Date of Award

2015

Document Type

Open Access Master's Thesis

Degree Name

Master of Science in Biological Sciences (MS)

Administrative Home Department

Department of Biological Sciences

Advisor 1

Thomas Werner

Committee Member 1

Rupali Datta

Committee Member 2

Hairong Wei

Abstract

Insect pesticide resistance has become a costly problem in the US. To make things more problematic, resistance to one pesticide is often associated with cross-resistance to other toxins, including pesticides that have not yet even been developed. In this study, we investigated a possible type of cross-resistance in the model fruit fly Drosophila melanogaster, some stocks of which are resistant to the very potent mushroom toxin α-amanitin. Because α-amanitin is solely produced by mushrooms, and because D. melanogaster does not feed on mushrooms in nature, the fruit flies should not be resistant to this toxin. In order to understand how this mushroom toxin resistance evolved, we first examined the physiological aspects of α-amanitin resistance in three D. melanogaster stocks, which were isolated in Asia half a century ago: Ama-KTT, Ama-MI, and Ama-KLM. We showed that all three fly stocks have not lost their α-amanitin resistance over time, even though they were maintained in the absence of selective pressure over the past ~1,200 generations. When we reared these flies on sub-lethal α-amanitin concentrations in our laboratory, the females unexpectedly doubled their fecundity. This effect could have dramatic ecological consequences by enabling D. melanogaster to invade the toxic mushroom niche. As a result of this potential niche switch, other mushroom-feeding flies could be driven to extinction. However, we also noted signs of developmental retardation and a shortened life span of the flies in response to increasing α-amanitin concentrations in the food, suggesting that the flies are not yet well adapted to the toxin. We next elucidated the molecular mechanisms causing α-amanitin resistance by performing a whole genome microarray study. Our data suggest that 1) cuticular proteins block α-amanitin from entering cells, 2) phase I and phase II detoxification enzymes modify α-amanitin to prepare it for excretion, 3) intracellular lipid particles sequester α-amanitin in the cytoplasm, and 4) peptidases cleave α-amanitin. Because the most highly up-regulated genes in our microarray study were Cytochrome P450/phase I detoxification genes that are known to detoxify pesticides, we speculate that the use of pesticides was the primary cause for the observed cross-resistance to the mushroom toxin α-amanitin.

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