KSEF RDE: Development and deployment of a non-toxic endophyte in tall fescue for forage

Tall fescue is the most widely planted forage grass in the United States, and a key pasture component in Kentucky. Its exceptional stress tolerance, stand longevity and productivity are largely attributable to the seedtransmitted, mutualistic symbiont (endophyte), Neotyphodium coenophialum. However, common strains of N. coenophialum produce ergot alkaloids that cause toxic symptoms in grazing livestock. The resulting fescue toxicosis causes close to $1 billion annual loss to U.S. livestock production. These anti-mammalian effects also impact ecological succession, posing environmental concerns, for example in reclaimed strip mines. Some nontoxic endophytes have recently been deployed in tall fescue cultivars, but may be less beneficial or less compatible with the plant. The ideal strain of N. coenophialum for tall fescue cultivars would confer the key benefits to the grass but would not produce ergot alkaloids. Toward this end, we identified the gene, dmaW, for the first step in ergot alkaloid biosynthesis, and determined that N. coenophialum has two active copies. We eliminated one dmaW copy by replacing it with a foreign marker gene. Furthermore, to obviate public and regulatory concerns, we developed and successfully tested a method to eliminate the foreign marker gene. We propose to apply these techniques to eliminate the second dmaW copy, and generate a non-ergot alkaloid endophyte with no foreign genes in the genome. Then, we will establish this modified endophyte in elite tall fescue breeding lines to develop new cultivars, and raise seeds of these cultivars to test their agronomic traits and suitability for pastured livestock. The proposal involves the following intellectual property of the University of Kentucky: patent 6,335,188 (issue date, 2002) for use and manipulation of dmaW, and three elite tall fescue breeding lines bred for late flowering, succulence, seedling vigor, yield potential, and disease resistance. The approach is also applicable to temperate forage grasses worldwide.