Subchronic toxicity study in rats evaluating herbicide-tolerant soybean DAS-68416-4.

A subchronic toxicity study was conducted in Wistar rats to evaluate the potential health effects of genetically modified (GM) herbicide-tolerant soybean DAS-68416-4. Rats were fed with diets containing toasted meal produced from GM soybean engineered with aad-12 and pat genes or containing non-GM soybean at a dose of 30.0, 15.0, or 7.5%,w/w% and 0% (control group) for 90 consecutive days. Animals were evaluated for general behavior, body weight gain, food consumption, food use efficiency, etc. At the middle and end of the study, blood and serum samples were collected for routine and biochemical assays. Internal organs were taken for calculating relative weights and doing histopathological examination. The rats were active and healthy without any abnormal symptoms during the entire study period. No biological differences in hematological or biochemical indices were detected. No histopathological changes were observed. Under the conditions of this study, herbicide-tolerant soybean DAS-68416-4 did not cause any treatment-related effects in Wistar rats following 90 days of dietary administration.

Quality of Soybean Products in Terms of Essential Amino Acids Composition.

The content of protein, moisture content and essential amino acids in conventional and genetically modified soybean grain and selected soybean products (soybean pâté, soybean drink, soybean dessert, tofu) was analyzed in this paper. The following comparative analysis of these products has not yet been carried out. No differences were observed in the amino acid profiles of soybeans and soybean products. The presence of essential amino acids was confirmed except for tryptophan. Its absence, however, may be due not to its absence in the raw material, but to its decomposition as a result of the acid hydrolysis of the sample occurring during its preparation for amino acid determination. Regardless of the type of soybean grain, the content of protein, moisture content and essential amino acids was similar (statistically insignificant difference). Thus, the type of raw material did not determine these parameters. There was a significant imbalance in the quantitative composition of essential amino acids in individual soybean products. Only statistically significant variation was found in genetically modified and conventional soybean pâté. Moreover, in each soy product their amount was lower irrespective of the raw material from which they were manufactured. Therefore, the authors indicate the necessity of enriching soybean products with complete protein to increase their nutritional value.

Subchronic toxicity study in rats evaluating genetically modified DAS-81419-2 soybean.

A 90-day feeding study in rats was conducted to evaluate the subchronic oral toxicity of genetically modified (GM) DAS-81419-2 soybean. Wistar rats were fed with diets containing toasted soybean meal produced from DAS-81419-2 soybean grain that expresses the Cry1F, Cry1Ac, and Pat proteins or containing conventional soybean at doses of 30.0%, 15.0%, 7.5%, or 0% (control group) for 90 consecutive days. The general behavior, body weight and food consumption were observed. At the middle and end of the experiment, blood, serum, and urine samples were collected for biochemical assays. At the conclusion of the study, the internal organs were weighed and histopathological examination was completed. The rats exhibited free movement and shiny coats without any abnormal symptoms or abnormal secretions in their noses, eyes, or mouths. There were no adverse effects on body weight in GM soybean groups and conventional soybean groups. No biological differences in hematological, biochemical, or urine indices were observed. No significant differences in relative organ weights were detected between the experimental groups and the control group. No histopathological changes were observed. Under the conditions of this study, DAS-81419-2 soybean did not cause any treatment-related effects in Wistar rats following 90 days of dietary administration.

Safety evaluation of DAS-44406-6 soybeans in Wistar rats.

A 90-day in-country feeding trial in Wistar rats was conducted at Tianjin Laboratory in China to assess toxicity of diets containing DAS-44406-6 soybean meal. There were no treatment-related changes observed when compared with the non-GM isoline control groups but histopathologically, 2 of 10 high-dose females were reported to show kidney lesions. However, these findings contrasted with the absence of any treatment-related kidney lesions in 3 separate 90-day toxicity studies previously conducted in Sprague Dawley rats. Strain difference is not expected in the kidney response, and based on the low incidence and contrary evidence from previous studies, it is likely that these lesions were of spontaneous origin, or artefactual. To determine that the lesions observed were not treatment-related in Wistar rats, a specific follow-up confirmatory study was conducted under Good Laboratory Practices (GLP) in the Wistar strain of rats following an identical study design to the Tianjin study. To increase the power of detecting effects, twice the number of animals per group (20/sex/group) were used, and no treatment-related kidney histopathological changes were observed. Based on these results and entire weight of evidence evaluation, it is concluded that the histopathological changes previously noted in the 2 female Wistar rats of Tianjin study were not treatment-related and that DAS-44406-6 soybeans are as safe as conventional non-GM soybeans.

Evaluation of 30% DAS-444Ø6-6 soybean meal in a subchronic rat toxicity study.

Event DAS-444Ø6-6 soybean is genetically modified (GM) to provide tolerance to 2,4-diclorophenoxyacetic acid (2,4-D), glyphosate, and glufosinate herbicides through expression of the AAD-12, 2mEPSPS, and PAT proteins, respectively. DAS-444Ø6-6 soybeans were evaluated for safety in subchronic rat feeding studies. The results from two previous subchronic rat feeding studies evaluating diets formulated with 20% inclusion of DAS-444Ø6-6 soybean meal (the latter also containing DAS-444Ø6-6 derived hulls and oil) did not show any treatment-related adverse effects. In 2017, to comply with recent guidance from EFSA, a third 90-day rat feeding study was conducted with Sprague-Dawley rats (16/sex/group) with diets formulated either with 15% or 30% w/w of toasted DAS-444Ø6-6 soybean meal. DAS-444Ø6-6 soybean hulls and oil were also added to the transgenic test diets at 1% or 2% w/w and 1.35% or 2.7%, respectively, for the low- and high-dose groups. No toxicologically significant effects were observed under the conditions of this study.

Epilithic biofilms as a discriminating matrix for long-term and growing season pesticide contamination in the aquatic environment: Emphasis on glyphosate and metabolite AMPA.

The indiscriminate use of pesticides represents high ecological risk in aquatic systems. Recently, the inclusion of epilithic biofilms as a reactive matrix has shown potential in diagnosing the health of water resources. The objective of this study was to use multiple matrices (water, suspended sediments, and biofilms) to discriminate contamination degrees in catchments with long and recent history of intensive pesticide use and to monitor growing season pesticides transfer to watercourses. Two catchments were monitored: one representative of "modern agriculture" in a subtropical environment, and another representative of recent agricultural expansion over the Pampa Biome in subtropical Brazil. Glyphosate and AMPA were accumulated in the biofilms and were detected at all sites and at all monitoring times, in concentrations ranging from 195 to 7673 µg kg-1 and from 225 to 4180 µg kg-1, respectively. Similarly, the fungicide tebuconazole has always been found in biofilms. The biofilms made it possible to discriminate the long-term history of pesticide use in the catchments and even to identify the influx pulses of pesticides immediately after their application to crops, which was not possible with active water sampling and even with suspended sediment monitoring. It is strongly recommended that, in regions with intensive cultivation of soybeans and other genetically modified crops, the presence of glyphosate and its metabolite AMPA be permanently monitored, a practice still very scarce in the literature.

Seed coat colour and structure are related to the seed dormancy and overwintering ability of crop-to-wild hybrid soybean.

The possible persistence of genetically modified (GM) crop-to-wild hybrid seeds in the soil seed bank is a major concern in risk assessment and is closely related to seed characteristics such as dormancy. In the present study, we generated F3 hybrids via crosses between GM soybean accessions and wild soybean and evaluated the dormancy, overwintering ability and inheritance of foreign genes in different-coloured hybrid seeds (yellow, green, brown and black). The results revealed that the 5-enolpyruvylshikimate-3-phosphate synthase transgene may have no influence on crop wild hybrid seed dormancy and overwintering ability, and the dormancy of the hybrid seeds was closely related to seed coat colour. F3 hybrid seeds with light colours (yellow and green) were relatively nondormant, while seeds that were dark (brown and black) in colour were relatively dormant. Moreover, the hybrid seeds that were dark in colour had a much stronger overwintering ability than the lighter-coloured seeds, with 21.33 % of the black seeds and 33.33 % of the brown seeds remaining viable after 240 days of soil burial. In contrast, almost all the F3 yellow and green seeds were no longer viable during winter. Scanning electron microscopy revealed that the lighter-coloured seeds had a thin palisade layer and very few surface deposits, while the darker-coloured seeds had a thicker palisade layer and a large area of honeycomb-like surface deposits similar to those of wild soybean seeds. Thus, the physical dormancy and overwintering ability of the darker-coloured seeds may be related to the seed coat. Our results suggest that transgenes of GM soybean might disperse into wild populations and persist in seed banks.

Effect of Soybean Protein Concentrate Preparation on Copy Numbers and Structural Characteristics of DNA from Genetically Modified Soybean.

To regulate the degradation of transgenic DNA and lay theoretical foundations for the rational utilization of genetically modified (GM) products, variations in copy numbers and structural characteristics of DNA from GM soybean event GTS 40-3-2 during soybean protein concentrate (SPC) preparation were evaluated. Results showed that defatting and the first ethanol extraction were key procedures inducing DNA degradation. After these two procedures, copy numbers of the lectin and cp4 epsps targets decreased by more than 4 × 108, occupying 36.88-49.30% of the total copy numbers from raw soybean. Atomic force microscopy images visually revealed the degradation of DNA that thinned and shortened during SPC preparation. Circular dichroism spectra suggested a lower helicity of DNA from defatted soybean kernel flour and a conformation transition of DNA from B-type to A-type after ethanol extraction. The fluorescence intensity of DNA decreased during SPC preparation, verifying the DNA damage along this preparation chain.

Comparative Effects of Traditional Versus Genetically Modified Soybean Oils on Colon Tumorigenesis in Mice.

Soybean oil, which has high abundance of linoleic acid (LA, 18:2ω-6), is the most commonly consumed edible oil. Recent studies support that a high dietary intake of LA is linked with increased risks of developing colonic inflammation and colon cancer. Here we studied the effects of the genetically modified Plenish® soybean oil, which has low abundance of LA as well as α-linolenic acid (ALA, 18:3ω-3), on development of azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced colon tumorigenesis in mice. Compared with a diet rich in traditional soybean oil, administration of a diet enriched with the Plenish oil has little impact on AOM/DSS-induced colon tumorigenesis, colonic infiltration of immune cells, expressions of inflammatory genes, and tumor markers. These results suggest that the traditional and the Plenish soybean oils have similar effects on development of AOM/DSS-induced colon cancer in mice.

Development of Multiplex PCR Coupled DNA Chip Technology for Assessment of Endogenous and Exogenous Allergens in GM Soybean.

Allergenicity assessment of transgenic plants and foods is important for food safety, labeling regulations, and health protection. The aim of this study was to develop an effective multi-allergen diagnostic approach for transgenic soybean assessment. For this purpose, multiplex polymerase chain reaction (PCR) coupled with DNA chip technology was employed. The study was focused on the herbicide-resistant Roundup Ready soya (RRS) using a set of certified reference materials consisting of 0, 0.1%, 0.5%, and 10% RRS. Technically, the procedure included design of PCR primers and probes; genomic DNA extraction; development of uniplex and multiplex PCR systems; DNA analysis by agarose gel electrophoresis; microarray development, hybridization, and scanning. The use of the asymmetric multiplex PCR method is shown to be very efficient for DNA hybridization with biochip probes. We demonstrate that newly developed fourplex PCR methods coupled with DNA-biochips enable simultaneous identification of three major endogenous allergens, namely, Gly m Bd 28K, Gly m Bd 30K, and lectin, as well as exogenous 5-enolppyruvyl shikimate-phosphate synthase (epsps) expressed in herbicide-resistant roundup ready GMOs. The approach developed in this study can be used for accurate, cheap, and fast testing of food allergens.

Development of a Graphene-Based Biosensor for Detecting Recombinant Cyanovirin-N.

We present a graphene-based biosensor selective to recombinant cyanovirin-N (rCV-N), an antiviral protein that has proven to be an effective microbicide to inhibit HIV replication. We modified the graphene monolayer devices with 1-pyrenebutanoic acid succinimidyl ester, which interacts with both graphene and the primary and secondary amines of antibodies. By monitoring the change in the electrical resistance of the device, we were able to detect rCV-N in solutions in the range of 0.01 to 10 ng/mL, and found that the detection limit was 0.45 pg/mL, which is much smaller than that obtained with currently available techniques. This is important for applications of this microbicide against HIV, since it may be produced at a large scale from soya bean seeds processed using the available industrial processing technologies. The sensor showed high sensitivity, selectivity, and reproducibility.

Electrochemical genoassays on gold-coated magnetic nanoparticles to quantify genetically modified organisms (GMOs) in food and feed as GMO percentage.

The integration of nanomaterials in the field of (bio)sensors has allowed developing strategies with improved analytical performance. In this work, ultrasmall core-shell Fe3O4@Au magnetic nanoparticles (MNPs) were used as the platform for the immobilization of event-specific Roundup Ready (RR) soybean and taxon-specific DNA sequences. Firstly, monodisperse Fe3O4 MNPs were synthesized by thermal decomposition and subsequently coated with a gold shell through reduction of Au(III) precursor on the surface of the MNPs in the presence of an organic capping agent. This nanosupport exhibited high colloidal stability, average particle size of 10.2 ±â€¯1.3 nm, and spherical shape. The covalent immobilization of ssDNA probe onto the Au shell of the Fe3O4@Au MNPs was achieved through a self-assembled monolayer (SAM) created from mixtures of alkane thiols (6-mercapto-1-hexanol and mercaptohexanoic acid). The influence of the thiols ratio on the electrochemical performance of the resulting electrochemical genoassays was studied, and remarkably, the best analytical performance was achieved for a pure mercaptohexanoic acid SAM. Two quantification assays were designed; one targeting an RR sequence and a second targeting a reference soybean gene, both with a sandwich format for hybridization, signaling probes labelled with fluorescein isothiocyanate (FITC), enzymatic amplification and chronoamperometric detection at screen-printed carbon electrodes (SPCE). The magnetogenoassays exhibited linear ranges from 0.1 to 10.0 nM and from 0.1 to 5.0 nM with similar detection limits of 0.02 nM and 0.05 nM for the event-specific (RR) and the taxon-specific (lectin) targets, respectively. The usefulness of the approach was demonstrated by its application to detect genetically modified organisms (GMOs) in feed and food.

On-site detection of stacked genetically modified soybean based on event-specific TM-LAMP and a DNAzyme-lateral flow biosensor.

Stacked genetically modified organisms (GMO) are becoming popular for their enhanced production efficiency and improved functional properties, and on-site detection of stacked GMO is an urgent challenge to be solved. In this study, we developed a cascade system combining event-specific tag-labeled multiplex LAMP with a DNAzyme-lateral flow biosensor for reliable detection of stacked events (DP305423× GTS 40-3-2). Three primer sets, both event-specific and soybean species-specific, were newly designed for the tag-labeled multiplex LAMP system. A trident-like lateral flow biosensor displayed amplified products simultaneously without cross contamination, and DNAzyme enhancement improved the sensitivity effectively. After optimization, the limit of detection was approximately 0.1% (w/w) for stacked GM soybean, which is sensitive enough to detect genetically modified content up to a threshold value established by several countries for regulatory compliance. The entire detection process could be shortened to 120min without any large-scale instrumentation. This method may be useful for the in-field detection of DP305423× GTS 40-3-2 soybean on a single kernel basis and on-site screening tests of stacked GM soybean lines and individual parent GM soybean lines in highly processed foods.

Evaluation of the safety of a genetically modified DAS-444Ø6-6 soybean meal and hulls in a 90-day dietary toxicity study in rats.

A 90-day sub chronic toxicity study was conducted in rats to evaluate the safety of genetically modified DAS-444Ø6-6 soybeans expressing herbicide tolerant proteins when compared with its conventional comparators (non-transgenic near isoline control soybean and three commercially available non-transgenic line control soybeans). Rats were given diets formulated with either 10% or 20% w/w of soybean meal and 1% or 2% hulls of DAS-444Ø6-6 soybean with an equivalent amount of hulls from an isoline non-transgenic control soybean for at least 90 days. In addition, three separate 20% w/w non-transgenic commercially available soybean varieties were also given to groups of rats to serve as reference controls. Animals were evaluated by cage-side and hand-held detailed clinical observations, ophthalmic examinations, body weights/body weight gains, feed consumption, hematology, prothrombin time, urinalysis, clinical chemistry, selected organ weights, and gross and histopathologic examinations. Under the conditions of this study, the genetically modified DAS-444Ø6-6 diets did not cause any treatment-related effects in rats following 90 days of dietary administration as compared with rats fed diets with soybean of isoline control or commercial reference controls and are considered equivalent to the diets prepared from conventional comparators.

[Impacts of genetically modified soybean leaf residues on Folsomia candida.] / è½¬åŸºå› å¤§è±†å¶ç‰‡æ®‹ä½“å¯¹ç™½ç¬¦è·³çš„å½±å“.

When the genetically modified soybean is planted in the field, the expression product of exogenous gene could be exposed in the soil ecosystem and bring potential risk to the soil fauna, with the form of leaves and other debris. A few of genetically modified soybeans developed by China independently were used in our study as materials. They were Phytophthora-resistant soybean harboring hrpZm gene (B4J8049), leaf-feeding insect-resistant soybean harboring Cry1C gene (A2A8001) and Leguminivora glycinivorella-resistant soybean harboring Cry1Iem gene (C802). By feeding Folsomia candida with the three genetically modified soybeans for continuous 60 days, the surviving rate, reproductive rate and changes on the body length of F. candida were studied. The results showed that all the three genetically modified soybeans of B4J8049, A2A8001 and C802 had no significant adverse effects on the growth of F. candida, as an environmental indicator organism. It was initially inferred that they were environmentally safe under short-term exposure, which provided basic data of ecological safety for their wide cultivation.

[Detection of transgenic components in animal feeds on Shanxi markets].

To assess the presence of genetically modified (GM) maize and soybean in a range of commercialized feed in Shanxi province of China in 2015, improved hexadecyltrimethy ammonium bromide (CTAB) method was used to extract DNA. The screening of packed feeds was carried out by qualitative PCR. Then positive feeds were unpacked and detected by the CaMV 35S promoter, NOS terminator, zSSIIb, Lectin and CryIA (b) genes. The identified maize and soybean events were confirmed by event-specific MON810 and GTS40-3-2. Results showed that 83.3% of the feeds was tested positive for GMOs, in which positive rates of maize, soybean, pig and layer feeds were 6.67%, 100%, 93.3% and 73.3%, respectively. The results of real-time PCR were consistent with qualitative PCR. These results indicated that commercialized GM feed had a wide positive product scope in Shanxi province of China. Further studies are necessary to study effects of feeding livestock and poultry with feed containing GM ingredients on animals and their products.

Performance of hybrid progeny formed between genetically modified herbicide-tolerant soybean and its wild ancestor.

Gene flow from genetically modified (GM) crops to wild relatives might affect the evolutionary dynamics of weedy populations and result in the persistence of escaped genes. To examine the effects of this gene flow, the growth of F1 hybrids that were formed by pollinating wild soybean (Glycine soja) with glyphosate-tolerant GM soybean (G. max) or its non-GM counterpart was examined in a greenhouse. The wild soybean was collected from two geographical populations in China. The performance of the wild soybean and the F2 hybrids was further explored in a field trial. Performance was measured by several vegetative and reproductive growth parameters, including the vegetative growth period, pod number, seed number, above-ground biomass and 100-seed weight. The pod setting percentage was very low in the hybrid plants. Genetically modified hybrid F1 plants had a significantly longer period of vegetative growth, higher biomass and lower 100-seed weight than the non-GM ones. The 100-seed weight of both F1 and F2 hybrids was significantly higher than that of wild soybean in both the greenhouse and the field trial. No difference in plant growth was found between GM and non-GM F2 hybrids in the field trial. The herbicide-resistant gene appeared not to adversely affect the growth of introgressed wild soybeans, suggesting that the escaped transgene could persist in nature in the absence of herbicide use.

Utilização do gráfico de Ghewhart como ferramenta analítica no controle da quantificação de soja geneticamente modificada em alimentos por PRC em tempo real

O Brasil regulamentou o plantio e a comercialização de soja geneticamente modificada pelo Decreto n º 4680 que estabelece o limite de 1% para a informação no rótulo dos produtos alimentares que contenham ou sejam constituídos por OGM. OINCQS é o laboratório oficial no Brasil que atua no controle pós-comercialização de soja transgênica nos alimentos. O gráfico de controle é uma representação de dados, desenvolvido por Shewhart para verificar se o processo analítico está sob controle estatístico e que o mesmo permanece sob controle ao longo do tempo. Este gráfico foi utilizado com os resultados da análise quantitativa de soja geneticamente modificada em alimentos e seu uso como instrumento analítico foi avaliado.

Soja geneticamente modificada em alimentos contendo farinha e preparados à base de farinha de trigo. Detecção e adequação à legislação de rotulagem / Genetically modified soybean in food containing wheat flour and wheat flour-based preparations. Detection and fitness to labeling legislation

A farinha de trigo é considerada como um dos principais ingredientes utilizados no preparo dos alimentos e, portanto, presente em praticamente todas as refeições dos brasileiros. Em 2000, o consumo per capita de farinha de trigo correspondia a 53 kg e estava distribuída da seguinte forma no mercado brasileiro: 47% padarias; 14% indústrias de massas; 20% consumo doméstico; 8% indústria de biscoito; 5% indústria de pães; e 6% em outros segmentos. O objetivo deste trabalho foi avaliar a presença de soja e soja Roundup Ready em amostras de farinhas de trigo e de preparados à base de farinha de trigo para a alimentação humana e verificar a adequação dos rótulos frente à legislação pela análise quantitativa por PCR em Tempo Real. O INCQS/FIOCRUZ analisou 16 amostras de farinhas de trigo de 10 marcas diferentes. Quinze (15) amostras apresentaram resultado positivo para o gene le1 da lectina, demonstrando a presença de soja nestes produtos, e doze (12) amostras apresentaram resultado positivo para o gene CP4 EPSPS, específico da soja RR ® , evidenciando a utilização de soja geneticamente modificada nestes produtos. Todas as amostras analisadas denominadas como preparados à base de farinha de trigo para a alimentação humana, conhecidos comercialmente como mistura para bolos, quiches, panquecas e bolinhos de chuva apresentaram resultados positivos para o gene específico da soja RR ® e deveriam conter no rótulo o símbolo estabelecido na Portaria nº 2658, de 22 de dezembro de 2003.