Elisabeth Waigmann

RNAi‐based GM plants: food for thought for risk assessors

RNA interference (RNAi) is an emerging technology that offers new opportunities for the generation of new traits in genetically modified (GM) plants. Potential risks associated with RNAi-based GM plants and issues specific to their risk assessment were discussed during an international scientific workshop (June 2014) organized by the European Food Safety Authority (EFSA). Selected key outcomes of the workshop are reported here.

90-day rodent feeding studies on whole GM food/feed: Is the mandatory EU requirement for 90-day rodent feeding studies on whole GM food/feed fit for purpose and consistent with animal welfare ethics?

Genetically modified (GM) plants and derived food and feed products are subject to risk assessment and regulatory approval before these can be marketed in the European Union (EU). This pre‐market risk assessment follows international standards to evaluate whether characteristics of the GM plant associated with the genetic modification are capable of causing adverse effects on human and animal health and the environment (hereafter referred to as hazards) [1]. Such hazards can arise directly from the novel trait intentionally introduced in the GM plant, or from unintended changes (Sidebar A). As unintended changes are, by their nature, generally unexpected, it is difficult to foresee them based on a specific risk hypothesis and test for them directly [2]. The identification of unintended changes in GM plants is therefore based on the molecular, agronomic/phenotypic and compositional characterisation of the GM plant (Sidebar B) [1], [2]. Any unintended changes identified are then assessed further for their potential to cause harm to human and animal health or the environment. > … safety assessment should focus on known hazards associated with the novel traits […] in order to evaluate plausible risks instead of unknown ones Since 8 December 2013, the Implementing Regulation (EU) No. 503/2013 provides detailed instructions for the risk assessment of GM food/feed in accordance with Regulation (EC) No. 1829/2003. As part of the toxicological assessment of GM plants, the Implementing Regulation now requires a mandatory 90‐day feeding study in rodents on whole GM food/feed in order “ to identify potential adverse effects on the whole GM food/feed, or address remaining uncertainties .” The mandatory nature of this requirement introduced a significant shift in the approach to 90‐day rodent feeding studies by necessitating such a study even when no specific risk hypothesis can be formulated, that is …

EFSA Guidelines on the environmental risk assessment of genetically modified animals in the EU: the process and risk assessment considerations

The scientific Panel of the European Food Safety Authority (EFSA) on genetically modified organisms (GMOs) developed guidelines for the environmental risk assessment of genetically modified (GM) animals that provide specific recommendations for GM fish, GM insects, GM mammals and GM birds. The guidelines advocate that the environmental risk assessment of GM animals is conducted on a case-by-case basis, in a scientifically sound and transparent manner, and follows common methodology for the identification, gathering and interpretation of the data. Besides specific considerations, the guidelines provide generic advice on the choice of comparators, the use of non-GM surrogates, the assessment of long-term effects, uncertainty analysis, modelling requirements and statistical principles. Three dedicated Working Groups of the EFSA GMO Panel were involved in the elaboration of the environmental risk assessment guidelines, which underwent a public consultation before finalisation. Relevant comments received from risk assessment bodies of EU Member States, the scientific community, stakeholders and the public were considered by the Working Groups. The Working Groups also took into account the external scientific reports on GM fish, GM insects, GM mammals and GM birds commissioned by EFSA. These reports provided background information by mapping relevant fields of expertise and identifying essential elements to be considered when performing an environmental risk assessment of GM animals.

Predictive Protein Toxicity and Its Use in Risk Assessment

In the EU novel proteins used in food or feed are assessed for their potential toxic effects in humans and livestock animals. The discovery of clear molecular features linked to the toxicity of a protein may be an important step towards the use of predictive protein toxicity in risk assessment.

Assessment of the potential integration of the DNA plasmid vaccine CLYNAV into the salmon genome

Abstract The European Commission mandated EFSA to review a new data package provided by the company Elanco, for the possible integration/non‐integration of the DNA plasmid vaccine CLYNAV into the genome of Atlantic salmon (Salmo salar) and to indicate whether EFSA agrees with the conclusions drawn by Elanco. The vaccine is injected into fish to confer protection against pancreas disease caused by the salmonid alphavirus. The majority of the experimental data provided by the company was for muscle tissue close to the injection site and for gonadal tissue. EFSA considers that the long persistence of DNA plasmid in muscle tissue close to the injection site and the potential heritability of an integration event in gonad cells support the focus of the assessment on both these tissues. The experimental data did not provide scientifically robust evidence for a true integration event. The company overall concluded that the likelihood of integration is negligible, based on considerations in the context of the companys environmental risk assessment, but did not provide a quantitative value for the rate of integration linked to the term ‘negligible’. It is therefore not possible to evaluate this statement specifically with regard to integration rates. EFSA notes that knowledge about homologous and non‐homologous integration predicts that integration could occur with certain frequency. Therefore, EFSA has constructed worst‐case scenarios leading to upper estimates for possible integration rates of the DNA plasmid vaccine into the Atlantic salmon genome. EFSA concludes that, based on the worst‐case scenarios described here and taking into account additional factors decreasing the likelihood of integration, the actual integration rate is likely to be orders of magnitude lower than the upper estimated integration rate calculated in the context of the worst‐case scenarios. With the available evidence, the actual integration rate cannot be estimated with more precision.

Annual post‐market environmental monitoring report on the cultivation of genetically modified maize MON 810 in 2016

Abstract Following a request from the European Commission, EFSA assessed the annual post‐market environmental monitoring (PMEM) report for the 2016 growing season of the Cry1Ab‐expressing maize event MON 810 provided by Monsanto Europe S.A. Partial compliance with refuge requirements was reported in Spain, as observed in previous years. EFSA reiterates the need to achieve full compliance in areas of high maize MON 810 adoption to delay resistance evolution, and therefore advocates increasing the level of compliance in such areas. Resistance monitoring data do not indicate a decrease in susceptibility to the Cry1Ab protein in the field corn borer populations tested in the 2016 season. However, EFSA identified some methodological and reporting limitations pertaining to resistance monitoring that need improvement in future PMEM reports. No complaints related to corn borer infestation of maize MON 810 were received via the farmer alert system during the 2016 cultivation season. EFSA encourages the consent holder to provide more information on this complementary resistance monitoring tool. The data on general surveillance do not indicate any unanticipated adverse effects on human and animal health or the environment arising from the cultivation of maize MON 810. EFSA reiterates its recommendations on the methodology and analysis of farmer questionnaires, and considers that future literature searches on maize MON 810 performed in the context of annual PMEM reports should follow the guidelines given in the 2017 EFSA explanatory note on literature searching. Moreover, EFSA encourages relevant stakeholders to implement a methodological framework that enables the use of existing networks in the broader context of environmental monitoring. EFSA concludes that no new evidence has been reported in the 2016 PMEM report that would invalidate previous EFSA evaluations on the safety of maize MON 810.

Guidance for the risk assessment of the presence at low level of genetically modified plant material in imported food and feed under Regulation (EC) No 1829/2003

Abstract This document provides guidance for the risk assessment under Regulation (EC) No 1829/2003 of the unintended, adventitious or technically unavoidable presence in food and feed of low level of genetically modified plant material intended for markets other than in the European Union. In this context, the presence at low level is defined to be maximum 0.9% of genetically modified plant material per ingredient. This guidance is intended to assist applicants by indicating which scientific requirements of Annex II of Regulation (EU) No 503/2013 are considered necessary for the risk assessment of the presence at low levels of genetically modified plant material in food and feed.

Risk assessment of new sequencing information on genetically modified soybean event 40‐3‐2

Abstract The GMO Panel has previously assessed genetically modified (GM) soybean 40‐3‐2 as a single event and as part of a two‐event stack, 305423 × 40‐3‐2. These soybean events were found to be as safe as their conventional counterparts and other appropriate comparators with respect to potential effects on human and animal health and the environment. On 4 April 2017, European Commission requested EFSA to analyse new nucleic acid sequencing and updated bioinformatics data for soybean event 40‐3‐2 and to indicate whether the conclusions of the GMO Panel on the previously assessed GM soybeans remain valid. The new sequencing data indicated that, the sequence of soybean event 40‐3‐2 as present in the stacked soybean 305423 × 40‐3‐2 contains an additional nucleotide in the 5′ flanking region of a 72 bp additional insert of CP4 EPSPS present in soybean 40‐3‐2. Re‐examination of the original sequencing data of the single soybean event 40‐3‐2 by the applicant, indicated that this additional nucleotide was already present in the original plant material used for the risk assessment of soybean event 40‐3‐2. Thus, with the exception of bioinformatics analyses, the studies performed for the risk assessment of the single event soybean 40‐3‐2 and the two‐event stack soybean 305423 × 40‐3‐2 remain valid. The updated bioinformatic analyses performed on the corrected sequence did not give rise to safety issues. Therefore, EFSA concludes, based on the information provided, that the original risk assessment of the soybean event 40‐3‐2 as a single and the stacked soybean 305423 × 40‐3‐2 remains valid.

Risk assessment of new sequencing information on genetically modified soybean event 305423

Abstract The GMO Panel has previously assessed genetically modified (GM) soybean 305423 as a single event and as part of a two‐event stack, 305423 × 40‐3‐2. These soybean events were found to be as safe as their conventional counterparts and other appropriate comparators with respect to potential effects on human and animal health and the environment. On 23 February 2017, European Commission requested EFSA to analyse new nucleic acid sequencing data and updated bioinformatics data for soybean event 305423 and to indicate whether the previous conclusions of the GMO Panel on the previously assessed GM soybeans remain valid. The new sequencing data indicated a four base pair (bp) difference compared to the sequencing data originally provided: one bp located in the genomic 3′ flanking region, two bp located in a gene silencing cassette and one bp in a partial promoter. These bp reported as differences in the new nucleic acid sequencing data on soybean event 305423 were already present in the original plant material used for the risk assessment. Thus, with the exception of bioinformatics analyses, including an off‐target search with the dsRNA expression cassette, the studies performed for the risk assessment of the single event soybean 305423 and the two‐event stack soybean 305423 × 40‐3‐2 remain valid. The new sequencing data and the bioinformatic analyses performed on the new sequence including the RNAi off‐target search, did not give rise to safety issues. Therefore, EFSA concludes that the original risk assessment of the single soybean event 305423 and the two‐event stack soybean 305423 × 40‐3‐2 remains valid.