A Framework for Postrelease Environmental Monitoring of Second‐generation Crops with Novel Traits

In preparation for a legal implementation of EU-regulation 1829/2003, the Norwegian Scientific Committee for Food Safety (VKM) has been requested by the Norwegian Directorate for Nature Management to conduct final environmental risk assessments for all genetically modified organisms (GMOs) and products containing or consisting of GMOs that are authorized in the European Union under Directive 2001/18/EC or Regulation 1829/2003/EC. The assignment includes a scientific environmental risk assessment of oilseed rape T45 (Reference EFSA/GMO/UK/2005/25) from Bayer CropScience for food and feed uses, import and processing. Oilseed rape T45 has previously been risk assessed by the VKM Panel on Genetically Modified Organisms (GMO), commissioned by the Norwegian Food Safety Authority related to the EFSAs public hearing in 2007 (VKM 2007a).
 Food additives produced from T45 oilseed rape were notified in the EU as existing food additives within the meaning of Article 8 (1)(b) of Regulation 1829/2003, authorized under Directive 89/10/EEC (Community Register 2005). Feed materials produced from T45 were also notified as existing feed products containing, consisting of or produced from T45 according to Articles 8 and 20 of Regulation (EC) No 1829/2003 in 2003. 
 A notification for placing on the market of T45 according to the Directive 2001/18/EC was submitted in March 2004 (C/GB/04/M5/4), covering import and processing of T45 into food and feed. The application was further transferred into Regulation (EC) No 1829/2003 in November 2005 (EFSA/GMO/UK/2005/25). An application for renewal of authorisation for continued marketing of food additives and feed materials produced from T45 oilseed rape was submitted under Regulation (EC) No 1829/2003 in 2007 (EFSA/GMO/RX/T45). The EFSA GMO Panel performed one single comprehensive risk assessment for all intended uses of genetically modified oilseed rape T45, and issued a comprehensive scientific opinion for both applications submitted under Regulation (EC) No 1829/2003. The scientific opinion was published in January 30 2008 (EFSA 2008), and food and feed products containing or produced from oilseed rape T45 was approved by Commission Decision 26 March 2009 (Commission Decision 2009/184/EC). 
 The oilseed rape T45 is however currently being phased out (EU-COM 2009). The commercialisation of T45 oilseed rape seeds in third countries was stopped after the 2005 planting season and stocks of all oilseed rape T45 lines have been recalled from distribution and destroyed. The applicant commits not to commercialize the event in the future and the import will therefore be restricted to adventitious levels in oilseed rape commodity. Thus the incidence of oilseed rape T45 in the EU is expected to be limited.
 The environmental risk assessment of the oilseed rape T45 is based on information provided by the notifier in the application EFSA/GMO/UK/2005/25 and EFSA/GMO/RX/T45, and scientific comments from EFSA and other member states made available on the EFSA website GMO Extranet. The risk assessment also considered other peer-reviewed scientific literature as relevant. 
 The VKM GMO Panel has evaluated T45 with reference to its intended uses in the European Economic Area (EEA), and according to the principles described in the Norwegian Food Act, the Norwegian Gene Technology Act and regulations relating to impact assessment pursuant to the Gene Technology Act, Directive 2001/18/EC on the deliberate release into the environment of genetically modified organisms, and Regulation (EC) No 1829/2003 on genetically modified food and feed. The Norwegian Scientific Committee for Food Safety has also decided to take account of the appropriate principles described in the EFSA guidelines for the risk assessment of GM plants and derived food and feed (EFSA 2006, 2011a), the environmental risk assessment of GM plants (EFSA 2010), the selection of comparators for the risk assessment of GM plants (EFSA 2011b), and for the post-market environmental monitoring of GM plants (EFSA 2006, 2011c). 
 The scientific risk assessment of oilseed rape T45 include molecular characterisation of the inserted DNA and expression of target proteins, comparative assessment of agronomic and phenotypic characteristics, unintended effects on plant fitness, potential for horizontal and vertical gene transfer, and evaluations of the post-market environmental plan.
 In line with its mandate, VKM emphasised that assessments of sustainable development, societal utility and ethical considerations, according to the Norwegian Gene Technology Act and Regulations relating to impact assessment pursuant to the Gene Technology Act, shall not be carried out by the Panel on Genetically Modified Organisms. 
 The glufosinate ammonium-tolerant oilseed rape transformation event T45 (Unique Identfier ACSBNØØ8-2) was developed by Agrobacterium-mediated transformation of protoplast from the conventional oilseed rape cultivar “AC Excel”. T45 contains a synthetic version of the native pat gene isolated from the bacteria Streptomyces viridochromogenes, strain Tü 494. The inserted gene encodes the enzyme phosphinothricin acetyltransferase (PAT), which confers tolerance to the herbical active substance glufosinate ammonium. The PAT enzyme detoxifies glufosinate-ammonium by acetylation of the L-isomer into N-acetyl-L-glufosinate ammonium (NAG) which does not inhibit glutamine synthetase and therefore confers tolerance to the herbicide.
 Glufosinate ammonium-tolerant oilseed rape transformation event T45 has been conventionally bred into an array of spring-type oilseed rape varieties.
 Molecular characterization:
 The molecular characterisation data established that only one copy of the gene cassette is integrated in the oilseed rape genomic DNA. Appropriate analysis of the integration site including sequence determination of the inserted DNA and flanking regions and bioinformatics analysis have been performed. Bioinformatics analyses of junction regions demonstrated the absence of any potential new ORFs coding for known toxins or allergens. The genetic stability of transformation event T45 was demonstrated at the genomic level over multiple generations by Southern analysis. Segregation analysis shows that event T45 is inherited as dominant, single locus trait. Phenotypic stability has been confirmed by stable tolerance to the herbicide for T45 lines and varieties derived from the event grown in Canada since 1993. 
 Oilseed rape transformation event T45 and the physical, chemical and functional characteristics of the proteins have previously been evaluated by The VKM Panel on Genetically Modified Organisms, and considered satisfactory (VKM 2007a).
 Comparative assessment:
 Based on results from comparative analyses of data from field trials located at representative sites and environments in Canada in 1995-1997, it is concluded that oilseed rape T45 is agronomically and phenotypically equivalent to the conventional counterpart and commercial available reference varieties, with the exception of maturity and the herbicide tolerance conferred by the PAT protein. The field evaluations support a conclusion of no phenotypic changes indicative of increased plant weed/pest potential of event T45 compared to conventional oilseed rape. Furthermore, the results demonstrate that in-crop applications of glufosinate herbicide do not alter the phenotypic and agronomic characteristics of event T45 compared to conventional oilseed rape.
 Environmental risk:
 According to the applicant, the event T45 has been phased out, and stocks of all oilseed rape T45 lines have been recalled from distribution and destroyed since 2005. However, since future cultivation and import of oilseed rape T45 into the EU/EEA area cannot be entirely ruled out, the environmental risk assessment consider exposure of viable seeds of T45 through accidental spillage into the environment during transportation, storage, handling, processing and use of derived products.
 Oilseed rape is mainly a self-pollinating species, but has entomophilous flowers capable of both self- and cross-pollinating. Normally the level of outcrossing is about 30%, but outcrossing frequencies up to 55% are reported. 
 Several plant species related to oilseed rape that are either cultivated, occurs as weeds of cultivated and disturbed lands, or grow outside cultivation areas to which gene introgression from oilseed rape could be of concern. These are found both in the Brassica species complex and in related genera. A series of controlled crosses between oilseed rape and related taxa have been reported in the scientific literature. Because of a mismatch in the chromosome numbers most hybrids have a severely reduced fertility. Exceptions are hybrids obtained from crosses between oilseed rape and wild turnip (B. rapa ssp. campestris) and to a lesser extent, mustard greens (B. juncea), where spontaneously hybridising and transgene introgression under field conditions have been confirmed. Wild turnip is native to Norway and a common weed in arable lowlands.
 There is no evidence that the herbicide tolerant trait results in enhanced fitness, persistence or invasiveness of oilseed rape T45, or hybridizing wild relatives, compared to conventional oilseed rape varieties, unless the plants are exposed to herbicides with the active substance glufosinate ammonium. Glufosinate ammonium-containing herbicides have been withdrawn from the Norwegian market since 2008, and the substance will be phased out in the EU in 2017 for reasons of reproductive toxicity.
 Accidental spillage and loss of viable seeds of T45 during transport, storage, handling in the environment and processing into derived products is, however, likely to take place over time, and the establishment of small populations of oilseed rape T45 cannot be excluded. Feral oilseed rape T

Assessment of new scientific elements supporting the prolongation of prohibition of the placing on the market of genetically modified oilseed rape GT73 for food and feed purposes in Austria

Assessment of new scientific elements supporting the prolongation of prohibition of the placing on the market of genetically modified oilseed rape Ms8, Rf3 and Ms8×Rf3 for food and feed purposes in Austria

Environmental Risk Assessment of Glufosinate-Tolerant Genetically Modified Oilseed Rape MS8, RF3 and MS8 x RF3 for Import, Processing and Feed Uses under Directive 2001/18/EC (Notification C/BE/96/01)

Food, Feed and Environmental Risk Assessment of Glufosinatetolerant Genetically Modified Oilseed Rape T45 for Food and Feed Uses, Import and Processing Under Regulation (EC) No 1829/2003 (Application EFSA/GMO/UK/2005/25)

Crop Ferality and Volunteerism

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.

The environmental risk assessment of the herbicide tolerant genetically modified oilseed rape MON 88302 (Reference EFSA/GMO/BE/2011/101) has been performed by the Panel on Genetically Modified Organisms (GMO) of the Norwegian Scientific Committee for Food Safety (VKM). VKM has been requested by the Norwegian Directorate for Nature Management and the Norwegian Food Safety Authority to issue a preliminary scientific opinion on the safety of the genetically modified oilseed rape MON 88302 (Unique identifier MON-88Ø2-9) for food and feed uses, import and processing, and submit relevant scientific comments or questions to EFSA on the application EFSA/GMOBE/2011/101. 
 The environmental risk assessment of the MON 88302 is based on information provided by the applicant in the application EFSA/GMO/BE/2011/101, and scientific comments from EFSA and other member states made available on the EFSA website GMO Extranet. The risk assessment also considered peer-reviewed scientific literature as relevant. 
 The VKM GMO Panel has evaluated MON 88302 with reference to its intended uses in the European Economic Area (EEA), and according to the principles described in the Norwegian Food Act, the Norwegian Gene Technology Act and regulations relating to impact assessment pursuant to the Gene Technology Act, Directive 2001/18/EC on the deliberate release into the environment of genetically modified organisms, and Regulation (EC) No 1829/2003 on genetically modified food and feed. The Norwegian Scientific Committee for Food Safety has also decided to take account of the appropriate principles described in the EFSA guidelines for the risk assessment of GM plants and derived food and feed (EFSA 2006, 2011a), the environmental risk assessment of GM plants (EFSA 2010), the selection of comparators for the risk assessment of GM plants (EFSA 2011b), and for the post-market environmental monitoring of GM plants (EFSA 2006, 2011c). 
 The scientific risk assessment of oilseed rape MON 88302 include molecular characterisation of the inserted DNA and expression of target proteins, comparative assessment of agronomic and phenotypic characteristics, unintended effects on plant fitness, potential for horizontal and vertical gene transfer, and evaluations of the post-market environmental plan.
 In line with its mandate, VKM emphasized that assessments of sustainable development, societal utility and ethical considerations, according to the Norwegian Gene Technology Act and Regulations relating to impact assessment pursuant to the Gene Technology Act, shall not be carried out by the Panel on Genetically Modified Organisms. The GMO Panel has therefore not considered possible health and environmental effects of cultivation and processing of oilseed rape MON 88302 outside the EU/EEA area.
 The genetically modified oilseed rape MON 88302 was developed to provide tolerance to the herbical active substance glyphosate by the introduction of a gene coding for the enzyme 5enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium tumefaciens, strain CP4 (CP4 EPSPS). Glyphosate is a non-selective herbicide and is normally phytotoxic to a broad range of plants. Its mode of action occurs by binding to and inactivating the EPSPS protein, which is a key enzyme in the shikimate pathway that leads to the biosynthesis of the aromatic amino acids tyrosine, tryptophan and phenylalanine. The disruption of this pathway and the resulting inability to produce key amino acids prevents growth and ultimately leads to plant death. 
 Molecular characterization:
 The VKM Panel on Genetically Modified Organisms find the conclusion that no major section of the T-DNA plasmid backbone is inserted in MON88302 oilseed rape justified. We also find it justified that there is only one major T-DNA insert in MON88302. 
 Comparative assessment:
 Based on results from comparative analyses of data from field trials located at representative sites and environments in the USA, Canada and Chile, it is concluded that oilseed rape MON 88302 is agronomically and phenotypically equivalent to the conventional counterpart and commercial available reference varieties, with the exception of the herbicide tolerance conferred by the CP4 EPSPS protein. The field evaluations support a conclusion of no phenotypic changes indicative of increased plant weed/pest potential of MON 88302 compared to conventional oilseed rape. Furthermore, the results demonstrate that in-crop applications of glyphosate herbicide do not alter the phenotypic and agronomic characteristics of MON 88302 compared to conventional oilseed rape.
 Evaluations of environmental interactions between genetically modified oilseed rape MON 88302 and the biotic and abiotic environment, and studies of seed dormancy, seed germination, pollen morphology and viability indicates no unintended effects of the introduced trait on these characteristics in MON 88302 oilseed rape. 
 Environmental risk:
 Considering the scope of the application EFSA/GMO/BE/2011/101, excluding cultivation purposes, the environmental risk assessment is limited to exposure through accidental spillage of viable seeds of MON 88302 into the environment during transportation, storage, handling, processing and use of derived products.
 Oilseed rape is mainly a self-pollinating species, but has entomophilous flowers capable of both self- and cross-pollinating. Normally the level of outcrossing is about 30 %, but outcrossing frequencies up to 55 % are reported. 
 Several plant species related to oilseed rape that are either cultivated, occurs as weeds of cultivated and disturbed lands, or grow outside cultivation areas to which gene introgression from oilseed rape could be of concern. These are found both in the Brassica species complex and in related genera. A series of controlled crosses between oilseed rape and related taxa have been reported in the scientific literature. Because of a mismatch in the chromosome numbers most hybrids have a severely reduced fertility. Exceptions are hybrids obtained from crosses between oilseed rape and wild turnip (B. rapa ssp. campestris) and to a lesser extent, mustard greens (B.juncea), where spontaneously hybridising and transgene introgression under field conditions have been confirmed. Wild turnip is native to Norway and a common weed in arable lowlands.
 There is no evidence that the herbicide tolerant trait results in enhanced fitness, persistence or invasiveness of oilseed rape MON 88302, or hybridizing wild relatives, compared to conventional oilseed rape varieties, unless the plants are exposed to glyphosate-containing herbicides.
 However, accidental spillage and loss of viable seeds of MON 88302 during transport, storage, handling in the environment and processing into derived products is likely to take place over time, and the establishment of small populations of oilseed rape MON 88302 on locations where glyphosate is frequently applied to control weeds e.g. on railway tracks, cannot be excluded. Feral oilseed rape MON 88302 arising from spilled seed could theoretically pollinate conventional crop plants if the escaped populations are immediately adjacent to field crops, and shed seeds from cross-pollinated crop plants could emerge as GM volunteers in subsequent crops. However, both the occurrence of feral oilseed rape resulting from seed import spills and the introgression of genetic material from feral oilseed rape populations to wild populations are likely to be low in an import scenario. Apart from the glyphosate tolerance trait, the resulting progeny will not possess a higher fitness and will not be different from progeny arising from cross-fertilisation with conventional oilseed rape varieties.
 The VKM GMO Panel concludes that this route of gene flow would not introduce significant numbers of transgenic plants into agricultural areas or result in any environmental consequences in Norway.
 The environmental risk assessment will be completed and finalized by the VKM Panel on Genetically Modified Organisms when requested additional information from the applicant is available.

Food/Feed and Environmental Risk Assessment of Genetically Modified Glufosinate-tolerant Oilseed Rape MS8, RF3 and MS8 x RF3 for Import, Processing and Feed Uses under Directive 2001/18/EC (Notification C/BE/96/01)

Confined field trials (CFT) of genetically engineered (GE) crops are used to generate data to inform environmental risk assessments (ERA). ERAs are required by regulatory authorities before novel GE crops can be released for cultivation. The transportability of CFT data to inform risk assessment in countries other than those where the CFT was conducted has been discussed previously in an analysis showing that the primary difference between CFT locations potentially impacting trial outcomes is the physical environment, particularly the agroclimate. This means that data from trials carried out in similar agroclimates could be considered relevant and sufficient to satisfy regulatory requirements for CFT data, irrespective of the country where the CFTs are conducted. This paper describes the development of an open-source tool to assist in determining the transportability of CFT data. This tool provides agroclimate together with overall crop production information to assist regulators and applicants in making informed choices on whether data from previous CFTs can inform an environmental risk assessment in a new country, as well as help developers determine optimal locations for planning future CFTs. The GEnZ Explorer is a freely available, thoroughly documented, and open-source tool that allows users to identify the agroclimate zones that are relevant for the production of 21 major crops and crop categories or to determine the agroclimatic zone at a specific location. This tool will help provide additional scientific justification for CFT data transportability, along with spatial visualization, to help ensure regulatory transparency.

The environmental risk assessment (ERA) of genetically modified (GM) crops in Japan requires collecting data from a comparative study of a GM and non-GM control in an in-country confined field trial (CFT). This in-country CFT requirement is used to address concerns that differences in the local environmental conditions may lead to differences in growth and/or risks of GM crops. However, this requirement for in-country CFT has recently been exempted for certain GM maize and GM cotton traits, and instead CFT data from other countries are used to inform the ERA of these GM events. However, in-country CFTs continue to be required for GM B. napus. Our objective is to assess whether using B. napus as a host crop increases the potential for differences between GM B. napus and conventional B. napus that may have an impact on biodiversity occurring only under the Japanese environment. In this paper agronomic data was compiled from seven local CFTs of GM B. napus events to assess the potential for differences between GM and non-GM B. napus for three key areas; competitiveness, potential to produce harmful substances, and outcrossing. Considering these elements, the need for conducting CFTs locally for ERA of future GM B. napus traits is discussed. The assessment concluded that conducting CFT locally is not necessary for GM B. napus events if traits do not bring competitive advantage or produce harmful substances only under Japanese environment.

It is commonly held that confined field trials (CFTs) used to evaluate the potential adverse environmental impacts of a genetically engineered (GE) plant should be conducted in each country where cultivation is intended, even when relevant and potentially sufficient data are already available from studies conducted elsewhere. The acceptance of data generated in CFTs “out of country” can only be realized in practice if the agro-climatic zone where a CFT is conducted is demonstrably representative of the agro-climatic zones in those geographies to which the data will be transported. In an attempt to elaborate this idea, a multi-disciplinary Working Group of scientists collaborated to develop a conceptual framework and associated process that can be used by the regulated and regulatory communities to support transportability of CFT data for environmental risk assessment (ERA). As proposed here, application of the conceptual framework provides a scientifically defensible process for evaluating if existing CFT data from remote sites are relevant and/or sufficient for local ERAs. Additionally, it promotes a strategic approach to identifying CFT site locations so that field data will be transportable from one regulatory jurisdiction to another. Application of the framework and process should be particularly beneficial to public sector product developers and small enterprises that develop innovative GE events but cannot afford to replicate redundant CFTs, and to regulatory authorities seeking to improve the deployment of limited institutional resources.

Sources of genetic diversity in feral oilseed rape (Brassica napus) populations

Aim. The aim of the work was to compare the polymorphism of commercial varieties and populations of B. napus growing outside cultivation to assess the genetic diversity of feral rapeseed populations in Belarus. Methods. The study assessed genetic diversity according to the data of 7 microsatellite loci genotyping. Results. The results indicate a greater genetic diversity in feral oilseed rape populations. An analysis of the structure of the genotypes distribution in the STRUCTURE software showed the division into three clusters – commertial varieties, feral populations and samples of B. rapa. Conclusions. The established genetic divergence between feral populations and commercial varieties indicates that feral oilseed rape is able to maintain persistent populations in Belarus. In practice, this should be taken into account when assessing the environmental risk when transgenic rape is released into the environment. And in the cultivation of transgenic rapeseed, special attention should be paid to measures to prevent the occurrence of its free-growing populations. Keywords: oilseed rape, feral populations, microsatellite loci, genetic diversity.

<abstract> <p>The miraculin transgenic tomato is a genetically modified (GM) crop that can be used as an alternative for low calories food and a natural non-sugar sweetener. Before the release and distribution, transgenic crop needs to go through an environmental risk assessment (ERA) as a backbone to achieve biosafety. Comparative analysis is a general principle of ERA to identify differences between transgenic crop and its non-transgenic counterpart which may indicate substantial equivalence and unintended effects. This experiment was aimed to compare the agronomic, compositional, and physiological characteristics of miraculin transgenic tomato cv. Moneymaker with non-transgenic tomato. The data obtained were plant height, stem diameter, relative growth rate, chlorophyll content, stomatal conductance, days to 50% flowering, days to fruit maturity, a number of flowers per cluster, a number of fruits per cluster, a number of fruits per plant, fruit weight, fruit diameter, harvest index, total dissolved solids, fructose, glucose, and sucrose contents, and total carotenoids, lycopene, and β-carotene contents. This study found that there were no significant differences between miraculin transgenic and non-transgenic tomato in all variables observed. It suggests that miraculin transgenic tomato is equivalent to its counterpart and unintended effects are not detected as.</p> </abstract>