Development of Genetic Markers and Genetic Analysis of Brugia malayi Susceptibility in the Mosquito Aedes aegypti

The mosquito, Aedes aegypti, is responsible for transmission of dengue and yellow fever and serves as an excellent model vector for transmission of malaria and lymphatic filariasis. Understanding the genetics of vector competence could allow for the development of vector-based control strategies that would alleviate mortality and morbidity worldwide. In the first study we developed genetic markers based upon chromosomal regions conserved throughout the Culicidae. We designed 35 PCR primer pairs based upon orthologous exons in Aedes aegypti and Drosophila melanogaster or Anopheles gambiae. Twenty-three of the primers yielded a single PCR product in at least one Dipteran in addition to A. aegypti when screened with genomic DNA from seven Dipterans including five mosquito species. Eight of the primers amplified a single PCR product in only A. aegypti while four primer pairs gave no PCR product in any species. The 23 successful CATS primer pairs gave broad genome coverage in A. aegypti and demonstrate an efficient strategy for developing comparative anchor marker loci for any species of Culicidae. In the second study we isolated and identified microsatellite sequences from multiple genomic libraries for A. aegypti. We identified seven single-copy simple microsatellites from three plasmid libraries enriched for (GA)n, (AAT)n and (TAGA)n motifs from A. aegypti. In addition, we identified 5 single-copy microsatellites from an A. aegypti cosmid library. Seven of these microsatellite markers were polymorphic and were genetically mapped in a segregating F1 intercross population. These markers greatly increase the number of microsatellite markers available for A. aegypti and provide additional tools for studying genetic variability of mosquito populations. Additionally, most A. aegypti microsatellites are closely associated with repetitive elements, accounting for our and other researchers limited success in developing an extensive panel of microsatellite marker loci. In the third aim of this study we employed single nucleotide polymorphism (SNP) markers to analyze an advanced intercross line for quantitative trait loci associated with Brugia malayi susceptibility. Using single marker analysis of variance and linear regression we identified a single QTL at LF178 associated with susceptibility when evaluated as a quantitative phenotype and we identified one QTL for susceptibility when the phenotype was treated as binary trait.