<p>Major drawbacks of the use of chemical insecticides for the control of insect pests are their expense, their persistence in and hazardous effects on the environment, and their escalating rates of application because of decreasing effectiveness. Therefore, over the last two decades efforts have increased to investigate possibilities of biological insect control like the use of predators, parasites, insect viruses or feromones. Likewise, increased utilization of the microbial insect pathogen <em>Bacillus</em><em>thuringiensis</em> can be regarded as a form of biological insect control. <em>B. thuringiensis is</em> characterized by its ability to produce crystalline inclusions during sporulation which form the major toxic determinant of this bacterium. Most <em>B. thuringiensis</em> strains are active against larvae of lepidopteran species (butterflies), but some show toxicity against larvae of dipteran (flies) or coleopteran (beetles) species. Not only strains, but also different crystal proteins occurring in a single crystal may vary in insecticidal spectra. Sprays based on <em>B. thuringiensis</em> spore/crystal preparations have been used for over thirty years as biological insecticides. Recently, the cloning of crystal protein genes and their expression in transgenic plants and micro-organisms has provided powerful alternative strategies for the protection of crops against insect damage. These additional potential applications resulted in an increased interest in this bacterium and its crystal proteins in recent years.<p>This thesis endeavours to contribute to our understanding of the entomocidal activity of <em>B. thuringiensis,</em> and more in particular of its crystal proteins. Studies are described to identify crystal proteins with new insecticidal spectra, to gain more insight into the mode of action of crystal proteins, and to examine some of their potential applications.<p>Chapter 1 reviews the mode of action of the crystal proteins. It presents a survey of histological and physiological changes observed in the insect and <em>in vitro</em> cultured cells induced by crystal proteins. In addition, investigations using isolated midgut cells, brush border membrane vesicles and artificial phospholipid vesicles are summarized. Finally, conclusions concerning the molecular basis of the mode of action of both <em>Lepidoptera</em> and <em>Diptera</em> specific crystal proteins are discussed.<p>In chapter 2, an analysis of crystal protein genes and their proteins, in particular those occurring in <em>B. thuringiensis</em> serotype <em>entomocidus</em> 60.5, <em>is</em> presented. The nucleotide sequence of a gene isolated from this bacterial strain, the reference type of <em>cryIC</em> , is presented together with its deduced amino acid sequence. Furthermore, several conserved features revealed by an analysis of the amino acid sequences of all crystal proteins are evaluated.<p>In chapter 3, domain-function studies on the crystal proteins are described using hybrid crystal proteins based on CryIA(b) and CryIC constructed in order to improve our understanding of the mode of action of crystal proteins, especially of its molecular basis.<p>Chapter 4 describes the analysis of a translation fusion product of two different insecticidal crystal protein genes. Based on the presented results an alternative strategy for the introduction of more effective insect resistance in transgenic organisms using <em>B. thuringiensis</em> crystal protein genes is proposed.<p>In chapter 5, the generation of insect resistant transgenic plants using the crystal protein genes <em>cryIA(b)</em> and <em>cryIC</em> , <em>is</em> described.<p>Finally, in chapter 6, the three-dimensional structure of one of the crystal proteins, which recently became available, is reviewed as well as alternative strategies for <em>B.</em><em>thuringiensis</em> applications to prevent crystal protein resistance in insect populations, all in relation to the results presented in this thesis.
|Qualification||Doctor of Philosophy|
|Award date||19 Jun 1992|
|Place of Publication||S.l.|
|Publication status||Published - 1992|
- plant protection