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Biochemistry

Protein insecticide

A new vegetal toxin is efficient in combating agricultural pests

Farmers throughout the world annually lose billions of US dollars from attacks by various pests on their crops. In Brazil, some studies have revealed that around 30% of the main crops are decimated by insects and other pests, such as ticks and fungi. In order to overcome this problem, the farmers have at their disposal a vast arsenal of chemical insecticides, bio-insecticides and more recently the so-called protein pesticides. When inserted into the plants’ genetic material – which in this case then becomes a genetically modified organism (GMO) -, they act as a bio-pesticide, killing determined types of insects, fungi and even viruses. The protein toxins most widely used today are produced from bacteria of the Bacillus thuringiensis species. Known as BT toxins, they are largely used in the growing of corn, cotton and potatoes in the United States, and in other countries that allow the use of transgenics. In spite of the efficiency in the control of pests, the BT toxins have limited action, because they offer protection only against a restricted number of insects, at times a single species of beetle, moth, butterfly or mosquito.

Advances in research with these bio-pesticides has led a group of researchers from the Toxic Proteins Laboratory (Laprotox) of the Federal University of Rio Grande do Sul (UFRGS) to discover a new type of insecticide protein whose action is wider and safer that that of the BT toxin. One is dealing with a protein group known as the ureases and produced by the plants themselves, as an insecticide activity against beetles of the type dry rot, thumbtacks and fleas, insects that are immune to BT toxin. These animals are pests imported into Brazilian agriculture. For example, the Soya bean plant is attacked by the Southern Green Stink Bug (Nezara viridula), cotton by the Peruvian Red Cotton Stainer (Dysdercus peruvianus) and the string bean by the dry rot beetle Callusobruchos maculatus. “The fleas, as well as eating the plants, are vectors for diseases”, says the biomedical doctor Célia Carlini, the Laprotox coordinator and the researcher responsible for this new development. The protein toxin is also efficient in combating urban pests, such as termites and cockroaches. To this end, the product can be sprayed into the environments of these insects or is available within traps.

In planted fields, the protein needs to be inserted into the plant that will make it into a genetically modified organism. For this to happen the researchers developed an artificial gene that codifies only the part of the protein that is toxic to the insect. After injection the urease, enzymes of the cathepsin type of the insect itself breakdown the protein and a fragment passes through the intestinal cell membrane attacking tissue, as yet unidentified by the researchers, and leading the animal to its death. “Our protein presents a great advantage in relation to the aspect of biosafety when compared with the BT toxin”, says Célia. “It is present in food that are consumed daily by the population, such as beans and plants of the cucurbitaceous type, such as melon, watermelon, turnip and cucumber. This is a sign that it will not do any harm to our organism.” Thus, plants such as Soya bean genetically modified with the insect protein will not cause any harm to human beings.

Two patents
The research from the Rio Grande do Sul group has resulted in two patents, both associated with the production process of the protein fragment with toxic action. This fragment is a peptide, characterized by being a compound formed by a chain of 81 amino acids. The researchers have given to it the name jaburetox, a word formed from the jack bean – the vegetable in which the protein was identified for the first time -, along with the words urease and toxin. As the group knew that the toxic power of the protein was in the fragment formed during the insect’s digestion, the next decision was to attempt to produce this molecule in the laboratory. Their result was surprising: studies showed that the jaburetox would act upon a much greater number of agricultural plants and against many more insects, including those immune to BT toxin. “Our two patents cover all of the production processes of this peptide, be it from the means of purification of the ureases and the insect enzymes or from the way for the construction of an artificial gene, which teaches the cell how to manufacture this peptide. This artificial gene can be introduced into a bacterium or yeast, for the production of the peptide in a test tube, or in a plant, within a transgenic process”, explains  Célia.

The discovery of the jaburetox peptide, which could transform itself into an important ally in the fight against pests in the field, is the crowning moment of more than two decades of research by the biomedical doctor Célia Carlini, a specialist in molecular biology with emphasis on the chemistry of proteins. “Our research began back in 1981 with a study on the canatoxin protein (CNTX), found in Jack Bean (Canavalia ensiformis). This type of bean is not edible, but is largely used in green manure. Highly productive and with a high protein value, it is planted in fields with the finality of incorporating nitrogen into the soil”, the researcher explained. From 1998 until 2005, Dr. Célia has been receiving financing coming from various programs of the Coordination for Personnel Training at Tertiary Level (Capes), from the National Council for Scientific and Technological Development (CNPq) and from the Research Support Foundation of the State of Rio Grande do Sul (Fapergs).

Known family
“During my master’s degree, I isolated canatoxin and went on to study its action mechanism. In 1997, during my post doctorate studies, at the Center for Insect Science of the University of Arizona, in the United States, I discovered that it had insecticide activity.” In a study involving eight different insects, she showed that it was efficient in combating the dry rot of the string bean and the Rhodnius prolixus bug. Nevertheless, it was only in 2001 that the researcher and her team discovered that the canatoxin belonged to a family of proteins very well known in plants called the ureases. “This was an important discovery, since we verified that there exists a much greater offer for the genes, because the urease protein is found in various vegetables. At the same time, we became more comfortable as to its biosafety, because these plants are edible, contrary to the jack bean, the original source of canatoxin.”

The laboratory tests, according to the researcher, have already revealed that the peptide is non-toxic in rats and mice. “The first transgenic plants can already be found in laboratory tests and the results should come out by the end of the year. We are testing transgenic tobacco and studying the action of the toxin on Manduca sexta lizards and fleas of the species Trips tabaci, which are tobacco plant pests… At the same time, the group, who have already published more than fifty scientific papers in national and international magazines, are in contact with researchers from the Brazilian Agricultural Research Corporation (Embrapa – Genetic Resources and Biotechnology) and the UFRGS specialists in cotton and Soya bean transgenics in order to carry out experiments with the peptide on these plants, and with the Luiz de Queiroz Upper School of Agriculture, of the University of Sao Paulo, in Piracicaba (SP), in order to test the discovery on sugarcane. In the case where the results demonstrate that the protein has toxic action against the pests that attack these crops, the next step will be to find a partner to initiate the commercial production of this new bio-pesticide.

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