Koller, F. and Cieslak, M. (2023) → A perspective from the EU: unintended genetic changes in plants caused by NGT—their relevance for a comprehensive molecular characterisation and risk assessment. Front. Bioeng. Biotechnol. 11:1276226. doi: 10.3389/fbioe.2023.1276226
– October 2023 – Several regions in the world are currently holding discussions in regard to the regulation of new genomic techniques (NGTs) and their application in agriculture. The European Commission, for instance, is proposing the introduction of specific regulation for NGT plants. Various questions need to be answered including e.g., the extent to which NGT-induced intended and unintended genetic modifications must be subjected to a mandatory risk assessment as part of an approval procedure. This review mostly focuses on findings in regard to unintended genetic changes that can be caused by the application of NGTs. → read more
Koller, F., Schulz, M., Juhas, M., Bauer-Panskus, A., Then, C. (2023) → The need for assessment of risks arising from interactions between NGT organisms from an EU perspective. Environ Sci Eur 35, 27 (2023). doi: 10.1186/s12302-023-00734-3
New genomic techniques (NGTs) allow new genotypes and traits to be developed in different ways and with different outcomes compared to previous genetic engineering methods or conventional breeding (including non-targeted mutagenesis). EU GMO regulation requires an assessment of their direct and indirect effects that may be immediate, delayed or cumulative. Such effects may also result from the interactions of NGT organisms simultaneously present in a shared receiving environment or emerge from a combination of their traits. This review elaborates such potential interactions based on a literature review and reasoned scenarios to identify possible pathways to harm.
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Kawall, K. (2021) → The Generic Risks and the Potential of SDN-1 Applications in Crop Plants. Plants 2021, 10, 2259. doi: 10.3390/plants10112259
The publication focuses on so-called SDN-1 ‘gene scissor’ applications, such as CRISPR/Cas, that are used to make small genetic changes in the genome of target organisms. The applications can, however, lead to significant changes in plant metabolism and constituents, especially if they are made repeatedly and in combination. The publication provides an overview of complex SDN-1 applications (i.e. multiplexing and alteration of multiple gene variants) as well as single point mutations in market-oriented crops. For this purpose, the current study analysed a dataset previously published by Modrzejewski et al. 2020, and divided induced SDN-1 changes into three categories, i.e. multiplexing, the alteration of multiple gene variants and the alteration of single genes. It was found that SDN-1 applications knocked out single genes in slightly more than half of the studies examined. Among them is the ‘CRISPR tomato’ which has an increased content of an antihypertensive ingredient (i.e. GABA) and is already approved in Japan. This latter case highlights the fact that conventional breeding was not able to change the relevant genes in the same way that was possible using gene scissors. More complex SDN-1 applications were carried out in other studies, e.g. the silencing of several different genes.
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Kawall, K. (2021) → Genome-edited Camelina sativa with a unique fatty acid content and its potential impact on ecosystems. Environmental Sciences Europe 33 (1):38. doi:10.1186/s12302-021-00482-2
The study provides an overview of unwanted and unexpected effects that a release of genome-edited plants can have on ecosystems. These result from the intended traits introduced by genome editing processes, such as CRISPR/Cas gene scissors, that can influence various metabolic processes. CRISPR/Cas has greatly increased the possibilities and the speed with which the genome of plants can be changed, regardless of whether additional genes are integrated into the genome. Even small genetic changes that may be induced several times and in combination with so-called SDN-1 applications, can significantly change metabolic pathways and ingredients.
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Kawall, K., Cotter, J., Then, C. (2020) →Broadening the GMO risk assessment in the EU for genome editing technologies in agriculture. Environ Sci Eur 32, 106. doi: 10.1186/s12302-020-00361-2
The study describes the risks associated with the use of new genetic engineering methods in plants and animals. On the one hand, these are unintended changes in the genome triggered by the genetic engineering process and, on the other hand, intended interventions resulting from genome editing that are also associated with risks.
The study gives an overview of genome editing techniques and describes unintended effects that are specific to their application. Such unintended effects can be so-called “off-target effects”, i.e. unwanted changes in other parts of the genome than were actually intended. In addition, “on-target effects” can occur, i.e. where unintentional changes occur in or near the target region of the genome. This includes, for example, the unintentional incorporation of additional DNA sequences or unintended rearrangements of the genome. In many cases, the methods of old and new genetic engineering are still combined to introduce the gene scissors into the cells. This is why unwanted effects arising from these older genetic engineering processes can also occur.
The new genome editing methods allow the genome to be changed to a degree that was previously hardly possible. Even though the changes introduced into the genome are often quite small, collectively they result in target organisms with completely new genetic combinations. It does not matter whether additional genes are introduced into the genome. The intended changes can have a significant impact on metabolic pathways and ingredients. That is why the new properties must be assessed carefully for risks even if no new genes are inserted.
Then, C., Kawall, K., Valenzuela, N. (2020) → Spatio-temporal controllability and environmental risk assessment of genetically engineered gene drive organisms from the perspective of EU GMO Regulation Integr Environ Assess Manag. doi:10.1002/ieam.4278
Gene drive organisms pass on genetically engineered elements to their offspring more frequently than would be the case with natural rules of inheritance. The risk assessment of such gene drive organisms poses a particular challenge since subsequent generations of the genetically modified organisms can exhibit properties not observed or intended in previous generations. Such unintended effects can result from the hybridization of the gene drive construct with natural populations and/or can be triggered by changing environmental conditions. This is particularly relevant for gene drive elements which have invasive properties and typically take dozens of generations to achieve the desired effect. Under such circumstances, these so-called “next generation effects” can significantly increase spatial and temporal complexity, which is associated with a high degree of uncertainty in risk assessment. To solve these problems, we propose the introduction of a new, additional step in the risk assessment of gene drive organisms that takes the following three criteria into account: the biology of the target organisms, their naturally occurring interactions with the environment (biotic and abiotic) and the intended biological properties introduced into the organisms using genetic engineering techniques. These three criteria are merged in an additional step within risk assessment to assess the spatial-temporal controllability of gene drive organisms. The latter can be used to define so-called “cut-off” criteria in the risk analysis: if it is likely that gene drive organisms will escape spatial-temporal controllability, the risk assessment cannot be considered sufficiently reliable because it would be inconclusive. In such circumstances, the release of the gene drive organisms into the environment would not be compatible with the precautionary principle.
Bauer-Panskus, A., Miyazaki, J., Kawall, K., Then, C. (2020). → Risk assessment of genetically engineered plants that can persist and propagate in the environment Environ Sci Eur 32, 32. doi: 10.1186/s12302-020-00301-0
New challenges arise in risk assessment when genetically engineered (GE) plants can persist and propagate in the environment as well as produce viable offspring. Next generation effects can be influenced by heterogeneous genetic backgrounds and unexpected effects can be triggered in interaction with environmental conditions. Consequently, the biological characteristics of the original events cannot be regarded as sufficient to conclude on hazards that may emerge in following generations. Potential hazards identified by the European Food Safety Authority (EFSA) include exacerbating weed problems, displacement and even extinction of native plant species. However, it is concerning that EFSA only takes into account the characteristics of the original events, leaving aside issues of unexpected and unintended effects in following generations that emerge from spontaneous propagation and gene flow. From our review of the available publications and the analysis of risk assessment that was carried out, we conclude that the risk assessment of GE organisms able to persist and spontaneously propagate in the environment actually suffers from a high degree of spatio-temporal complexity causing many uncertainties. To deal with this problem, we recommend establishing ‘cut-off criteria’ in risk assessment that include factual limits of knowledge. It is proposed that these criteria are applied in a specific step within risk assessment, i.e. ‘spatio-temporal controllability’ that uses well-defined biological characteristics to delineate some of the boundaries between known and unknowns. This additional step in risk assessment will foster robustness in the process and can substantially benefit the reliability and overall conclusiveness of risk assessment and decision-making on potential releases.
Kawall, K. (2019). → New Possibilities on the Horizon: Genome Editing Makes the Whole Genome Accessible for Changes Front. Plant Sci. 10:525. doi: 10.3389/fpls.2019.00525
The emergence of new genome editing techniques, such as site-directed nucleases, clustered regulatory interspaced short palindromic repeats (CRISPRs)/Cas9, transcription activator-like effector nucleases (TALENs) or zinc finger nucleases (ZFNs), has greatly increased the feasibility of introducing any desired changes into the genome of a target organism. The ability to target a Cas nuclease to DNA sequences with a single-guide RNA (sgRNA) has provided a dynamic tool for genome editing and is naturally derived from an adaptive immune system in bacteria and archaea. CRISPR/Cas systems are being rapidly improved and refined, thereby opening up even more possibilities. Classical plant breeding is based on genetic variations that occur naturally and is used to select plants with improved traits. Induced mutagenesis is used to enhance mutational frequency and accelerate this process. Plants have evolved cellular processes, including certain repair mechanisms that ensure DNA integrity and maintenance of distinct DNA loci. The focus of this review is on the characterization of new potentials in plant breeding through the use of CRISPR/Cas systems that eliminate natural limitations in order to induce thus far unachievable genomic changes.