Weight of the evidence: independent research projects confirm industry conclusions on the safety of insect-protected maize MON 810.

The cumulative weight of the evidence demonstrates the safety and equivalence of genetically engineered (GE) crops compared to the conventional varieties from which they have been derived. Confirmatory toxicology and animal nutrition studies have nevertheless become an expected/mandated component of GE crop safety assessments, despite the lack of additional value these studies provide for product safety assessment. Characterization and safety data (e.g. trait protein safety; molecular, compositional, and agronomic/phenotypic assessments), and animal feeding studies form a weight of the evidence supporting the safety of insect-protected maize MON 810. Independent animal testing has recently confirmed the lack of MON 810 toxicity in subchronic and chronic toxicity studies. These results could have been predicted from the available safety data. Animal testing of GE crops should be supported by testable scientific hypotheses and testing should be consistent with ethical obligations to reduce, refine, and replace (3Rs) animal testing when possible.

Safety assessment of dicamba mono-oxygenases that confer dicamba tolerance to various crops.

Dicamba tolerant (DT) soybean, cotton and maize were developed through constitutive expression of dicamba mono-oxygenase (DMO) in chloroplasts. DMO expressed in three DT crops exhibit 91.6-97.1% amino acid sequence identity to wild type DMO. All DMO forms maintain the characteristics of Rieske oxygenases that have a history of safe use. Additionally, they are all functionally similar in vivo since the three DT crops are all tolerant to dicamba treatment. None of these DMO sequences were found to have similarity to any known allergens or toxins. Herein, to further understand the safety of these DMO variants, a weight of evidence approach was employed. Each purified DMO protein was found to be completely deactivated in vitro by heating at temperatures 55 °C and above, and all were completely digested within 30 s or 5 min by pepsin and pancreatin, respectively. Mice orally dosed with each of these DMO proteins showed no adverse effects as evidenced by analysis of body weight gain, food consumption and clinical observations. Therefore, the weight of evidence from all these protein safety studies support the conclusion that the various forms of DMO proteins introduced into DT soybean, cotton and maize are safe for food and feed consumption, and the small amino acid sequence differences outside the active site of DMO do not raise any additional safety concerns.

Corn rootworm-active RNA DvSnf7: Repeat dose oral toxicology assessment in support of human and mammalian safety.

Genetically modified (GM) crops have been developed and commercialized that utilize double stranded RNAs (dsRNA) to suppress a target gene(s), producing virus resistance, nutritional and quality traits. MON 87411 is a GM maize variety that leverages dsRNAs to selectively control corn rootworm through production of a 240 base pair (bp) dsRNA fragment targeting for suppression the western corn rootworm (Diabrotica virgifera virgifera) Snf7 gene (DvSnf7). A bioinformatics assessment found that endogenous corn small RNAs matched ∼450 to 2300 unique RNA transcripts that likely code for proteins in rat, mouse, and human, demonstrating safe dsRNA consumption by mammals. Mice were administered DvSnf7 RNA (968 nucleotides, including the 240 bp DvSnf7 dsRNA) at 1, 10, or 100 mg/kg by oral gavage in a 28-day repeat dose toxicity study. No treatment-related effects were observed in body weights, food consumption, clinical observations, clinical chemistry, hematology, gross pathology, or histopathology endpoints. Therefore, the No Observed Adverse Effect Level (NOAEL) for DvSnf7 RNA was 100 mg/kg, the highest dose tested. These results demonstrate that dsRNA for insect control does not produce adverse health effects in mammals at oral doses millions to billions of times higher than anticipated human exposures and therefore poses negligible risk to mammals.

Adaptation of the ToxRTool to Assess the Reliability of Toxicology Studies Conducted with Genetically Modified Crops and Implications for Future Safety Testing.

To determine the reliability of food safety studies carried out in rodents with genetically modified (GM) crops, a Food Safety Study Reliability Tool (FSSRTool) was adapted from the European Centre for the Validation of Alternative Methods' (ECVAM) ToxRTool. Reliability was defined as the inherent quality of the study with regard to use of standardized testing methodology, full documentation of experimental procedures and results, and the plausibility of the findings. Codex guidelines for GM crop safety evaluations indicate toxicology studies are not needed when comparability of the GM crop to its conventional counterpart has been demonstrated. This guidance notwithstanding, animal feeding studies have routinely been conducted with GM crops, but their conclusions on safety are not always consistent. To accurately evaluate potential risks from GM crops, risk assessors need clearly interpretable results from reliable studies. The development of the FSSRTool, which provides the user with a means of assessing the reliability of a toxicology study to inform risk assessment, is discussed. Its application to the body of literature on GM crop food safety studies demonstrates that reliable studies report no toxicologically relevant differences between rodents fed GM crops or their non-GM comparators.

The food and environmental safety of Bt crops.

Bacillus thuringiensis (Bt) microbial pesticides have a 50-year history of safety in agriculture. Cry proteins are among the active insecticidal ingredients in these pesticides, and genes coding for Cry proteins have been introduced into agricultural crops using modern biotechnology. The Cry gene sequences are often modified to enable effective expression in planta and several Cry proteins have been modified to increase biological activity against the target pest(s). Additionally, the domains of different but structurally conserved Cry proteins can be combined to produce chimeric proteins with enhanced insecticidal properties. Environmental studies are performed and include invertebrates, mammals, and avian species. Mammalian studies used to support the food and feed safety assessment are also used to support the wild mammal assessment. In addition to the NTO assessment, the environmental assessment includes a comparative assessment between the Bt crop and the appropriate conventional control that is genetically similar but lacks the introduced trait to address unintended effects. Specific phenotypic, agronomic, and ecological characteristics are measured in the Bt crop and the conventional control to evaluate whether the introduction of the insect resistance has resulted in any changes that might cause ecological harm in terms of altered weed characteristics, susceptibility to pests, or adverse environmental impact. Additionally, environmental interaction data are collected in field experiments for Bt crop to evaluate potential adverse effects. Further to the agronomic and phenotypic evaluation, potential movement of transgenes from a genetically modified crop plants into wild relatives is assessed for a new pest resistance gene in a new crop. This review summarizes the evidence for safety of crops containing Cry proteins for humans, livestock, and other non-target organisms.

A 28-day oral toxicity evaluation of small interfering RNAs and a long double-stranded RNA targeting vacuolar ATPase in mice.

New biotechnology-derived crop traits have been developed utilizing the natural process of RNA interference (RNAi). However, plant-produced double stranded RNAs (dsRNAs) are not known to present a hazard to mammals because numerous biological barriers limit uptake and potential for activity. To evaluate this experimentally, dsRNA sequences matching the mouse vATPase gene (an established target for control of corn rootworms) were evaluated in a 28-day toxicity study with mice. Test groups were orally gavaged with escalating doses of either a pool of four 21-mer vATPase small interfering RNAs (siRNAs) or a 218-base pair vATPase dsRNA. There were no treatment-related effects on body weight, food consumption, clinical observations, clinical chemistry, hematology, gross pathology, or histopathology endpoints. The highest dose levels tested were considered to be the no observed adverse effect levels (NOAELs) for the 21-mer siRNAs (48 mg/kg/day) and the 218 bp dsRNA (64 mg/kg/day). As an additional exploratory endpoint, vATPase gene expression, was evaluated in selected gastrointestinal tract and systemic tissues. The results of this assay did not indicate treatment-related suppression of vATPase. The results of this study indicate that orally ingested dsRNAs, even those targeting a gene in the test species, do not produce adverse health effects in mammals.

Purification, characterization and safety assessment of the introduced cold shock protein B in DroughtGard maize.

DroughtGard maize was developed through constitutive expression of cold shock protein B (CSPB) from Bacillus subtilis to improve performance of maize (Zea mays) under water-limited conditions. B. subtilis commonly occurs in fermented foods and CSPB has a history of safe use. Safety studies were performed to further evaluate safety of CSPB introduced into maize. CSPB was compared to proteins found in current allergen and protein toxin databases and there are no sequence similarities between CSPB and known allergens or toxins. In order to validate the use of Escherichia coli-derived CSPB in other safety studies, physicochemical and functional characterization confirmed that the CSPB produced by DroughtGard possesses comparable molecular weight, immunoreactivity, and functional activity to CSPB produced from E. coli and that neither is glycosylated. CSPB was completely digested with sequential exposure to pepsin and pancreatin for 2 min and 30 s, respectively, suggesting that CSPB will be degraded in the mammalian digestive tract and would not be expected to be allergenic. Mice orally dosed with CSPB at 2160 mg/kg, followed by analysis of body weight gains, food consumption and clinical observations, showed no discernible adverse effects. This comprehensive safety assessment indicated that the CSPB protein from DroughtGard is safe for food and feed consumption.