetically modified crops require a thorough molecular characterization of the novel DNA as inserted into the plant that is intended for commercialization, as well as a comparison of agronomic and nutritional characteristics of the genetically modified to the non-modified counterpart. These characterization data are used to identify any unintended changes in the inserted DNA or in the modified plant that would require assessment for safety in addition to the assessment of the intended modification. An unusual case of an unintended effect discovered from the molecular characterization of a genetically modified late blight resistant potato developed for growing in Bangladesh and Indonesia is presented here. Not only was a significant portion of the plasmid vector backbone DNA inserted into the plant along with the intended insertion of an R-gene for late blight resistance, but the inserted DNA was split into two separate fragments and inserted into two separate chromosomes. One fragment carries the R-gene and the other fragment carries the NPTII selectable marker gene and the plasmid backbone DNA. The implications of this for the food safety assessment of this late blight resistant potato are considered.

     

Engineering crops, a deserving venture.

Plant transformation has had a deep impact on several aspects of basic and applied research. Genetic transformation has offered new opportunities compared to traditional breeding practises since it allows the integration into a host genome of specific sequences leading to a strong reduction of the casualness of gene transfer. One of the first target areas was plant protection against pests, pathogens and environmental stresses while the recent plant engineering programs are aimed at increasing food quality, in particular at increasing nutritional characteristics of food crops. Moreover, transgenic plants, tissue or cell cultures represent an attractive biological system for producing heterologous proteins since they offer economic and qualitative benefits. High yield production can be obtained and large-scale commercial production will take advantage of the existing infrastructure for crop cultivation, processing and storage. There are also qualitative benefits since protein synthesis secretion and post-translational modifications are similar in plants and animal cells. There are no human viral pathogens harboured by plants: thus, especially for pharmaceuticals, plants represent the safer production system. Plant transformation has become an essential instrument also for basic research, in particular for the functional characterisation of genes identified by sequencing of whole genomes. Large collections of insertion mutants have been obtained in the model plant Arabidopsis to provide a high level of genome saturation that means 95% chance of inactivating any gene at least once. To instil greater public confidence in modern plant biotechnology recent advances have already been made to overcome the potential risks for human health and environment.     

Genes in food--why the furore?

Although unprecedented and perhaps unique in its irrationality, the recent furore over genetically modified (GM) food holds extremely important lessons for scientists. Some sections of the media undoubtedly bear a heavy responsibility for giving the expression 'GM' threatening connotations that are quite unwarranted. However, influential contributions to the hysteria have come from a surprising range of other sources, including some scientists. The research community has failed in its responsibility to society in three ways. Firstly, plant scientists did not appreciate that certain techniques (such as the use of antibiotic resistance genes as markers during plant transformation) would inevitably provoke public consternation. As a result, they took no steps to address such concerns. Secondly, researchers overlooked, minimized or in some cases simply dismissed the significance of public fears that they were 'interfering with Nature' or 'playing God'. Thirdly, plant breeders apparently saw no need to take pro-active measures with regard to the media and public in placing potential environmental and nutritional benefits of GM crops on the agenda in a positive fashion. Partly because of this failure, GM food is now firmly established in the public mind as wholly objectionable. One measure of how far we have travelled down that road is that it hardly matters any more whether objections are based on alleged environmental risks of cultivating GM crops or alleged toxicological hazards of eating them. 'Genetically modified organism', like 'radioactivity', has become an odious, generic shibboleth. Given that millions of people throughout the world are already benefiting from pharmaceuticals made by GM organisms, this is bizarre.     

CRISPR/Cas‐based tools for the targeted control of plant viruses

Plant viruses are known to infect most economically important crops and pose a major threat to global food security. Currently, few resistant host phenotypes have been delineated, and while chemicals are used for crop protection against insect pests and bacterial or fungal diseases, these are inefficient against viral diseases. Genetic engineering emerged as a way of modifying the plant genome by introducing functional genes in plants to improve crop productivity under adverse environmental conditions. Recently, new breeding technologies, and in particular the exciting CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR‐associated proteins) technology, was shown to be a powerful alternative to engineer resistance against plant viruses, thus has great potential for reducing crop losses and improving plant productivity to directly contribute to food security. Indeed, it could circumvent the “Genetic modification” issues because it allows for genome editing without the integration of foreign DNA or RNA into the genome of the host plant, and it is simpler and more versatile than other new breeding technologies. In this review, we describe the predominant features of the major CRISPR/Cas systems and outline strategies for the delivery of CRISPR/Cas reagents to plant cells. We also provide an overview of recent advances that have engineered CRISPR/Cas‐based resistance against DNA and RNA viruses in plants through the targeted manipulation of either the viral genome or susceptibility factors of the host plant genome. Finally, we provide insight into the limitations and challenges that CRISPR/Cas technology currently faces and discuss a few alternative applications of the technology in virus research.     

The challenges facing scientists in the development of foods in Europe using biotechnology

The public's major concern over the introduction of genetic engineering into the food chain focuses on potential health risks. Proving that a food is safe is an impossible goal since there will always be a risk associated with eating food. Diets that are natural in every sense of the word pose risks but the general public believe that they are inherently less risky. The differences in risk between foods that are natural, as opposed to foods that are produced by the application of technology, are likely to be minute. Nor does it follow that they all lie in favour of `natural' food. The fact is that all foods pose a balance of risks and benefits but the scientific method has so far not been applied to its measurement. Only risk is emphasised and estimated, albeit with many conservative assumptions, often resulting in an emphasis on minute risk. Conventional plant products are not subject to the rigorous risk evaluations that apply to genetically modified plants. So the outcome is that the public only receive part of the information and this is the emphasis on risk of the `artificial' food. The only future for foods produced by biotechnology in Europe is if the public are persuaded of the real health benefits that will result from its application. Given the present state of nutritional knowledge, and the limited resources that are given to it, there are only a few clear examples of where nutritional improvement of plant food would bring really significant benefits. The paper highlights these with examples and indicates where further nutritional research will be required before other targets for improvement can be realised.     

Evaluation of a non-targeted "omic" approach in the safety assessment of genetically modified plants

Genetically modified plants must be approved before release in the European Union, and the approval is generally based upon a comparison of various characteristics between the transgenic plant and a conventional counterpart. As a case study, focusing on safety assessment of genetically modified plants, we here report the development and characterisation of six independently transformed ARABIDOPSIS THALIANA lines modified in the flavonoid biosynthesis. Analyses of integration events and comparative analysis for characterisation of the intended effects were performed by PCR, quantitative Real-time PCR, and High Performance Liquid Chromatography. Analysis by cDNA microarray was used as a non-targeted approach for the identification of potential unintended effects caused by the transformation. The results revealed that, although the transgenic lines possessed different types of integration events, no unintended effects were identified. However, we found that the majority of genes showing differential expression were identified as stress-related genes and that environmental conditions had a large impact on the expression of several genes, proteins, and metabolites. We suggest that the microarray approach has the potential to become a useful tool for screening of unintended effects, but state that it is crucial to have substantial information on the natural variation in traditional crops in order to be able to interpret "omics" data correctly within the framework of food safety assessment strategies of novel plant varieties, including genetically modified plant varieties.     

Nutritional consequences of the transformation from subsistence to commercial agriculture in Tabasco,Mexico

Recent changes in the area of the Plan Chontalpa in Tabasco have greatly reduced the production of subsistence crops by rural families, resulting in decreased crop diversity and a concomitant increase in the degree of dependence on outside sources of food. Results from a nutrition survey of 149 families demonstrate that dietary diversity, dietary quality, and nutritional status of preschool children are negatively associated with lower crop diversity and increased dependence on purchased foods. Dietary deterioration is illustrated by the negative relationship found between nutritional status and increased sugar consumption. The assumption that a rise in income accompanying the adoption of commercial production will automatically lead to improved nutrition is challenged: income levels were not found to be consistently related to nutritional status. Children of families that have converted to cattle production, despite greater land availability and family incomes, do not have improved nutritional status. In the study area, where wages are low and food prices are very high, the value of a higher degree of self-sufficiency in food is recognized, yet families continue to switch to cash crops due to the environmental, economic, and time constraints imposed by the system of commercial agriculture in which they participate. The solution is not to return to traditional subsistence farming, however, but to determine under what conditions a more progressive form of agricultural change can occur.     

转基因植物中标记基因的剔除

在目前的植物转化系统中,要求在关注基因或目的基因转入细胞时,同时有标记基因存在.标记基因主要是抗生素或除草剂的抗性基因.借标记基因的表达可以将转化细胞从大量的未转化细胞中筛选出来,但标记基因的继续存在,特别是在转基因食品中,是人们广泛关注的问题.培育无标记基因的转基因植株已成为植物生物工程研究中的新课题.该文介绍了剔除标记基因的两种方法:分离剔除和重组剔除,并对近年来这两种方法在培育无标记基因的转基因植物中的应用和进展作了介绍.     

Assessment of the food safety issues related to genetically modified foods

International consensus has been reached on the principles regarding evaluation of the food safety of genetically modified plants. The concept of substantial equivalence has been developed as part of a safety evaluation framework, based on the idea that existing foods can serve as a basis for comparing the properties of genetically modified foods with the appropriate counterpart. Application of the concept is not a safety assessment per se, but helps to identify similarities and differences between the existing food and the new product, which are then subject to further toxicological investigation. Substantial equivalence is a starting point in the safety evaluation, rather than an endpoint of the assessment. Consensus on practical application of the principle should be further elaborated. Experiences with the safety testing of newly inserted proteins and of whole genetically modified foods are reviewed, and limitations of current test methodologies are discussed. The development and validation of new profiling methods such as DNA microarray technology, proteomics, and metabolomics for the identification and characterization of unintended effects, which may occur as a result of the genetic modification, is recommended. The assessment of the allergenicity of newly inserted proteins and of marker genes is discussed. An issue that will gain importance in the near future is that of post-marketing surveillance of the foods derived from genetically modified crops. It is concluded, among others that, that application of the principle of substantial equivalence has proven adequate, and that no alternative adequate safety assessment strategies are available.     

Evolving challenges and strategies for fungal control in the food supply chain

Fungi that spoil foods or infect crops can have major socioeconomic impacts, posing threats to food security. The strategies needed to manage these fungi are evolving, given the growing incidence of fungicide resistance, tightening regulations of chemicals use and market trends imposing new food-preservation challenges. For example, alternative methods for crop protection such as RNA-based fungicides, biocontrol, or stimulation of natural plant defences may lessen concerns like environmental toxicity of chemical fungicides. There is renewed focus on natural product preservatives and fungicides, which can bypass regulations for ‘clean label’ food products. These require investment to find effective, safe activities within complex mixtures such as plant extracts. Alternatively, physical measures may be one key for fungal control, such as polymer materials which passively resist attachment and colonization by fungi. Reducing or replacing traditional chlorine treatments (e.g. of post-harvest produce) is desirable to limit formation of disinfection by-products. In addition, the current growth in lower sugar food products can alter metabolic routing of carbon utilization in spoilage yeasts, with implications for efficacy of food preservatives acting via metabolism. The use of preservative or fungicide combinations, while involving more than one chemical, can reduce total chemicals usage where these act synergistically. Such approaches might also help target different subpopulations within heteroresistant fungal populations. These approaches are discussed in the context of current challenges for food preservation, focussing on pre-harvest fungal control, fresh produce and stored food preservation. Several strategies show growing potential for mitigating or reversing the risks posed by fungi in the food supply chain.     

Assuring the safety of genetically modified (GM) foods: the importance of an holistic,integrative approach

Genes change continuously by natural mutation and recombination enabling man to select and breed crops having the most desirable traits such as yield or flavour. Genetic modification (GM) is a recent development which allows specific genes to be identified, isolated, copied and inserted into other plants with a high level of specificity. The food safety considerations for GM crops are basically the same as those arising from conventionally bred crops, very few of which have been subject to any testing yet are generally regarded as being safe to eat. In contrast a rigorous safety testing paradigm has been developed for GM crops, which utilises a systematic, stepwise and holistic approach. The resultant science based process, focuses on a classical evaluation of the toxic potential of the introduced novel trait and the wholesomeness of the transformed crop. In addition, detailed consideration is given to the history and safe use of the parent crop as well as that of the gene donor. The overall safety evaluation is conducted under the concept known as substantial equivalence which is enshrined in all international crop biotechnology guidelines. This provides the framework for a comparative approach to identify the similarities and differences between the GM product and its comparator which has a known history of safe use. By building a detailed profile on each step in the transformation process, from parent to new crop, and by thoroughly evaluating the significance from a safety perspective, of any differences that may be detected, a very comprehensive matrix of information is constructed which enables the conclusion as to whether the GM crop, derived food or feed is as safe as its traditional counterpart. Using this approach in the evaluation of more than 50 GM crops which have been approved worldwide, the conclusion has been that foods and feeds derived from genetically modified crops are as safe and nutritious as those derived from traditional crops. The lack of any adverse effects resulting from the production and consumption of GM crops grown on more than 300 million cumulative acres over the last 5 years supports these safety conclusions.     

Molecular Strategies for Addressing Gene Flow Problems and Their Potential Applications in Abiotic Stress Tolerant Transgenic Plants

Transgenic plant technology provides a powerful tool to improve abiotic stress tolerance of crop plants. However, introgression of stress tolerance genes into weedy relatives may increase the potential for persistence and invasiveness, resulting in undesirable ecological consequences. A variety of gene confinement strategies have been developed to reduce unwanted transgene movement. In this review, we discuss some of these strategies, such as male and female sterility, GeneSafe?, parthenocarpy, chloroplast transformation and gene deletor technologies. In the case of the gene deletor technology, all transgenes from pollen, seeds, fruits or other organs may be eliminated once the transgene functions are no longer needed at the stage when the presence of the transgene becomes a cause for ecological or public concern. The gene deletor and other technologies can be useful to reduce unintended dispersal of stress tolerance genes and thus may facilitate commercialization of transgenic crops with enhanced tolerance to abiotic stresses.     

Large‐scale molecular genetic analysis in plant‐pathogenic fungi: a decade of genome‐wide functional analysis

Plant‐pathogenic fungi cause diseases to all major crop plants world‐wide and threaten global food security. Underpinning fungal diseases are virulence genes facilitating plant host colonization that often marks pathogenesis and crop failures, as well as an increase in staple food prices. Fungal molecular genetics is therefore the cornerstone to the sustainable prevention of disease outbreaks. Pathogenicity studies using mutant collections provide immense function‐based information regarding virulence genes of economically relevant fungi. These collections are rich in potential targets for existing and new biological control agents. They contribute to host resistance breeding against fungal pathogens and are instrumental in searching for novel resistance genes through the identification of fungal effectors. Therefore, functional analyses of mutant collections propel gene discovery and characterization, and may be incorporated into disease management strategies. In the light of these attributes, mutant collections enhance the development of practical solutions to confront modern agricultural constraints. Here, a critical review of mutant collections constructed by various laboratories during the past decade is provided. We used Magnaporthe oryzae and Fusarium graminearum studies to show how mutant screens contribute to bridge existing knowledge gaps in pathogenicity and fungal–host interactions.     

The nutritional value of feeding on crops: Diets of vervet monkeys in a humanized landscape

Anthropogenic influences have dramatically altered the environments with which primates interact. In particular, the introduction of anthropogenic food sources to primate groups has implications for feeding behaviour, social behaviour, activity budgets, demography and life history. While the incorporation of anthropogenic foods can be beneficial to primates in a variety of nutritional ways including increased energetic return, they also carry risks associated with proximity to humans, such as risk of being hunted, disease risk and risk of conflict. Given such risks, we initiated a 3‐year study where we sought to understand the underlying nutritional motivations for anthropogenic food resource use by vervet monkeys (Cercopithecus aethiops) in the humanized matrix surrounding the Nabugabo Field Station in central Uganda. Feeding effort, defined as proportion of feeding scans spent on anthropogenic food, was not associated with ripe fruit availability nor with crop availability as determined by phenological monitoring. Likewise, there was no difference in the protein, fibre, or lipid composition of crop food items compared to wild food items. Individuals spent less time feeding overall in months over the 3 years with a higher proportion of time spent feeding on crop foods, suggesting a potential benefit in terms of accessibility (reduction in the proportion of activity budget devoted to feeding).