University of Cincinnati

UNIVERSITY OF CINCINNATI Date: 15-Dec-2009 I, Aaron Greene , hereby submit this original work as part of the requirements for the degree of: Master of Science in Biological Sciences It is entitled: Heritable Behavioral Resistance to Natural and Novel Ectoparasites in Drosophila melanogaster Student Signature: Aaron Greene This work and its defense approved by: Committee Chair: Michal Polak, PhD Michal Polak, PhD Iain Cartwright, PhD Iain Cartwright, PhD Stephanie Rollmann, PhD Stephanie Rollmann, PhD 5/28/2010 863 Heritable Behavioral Resistance to Natural and Novel Ectoparasites in Drosophila melanogaster A thesis submitted to the Graduate School of the University of Cincinnati In partial fulfillment of the requirements for the degree of MASTER OF SCIENCE In the Department of Biological Sciences of the College of Arts and Sciences By AARON VILAS GREENE B.S. Biological Sciences Mansfield University, November 2001 Committee Chair: Dr. Michal Polak 34 Abstract Ectoparasites affect many different organisms, are naturally abundant, and have been shown to decrease fitness and drive host evolution. However, few studies have estimated heritable variation in resistance to ectoparasitism, and none have tested the effects of those defenses on other ectoparasites. The threat of parasitism can be so costly that potential hosts develop several lines of defense to protect themselves from parasitism. The ability to resist novel enemies, such as ectoparasites, has been shown to carry fitness costs. However, possessing traits that effectively resist or limit the threat of multiple enemies potentially could benefit the host by minimizing the costs associated with resistance. The current paper details the presence of behavioral resistance potentially utilized by Drosophila melanogaster to defend against ectoparasitic mites . This study reports the results of artificial selection for increased resistance in Drosophila melanogaster to the ectoparasitic mite, Macrocheles subbadius , which is a known enemy of some Drosophila species. My work also investigates the effectiveness of the selected resistance against the natural enemy Gamasodes queenslandicus . Selection was applied to the pre-attachment phase, thereby targeting behavioral defensive traits. Realized heritability ( h2) of resistance against Macrocheles is estimated at 0.06 (SE 0.015). Results also demonstrate that flies selected for increased resistance to M. subbadius also have improved resistance to G. queenslandicus. This study demonstrates the evolutionary potential of generalized behavioral defenses against ectoparasite attack in D. melanogaster . i Acknowledgements I would like to first thank my advisor, Dr. Michal Polak for his guidance and support. I would also like to thank my committee members Dr’s Stephanie Rollmann and Iain Cartwright who both offered their invaluable expertise and advice. The Department of Biological Sciences at the University of Cincinnati provided financial support for this project. I would also like to thank my lab mates, Brooke Hamilton, Karl Grieshop and Dr. Arash Rashed for their support and insightful talks throughout the course of my research. Lastly, I would like to thank my family and friends who gave me support and encouragement throughout this process. ii Table of Contents Abstract……………………………………………….……………………………………….. i Acknowledgements …………………………….….………………………………………… ii Introduction …………………………………………………………………………………… 1 Thesis Overview………………………………………………………………………. 4 Methods ……………………………………………………………………………………… 5 Collecting Organisms and culturing………………………………………………..... 5 Resistance Behaviors…………………………………………………………………. 6 Selection Lines………………………………………………………………………… 7 Response to Selection and Heritability…………………………………………….... 9 Resistance against Gamasodes ……………………………………………………... 10 Fly Mortality within Chambers………………………………………………………... 10 Mechanisms of Defense…………………………………………………………….... 11 Statistical Analyses……………………………………………………………………. 12 Results …………………………………………..…………………………………………….. 14 Resistance Behaviors………………………………………………………………… 14 Resistance to Selection and Heritability............................................................... 16 Fly Mortality within Chambers……………………………………………………….. 17 Mechanisms of Defense……………………………………………………………... 18 Discussion …………………………………………………………………………………….. 18 Resistance Behaviors…………………………………………………………………. 19 Response to Selection and Heritability……………………………………………… 21 Mechanisms of Defense………………………………………………………………. 22 Resistance against Gamasodes……………………………………………………... 23 Conclusions …………………………………………………………………………………... 26 References ……………………………………………………………………………………... 28 iv List of Tables and Figures Figures: Figure 1: Infestation Chamber…………………………………………………………… 32 Figure 2: Flick frequency…………………………………………………………………. 33 Figure 3: Decamp frequency…………………………………………………………….. 33 Figure 4: Prying Frequency……………………………………………………………… 34 Figure 5: Rolling Frequency……………………………………………………………... 34 Figure 6: Divergence in Resistance through 13 Generations………………………... 35 Figure 7: Contrast in Resistance to M. subbadius ……………………………………... 36 Figure 8: Contrast in Resistance to G. queenslandicus ……………………………….. 37 Figure 9: Estimated Heritability of Resistance………………………………………….. 38 Tables: Table 1: Incidences of Resistance Behaviors…………………………………………. 39 Table 2: Mortality Rates within No Mite Chambers……………………………………. 40 Table 3: Resistance Assays to M. subbadius ………………………………………….. 41 Table 4: Resistance Assays to G. queenslandicus……………………………………. 43 v INTRODUCTION : Parasites are ubiquitous in the environment, and affect many organisms at some stage of their life (Price 1980). Parasites are defined as organisms that live on (ectoparasites) or in (endoparasites) other organisms and that exploit their hosts as a resource for their own nutrition, longevity, and reproduction (Sheldon and Verhulst 1996). During their interaction with hosts, parasites can decrease host fitness (Price 1980; Ewald 1994) and through the selection they impose, can drive rapid host evolution and alter host population dynamics and host population genetic structure (e.g. Burdon 1980; Dwyer et al. 1980; May and Anderson 1983). Predicting host evolutionary response to the effects of parasite – imposed selection on the evolution of a host population requires an understanding of the heritable genetic variation in host defensive traits (Endler 1986; Polak 2003). One approach to examining the heritable nature of parasite resistance in insects is the use of artificial selection experiments. The central idea behind using artificial selection in this context is that a significant response to artificial selection on a particular trait demonstrates the presence of additive genetic variation for that trait in the original (base) population (Falconer and Mackay 1996). Several examples of artificial selection for increased resistance against parasites exist in the literature. The mosquito, Anopheles gambiae , has been used to show significant response to selection for resistance against the malaria parasite, Plasmodium (Collins et al. 1986). Kraaijeveld & Godfray (1997) successfully selected for resistance against two species of parasitoid wasps in Drosophila melanogaster . More recently, D. nigrospiracula was successfully selected for increased resistance to the ectoparasitic mite, Macrocheles subbadius 1 (Polak 2003). Thus, the evidence suggests that there may often be genetic variation for resistance in natural populations, but little is known about the heritable genetic basis of resistance against ectoparasitism. Most of our understanding of parasite resistance comes from studying endoparasites (e.g. parasitoids). However, ectoparasites make an ideal model for use in this field for several reasons. First, ectoparasites are abundant in natural communities (Marshall 1981), and they have been shown to damage a variety of host fitness traits (Forbes and Baker 1991; Polak 1996). Ectoparasites also are known vectors of many parasitic diseases of animals, including humans, and so can have significant economic, veterinary and medical consequences (Lehmann 1993; Marshall 1981). Perhaps the most advantageous practical aspect of utilizing ectoparasites in ecological and evolutionary studies is the benefit of visual confirmation of parasitism. In systems utilizing other forms of parasites such as parasitoids and entomophagic nematodes (i.e., endoparasites), infection determination often requires the need to sacrifice the host. Two common behavioral strategies animals use to prevent infestation and minimize the cost of ectoparasite infestation are active avoidance and removal of ectoparasites (Hart 1994). In bats, for example, studies have shown that increased grooming behavior occurs in species affected by higher densities of ectoparasites (ter Hofstede and Fenton 2005). There have been very few studies to determine the resistance behavior of an invertebrate host to a potential ectoparasite. However, the honeybee, Apis mellifera, has been observed to express self-grooming and allogrooming behavior (Peng et al. 1987) in populations affected by the parasitic mite, 2 Varroa destructor . In addition, worker bees have been shown to identify drone brood cells infected with the ectoparasites, expose the cell, and destroy the mites (Martin et al. 2001), or remove the infected larvae from the hive (Boecking and Spivak 1999). The general aim of my project is to evaluate the evolutionary potential of resistance to ectoparasitic mites in Drosophila melanogaster Meigen. This study conducts an artificial selection

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