2010 Annual Report
Objective 2: Microarray technology and 454FLX sequencing will reveal gene expression in the larval Hessian fly during compatible and incompatible interactions with wheat. The morphology of midgut and salivary gland tissues will be examined by transmission electron microscopy. Comparative transcriptomics will identify Hessian fly genes involved in parasitism of wheat. The role of Hessian fly genes in host susceptibility or resistance will be assessed through RNAi knockdown.
Objective 3: Microsatellite markers will be used to assess heterogeneity and gene flow in Hessian fly populations. Changes in allelic variation will assist in assessing the risks to deployed resistance. Differentiation at different geographic scales will be assessed by Fst and Rst values. Estimation of effective population size (Ne) will be used to measure the strength of genetic drift in populations.
Objective 2 – Expression analysis of genes encoding secreted salivary gland proteins (SSGPs) has been conducted in collaboration with USDA-ARS scientists at Manhattan, KS using a Hessian fly Affymetrix array. Results from the array analysis have been validated by quantitative real-time PCR (qPCR) for selected SSGPs. Initial analysis involved three laboratory lines of Hessian fly (vH9, vH13, and white eye) and two populations (Israel and Alabama). We are continuing microinjection of double stranded RNA (dsRNA) into embryos for RNA interference (RNAi) in 1st-instar larvae. A tobacco protoplast system is being used to discover the target sites of SSGPs in plant cells and putative function of SSGPs. Gene expression in different compatible/incompatible interactions is being documented by 454 sequencing. A new research direction involving a bioassay for Hessian fly larvae has been developed that was not available when the objectives and milestones for the Project Plan were developed.
Objective 3 – The microsatellites that have been chosen for use in population studies have been characterized and mapped to the polytene chromosomes. These microsatellites have been used to analyze population structure globally by assessing gene flow between populations. In addition, a microgeographic study with populations (third year collections) in Alabama is currently underway to gain better understanding of gene flow within small geographic distances. Currently, we have a few samples from Alabama from this year, but are awaiting more samples in order to finish our analysis.
Shukle, R.H., Subramanyam, S., Williams, C.E. 2010. Ultrastructural Changes in the Midgut of Hessian Fly Larvae Feeding on Resistant Wheat. Journal of Insect Physiology. 56:754-760.
Kosma, K., Nemacheck, J.A., Jenks, M., Williams, C.E. 2010. Changes in the Properties of Wheat Leaf Cuticle During Interactions with Hessian Fly. Plant Journal. 63:31-43.
Zhang, S., Shukle, R. H., Mittapalli, O., Zhu, Y. C., Reese, J. C., Wang, H., Hua, B.Z., Chen, M.S. 2010. The gut transcriptome of a gall midge, Mayetiola destructor. Journal of Insect Physiology. 56:1198-1206.
Liu, X, Williams C. E., Nemacheck, J. A., Wang, H., Subramanyam, S., Zheng, C., Chen, M.-S. 2010. Reactive Oxygen Species are Involved in Plant Defense Against a Gall Midge. Plant Physiology. 152:985-999.