<p style="text-align: justify;">Potato (<i>Solanum tuberosum</i> L.) is grown commercially in over 150 countries with a combined harvest of over 315 million metric tonnes in 2006. The major producers of potatoes are China, Russia, India, the United States, Ukraine, Poland and Germany. Potatoes are the fourth most important food crop in the world, providing more edible food than the combined world output of fish and meat. They are grown for the fresh and processed food industries, especially the frozen food sector. In North America, potato tubers are used primarily for French fries, chips, and dehydrated flakes. Other food uses of the crop include consumption of fresh tubers, and in the production of flour, starch and alcohol. Colorado potato beetle (CPB; <i>Leptinotarsa decemlineata</i> [Say]) is the most destructive insect pest of potatoes in North America. The adult and all larval stages feed primarily on foliage and occasionally on stems. When the population of beetles is high, plants can be completely defoliated. Extensive feeding at any time during the growing season can reduce yield, as a reduction in leaf surface area decreases the plant&rsquo;s ability to produce and store nutrients, which affects tuber size and number. Commercial production of potatoes is nearly impossible without using insecticides to control CPB. Thirty-four percent of total insecticide use on potatoes is for control of CPB, more than used on any other insect potato pest. There are several insecticide classes that are available for CPB control including organophosphates, carbamates, pyrethroids, chlorinated hydrocarbons, insect growth regulators, chloronicotinyl, spinosads and abamectins. Colorado potato beetle has shown a tremendous ability to develop resistance to insecticides, including the arsenicals, organochlorines, carbamates, organophosphates, and pyrethroids. Cross-resistance to organophosphates and carbamates, and multiple resistance to organophosphates, carbamates, and pyrethroids has also been reported. Potato leafroll virus (PLRV) is a spherical RNA virus belonging to the luteovirus group and is transmitted in a persistent manner by aphids, primarily the green peach aphid (<i>Myzus persicae</i>). When an aphid picks up the virus from an infected plant, the virus circulates through the aphid, entering the salivary glands. The virus is then transmitted when the aphid probes (feeds on) other potato plants. Although acquisition of the virus by an aphid is rapid (few minutes of feeding), it generally requires 24-48 hours before the aphid can transmit it to another plant. This delay provides a window of opportunity to limit virus spread through the timely application of insecticides for green peach aphid control. Aphids are the only vector of PLRV transmission and infectious aphids remain so throughout their lifespan. The symptoms of PLRV infection include rolling upward of the top leaves and the presence of net necrosis (small brown strands of discoloured tissue extending throughout the stem end of the tuber after a month in storage) in the tubers of some cultivars. The transgenic Russet Burbank NewLeaf&reg; Plus potato lines, RBMT21-129, RBMT21-350 and RBMT22-082, were produced using recombinant DNA techniques and contain two novel genes, whose individual expression results in resistance to attack by CPB and resistance to infection by PLRV. Resistance to attack by CPB was accomplished by introducing the <i>cry3A</i> gene from <i>Bacillus thuringiensis</i> subsp. <i>tenebrionis</i>, which encodes an insecticidal crystalline Cry3A delta-endotoxin protein. The insecticidal activity of Cry3A protein is due to its selective binding to specific sites localized on the brush border midgut epithelium of susceptible insect species. Following binding, cation-specific pores are formed that disrupt midgut ion flow and thereby cause gut paralysis, ultimately leading to bacterial sepsis and death. Delta-endotoxins, such as the Cry3A protein expressed in CPB resistant potato lines, exhibit highly selective insecticidal activity against a narrow range of coleopteran insects such as CPB, elm leaf beetle and yellow mealworm. Their specificity of action is directly attributable to the presence of specific receptors in the target insects. There are no receptors for delta-endotoxins of <i>B. thuringiensis</i> on the surface of mammalian intestinal cells, therefore, livestock animals and humans are not susceptible to these proteins. Introducing DNA sequences corresponding to the ORF-1 and ORF-2 regions from PLRV conferred resistance to PLRV infection. These two ORFs encode the putative viral helicase and replicase domains that are required for viral RNA synthesis. The introduced viral sequences do not result in the formation of any infectious particles, nor does their expression result in any disease pathology. These transgenic potato cultivars exhibit the trait of resistance to infection and subsequent disease caused by PLRV through an incompletely understood process that has been termed &quot;replicase-mediated resistance&quot;, which may involve silencing of viral gene translation. The transgenic NewLeaf&reg; Plus potato lines were tested in field trials in the United States and Canada. Data collected from these trials demonstrated these potato lines grew normally and exhibited the expected morphology, reproductive, and physical characteristics of &lsquo;Russet Burbank&rsquo; potatoes. Susceptibility to diseases and insects, other than CPB and PLRV, remained unchanged. Transgenic NewLeaf&reg; Plus potato lines did not pose a plant pest risk, or exhibit enhanced weediness potential. Dietary toxicity studies were performed using the Cry3A protein on four beneficial insects (honeybee, ladybird beetle, green lacewing and parasitic wasp) in addition to nontarget species such as earthworms, mice and bobwhite quail. The results demonstrated that Cry3A did not negatively affect beneficial or nontarget organisms under study. These NewLeaf&reg; Plus potato lines were not expected to impact on threatened or endangered species. Generally, varieties of <i>S. tuberosum</i> are capable of crossbreeding with each other. However, transgenic NewLeaf&reg; Plus potato lines were derived from the male sterile &lsquo;Russet Burbank&rsquo; potato, which does not produce pollen. As a result, it is not possible for these transgenic potatoes to cross-pollinate other potato varieties or related species. As a rule, successful hybridization between potatoes and Solanum species is only possible with other tuber-bearing species. In Canada, there are no tuber producing wild relatives of Solanum. In the United States, tuber-bearing Solanum species include <i>S. jamesii, S. fendleri</i>, and <i>S. pinnatisectum</i>; however, the possibility of cultivated potato crossing with these species is remote because of geographical isolation and other biological barriers to natural hybridization. No natural hybrids have been observed between these species and cultivated <i>S. tuberosum</i>. Regulatory authorities in Canada and the United States have mandatory requirements for developers of <i>Bt</i> potatoes to implement specific Insect Resistant Management (IRM) Programs. The potential exists for <i>Bt</i>-resistant CPB populations to develop as acreages planted with transgenic CPB-resistant potatoes expand. Hence, these IRM programs are designed to reduce this potential and prolong the effectiveness of plant-expressed <i>Bt</i> toxins, and the microbial <i>Bt</i> spray formulations that contain these same toxins. The food and livestock feed safety of NewLeaf&reg; Plus potatoes was established based on several standard criteria. As part of the safety assessment, the nutritional composition of these transgenic potatoes was found to be equivalent to conventional potatoes as shown by the analyses of key nutrients including proximates (<i>e.g.</i>, total protein, moisture, fat, ash, crude fibre, carbohydrates and calorie content), total solids, sugars, vitamin C, soluble protein, and natural glycoalkaloid toxicants. The glycoalkaloids, solanine and chaconine, are naturally occurring toxicants found in potato tubers, particularly green tubers that have been exposed to sunlight. Analyses of total glycoalkaloid (TGA) levels in each of the transgenic lines demonstrated that in each case the levels were within the standard levels previously established for potatoes. Neither of the Cry3A nor PLRV replicase proteins shares amino acid sequence homology with any known protein toxins. Previous studies on the acute oral toxicity of Cry3A protein have established that this protein does not result in any adverse effects when fed to laboratory mice at doses up to 5220 mg/kg body weight. Toxicity testing was not required for the PLRV replicase because of the long history of exposure of human beings to this protein through the consumption of PLRV-infected potatoes. The Cry3A and PLRV replicase proteins do not possess characteristics typical of known protein allergens. There were no regions of homology when the amino acid sequences of these introduced proteins were compared to the amino acid sequences of known protein allergens, and unlike known protein allergens, the Cry3A protein was rapidly degraded by simulated gastric or intestinal fluids. It was determined that these two proteins were highly unlikely to be allergenic.</p>

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