Evaluation of Diverse Endophyte Strains for Effects on Tall Fescue Drought Tolerance Mioko Tamura and C.P. West Crop, Soil & Environmental Sciences, Univ. of Arkansas, Fayetteville, AR Results Discussion Greenhouse Trial In the greenhouse trial (Table 1), comparisons can only be made within a run. In Run 1, strain 22 resulted in the highest survival rate, but not different from E-. High survival in strain 22 was associated with high water content, suggesting a slow rate of desiccation. Strain 22 also had the highest regrowth rate. Survival of E+ was unexpectedly low. In Run 2 (Table 1), no novel strains caused survival greater than E- or as well as E+, except strain 25. All novel strains showed regrowth numerically intermediate between E- and E+. Field Trial In the field trial, mild drought stress occurred in June, which was drier and warmer than normal (Table 2). July and August sampling periods were wetter than normal. Therefore, drought stress did not develop enough to rigorously test the novel strains for their effects on host growth and survival. Relative densities (Table 3) showed that plant density increased over the summer (values >1) due to the favorable growing conditions. Endophyte strains varied in hexose and sucrose (Table 4) concentrations. In the July sampling (reflecting June growing conditions), strain 22 tended to have the lowest hexose and highest sucrose levels. Correlations between carbohydrate levels and drought survival were prevented by growing conditions that were too favorable for inducing plant mortality. In conclusion, strain 22 showed some potential for superior drought tolerance enhancement of its host in the greenhouse trial; however, this could not be corroborated in the field because of unusually wet conditions. The field screening trial will be repeated in Introduction Tall fescue, an important cool-season forage grass, is susceptible to summer drought stress in southeastern U.S., thereby reducing persistence. The plant naturally possesses a mutualistic fungal endophyte (Neotyph- odium coenophialum). The endophyte typically produces ergot alkaloids, which are toxic to animals; however, it is also shown to enhance host drought tolerance. Recent research has identified nontoxic, ‘novel’ strains of endophyte that lack ergot alkaloids. Such strains have potential use in tall fescue cultivars for protecting the plant from drought stress. We need to test such strains in an adapted tall fescue population to identify which ones benefit host drought tolerance. We hypothesize that beneficial strains would pro- mote greater soluble carbohydrate levels in the surviving tiller base as a mechanism enhancing viability of tiller meristems over endophyte-free plants. Objectives  Screen nontoxic endophyte strains for enhanced host drought tolerance in greenhouse and field trials.  Investigate the association between soluble carbohydrate accumulation in tiller bases and persistence under drought in the field. Methods  A collection of novel endophyte strains (numbered 15-32), which lack toxic alkaloid production were inoculated in cv. Kentucky-31. Greenhouse Trial  10 nontoxic strains – separated into two runs of 5 strains each 2 controls: E- = endophyte-free; E+ = infected with toxic endophyte.  PVC tubes (15 × 76 cm) filled with pasteurized sand, transplanted with 3 plants/pot. 2 pots/strain/block; total of 5 blocks.  Drought was imposed by withholding water for at least 4 wks.  Blocks were sequentially rewatered to 80% capacity over 3-4 wks to terminate drought over a range of severities (Fig. 1).  Before rewatering, plants were clipped to 2-cm stubble, which was sampled for tiller-base water content. Regrowth measured after 1 wk. Field Trial (Fig. 2)  16 nontoxic strains and 2 controls, E- and E+; 4 replicates.  Biomass: hand-clipped to 8-cm stubble in nonirrigated zone of plots.  Tiller density: visually scored as 1-10 along a 40-cm length of row, with 10 = fully occupied row.  Total hexose (glucose + fructose) and sucrose contents in 8-cm tiller bases. Extracted ground samples with 80% ethanol (Zhao et al. 2008). For additional information, contact: Mioko Tamura, or Dr. Chuck West, Acknowledgement This research was supported by USDA-ARS Specific Cooperative Agreement through the Dale Bumpers Small Farm Research Center, Booneville AR. Reference Zhao, D., C.T. MacKown, P.J. Starks, and B.K. Kindiger Interspecies variation of forage nutritive value and nonstructural carbohydrates in perennial cool-season grasses. Agron. J. 100: Fig. 1. PVC pots in the greenhouse. Table 1. Survival rate, water content, and regrowth length of tillers for each strain in the greenhouse experiment; means of 2 replications and 5 blocks. Run 1Run 2 EndophyteWater EndophyteWater strain Survival content Regrowth strainSurvival content Regrowth %g g¯¹ DWmm %g g¯¹ DWmm E -29ab†0.27ab19bE -30b0.19ab47b E +20b0.25ab7bE +57a0.25a79a 1719b0.26ab8b 23 b0.16b61ab 1932ab0.27ab25ab2433b0.20ab78a 20 b0.23b15b2540ab0.21ab75ab 2132ab0.28ab22b2634b0.23a62ab 2247a0.34a37a2730b0.20ab67ab LSD LSD † Means followed by the same letter are not significantly different by Tukey (α=0.05). Table 4. Hexose (glucose + fructose) and sucrose concentrations in the field tiller bases for selected endophyte strains on the first day of each month indicated; means of 4 replications. EndophyteHexoses Sucrose strainJulyAugustSeptember JulyAugustSeptember –––––––––––––––––––––––––––––– mg g ¯¹ –––––––––––––––––––––––––––– E-4.4b†1.2ab b15.4a14.6 E+5.3a1.7a4.29.7c14.5ab c1.3ab b13.1b c0.8b a14.8ab b1.2ab——11.5ab14.7ab—— 253.3c1.3ab——11.5ab14.4ab—— LSD † Means followed by the same letter are not significant by Tukey (α=0.05). Fig. 2. Field trial showing 2 reps. Table 2. Monthly rainfall and mean temperature of June - August 2009 Month Mean temperature Departure from normal † Monthly rainfall Departure from normal —————⁰C ——————————mm ————— June July August cm Nonirrigated sampling zone N. coenophialum; E. Bernard Results Table 3. Cumulative biomass yield (1 June – 1 September) and actual and relative tiller density of row segments; means of 4 replications. EndophyteBiomassTiller density†Relative density† strain yield on 1 SeptemberSeptember/June kg ha -1 E-3500‡9.5ab1.45a § E ab1.06bc ab1.21b b1.23ab ab1.18bc b1.03bc ab0.97c ab1.18bc b1.15bc a1.12bc ab1.18bc ab1.09bc ab1.14bc ab1.10bc ab1.04bc ab1.13bc LSD † Density was scored as in 0.5 unit intervals; 10 defined as full cover. Relative density was calculated as ratio of September/June tiller density. ‡ There was no significant difference among strains for biomass yield by Tukey (P>0.05). § Means followed by the same letter are not significantly different by Tukey (α=0.05). 15-cm †Departure from normal, average value over 29 yr ( ).