<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.</p>

<p style="text-align: justify;">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.</p>

<p style="text-align: justify;">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 Colorado potato beetle 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.<br />
The transgenic potato lines BT6, BT10, BT12, BT16, BT17, BT18, and BT23 were genetically engineered to resist CPB by producing their own insecticide. These lines were developed by introducing the <i>cry3A</i> gene, isolated from the common soil bacterium <i>Bacillus thuringiensis</i> subspecies tenebrionis (Btt), into the potato cultivar &lsquo;Russet Burbank&rsquo; by <i>Agrobacterium</i>-mediated transformation. The <i>cry3A</i> gene produces the insect control protein Cry3A, a delta-endotoxin.<br />
The Cry3A protein produced by BT6, BT10, BT12, BT16, BT17, BT18, and BT23 is identical to that found in nature and in commercial <i>Bt</i> spray formulations. Cry proteins, of which Cry3A is only one, act by selectively binding to specific sites localized on the lining of the midgut of susceptible insect species. Following binding, pores are formed that disrupt midgut ion flow causing gut paralysis and eventual death due to bacterial sepsis. Cry3A is insecticidal only when eaten by the larvae of coleopteran insects such as Colorado potato beetle and its specificity of action is directly attributable to the presence of specific binding sites in the target insects. There are no binding sites 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.</p>

<p style="text-align: justify;">Transgenic potato lines BT6, BT10, BT12, BT16, BT17, BT18, and BT23 were tested in field trials in the United States (1991-1994) and Canada (1992-1995). Data collected from these trials demonstrated these potato lines were not different from conventional Russet Burbank. They grew normally and exhibited the expected morphology, reproductive and physiological characteristics of potatoes. Susceptibility to diseases and insects, other than CPB, remained unchanged and the transgenic potato lines did not exhibit enhanced weediness potential.</p>

<p style="text-align: justify;">Dietary toxicity studies were performed using the Cry3A protein on four beneficial insects (honeybee, ladybird beetle, green lacewing and parasitic wasp), and eight non-target insect species (southern corn rootworm, yellow fever mosquito, green peach aphid, European corn borer, tobacco hornworm, corn earworm, tobacco budworm and German cockroach). No negative effects were observed, except for slightly higher mortality and reduced honeydew production of green peach aphids, which as vectors of damaging potato viruses, are normally controlled in potato fields by chemical means. BT6, BT10, BT12, BT16, BT17, BT18, and BT23 were not expected to impact on threatened or endangered species.</p>

<p style="text-align: justify;">Generally, varieties of <i>S. tuberosum</i> are capable of cross-breeding with each other, but genetic exchange with other Solanum species is usually unsuccessful. In Canada there are no tuber producing wild relatives of Solanum. In the United States, tuber-bearing Solanum species include <i>S. jamesii</i>, <i>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>.</p>

<p style="text-align: justify;">Gene transfer from BT6, BT10, BT12, BT16, BT17, BT18, and BT23 to other potato cultivars is unlikely. Multiple barriers, such as male sterility, prevent pollen production in the CPB-resistant potato lines as the transgenic lines are derived from the male sterile cultivar &#39;Russet Burbank&#39;. Cross-pollination between BT6, BT10, BT12, BT16, BT17, BT18, and BT23 and male-fertile potato cultivars and subsequent seed production is restricted due to limited pollen dispersal in potatoes.</p>

<p style="text-align: justify;">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.</p>

<p style="text-align: justify;">The food and livestock feed safety of transformed potato lines BT6, BT10, BT12, BT16, BT17, BT18, and BT23 was established based on several standard criteria. As part of the safety assessment, the nutritional composition of the seven CPB-resistant lines was found to be equivalent to conventional potatoes as shown by the analyses of key nutritional parameters including protein, fat, ash, total dietary fibre, carbohydrates, calories, and micronutrients and trace elements (<i>e.g.</i>, thiamine, niacin, riboflavin, vitamin C, calcium, iron and zinc). There were significant differences in carbohydrate, fat and fibre although in each case the measured concentration was within the range normally reported for potatoes. It was concluded that the consumption of products from BT6, BT10, BT12, BT16, BT17, BT18, and BT23 potatoes has no significant impact on the nutritional quality of the Canadian and American food supply.</p>

<p style="text-align: justify;">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 levels in each of the transgenic lines demonstrated that in each case the levels were within the standards previously established for potatoes. The potential toxicity of the Cry3A protein was also examined: in acute oral toxicity studies with mice Cry3A was found to have no negative effects, and the amino acid sequence of Cry3A was not homologous with those of known mammalian protein toxins.</p>

<p style="text-align: justify;">The potential for allergenicity of the Cry3A protein was assessed based upon the physiochemical properties of known food allergens, such as stability to acid and/or proteolytic digestion, heat stability, and glycosylation. Cry3A did not demonstrate any characteristics normally associated with food allergens. Unlike known protein allergens, which are normally resistant to digestion, Cry3A was rapidly inactivated and degraded when subjected to typical mammalian acidic stomach conditions. Additionally, the Cry3A protein has a history of safe use as demonstrated by its application in microbial <i>Bt</i> spray formulations in agriculture and forestry for more than 30 years with no evidence of adverse effects. These facts, combined with the lack of amino acid sequence homology between Cry3A protein and known allergens, were sufficient to provide with reasonable certainty that Cry3A has no allergenic potential.</p>
