The expression of Bt Cry1Ac in transformed cotton Bt Cry1Ac under abiotic stress

Abstract

Bacillus thuringiensis (Bt) is a gram-positive common soil bacterium that produces crystals (Cry) containing proteins that are toxic to certain insects, in particular larvae of Lepidoptera and Diptera. The Bt toxin in the past has been widely used as a bioactive compound for the biological control of mainly lepidopteran pests. Most recently a variety of crops, including cotton and maize, have been genetically modified to express a Bt toxin to confer resistance to lepidopteran pests. However, the effect of abiotic environmental stress, such as drought and heat, which are typical for Africa, on Bt toxin expression in a genetically modified crop has so far not been fully evaluated. This study focuses on the expression and stability of the Cry1Ac insecticidal protein from Bacillus thuringiensis in genetically modified cotton plants under drought and heat stress. These include the physiological and biochemical characterization of the expressed Bt toxin gene under drought stress as well as the biological activity against first-instar larvae of the African cotton bollworm Helicoverpa armigera (Lepidoptera: Noctuidae). Non-genetically modified cotton (Gossypium hirsutum cv. Opal), as well as genetically modified cotton (cv. Nuopal) expressing the Bt toxin Cry1Ac, were exposed to drought and heat stress. Drought stress was induced by withholding watering plants until the soil moisture content reached 25- 30 % of field capacity. Non-stressed control plants were watered and soil moisture content to 80-100 % of field capacity was maintained. For heat stress, plants were grown at 38 to 32 DC during the day and night, respectively, whereas control plants were grown in a growth cabinet at a 28/25 DC day/night cycle. For growth analysis plants were harvested every second week after planting. At each harvest, different parts of the plant were collected and their fresh and dry weight determined. For biochemical analysis and determining biological activity against first-instar larvae of H. armigera, two types of experiments were carried out, the first experiment four weeks after treatment induction and the second experiment eight weeks after treatment induction. Different plant material (leaves, flowers and immature green bolls) were used for Bt detection as well as for determining biological activity against first-instar larvae of H. armigera. Under drought stress conditions a reduction in leaf area and leaf dry weight were found in both Bt toxin expressing and non-expressing cotton plants, but no significant difference in physiological performance between Bt-expressing and non-expressing cotton plants was found. This study shows that the Bt toxin (Cry1Ac) level decreases in senescent plants and that drought stress did not affect the growth and development of genetically modified Bt plants when compared to non-Bt plants. Although the expression of Bt toxin (Cry1Ac) in Bt cotton plants decreased under drought stress no effect on the efficacy of the toxin against H. armigera was observed. In addition, no significant decrease of Bt toxin content was found in Bt cotton leaves after exposure to heat stress when compared to leaves from nonheat stressed plants.