2012 Annual Report 1a.Objectives (from AD-416): The goal of this project is to isolate and characterize strains of fungi (Ambrosiella spp.) associated with ambrosia beetles. Objectives include. 1)Identify and assess genetic diversity among strains isolated from Xylosandrus crassiusculus and X. germanus and. 2)Determine impact of fungal diversity on beetle development within various hosts and substrates. 1b.Approach (from AD-416): Ambrosia beetles (Xylosandrus spp. and others) are pests of a wide array of woody plants used in the nursery industry. They depend, for their survival, on their own cultivation of a fungus (Ambrosiella spp. and others). Fungal strains isolated from Xylosandrus spp. will be identified and assessed through sequencing of 18S, 28S and ITS regions of ribosomal DNA from X. crassiusculus and X. germanus. Brood development and symbiont growth will be evaluated for each fungal genotype on representative hosts or diet substrates. Fecundity of beetles within galleries will be evaluated. 3.Progress Report: The 18S, 28S, and ITS region of the ribosomal DNA from representative isolates have been sequenced to identify fungal species and data are currently being analyzed. Multilocus gene screening is also currently underway to assess intraspecific diversity. We are evaluating the beta–tubulin, IGS, translation elongation factor1, and mtSSU genes as possible markers for population genetics.