Regulatory approaches for genome edited agricultural plants in select countries and jurisdictions around the world.

Genome editing in agriculture and food is leading to new, improved crops and other products. Depending on the regulatory approach taken in each country or region, commercialization of these crops and products may or may not require approval from the respective regulatory authorities. This paper describes the regulatory landscape governing genome edited agriculture and food products in a selection of countries and regions.

Allergenicity Assessment of Novel Food Proteins: What Should Be Improved?

Allergenicity prediction is one of the most challenging aspects in the safety assessment of foods derived from either biotechnology or novel food proteins. Here we present a bottom-up strategy that defines a priori the specific risk assessment (RA) needs based on a database appropriately built for such purposes.

[Novel food sources: from GMO to the broadening of Russia's bioresource base].

The modern strategy of humanity food providing is aimed at finding the exit from the food crisis in the shortest possible time, by the end of XXI century food and feed production should increase by at least 70%. These tasks solution implies not only the use of science-oriented technologies, but also the expansion of the food base by means of novel food sources, which don't have a history of safe use. In the Russian Federation the formation of novel food's safety assessment approaches is regulated at the state level and is the most important requirement for the possibility of usage. Russian experience of the second half of the XX century in the area of novel food sources' biomedical research unites two stages. The first of them dates back to the middle of the 1960s', when the Soviet scientists, in particular, the workforce of the Institute of Nutrition of the USSR Academy of Medical Sciences, under the leadership of Academician A.A. Pokrovskii, have developed the evaluation approaches of the biological value and safety of microbial synthesized protein. The second stage of the safety assessment research development was the work with the genetically modified organisms of plant origin (GMO), that begun in the middle of the 1990s'. Since the moment of formation in 1995-1996, 9 methodical guidelines that regulate methods of safety assessment and control over GMO have been developed. Comprehensively formed by 2020, safety assessment system has been used in the framework of 27 GMO lines state registration that passed a whole cycle of medical and biological research and were allowed for use in nutrition of the population of the Eurasian Economic Union. Within the framework of these research a considerable amount of factual material has been accumulated, a regulatory and methodological basis has been built, and a substantial background for further fundamental and applied scientific research in the field of development and safety assessment of novel food has been created.

Consideration of familiarity accumulated in the confined field trials for environmental risk assessment of genetically modified soybean (Glycine max) in Japan.

To date, there have been 160 regulatory approvals for environmental safety in Japan for the major genetically modified (GM) crops, including corn, soybean, canola and cotton. Confined field trials (CFTs) have been conducted in Japan for all single events, which contain various traits. The accumulated information from these previously conducted CFTs, as well as the agronomic field study data from other countries, provides a rich source of information to establish "familiarity" with the crops. This familiarity can be defined as the knowledge gained through experience over time, and used to inform the environmental risk assessments (ERA) of new GM crops in Japan. In this paper, we compiled agronomic data from the CFTs performed in Japan for 11 GM soybean events which obtained food, feed and environmental safety approvals from regulatory agencies in Japan. These CFTs were conducted by multiple developers according to Japan regulations to support the ERA of these GM soybean, covering standard measurement endpoints evaluated across developers in Japan. With this dataset, we demonstrate how familiarity gained from the CFTs of GM soybeans in Japan can be used to inform on the ERA of new GM soybean events. By leveraging this concept of familiarity, we discuss potential enhancements to the ERA process for GM soybean events in Japan.

No country for new seeds: food and environmental security implications of Mexico's intended ban on GM crops.

An intended generalized ban on the "introduction and use of transgenic seeds" has been announced by the Mexican government, which, unlike the other agricultural programs under this administration, lacks a budget and rules of operation. In this policy brief we consider scenarios of implementing such an intended ban, de-regulating the use of genetically modified crops, or an intermediate scenario under the existing biosafety regulations. We recommend maintaining the status quo given its potential contributions to food and environmental security, in addition to a better economic outlook. However, a greater impulse needs to be given by the federal government to foment the domestic development of GM crops that are pertinent for Mexican agri-food needs, in addition to funding the development and implementation of various agro-ecological practices that increase biodiversity in food production units.

EFSA Genetically Engineered Crop Composition Equivalence Approach: Performance and Consistency.

The European Food Safety Authority (EFSA) oversees the safety assessment of genetically engineered (GE) crops in the European Union and has developed a study design and statistical approach for assessing the compositional equivalency between a GE crop and the corresponding non-GE crop on the basis of the results from a small number of concurrently grown reference lines. Confidence limits around the differences in mean analyte composition between the GE variety and the reference lines are compared with equivalence limits on the basis of the variability of the reference lines. Here, we evaluated the performance and consistency of the equivalence conclusions using a non-GE variety that is, by definition, equivalent to the non-GE crop. Using this approach across the same analytes with the same non-GE variety, it was found that equivalence could not be concluded for 19.7, 22.9, 25.4, and 53.5% of the analytes in four separate studies. In addition, equivalency conclusions for the same analyte often differed from study to study. These results call into question the consistency and value of this approach in the risk assessment of GE crops.

Comparative analysis of genetically-modified crops: Part 1. Conditional difference testing with a given genetic background.

The European Food Safety Authority (EFSA) mandates two sets of statistical tests in the comparative assessment of a genetically-modified (GM) crop: difference testing to demonstrate whether the GM crop is different from its appropriate non-traited control; and equivalence testing to demonstrate whether it is equivalent to conventional references with an history-of-safe-use. The equivalence testing method prescribed by EFSA confounds the so-called GM trait effect with genotypic differences between the reference varieties and non-traited control. Critically, these genotypic differences, which we define as a 'control background effect', are the result of conventional plant breeding. Thus, the result of EFSA equivalence testing often has little or nothing to do with the GM trait effect, which should be the sole focus of the comparative assessment. Here, an integrated method is introduced for both difference and equivalence testing that considers the differences of the three genotype groups (GM, control, and references) as a two-dimensional random variable. A novel statistical model is proposed, called the trait model, that treats the effects of the GM and control materials as fixed for their difference, and as random for their common background. For significance testing, the covariance structure of the three genotype groups is utilized to decompose the differences into the trait effect and the control background effect. The trait difference is then derived as a conditional mean, given the background effect. The comparative assessment can then focus on the conditional mean difference, which is independent of the control background effect. Furthermore, the trait model is flexible enough to include various types of genotype-by-environment (G×E) interactions inherent to the experimental design of the trial. Numerical evaluations and simulations show that this new method is substantially more efficient than the current EFSA method in reducing both Type I and Type II errors (protecting both the consumer and producer risk) after the background effect is removed from the test statistic, and successfully addresses two major criticisms (i.e. statistical model lack of G×E, and study-specific equivalence criterion) that have been raised.

The GMO90+ Project: Absence of Evidence for Biologically Meaningful Effects of Genetically Modified Maize-based Diets on Wistar Rats After 6-Months Feeding Comparative Trial.

The GMO90+ project was designed to identify biomarkers of exposure or health effects in Wistar Han RCC rats exposed in their diet to 2 genetically modified plants (GMP) and assess additional information with the use of metabolomic and transcriptomic techniques. Rats were fed for 6-months with 8 maize-based diets at 33% that comprised either MON810 (11% and 33%) or NK603 grains (11% and 33% with or without glyphosate treatment) or their corresponding near-isogenic controls. Extensive chemical and targeted analyses undertaken to assess each diet demonstrated that they could be used for the feeding trial. Rats were necropsied after 3 and 6 months. Based on the Organization for Economic Cooperation and Development test guideline 408, the parameters tested showed a limited number of significant differences in pairwise comparisons, very few concerning GMP versus non-GMP. In such cases, no biological relevance could be established owing to the absence of difference in biologically linked variables, dose-response effects, or clinical disorders. No alteration of the reproduction function and kidney physiology was found. Metabolomics analyses on fluids (blood, urine) were performed after 3, 4.5, and 6 months. Transcriptomics analyses on organs (liver, kidney) were performed after 3 and 6 months. Again, among the significant differences in pairwise comparisons, no GMP effect was observed in contrast to that of maize variety and culture site. Indeed, based on transcriptomic and metabolomic data, we could differentiate MON- to NK-based diets. In conclusion, using this experimental design, no biomarkers of adverse health effect could be attributed to the consumption of GMP diets in comparison with the consumption of their near-isogenic non-GMP controls.

Variability of Crops' Compositional Characteristics: What Do Experimental Data Show?

Common pillar across the risk assessment strategies implemented worldwide for genetically modified plants is the comparison of their compositional profile to that of conventional counterparts deemed safe. If differences are observed, those that cannot be attributed to natural variation are further evaluated for their safety relevance. This principle is clear, but its implementation is challenging. Here we first discuss the difficulties of estimating natural variation of crop-specific compositional end-points and the various attempts made, together with their advantages and limitations. Second we present the empirical distribution curves of compositional end-points for two crops bearing a large commercial interest worldwide, maize and soybean. These curves provide novel information on end-point specific variability relevant for further progressing in the risk assessment process.

Humoral and cellular immune response in Wistar Han RCC rats fed two genetically modified maize MON810 varieties for 90 days (EU 7th Framework Programme project GRACE).

The genetically modified maize event MON810 expresses a Bacillus thuringiensis-derived gene, which encodes the insecticidal protein Cry1Ab to control some lepidopteran insect pests such as the European corn borer. It has been claimed that the immune system may be affected following the oral/intragastric administration of the MON810 maize in various different animal species. In the frame of the EU-funded project GRACE, two 90-day feeding trials, the so-called studies D and E, were performed to analyze the humoral and cellular immune responses of male and female Wistar Han RCC rats fed the MON810 maize. A MON810 maize variety of Monsanto was used in the study D and a MON810 maize variety of Pioneer Hi-Bred was used in the study E. The total as well as the maize protein- and Cry1Ab-serum-specific IgG, IgM, IgA and IgE levels, the proliferative activity of the lymphocytes, the phagocytic activity of the granulocytes and monocytes, the respiratory burst of the phagocytes, a phenotypic analysis of spleen, thymus and lymph node cells as well as the in vitro production of cytokines by spleen cells were analyzed. No specific Cry1Ab immune response was observed in MON810 rats, and anti-maize protein antibody responses were similar in MON810 and control rats. Single parameters were sporadically altered in rats fed the MON810 maize when compared to control rats, but these alterations are considered to be of no immunotoxicological significance.

Development of Certified Matrix-Based Reference Material as a Calibrator for Genetically Modified Rice G6H1 Analysis.

The accurate monitoring and quantification of genetically modified organisms (GMOs) are key points for the implementation of labeling regulations, and a certified reference material (CRM) acts as the scaleplate for quantifying the GM contents of foods/feeds and evaluating a GMO analytical method or equipment. Herein we developed a series of CRMs for transgenic rice event G6H1, which possesses insect-resistant and herbicide-tolerant traits. Three G6H1 CRMs were produced by mixing seed powders obtained from homozygous G6H1 and its recipient cultivar Xiushui 110 at mass ratios of 49.825%, 9.967%, and 4.986%. The between-bottle homogeneity and within-bottle homogeneity were thoroughly evaluated with consistent results. The potential DNA degradation in transportation and shelf life were evaluated with an expiration period of at least 12 months. The property values of three CRMs (G6H1a, G6H1b, G6H1c) were given as (49.825 ± 0.448) g/kg, (9.967 ± 1.757) g/kg, and (4.986 ± 1.274 g/kg based on mass fraction ratio, respectively. Furthermore, the three CRMs were characterized with values of (5.01 ± 0.08)%, (1.06 ± 0.22)%, and (0.53 ± 0.11)% based on the copy number ratio using the droplet digital PCR method. All results confirmed that the produced G6H1 matrix-based CRMs are of high quality with precise characterization values and can be used as calibrators in GM rice G6H1 inspection and monitoring and in evaluating new analytical methods or devices targeting the G6H1 event.

Genetically Engineered Resveratrol-Enriched Rice Inhibits Neuroinflammation in Lipopolysaccharide-Activated BV2 Microglia Via Downregulating Mitogen-Activated Protein Kinase-Nuclear Factor Kappa B Signaling Pathway.

Resveratrol, a natural stilbenoid, is produced by several plants, especially grape vines. Its strong potency against obesity, metabolic disorders, vascular disease, inflammation, and various cancers has already been reported. Large amounts of wine or grapes need to be consumed to obtain the amount of resveratrol required for biological activity. Pure resveratrol at concentrations as low as 10 µM induces cytotoxicity to normal cells. To overcome these limitations, we prepared genetically modified resveratrol-enriched rice (RR). We previously reported the strong antiaging potential of RR against ultraviolet B/reactive oxygen species-induced toxicity in normal human dermal fibroblasts (NHDF). As aging is characterized by neuroinflammation and neurodegeneration, we further evaluated the role of RR against LPS-induced neuroinflammation. RR inhibited nitric oxide production and the expression of inflammatory proteins such as iNOS and COX-2. RR significantly modulated mitogen-activated protein kinase signaling, activator protein AP-1 signaling, and nuclear factor kappa B (NF-κB) mediated transcription of inflammatory proteins via inhibition of NF-κB translocation, IkB phosphorylation, and proinflammatory cytokine productions such as interleukin IL-6, IL-1ß, tumor necrosis factor alpha (TNF-α), and prostaglandin E2 (PGE2). These findings show that the strong antineuroinflammatory effects of RR can be beneficial for aging-mediated neurodegenerative conditions as well as disorders of the central nervous system caused by neuroinflammation.

Characterization of GMO or glyphosate effects on the composition of maize grain and maize-based diet for rat feeding.

INTRODUCTION: In addition to classical targeted biochemical analyses, metabolomic analyses seem pertinent to reveal expected as well as unexpected compositional differences between plant genetically modified organisms (GMO) and non-GMO samples. Data previously published in the existing literature led to divergent conclusions on the effect of maize transgenes on grain compositional changes and feeding effects. Therefore, a new study examining field-grown harvested products and feeds derived from them remains useful. OBJECTIVES: Our aim was to use a metabolomics approach to characterize grain and grain-based diet compositional changes for two GMO events, one involving Bacillus thuringiensis toxin to provide insect resistance and the other one conferring herbicide tolerance by detoxification of glyphosate. We also investigated the potential compositional modifications induced by the use of a glyphosate-based herbicide on the transgenic line conferring glyphosate tolerance. RESULTS: The majority of statistically significant differences in grain composition, evidenced by the use of 1H-NMR profiling of polar extracts and LC-ESI-QTOF-MS profiling of semi-polar extracts, could be attributed to the combined effect of genotype and environment. In comparison, transgene and glyphosate effects remained limited in grain for the compound families studied. Some but not all compositional changes observed in grain were also detected in grain-based diets formulated for rats. CONCLUSION: Only part of the data previously published in the existing literature on maize grains of plants with the same GMO events could be reproduced in our experiment. All spectra have been deposited in a repository freely accessible to the public. Our grain and diet characterization opened the way for an in depth study of the effects of these diets on rat health.

An overview of genetically modified crop governance, issues and challenges in Malaysia.

The application of agricultural biotechnology attracts the interest of many stakeholders. Genetically modified (GM) crops, for example, have been rapidly increasing in production for the last 20 years. Despite their known benefits, GM crops also pose many concerns not only to human and animal health but also to the environment. Malaysia, in general, allows the use of GM technology applications but it has to come with precautionary and safety measures consistent with the international obligations and domestic legal frameworks. This paper provides an overview of GM crop technology from international and national context and explores the governance and issues surrounding this technology application in Malaysia. Basically, GM research activities in Malaysia are still at an early stage of research and development and most of the GM crops approved for release are limited for food, feed and processing purposes. Even though Malaysia has not planted any GM crops commercially, actions toward such a direction seem promising. Several issues concerning GM crops as discussed in this paper will become more complex as the number of GM crops and varieties commercialised globally increase and Malaysia starts to plant GM crops. © 2017 Society of Chemical Industry.

Food and Feed Safety of Genetically Engineered Food Crops.

The first genetically engineered (GE) food crop (tomato) was introduced in 1995, followed by the successful development and commercial release of maize, soybeans, cotton, canola, potatoes, papaya, alfalfa, squash, and sugar beets with specific new genetic traits. Even though the safety of every new GE crop has been evaluated by various regulatory authorities throughout the world prior to its commercial release, the ongoing public debate about the safety of food and feed derived from GE plants has not abated. Such debates often overshadow an important fact that all crops used as human food or animal feed include varieties that have been developed through conventional breeding and selection over hundreds or thousands of years, or through intentional but random mutagenesis. Developing food crops through such breeding practices result in large-scale genomic changes in the resulting crops, and these genomic changes do not undergo molecular characterization. In contrast, new GE crops are developed using well-characterized DNA fragments and the resulting crops are tested and evaluated with much greater scrutiny. This document reviews the safety data and information of GE crops and foods obtained from them.

CRISPR is knocking on barn door.

Genome modification at specific loci in livestock species was only achievable by performing homologous recombination in somatic cells followed by somatic cell nuclear transfer. The difficulty and inefficiency of this method have slowed down the multiple applications of genome modification in farm animals. The discovery of site-specific endonucleases has provided a different and more direct route for targeted mutagenesis, as these enzymes allow the ablation (KO) or insertion (KI) of specific genomic sequences on a single step, directly applied to zygotes. Clustered regularly interspaced short palindromic repeats (CRISPR), the last site-specific endonuclease to be developed, is a RNA-guided endonuclease, easy to engineer and direct to a given target site. This technology has been successfully applied to rabbits, swine, goats, sheep and cattle, situating genome editing in livestock species at an attainable distance, thereby empowering scientist to develop a myriad of applications. Genetically modified livestock animals can be used as biomodels to study human or livestock physiology and disease, as bioreactors to produce complex proteins, or as organ donors for transplantation. Specifically on livestock production, genome editing in farm animals may serve to improve productive genetic traits, to improve various animal products, to confer resistance to diseases or to minimize the environmental impact on farming. In this review, we provide an overview of the current methods for site-specific genome modification in livestock species, discuss potential and already developed applications of genome edition in farm animals and debate about the possibilities for approval of products derived from gene-edited animals for human consumption.