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Reinforced Safety

GMOs developed by Embrapa will be assessed by Biosafety Network

MIGUEL BOYAYANGenetically modified potatoes and beans are a target for the Biosafety NetworkMIGUEL BOYAYAN

The Brazilian Agricultural Research Corporation (Embrapa) is getting ready to start to plant a genetically modified papaya that is resistant to the papaya ringspot virus, or PRS, a disease that reduces the quality and quantity of the fruits of the papaya tree. It is the first genetically modified organism (GMO) developed by Embrapa with an authorization for fields tests: The Research Area Operating License (Loape in the Portuguese acronym), a document issued by the Brazilian Institute for the Environment and Renewable Natural Resources (Ibama) that authorizes controlled planting, was obtained in October. It will also be one of the first products to be analyzed and tested by the GMO Biosafety Network, created by Embrapa in the middle of last year, with the objective of qualifying human resources, and the institution itself, in research with transgenics.

In the case of papaya, several strategies for controlling this virosis have already been used without any success. Even so, Brazil is a leader in the world market, with a production of something around 1.70 million tons of papaya in 2000, which corresponds to a little more than one third of the plants cultivated in the other countries. With the papaya tree immune to the virus, the plant’s productivity and the earnings for the producers may be even greater.In a laboratory environment and plant house, Embrapa has developed small papaya cuttings in which genes resistant to the virus were introduced. Preliminary tests have revealed that the plants have immunity to ringspot. Now, with the authorization from Ibama, the transgenic papaya will have its performance assessed in the field, more precisely in Cruz das Almas, in the state of Bahia, in an area guarded 24 hours a day, protected by an electrified fence, and accompanied by a rigid biosafety protocol. “If there is any problem, we will have a way of controlling it”, says Deise Maria Fontana Capalbo, a researcher with Embrapa Environment.

Cultivation in the field
In Cruz das Almas, the researchers are going to assess the influence of the transgenic papaya on the organisms in the soil as a whole, as well as the quality and the properties of the leaves and of the fruit. “In the first year, we will be testing the resistance of the plant to the conditions of the soil and, in the second, the field conditions strictly speaking”, Deise Capalbo explains. The risks that the GMO may contaminate crops of conventional papaya in adjacent areas are low, as papaya has no pollen and for that reason there is no possibility of gene escape to other cultivars, she argues. “It is as the plant had been vaccinated, and the envelope of the virus does not express itself in the reproductive part”, she clarifies.

It is, however, fundamental to assess its impact on other organisms and on the ecosystem. When an insecticide is used to kill the aphids that transmit the ringspot virus, for example, there is the risk of the product also killing some insects that fly around the papaya, denominated by the specialists as non-target organisms. “We will be assessing whether, in the case of the papaya, the insect that flies around the tree will or will not be affected. The cost-benefit ratio has to be taken into account, to see whether the risk of affecting the insect is worthwhile”, Deise Capalbo ponders. “If there is no detriment to the function and not just to the number of individuals in that ecosystem it may perhaps be possible to tolerate a situation of a few losses of the non-target organisms”, she rationalizes.

The researchers will also investigate the nutritional safety of the genetically modified fruit. A series of tests and proofs such as those for substantial equivalence (SE), molecular and agronomic characteristics of the product, amongst others to assess the nutritional qualities of the plant and to compare them with those of the conventional plant. “If the two plants do not have identical properties and characteristics, the project needs reassessing”, explains Deise Capalbo. The assessment of the impact of the GMO on the environment and its nutritional safety will follow the rules of the Law on Biosafety, guidelines of the National Technical Biosafety Commission (CTNBio), international standards and protocols drawn up by Embrapa itself. All this work will be followed up and analyzed by Embrapa’s Biosafety Network. Working in a network has the advantage of standardizing communication between researchers, allowing a quick revision of procedures and analyses, incorporating new data to the investigation, besides expanding significantly contact with similar international scientific ventures. “Our concern is to give answers to basic issues about GMOs”, she says.

The network is made up of 120 researchers linked to 12 of the company’s research centers, besides five universities: the federal universities of Minas Gerais, Rio de Janeiro, Fluminense, Viçosa and Bahia, as well as the universities of Brasilia, Feira de Santana, the State University of Campinas (Unicamp) and the Júlio de Mesquita Filho State University (Unesp), in Jaboticabal. To fund the Biosafety Network, Embrapa sought funds from the Financier of Studies and Projects (Finep). The project is budgeted at R$ 9 million, which will be invested in the course of four years, explains Deise Capalbo, one of the coordinators of the project. The project for the network was conceived four years ago, at the same time that CTNBio authorized the sales of the Roundup Ready soybeans, from Monsanto. At the time, Embrapa was speeding up the development of research in the laboratory of plants resistant to insects and chemical herbicides, and the decision from CTNBio opened up a prospect for future marketing. The network was approved and started to work in the middle of last year. The first step was to select the products that would be assessed, starting with those that were closest to reaching the market. Besides papaya, the choices fell on a bean that was resistant to golden mosaic, a potato that is resistant to the virus Y virus, and Bt cotton that is resistant to insects. In a second stage, the laboratories that ought to take part in the network were identified.

There is no variety of beans in Brazil with immunity to golden mosaic (BGMV in Phaseolus sp.), which puts the researchers before the challenge of seeking alternatives for the producer. Initial studies under controlled conditions indicated that the genetic modification developed by researchers from Embrapa have a potential for solving the problem. In the case of the potato, and using recombinant DNA technology, the researchers a clone of the Achat cultivar, transformed with the CP-PVY (coat protein of the potato virus Y) gene, with a high resistance to the disease that degenerates the tubercle. As the Achat already shows a high tolerance to the Potato Leaf Roll Virus (PLRV), combining it with resistance to the PVY will make it possible for the producers to keep productivity high, the researchers suppose. They do not expect any negative environmental impacts from gene flow (the transfer of genes from one plant to another), since the genetically modified potato does not flower. As the inserted gene codifies a protein that is already widely consumed in tubercles naturally infected by virus, it is very probable too that no problems will occur deriving from its consumption by human beings or animals.

The researches with cotton are still at an initial stage. Embrapa has not yet got ready the modified plant with the gene of Bacillus thuringiensis (Bt) which is expected to make it immune to the attack of the caterpillar or weevil. “We already know the gene, but we have to introduce it into the plant, to get some small seedlings, before confirming whether the cotton is resistant”, says Deise Capalbo. But, in the case of cotton, there is a great concern with regard to the safety of this transgenic for human beings and non-target organisms, in the jargon of the researchers. “Cotton pollinizes, and that is why there is the possibility of gene escape and gene flow via soil or pollen”, Deise Capaldo points out. This problem can be particularly serious in Brazil, a country where native cotton plants exist. That is why, before taking planting to the field, the area has to be defined where one can plant without risks for the autochthonous varieties of the plant. “We have to carry out a mapping of the areas where there are non-transgenic plants or significant varieties, which have to be preserved”, she explains. More than this: the habits of the insects need to bestudied, the distance they are capable of covering, and even the conditions for agricultural handling. “We have now begun studies with bees, an insect that pollinizes cotton”, she reveals.

Bean database
The 120 researchers of the Network are going to accompany the evolution and analysis of the papaya crop in Cruz das Almas and the development of the researches into genetically modified potatoes, beans and cotton. In the case of nutritional safety, the first initiative is to create a database with detailed information on the conventional varieties of the modified plants, according to Edson Watanabe, a researcher with Embrapa Food Technology. Ascertaining the substantial equivalence of the genetically modified product, in relation to its conventional analog, constitutes one of the first steps in the assessment of the nutritional safety of a GMO. “The expectation is that the GMO will be equivalent to and just as safe as its conventional analog”, Watanabe explains. To start with, the centesimal composition of the transgenic is being studied proteins, lipids, carbohydrates etc. and the key food components.

In the case of soybeans, this component is, at least up until this moment, isoflavone. “But it is different with papaya”, Watanabe points out, recalling that there is not any single model for analysis, which should be determined case by case. In the comparisons between the two products, resource can be made to international databases, but for the analysis of Brazilian plants, domestic protocols have to be developed. Embrapa Food Technology, for example, has now started to create a database with information on the various varieties of conventional beans, cultivated in Brazilian territory. The safety tests also include examinations of toxicity. In the case of soybeans and corn two kinds of grains normally used as a basis for animal feed, the methodology used to assess the nutritional safety of transgenic products, already approved by the United States, is acute gavage (feeding by mouth) on ice, carried out by means of the injection of the protein expressed by the new gene directly into the rodents stomachs. In another kind of examination, the rodents are fed over 90 days with feeds produced from transgenic grains and assessed by the team of researchers. “Other proposals exist, though they have not been validated”, Watanabe observes.

The possibility that the transgenic product could cause allergy also has to be looked into. Bioinformatics makes it possible to get information on the sequence of amino acids of the expressed protein, comparing it with those from the international protein databases, including the allergenic ones. “Digestion by pepsin (a digestive enzyme produced by the stomach) is also measured, since allergenic proteins are more resistant to digestion”, he says. When it is a question of processed foods like the oil and bran from soybeans, for example, there needs to be an assessment of the product throughout the whole industrial process. And in the case of feed, the performance of the animal fed with the product is assessed. Verifying the substantial equivalence also includes studies like the molecular and agronomic characterization of the product, to observe the size and yield of the plants. “One has to know how many copies of plasmid were inserted into the plant, where these copies were inserted, and whether the genetic modification is stable, amongst other factors”, is the explanation given by Watanabe.

Good practices
Besides the Biosafety Network, Embrapa is investing in the accreditation of its laboratories researching into transgenics. By the end of the year, Embrapa Food Technology, in Rio de Janeiro, should present to the National Institute of Metrology, Standardization and Industrial Quality (Inmetro the necessary documents for the ISO 17025 accreditation in laboratory analyses. Next year, “if the transgenics reach Embrapa Food Technology, accreditation will also be requested of Inmetro for Good Laboratory Practices (GLP)”, according to Rosemar Antoniassi, who supervises Embrapa’s Quality System. Since 1998, reforms have been carried out in the infrastructure and equipment acquired to expand the capacity for analyzing, within the GLP standards. “We have been investing in storage and disposal. The important thing in GLP is to adopt procedures for documentation and for records of research into food and environmental safety”, she underscores. “All the steps in the research must be reported”, Rosemar Antoniassi observes. To do so, training was carried out, involving researchers from the various areas of research. Accreditation is applied for case by case, for each one of the GMOs to be assessed by Embrapa, and is granted after a careful audit by Inmetro.