IPM MANAGEMENT IN COTTON FIELDS & FIELD CROPS Transgenic Cotton & Corn with Bt (Bacillus thuringiensis) Bacillus thuringiensis Bt was first discovered in 1901 by Japanese biologist Ishiwatari. In Germany 1911, Bt was isolated by Berliner in flour moth caterpillars, as the cause of a disease called Schlaffsucht In 1976, Zakharyan reported the presence of a plasmid in a strain of Bt involved in endospore insecticidal and crystal formation, called crystal proteins or Cry (proteins) toxins. Cry toxins belong to a large family known as PFT PORE FORMING TOXINS (PFT) PFT are important virulence factors produced by bacteria to kill eukaryotic cells by forming holes in the cellular membrane. They represent a diverse group of proteins with a wide range of target cells. The mode of action of PFT involves: (a) Receptor recognition. (b) Activation by proteases (protein enzymes). (c) Aaggregation into oligomericstructures (molecule that consists of a few monomer units). (d) Insertion into the membrane to form ionic pores. BT CRY TOXIN – MOD OF ACTION The toxin core travels across the peritrophic matrix Cry toxins interact with specific receptors called cadherin, located on the host cell surface, the brush border membrane of the gut cells. Activated by host proteases following receptor binding . A new pre-pore oligomeric structure is formatted. The new formation is insert into the membrane to form ionic pores. Pores lead to osmotic cell lysis. Bt as an Insecticide Bt is a naturally-occurring soilborne bacterium that is found worldwide. A unique feature of this bacterium: its production of crystal-like proteins (Cry) and Cytolitic (Cyt) protein families, that selectively kill specific groups of insects: Lepidoptera, Coleoptera, Diptera and also other invertebrates such as nematodes Cry proteins are delta endotoxins, insect stomach poisons that must be eaten to kill insects. Once eaten, insects stop feeding within 2 hours of a first bite and, if enough toxin is eaten, die within 2-3 days. For more than 30 years, various formulations of Bt have been used successfully against insect pests on a variety of crops. Δ-ENDOTOXINS Historically, B.t. toxins have been divided into two groups on the basis of target specificity: (a) Insect-specific Cry (crystal) proteins (b) Generally cytolytic Cyt proteins. δ-endotoxins are a super family of proteins that occur as crystalline inclusions in the spore-forming bacterium Bacillus thuringiensis. Considered environmentally safe insecticides. The Cry proteins are the group that is mostly being used in commercial insecticide preparations. In 1981, scientists cloned the first Bt toxin gene, which led quickly to the development of the first transgenic Bt plants in the mid1980s. There are several strains of Bt, each with differing Cry proteins. Scientists have identified more than 60 Cry proteins. Other Types of Bt Toxins Other types of Bt toxins are called Vegetative Insecticidal Proteins, or VIPs. VIPs are also considered relatively safe for nonpest species. Other classes of toxins produced by Bt have a broader spectrum of toxicity. Transgenic Bt Plants A transgenic Bt plant produces delta endotoxin or Vips controlled by a single gene in the bacteria. A modified version of this gene can be placed in plants. Plants containing this gene can produce delta endotoxin or VIP and therefore be toxic to insects that are susceptible to that form of the protein. Plant geneticists create Bt crops by inserting selected exotic DNA into the corn plant's own DNA. Seed companies select elite hybrids for the Bt transformation in order to retain important agronomic qualities for yield, harvest ability and disease resistance The Essential Global Needs leading Towards Production of Bt Crops The need to increase food production to meet the food and feed demands of the world, requiring higher production, particularly in developing countries in Asia, Africa and Latin America. This demand has to meet primarily through yield increases on existing cultivated lands in order to be environmentally sustainable and cost effective. How safe is Bt and Bt Crops? The EPA U.S. (Environmental Protection Agency), considered 20 years of human and animal safety data before registering Bt corn. Bt proteins are not toxic to people, domestic animals, fish, or wildlife; and they have no negative impacts on the environment. The Food and Drug Administration (FDA) exempts Bt Cry proteins from residue analyses because of Bt's history of safety and because these proteins degrade rapidly. Bt hybrids Genetic Package Three primary components of the genetic package inserted into the crop include: A. Protein gene. Bt genes, modified for improved expression in crops, produce Cry proteins. B. Promoter. A DNA sequence that regulates where, when, and to what degree, an associated gene is expressed. C. Genetic marker. The presence of a genetic marker allows seed companies to identify successful transformations. The Promoter Some promoters limit protein production to specific parts of the plant. Example: Some promoters produce protein throughout leaves, green tissue and pollen, whereas others produce protein throughout the plant. Genetic marker. Current examples of markers include genes for herbicide resistance or antibiotic resistance. The genetic package is inserted into plant through a variety of plant transformation techniques Successful transformations, called "events," vary in the components of the genetic package and where this DNA is inserted into the corn DNA. The insertion site may affect Bt protein production and could affect other plant functions. Consequently, seed companies carefully scrutinize transformation events to ensure adequate production of Bt protein and no negative effects on agronomic traits. Transgenic Bt Crops, Advantages Non-toxic to vertebrates unlike synthetic pesticides Very specific to particular insect pests. Reduction in sprays per season, with synthetic pesticides Very specific to particular insect pests. Suitable for small-scale farmers, no equipment and pesticide knowledge are needed Benefits both the environment and labourers’ health, Increase incomes through higher yields of healthier grain. Global Area of Biotech Crops, 1996 to 2010 (Million Hectares) Bt Issues There is some evidence that the prolonged use of Bt crops can create emergence of resistant insect strains or elevation in populations of insects that are naturally resistant; these would likely have emerged with the use of Bt alone. How does resistance develop? Many factors contribute to the development of resistance: Widespread use of Bt crops High season-long mortality Two or more generations per year. The lack of “refuges” – small crop areas that are not GM. Insect resistance management (IRM) high dose and refuge strategy assumes resistance is recessive. Many susceptible moths (SS) are produced in refuge maize that mate with rare resistant (RR) moths. Mating of resistant (RR) and susceptible (SS) moths produces heterozygous (RS) moths that die when they feed on high-dose Bt maize. This strategy dilutes resistance genes and delays or prevents the evolution of resistance to Bt maize. © 2012 Nature Education All rights reserved. Avoiding/Delaying appearance of resistance Farmers who plant crops with Bt trait(s) must also plant a refuge area - a block or strip of crops without the Bt gene. Insects who feed on that crop remain susceptible to Bt technology. When they then mate with the rare insects that survive after feeding on Bt corn, they produce offspring that is susceptible to the technology. Refuge Configurations Farmers Can Use Different Refuge Requirements The EPA requires a corn field that contains insect-protected (or Bt) crops with a single modeof-action against corn rootworm to be planted with 20% non-Bt corn in corn-growing areas (the U.S. Corn Belt) and 50% non-Bt in cotton-growing areas (the U.S. Cotton Belt). New seed technologies that have multiple Bt genes, or modes-of-action, provide additional protection. This enabled the EPA to reduce the size of structured. The right amount of refuge seed is included in the bag Farmers simply put the complete seed bag in the planter and plant both the Bt crop and the refuge For cotton growers, there was a lot of pressure from pests before the introduction of Bt cotton in 1996. Due to synthetic pesticide resistance, farmers were losing much of their cotton to tobacco budworms, cotton bollworms, and pink bollworms. Tobacco Budworm, Heliothis virescens Cotton Ballworm Helicoverpa armigera Cotton Ballworm Helicoverpa zea The Pink Bollworm Pectinophora gossypiella Benefits of Bt Cotton China , 2007 Green bolls (100/field) were sampled from the uppermost internodes within adjacent fields of Bt (Deltapine 33B) and non-Bt (Hyperformer HS 44) cotton experiencing severe pink bollworm pressure late in the growing season. Average top crop lint yield reductions ranging from 30 to 70% were observed in the uppermost bolls of the non-Bt cotton variety. Average top crop lint yield reductions ranging from 0 to 40% were observed in the uppermost bolls of the transgenic Bt cotton variety. . YI E L D S O F BT C O T T O N V E R S U S CONVENTIONAL COTTON IN CHINA (KG/HA) . Benefits of Bt Cotton in India, 2011 Pesticide use has been cut by at least half, a new study shows. The use of Bt cotton helps to avoid at least 2.4 million cases of pesticide poisoning in Indian farmers each year, saving US$14 million in annual health costs. (The current study, published online in the journal Ecological Economics) India is now the world’s biggest producer of Bt cotton with an estimated 23.2 million acres planting in 2010. Insects Evolving Resistance Scientists have confirmed incidents of insects evolving resistance to Bt toxins in the field to date: Bt cotton in India (2010) and US (2008). Field monitoring in parts of Gujarat has discovered that the Bt crop is no longer effective against the pink bollworm pest there. GM Cotton May Affect Biomass A study concluded in India 2011, concluded: A decade of planting with GM cotton, or any GM crop with Bt genes could lead to total destruction of soil organisms, “leaving dead soil unable to produce food.” GM Cotton May Affect Biomass Bacteria overall were reduced by 14 percent, while the total microbial biomass was reduced by 8.9 percent. Vital soil enzymes, which make nutrients available to plants, have also been drastically reduced. Acid phosphatase which contributes to the uptake of phosphates was lowered by 26.6 percent. Nitrogenase enzymes, which help fix nitrogen, were diminished by 22.6 percent. Bt Cotton Led Agri-revolution, 2009 Bt cotton adversely affected the farmers leading to suicides being committed, and loss of biodiversity. There are studies, which show that yields do not rise necessarily due to growing of Bt cotton. The quality of the cotton grown is poor in the case of Bt cotton. It provides little resistance to Pink bollworm. The economic advantage of growing Bt cotton is over-exaggerated. The seed companies are extracting huge royalties by selling the genetically modified seeds at higher prices to farmers. European corn borer Ostrinia nubilalis Southwestern Corn Borer, Diatraea grandiosella, Corn Earworm Helicoverpa zea Fall Armyworm Spodoptera exigua Black Cutworm Peridroma saucia Western corn rootworm (Diabrotica virgifera) The diamondback moth (Plutella xylostella), sometimes called cabbage moth Company Bt gene Dow Chemical CryIF + Cry34Ab1 + Co. + Cry35Ab1 + Monsanto C0ry3Bb1 + Cry1A.105+ Cry2Ab Syngenta Seeds Inc VIP and CryIAb Trade name Pests controlled or suppressed SmartStax European and southwestern corn borers, corn earworm, fall armyworm and corn rootworms Agrisure Viptera 3110 Corn borers, corn earworm, fall armyworm, black cutworm Refuge requirement 5% non-Bt refuge in , or immediately adjacent to Bt field 20% non-Bt refuge needed within 1/4 to 1/2 mile Resistance of Rootworm in Bt Cron Severe lodging caused by western corn rootworm larvae to Bt corn expressing the Cry3Bb1 protein (northwestern Illinois, August 16 2011). It is said that the Western corn rootworm (Diabrotica virgifera) causes one billion dollars worth of damage annually in the maizegrowing areas of the USA. The soil-dwelling larvae of the beetle eat the aerial roots, which quickly die back. Older larvae then penetrate the heart of the root and from there move further into the maize plants. Many infested plants collapse before harvest as a result of the root damage. If rootworms do become resistant to Bt corn, it “could become the most economically damaging example of insect resistance to a genetically modified crop in the U.S.,” said Bruce Tabashnik, an entomologist at the University of Arizona. “It’s a pest of great economic significance — a billion-dollar pest.” FIELD-EVOLVED INSECT RESISTANCE BT CROPS: A REVIEW TO After more than a decade since initial commercialization of Bt crops, most target pest populations remain susceptible, field-evolved resistance has been documented in some populations of three noctuid moth species: Spodoptera frugiperda (Armyworm) , Bt corn ,Puerto Rico, Busseola fusca (stem borer) to Cry1Ab in Bt corn in South Africa, Helicoverpa zea (Corn Earworm ) to Cry1Ac and Cry2Ab in Bt cotton in the southeastern United States. Factors delaying resistance include: Recessive inheritance of resistance, Abundant refuges of non-Bt host plants, Two-toxin Bt crops deployed separately from one-toxin Bt crops. Journal of Economic Entomology- 2009 BT TOXINS Scientists have modified the molecular structure of two Bt toxins, Cry1Ab and Cry1Ac, in order to overcome resistance. The novel toxins, Cry1AbMod and Cry1AcMod, are effective against five resistant insect species, such as the diamondback moth, the cotton bollworm, and the European corn borer. Cry1AbMod and Cry1AcMod can be used alone or in combination with other Bt toxins for plant protection New Bacteria Toxins Against Resistant Insect Pests Scientists have developed Bt toxins for the management of Bt resistance in European corn borer and other crop pests. B.t. VIP Toxins In addition to the endotoxins, B. thuringiensis also produces secreted insecticidal proteins during its vegetative growth stage, namely, vegetative insecticidal proteins (Vip). Vip toxins include two major groups : Vip1 and Vip2 The combination of Vip1 and Vip2 is highly insecticidal to an agriculturally important insect, the western corn rootworm (Diabrotica virgifera), but does not show any insecticidal activity for any lepidopteran insects VIP3 TOXIN The first-identified Vip3 toxin, Vip3Aa1, is highly insecticidal to several major lepidopteran pests of maize and cotton, including the fall armyworm Spodoptera frugiperda and the cotton bollworm Helicoverpa zea, but shows no activity against the European corn borer Ostrinia nubilalis, a major pest of maize. Vip3 contributes to the overall toxicity of B. t strains STRATEGIES TO IMPROVE THE INSECTICIDAL ACTIVITY OF CRY TOXINS FROM BACILLUS THURINGIENSIS (i) Chitinase that improves accessibility of the toxin to the epithelial membrane. (ii) Serine protease inhibitors that reduce degradation of Cry or of toxin-receptors. (iii) Introduction of intramolecular cleavage sites in the toxin that improves binding interaction. (iv) Introduction of more binding sites such as the CR12-MPE-peptide from cadherin receptor or the Cyt1A toxin. (v) Deletion of helix α-1 that induces toxinoligomerization and skip cadherin interaction. Peptides Vol: 30 589-595 2009 STRATEGIES TO IMPROVE THE INSECTICIDAL ACTIVITY OF CRY TOXINS FROM B.T Monsanto's GMO sweet corn is engineered to tolerate the herbicide Roundup, and to produce the insectkilling toxin Bt. These modifications have been shown to lead to some serious health problems in animals who eat them. How can genetically modified (GM) corn and its attributed changes in agricultural practices affect the agroecosystem’s biodiversity and the economic status and social life of the farmers? Philippines 2013 GMO IMPACT ON HUMANITY Genetic engineering may disrupt the precise sequence of a food's genetic code. Disturbs the functions of neighboring genes, Can give rise to potentially toxic or allergenic molecules or even alter the nutritional value of food produced. The Bt toxin used in GMO corn, was recently (2013), detected in the blood of pregnant women and their babies, with possibly harmful consequences. GMO Impact on livestock– Philippines 2013 Livestock, especially goats and cattle that eat GM corn leaves and stalks, become sickly, suffering from diarrhea. Carabaos and cows have been observed to lose weight and are sickly. In Caquilingan, Cordon, Isabela for example, five carabaos died in 2011. Farmers examined the carabaos’ stomachs, these were filled with undigested corn fodder. GMO May Cause Genetic Contamination A GMO crop, once released in the open, reproduces via pollination and interacts genetically with natural varieties of the same crop. A GMO crop produces ‘genetic contamination’. According to a study published in Nature, one of the world's leading scientific journals, Bt corn has contaminated indigenous varieties of corn tested in Oaxaca, Mexico. GMO Impact on Biodiversity GMO brought into natural surroundings may have a toxic or lethal impact on other living organisms. Bt corn destroyed the larvae of the monarch butterfly. Many other natural plant and animal life may be impacted in the same way. GMO Impact on Environment – Philippines 2013 Scarce vegetation, acidic soil, Silted rivers, only few fishes remain; Papaya fruits are deformed Soil is easily eroded especially during the rainy season. Stones protrude out from the soil and the soil turns red and clayish. Corn leaves turn violet. Published 2013 by Magsasaka at Siyentipiko para sa Pagunlad ng Agrikultura MASIPAG The benefits of GMOs have been oversold by the companies, like Monsanto and Syngenta, that peddle them. It has been shown, however, that insects are fast developing resistance to Bt as well as to herbicides, resulting in even more massive infestation by the new superbugs. No substantial evidence exists that GM crops yield more than conventional crops. What genetically engineered crops definitely do lead to is greater use of pesticide, which is harmful both to humans and the environment. Public debates about transgenic crops should focus more on the health and environment al benefits which can be substantial.