* What are the natural selective, demographic and epidemiological pressures that drive evolution of insect immune systems?
We use molecular population genetic and comparative genomic analyses to determine rates of evolution in various components of the immune system in both Drosophila and Anopheles. These patterns of evolution lend insight into the mechanistic basis of host-pathogen interactions.
* What is the genetic basis for individual variation in immune performance? Why are some individuals of a species more adept at fighting infection than others, and what evolutionary forces maintain this variation? How is the immune system interconnected with other components of host physiology?
We use quantitative genetic mapping to discover which genes carry polymorphism in natural populations that results in individual differences in immune performance (this is analogous to mapping the genes for, say, heart disease in humans). We study the degree and manner in which environmental variation and pathogen diversity impact the genetic basis for resistance and may maintain variation for resistance in populations. Immune defense is physiologically demanding, and is often coupled with disruptions of other host processes such as basal metabolism, reproduction and circadian rhythmicity. We are studying the genetic connections between diverse physiological processes, and how these linkages may affect the evolution of immunity.
* What are the genetic and mechanistic bases for virulence in pathogenic bacteria?
We study the pathology and genetics of microbial pathogens of insects to determine the mechanisms and evolution of virulence. Much of our work in this area is focused on bacteria in the genus Providencia, which are also opportunistic infectors of humans.
We choose to perform some of this work in Drosophila because it is an experimentally tractable model system and because a wealth of experimental resources are available. We expect that the conclusions we can draw from Drosophila will be generalizable over insects. Our work in Anopheles mosquitoes is conducted in the context of transmission of human malaria.Malaria is a devastating disease, particularly in Africa, with hundreds of millions of clinical cases and nearly a million deaths reported each year. Better understanding of malaria interactions with the mosquito host may lead to more effective control of disease transmission. We study bacterial virulence to insects both as a model for clinical infections of humans and with an eye toward novel biological control strategies of medically and agriculturally relevant insect pests.
The evolutionary pressures on immunity genes can be determined through population genetic and molecular evolutionary analyses. Patterns of DNA sequence diversity within species can be used to identify genes that are subject to Darwinian natural selection. Comparing gene sequences of flies collected from different populations can help determine whether the immune system is adapted to local pathogens or environmental conditions. Genome comparisons across different species of Drosophila , mosquitoes, and other insects can be compared to identify genes that consistently experience strong directional selection and to establish patterns of genome evolution such as specific expansions of gene families.
We and other researchers in the field have found that pathogen pattern recognition proteins and antimicrobial peptides largely evolve under purifying selection within species, but that these genes are subject to rapid genomic duplication and deletion between species. In surprising contrast, intracellular signal transduction proteins, which are highly conserved in copy number across species, show very strong evidence of adaptive evolution at the amino acid sequence level (see Lazzaro 2008 for a review emphasizing Drosophila). These observations have led to the development of a model positing that rapid bursts of adaptive genomic evolution aid in defense against novel pathogens that may be encountered during ecological niche shifts, and that adaptation at the amino acid sequence level may be driven by co-evolution between insects and pathogens that have the capacity to interfere with immune signaling processes.
Over the shorter evolutionary term, adaptation of immune response genes to geographically local pathogens or evnironmental conditions may be revealed through genetic substructure in populations. We are currently testing this hypothesis in D. melanogaster through analysis of globally sampled flies. We are also following the evolutionary dynamics of candidate defense genes in geographically and ecologically distinct subpopulations of the primary sub-Saharan African malaria vector, A. gambiae. We anticipate that better understanding of the evolutionary genetics of mosquito defenses against malaria parasites may lead to improved disease control and treatment initiatives.
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A primary emphasis in the lab is determining the genetic basis for variation in immune quality among wild insects. We can most powerfully conduct these studies using genetically pure lines of D. melanogaster that are derived from flies captured in nature, which thus represent as a snapshot of variability in the wild population at the time of collection. The quality of the immune response can be measured in each genetic line in a variety of ways, including time to death, systemic bacterial load, and expression level of immune response genes after infection. We can then map these measurements of immune strength to specific genes in the immune system, we can determine whether same genetic polymorphisms determine resistance to distinct microbial pathogens, we can test the degree to which the quality of immune response is determined by epistatic interactions among genes, and we can evaluate the consistency of immune quality across varying environmental conditions.
Wild mosquito populations also harbor substantial genetic variation for resistance to the establishment of malaria parasites. We are collaborating with Ken Vernick and his lab at Institut Pasteur to conduct analogous mapping in west African populations of A. gambiae to identify resistance alleles.
Our studies of the quantitative genetic basis for natural variation in D. melanogaster resistance to bacterial infection have shown that, concordant with the molecular evolutionary data, the majority of the phenotypically important genetic variation maps to signal transduction and pathogen recognition genes. Molecular variation in antimicrobial peptide genes makes little contribution to phenotypic variation in immune performance, presumably because these are semi-redundant downstream endpoints in the immune system (Lazzaro et al. 2004). We have shown, however, only weak genetic correlations in resistance to infection by different bacteria (Lazzaro et al. 2006). My lab is currently in the midst of an ambitious experiment to determine the effective heritability of antibacterial immunocompetence by testing the relative contributions of additive and epistatic genetic variance in resistance. This work will have relevance in applied entomological settings and to a variety of theoretical models in evolutionary biology.
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| D. melanogaster can be manually infected by piercing the cuticle with a bacteria-coated pin. | Systemic bacterial load can be estimated by counting the number of viable bacteria sustained by infected flies. |
 | Wild fruit flies show show substantial genetic variation for the ability to resist bacterial infection. This plot shows the average number of bacteria recovered from 101 different lines of D. melanogaster that were given identical infections one day earlier. |
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Ultimately, it is essential to understand the ecology of infectious disease in insect populations in order to draw inference about evolutionary pressures on insect immune systems. My lab has therefore been engaged in a multi-year project to characterize the diversity of bacteria that infect wild D. melanogaster. The data we have collected so far reveal a surprisingly high taxonomic diversity of bacteria infecting D. melanogaster in nature, which may limit the potential for specific co-evolution between D. melanogaster and its bacterial pathogens. Most bacterial infections appear not to be caused by obligate pathogens, but result from opportunistic infections. This may place adaptive constraint on these microbes, if the evolution of enhanced infectivity compromises life in non-pathogenic settings. Nevertheless, many of these bacteria have interesting and effective virulence mechanisms that allow them to exploit insects as hosts. We now have many isolates in culture of many bacteria originally collected as infections of D. melanogaster and have been using these in studies both of D. melanogaster defense and the evolutionary and functional genetics of microbial virulence.
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| Hemolymph is extracted from D. melanogaster to test for the presence of infectious bacteria. | Bacteria isolated from a wild fly. |
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