2009 Annual Report 1a.Objectives (from AD-416) To identify endophytic microbes that enhance performance of native grasses. To evaluate performance of native grasses containing novel endophyte combinations in laboratory and field environments. 1b.Approach (from AD-416) 1. Identify individual endophytes inhabiting black grama, sand dropseed, and alkali sacaton, four-wing saltbush, and other species. 2. Cross inoculate black grama and other grass species with identified endophytes to evaluate endophyte effects on early plant vigor. Plant-endopyte combinations exhibiting significantly improved vigor will be selected for longer-term evaluations in greenhouse and field trials. 3. Analyze genetic variability of black grama with respect to reproductive strategies and success, while identifying plant-specific genetic markers. 4. Establishment of field trials and common garden studies to determine success of fungal endophyte transfers. These studies will be planned and conducted in cooperation with the BLM and may include test plots on BLM sites where remediation is desired. 3.Progress Report Endophytes associated with Atriplex canescens and Atriplex griffithsii have been analyzed. To accomplish this, new PCR primers were developed with improved specificity for Atriplex endophytes. A manuscript describing the endophytes identified is in preparation. Black grama (Bouteloua eriopoda) plants from distinct seed sources have been inoculated with endophytes from A. canescens or from black grama. Plants are under cultivation in the greenhouse. DNA will be extracted from these plants in the weeks to come, and will be subjected to tag-encoded pyrosequencing in order to simultaneously assess variability of the plant genotypes and associated endophytes. Field sites for common garden studies have been prepared on two BLM sites, and are on schedule for targeted planting in August of 2009. Because the common garden sites are located on remote, undeveloped site, an irrigation system based on hydrogel-filled vertical tubes was designed and evaluated. A manuscript describing the system has been submitted and accepted.