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GMO release: Scientific Sense? Dr. William H.L Stafford, Advanced Research Center for Applied Microbiology, Department of Biotechnology, University of the Western Cape. GMOs and breeding We have been modifying our food sources for thousands of years, selecting for favourable characteristics. These breeding techniques rely on fertlization by cross-pollination of the same species Biotechnology has provided us with the tools to engineer a a plant with any chosen characteristic. Species barriers can be crossed- we can take a gene from one organism and place it in another totally unrelated organism and create transgenics. Genetic engineering of plants GM crops >90% of GM crops are engineered with transgenes for herbicide resistance and insecticides. T-vector © Promoter © Marker gene © Desired gene © Terminator ENVIRONMENTAL RISKS Random insertion of transgene and high expression levels Genetic pleiotropic effects result from abnormal expression levels, position effects of inserted genes and secondary mutations. The CaMv is a strong promoter providing a high level of gene expression. All living organisms that interact with the transgenic plant are exposed to high levels of the expressed transgene. The insecticidal Bt-toxins, isolated from Bacillus thuringiensis, are often engineered into plants. Lacewings fed on aphid pests that had eaten Bt-maize took longer to develop and were two to three times more likely to die. (Birch, A.N.E., et al. 1997, Soft Fruit and Perennial Crops and Marvier, M. 2001. Ecology of transgenic crops. American Scientist) Insecticidal toxins from transgenic plants can kill beneficial (non-target) species and be passed higher up the food chain Effects on biodiversity and evolution Genes can spread from transgenic plants by ordinary cross pollination to nontransgenic plants of a related species, and also by horizontal gene transfer to unrelated species. The most obvious effects of cross-pollination already identified are in creating herbicide-tolerant, or insecticidal weeds and superweeds. Studies with oilseed rape (Brassica napus) have shown that the Bt gene can be passed on to a wild, weedier relative (Brassica rapa) (Halfhill, M.D., et al. 2002. Environmental Biosafety Research). Another hazard is the spread of the antibiotic resistance marker genes which are in a high proportion of transgenic plants. There are unpredictable physiological impacts on the organisms to which the genes and gene-constructs are spread and hence on the ecological environment. Horizontal gene transfer has been relatively rare in our evolutionary past, due to natural species barriers prevent gene exchange, and mechanisms which inactivate or break down foreign DNA Horizontal transfer from the transgenic plants may spread the novel genes and gene-constructs to unrelated speciesmicrobes in the soil, worms, insects reptiles, birds, small mammals and human beings. Agrobacterium vector system allows facilitated means of gene escape. Increased horizontal gene transfer in the soil rhizosphere and seeds have been noted (Sengelov et al. 2001. Current Microbiology.) The Ti plasmid of the Agrobacterium has mobile genetic elements elements to integrate into the plant DNA. This integration is random and subjected to further recombination… Gene transfer and escape Both the Ti vector and the CaMv promoter have palindromic sequence that are recombination hotspots and subjected to increase horizontal gene transfer (Kohli et al. 1999, Plant.J.) This recombination with other DNA can lead to: Reactivation of dormant viruses and the generation of new viruses Generation of new bacterial pathogens and the spread of drug and antibiotic resistance marker genes among pathogens Insertion of genes into cells of recipient species, mutagenesis and cancer transformation. Such elements have increased penetrance into the environment (especially with selection) and can outcross other varieties Outcrossing Transgenic A. thaliana plants were roughly 20 times more likely to outcross. These results show that genetic engineering can substantially increase the probability of transgene escape. (Bergelson, J. et al. 1998. Nature) Transgenic DNA introgressed into traditional maize landraces in Oaxaca, Mexico (Quist D and Chapela IH. Nature 2001) Horizontal gene transfer from one species to another may be a major factor in evolutionary change! (Syvanen, M. 1986. Trends In Genetics) Summary: environmental risks Increased resistance to herbicides, leading to superweed characteristics Increased spread of antibiotic resistance Reduced biodiversity due to outcrossing and selection Increased occurrence of cancers caused by insertional mutagenesis and position effect changes in gene expression. Unpredictable effects on genetic evolution and ecosystem function HEALTH RISKS: DNA stability DNA can persist in the soil where transform a range of organisms. Transgenes will be able to spread to bacteria and viruses, spreading antibiotic resistance genes among the pathogens. DNA is not completely broken down in the gut. Genes can spread from ingested transgenic plant material to bacteria in the gut and also to cells of the organism itself. Antibiotic resistant marker genes from genetically engineered bacteria can be transferred to indigenous bacteria (Netherwood T, et al. Technical report on the Food Standards Agency project G010008) Expected and unexpected toxicity Transgenic potatoes expressing GNA insecticide (“Snowdrop”, Galanthus nivalis, lectin) fed to rats resulted in increase in intestinal mucosal thickness and T-lymphocyte infiltration. (Erwin S.W.B.and Putzai A. 1999 Lancet) Monsanto's transgenic soya has a 26.7% increase in a trypsininhibitor and has been sown to inhibit the growth rate of male rats. This raises the possibility that transgenic soya is responsible for the reported recent increase in soya allergy. Human gene therapy experiments for severe combined immunodeficiency (SCID) caused by the absence of the enzyme adenosine deaminase (ADA) were halted by the FDA after a second treated child died of cancer. Required features for release of a GMO Efficient and specific gene targeting to cells Stable, single integration of gene at defined site Normal levels of expression of desired gene Proven safety X The GMO crops on the market fulfill none of these criteria Future of biotechnology It is important to distinguish between contained use of transgenic organisms and their release to the environment. It is vital that GMO crops are proven safe for through proper independent, long-term feeding trials and environmental impact assessments It is essential to monitor GMOs since they have been released and we need to observe the effects of this experiment References Ho, M.W., Meyer, H. and Cummins, J. (1998). The biotechnology bubble. The Ecologist 28(3), 146-153 Kohli A.,Griffiths S, Palacios N, Twyman R, Vain P, Laurie D and Christou P. (1999) Molecular characterization of transforming plasmid rearrangements in transgenic rice reveals a recombination hot spot in the CaMV 35S promoter and confirms the predominance of microhomology mediated recombination" Plant.J. 17,591-601. Sengelov G, Kristensen KJ, Sorensen AH, Kroer N, and Sorensen SJ. Effect of genomic location on horizontal transfer of a recombinant gene cassette between Pseudomonas strains in the rhizosphere and spermosphere of barley seedlings. Current Microbiology 2001, 42, 160-7. George A. Kowalchuk, Maaike Bruinsma and Johannes A. van Veen. Assessing responses of soil microorganisms to GM plants TRENDS in Ecology and Evolution Vol.18 No.8 August 2003 Bergelson, J., Purrington, C.B. and Wichmann, G. (1998). Promiscuity in transgenic plants. Nature 395, 25 Quist D and Chapela IH. (2001) Transgenic DNA introgressed into traditional maize landraces in Oaxaca, Mexico. Nature, 414, 541-3, 2001 Syvanen, M . 1986. Cross-species gene transfer: a major factor in evolution? Trends In Genetics pp 1—4 Hilbeck, A., Baumgartner, M., Fried, P.M. and Bigler, F. (1997). Effects of transgenic Bacillus thuringiensis-corn-fed prey on mortality and development time of immature Chrysoperla carnea (Neuroptera: Chrysopidae). Environmental Entomology Halfhill, M.D., R.J. Millwood, P.L. Raymer, and C.N.Stewart, Jr. 2002. Bt-transgenic oilseed rape hybridization with its weedy relative, Brassica rapa.Environmental Biosafety Research 1: 19-28. Birch, A.N.E., Geoghegan, I.I., Majerus, M.E.N., Hackett, C. and Allen, J. (1997). Interaction between plant resistance genes, pest aphidpopulation and beneficial aphid predators. Soft Fruit and Pernial Crops. October, 68-79. Hilbeck, A., Baumgartner, M., Fried, P.M. and Bigler, F. (1997). Effects of transgenic Bacillus thuringiensis-corn-fed prey on mortality and development time of immature Chrysoperla carnea (Neuroptera: Chrysopidae). Environmental Entomology 27, 480-487. Vaden V.S. and Melcher, U. (1990). Recombination sites in cauliflower mosaic virus DNAs: implications for mechanisms of recombination. Virology 177, 717-26 Halfhill, M.D., R.J. Millwood, P.L. Raymer, and C.N.Stewart, Jr. 2002. Bt-transgenic oilseed rape hybridization with its weedy relative, Brassica rapa.. Environmental Biosafety Research 1: 19-28. Lommel, S.A. and Xiong, Z. (1991). Recombination of a functional red clover necrotic mosaic virus by recombination rescue of the cell-to-cell movement gene expressed in a transgenic plant. J. Cell Biochem. 15A, 151; Greene, A.E. and Allison, R.F. (1994). Recombination between viral RNA and transgenic plant transcripts. Science 263, 1423-5; Wintermantel, W.M. and Schoelz, J.E. (1996). Isolation of recombinant viruses between cauliflower mosaic virus and a viral gene in transgenic plants under conditions of moderate selection pressure. Virology 223, 156-64. Sengelov G, Kristensen KJ, Sorensen AH, Kroer N, and Sorensen SJ. (2001) Effect of genomic location on horizontal transfer of a recombinant gene cassette between Pseudomonas strains in the rhizosphere and spermosphere of barley seedlings. Current Microbiology, 42, 160-7. Ewen S. W. B, and Pusztai,A. (1999) Effect of diets containing genetically modified potatoes expressing Galanthus nivalis lectin on rat small intestine Lancet 16 October MacKenzie, D. (1999). Gut reaction. New Scientist 30 Jan., p.4. Schubbert, R., Renz, D., Schmitz, B. and Doerfler, W. (1997). Foreign (M13) DNA ingested by mice reaches peripheral leukocytes, spleen and liver via the intestinal wall mucosa and can be covalently linked to mouse DNA. Proc. Natl. Acad. Sci. USA 94, 961-6. Netherwood T, Martin-Orue SM, O'Donnell AG, Gockling S, Gilbert HJ and Mathers JC. Transgenes in genetically modified Soya survive passage through the small bowel but are completely degraded in the colon. Technical report on the Food Standards Agency project G010008 "Evaluating the risks associated with using GMOs in human foods"- University of Newcastle.