Identifying Arabidopsis thaliana Defense Genes to Phloem-feeding Insects

Abstract

The whitefly (Bemisia tabaci) is a serious agricultural pest that afflicts a wide variety of ornamental and vegetable crop species. To enable survival on a great diversity of host plants, whiteflies must have the ability to avoid or detoxify numerous different plant defensive chemicals. Such toxins include a group of insect-deterrent molecules called glucosinolates (GSs), which also provide the pungent taste of Brassica vegetables such as radish and cabbage. In our BARD grant, we used the whitefly B. tabaci and Arabidopsis (a Brassica plant model) defense mutants and transgenic lines, to gain comprehensive understanding both on plant defense pathways against whiteflies and whitefly defense strategies against plants. Our major focus was on GSs. We produced transgenic Arabidopsis plants accumulating high levels of GSs. At the first step, we examined how exposure to high levels of GSs affects decision making and performance of whiteflies when provided plants with normal levels or high levels of GSs. Our major conclusions can be divided into three: (I) exposure to plants accumulating high levels of GSs, negatively affected the performance of both whitefly adult females and immature; (II) whitefly adult females are likely to be capable of sensing different levels of GSs in their host plants and are able to choose, for oviposition, the host plant on which their offspring survive and develop better (preference-performance relationship); (III) the dual presence of plants with normal levels and high levels of GSs, confused whitefly adult females, and led to difficulties in making a choice between the different host plants. These findings have an applicative perspective. Whiteflies are known as a serious pest of Brassica cropping systems. If the differences found here on adjacent small plants translate to field situations, intercropping with closely-related Brassica cultivars could negatively influence whitefly population build-up. At the second step, we characterized the defensive mechanisms whiteflies use to detoxify GSs and other plant toxins. We identified five detoxification genes, which can be considered as putative "key" general induced detoxifiers because their expression-levels responded to several unrelated plant toxic compounds. This knowledge is currently used (using new funding) to develop a new technology that will allow the production of pestresistant crops capable of protecting themselves from whiteflies by silencing insect detoxification genes without which successful host utilization can not occur. Finally, we made an effort to identify defense genes that deter whitefly performance, by infesting with whiteflies, wild-type and defense mutated Arabidopsis plants. The infested plants were used to construct deep-sequencing expression libraries. The 30- 50 million sequence reads per library, provide an unbiased and quantitative assessment of gene expression and contain sequences from both Arabidopsis and whiteflies. Therefore, the libraries give us sequence data that can be mined for both the plant and insect gene expression responses. An intensive analysis of these datasets is underway. We also conducted electrical penetration graph (EPG) recordings of whiteflies feeding on Arabidopsis wild-type and defense mutant plants in order to determine the time-point and feeding behavior in which plant-defense genes are expressed. We are in the process of analyzing the recordings and calculating 125 feeding behavior parameters for each whitefly. From the analyses conducted so far we conclude that the Arabidopsis defense mutants do not affect adult feeding behavior in the same manner that they affect immatures development. Analysis of the immatures feeding behavior is not yet completed, but if it shows the same disconnect between feeding behavior data and developmental rate data, we would conclude that the differences in the defense mutants are due to a qualitative effect based on the chemical constituency of the phloem sap.