Analysis of the host-pathogen proteomics of Israeli Acute Paralysis Virus in the honey bee using mass spectrometry

Recent declines in honey bee populations worldwide have spurred significant research into the impact of pathogens on colony health. The role of the Israeli Acute Paralysis Virus (IAPV) in hive mortality has become of particular concern since being correlated with colony losses, although the pathogenic mechanism used by IAPV remains largely unknown. To compound this problem, few molecular studies of the honey bee immune response exist. This lack of knowledge poses a significant barrier if we are to address the impact IAPV has on honey bee health. In this thesis, two routes of research were conducted to aid our understanding of the honey bee host-pathogen relationship. First, a cell culture system using honey bee hemocytes was established and optimized to address the lack of an available honey bee in vitro system, which has significantly delayed honey bee molecular research and especially host-pathogen studies. While hemocyte cultures were simple to create, cell division was not observed and attempts to immortalize the hemocytes through oncogene transfection were hindered by the fragility of these cells. Overall, hemocyte cultures are a useful tool for some experimental applications requiring medium numbers of cells and not involving substantial manipulation. Next, to investigate changes in host protein expression during IAPV infection, mass spectrometry-based quantitative proteomics was used to compare IAPV infected and healthy pupae. This approach applied stable isotope dimethylation labeling combined with multidimensional fractionation using strong cation exhcange to identify and quantify ~800 proteins over three time points. Proteins that were significantly changing during infection were determined and clustered into four distinct expression patterns. To infer functional roles of proteins, Drosophila homologues were obtained for each protein in the data set and the corresponding GO Terms used for functional analysis. Proteins involved in processes including translation and the ubiquitin-proteasome pathway, among others, were identified and future investigation of these pathways will be useful in identifying host proteins required for infection. This analysis represents an important first step towards understanding the honey bee host response to IAPV infection through the systems-level analysis of protein expression and demonstrates the utility of mass spectrometry-based proteomics in honey bee research.Science, Faculty ofGraduat