2012 Annual Report 1a.Objectives (from AD-416): The objective of this research is to increase understanding of the molecular basis of host resistance and insect virulence in the wheat/Hessian fly system. The long-term goal is to develop strategies for durable resistance to this important insect pest. 1b.Approach (from AD-416): The molecular basis of host resistance will be approached by cloning and characterizing a wheat resistance gene for Hessian fly known as Hdic from Triticum turgidum subsp. dicoccum. This resistance gene resides in a region of wheat chromosome 1A that contains at least 14 other resistance genes for Hessian fly. Candidate genes will be identified by fine mapping and sequencing of BAC contigs. Candidate genes will be tested initially by gene silencing using RNA interference. A cosmid library from the donor of Hdic will be constructed to isolate the resistance allele. Expression vectors will be constructed for the candidate gene and tested for ability to confer resistance to Hessian fly. The molecular basis of Hessian fly virulence/avirulence will be approached by determining the functions of secreted salivary gland proteins (SSGP) of Hessian fly in virulence or avirulence to wheat. Differential gene expression of SSGPs will be tested in different biotypes of Hessian fly using a custom-designed microarray. Candidate genes for virulence or avirulence effectors will be identified. Candidate genes will be tested by gene silencing using RNA interference. 3.Progress Report: Hessian fly is mainly controlled through host plant resistance. Therefore, understanding the mechanisms of wheat defense may provide useful information to improve wheat resistance to Hessian fly. One potential target for plant defense against insects is the digestive enzymes, particularly proteases, in the insect gut. This research took advantage of the availability of the Hessian fly genome sequence and systematically analyzed the composition and expression of all digestive proteases in the Hessian fly larval gut. Major putative digestive trypsins, chymotrypsins, and cysteine proteases were identified and their expression profiles among tissues and different developmental stages were determined. This work should provide a foundation for future research for utilization of plant protease inhibitors for management of this insect pest. One of the promising new means for insect control is based on small, non-coding RNA molecules known as small interfering RNAs (siRNAs) and microRNAs (miRNAs). Small RNAs can silence insect genes, resulting in the death of the insect. This work identified and characterized a large number of miRNAs for the first time in Hessian fly. Some of the miRNAs were found to be Hessian fly-specific and their expression was affected by host plant genotypes. Further research will be directed toward testing different delivery methods so that miRNAs can be tested for ability to control this insect pest.