Ten wether lambs were used to determine the effects of ergovaline consumption from endophyte infected tall fescue, on nutrient utilization and metabolism. Lambs were fed a diet of 23% endophyte free tall fescue seed (E-) and 77% concentrate from d -14 to -1 (adaptation phase). On d 0, five lambs were switched to an endophyte infected seed diet (E+) where they remained through d 14. Nutrient digestibilities tended to increase from adaptation through the acute (d 1 to 4) to subacute (d 10 to 14) phases when E- was fed. E+ digestibilities were highest (P < 0.05) in the acute phase. Lambs fed E+ had higher rectal temperatures in the acute (P < 0.01) and subacute phases (P < 0.05). Fecal recovery of ergovaline increased as day of collection increased in the acute and subacute phases. Lysergic acid fecal recovery increased with day of collection in the acute phase, but no effect was found in the subacute phase. Serum enzyme analyses did not indicate tissue damage from alkaloid consumption. These results demonstrate lambs try to adapt to endophyte infected fescue seed consumption through increased nutrient digestibilities and increased ergovaline and lysergic acid excretion.

The present investigation was designed to investigate the use of blessed thistle (Cnicus benedictus) in amelioration of fescue toxicosis in beef cattle. Twenty-seven crossbred Angus cows were blocked by age and weight into groups of three. Each group (n=9) was then randomly assigned to one of three dietary treatments: endophyte-free (EF), endophyte-infected (EI), or endophyte-infected with addition of cut and stemmed blessed thistle (EIBT). Each group was also randomly assigned to one of three 29-day replicates. Parameters measured during this trial included: urinary ergot alkaloid concentration (ng ergot alkaloids/mg creatinine), body weight (kg), rectal temperature (°C), serum progesterone (ng/ml), and serum prolactin (ng/ml). No effect of treatment (P>0.05) was detected in any of these parameters. A treatment x time interaction (P<0.0001) was observed for progesterone concentrations, likely a result of estrous cyclicity of cows. Results of this study suggest that we were unsuccessful in inducing fescue toxicity in test subjects assigned to EI and EIBT treatments. Minimal dietary ergovaline (1.72 μg kg-1) and cool ambient temperatures (-6.7°C to 23.3°C) likely contributed to this inability. Data from this research do suggest, however, that C. benedictus can be successfully incorporated into cattle diets. Further research will need to be performed to fully elucidate what, if any, benefits blessed thistle can have in the treatment of fescue toxicity.

A wild-type endophyte (Neotyphodium coenophialum [(Morgan-Jones & Gams) Glen Bacon & Hanlin]) that infects tall fescue [Lolium arundinaceum (Schreb.) Darbysh. = Schedonorous arundinaceum (Schreb.) Dumort.] imparts tolerances to moisture, heat, and grazing stresses, but also produces ergot alkaloids that adversely affect performance and physiology of cattle. Novel endophytes, developed by AgResearch Ltd. NZ, can sustain fescue persistence and productivity, but do not produce toxic ergot alkaloids. University of Kentucky Plant Breeder, T. D. Phillips Ph.D, developed a tall fescue experimental population (KYFA9301) for the upper transition zone. A 2-yr grazing experiment was conducted with steers to evaluate steer performance and physiology, and forage quality and productivity of KYFA9301 infected with AR584 novel endophyte (AR584) compared with KY31 wild-type endophyte (KY31), endophyte-free KYFA9301 (EF9301) and AR542-‘Jesup’ (MaxQ). Fescue-endophyte combinations were assigned to 1.0-ha pastures in a randomized complete block design with three replications. Pastures were grazed with variable stocking (four testers) from 6 May to 23 July 2008 (76 d), and 2 April to 25 June 2009 (84 d). Shrunk bodyweights were taken at initiation and termination of grazing each year. Average daily gains among MaxQ, AR584, and EF9301 were similar and were greater (P < 0.10) than KY31. Rectal and skin temperatures were collected three times each year at approximately days 28, 56, and study completion, along with blood collection for serum prolactin assay. Rectal and skin temperatures among AR584, MaxQ, and EF9301 were similar and were lower (P < 0.10) than KY31. Serum prolactin concentrations were similar among the three nontoxic varieties and higher (P < 0.10) than KY31. Forage collections were taken at 2 week intervals throughout the study each year and nutritive quality analysis were conducted through wet chemistry to determine forage acid detergent fiber (ADF), neutral detergent fiber (NDF), crude protein (CP), and In-Vitro Dry Matter Digestibility (IVDMD). Nutritive analyses indicated no differences between fescue-endophyte combinations with the exception of EF9301 having higher ADF concentrations (P = 0.031) than KY31 during the dry year of 2008. Results indicated steer performance and physiological responses for KYFA9301, with and without AR584 were enhanced compared to KY31 and similar to those for MaxQ.

Plants naturally produce primary and secondary metabolites. Primary metabolites are directly involved with plant growth and metabolic function. Plant secondary metabolites (PSM) were once thought of as metabolic waste products, and more recently viewed as toxins to herbivores. However, ongoing research shows that PSM are beneficial to herbivores at low doses, and PSM aid plants by attracting pollinators, recovering from injury, protecting from ultraviolet radiation, increasing drought tolerance, and aid in defense against pathogens, diseases, and herbivores. Plant secondary metabolites also influence soil nutrient cycling, and can increase the sustainability of agroecosystems. Endophyte-infected tall fescue (Festuca arundinacea Schreb.) , which contains ergovaline, and reed canarygrass (Phalaris arundinacea L.), containing gramine, were studied along with the legumes alfalfa (Medicago sativa L.) which contains saponins, and tannin-containing sainfoin (Onobrychis viciifolia Scop.) and birdsfoot trefoil (Lotus corniculatus L.). This dissertation researches (i) how planting configuration (monocultures vs. two-way mixtures) influences PSM and total N in plants, (ii) how cattle grazing forages containing PSM affects soil quality, nutrient cycling, and PSM, and (iii) how cattle manure from different diets, containing different PSM, influences soil nutrient cycling.

Les associations symbiotiques entre différentes espèces de champignon du genre Epichloë et des plantes fourragères sont à l’origine de la production de mycotoxines responsables, dans certains pays, de toxicoses du bétail et de pertes économiques. Le développement de d’E. coenophiala dans la fétuque, qui est associé à la production d’ergovaline, est responsable de « fescue foot disease » alors que la présence d’E. festucae var. lolii dans le ray-grass anglais, qui est accompagnée d’une production de lolitrem B, est responsable du « ryegrass staggers » et de la production d’ergovaline, dont les effets toxiques associés à la présence de lolitrème B sont mal documentés. Mon travail de thèse a consisté à explorer les effets de la distribution de différents fourrages de fétuque et ray-grass endophytés toxinogènes produits dans des conditions agricoles françaises, sur la santé et la production animale et les mécanismes d’action de l’ergovaline et du lolitrème B. / The symbiotic associations between different varieties of fungi of the Epichloë kind and forages are responsible for the production of mycotoxins which are responsible of toxicoses in livestock and economic losses in some countries. The development of E. coenophiala in tall fescue, which is associated with the production of ergovaline, is responsible for « fescue foot disease » while the presence of E. festucae var. lolii in ryegrass, which is accompanied by the production of lolitrem B, is responsible for the « ryegrass staggers ». and the production of ergovaline, whose the toxic effects associated in the presence of lolitrem B are poorly documented. My thesis was to explore the effects of the distribution of different toxic forages of endophyte-infected tall fescue or ryegrass produced in French agricultural conditions on health and animal production and the mechanisms of action of ergovaline and lolitrem B.

Les associations symbiotiques Epichloë- graminées fourragères les plus étudiées concernent le ray grass anglais (Lolium perenne) avec Epichloë festucae var. lolii et la fétuque élevée (L. arundinaceum) avec E. coenophiala. Ces champignons peuvent synthétiser des alcaloïdes toxiques pour le bétail tels que l’ergovaline et le lolitrème B. Mon travail de thèse a consisté à explorer les facteurs en lien avec la production de mycotoxines dans les graminées. Pour cela différentes études ont été conduites sur la fétuque et le ray grass afin de révéler i) la présence de souches endophytées toxinogènes dans le Sud Ouest de la France et ii) l’influence des conditions climatiques françaises sur la synthèse d’ergovaline et de lolitrème B dans des variétés fourragères connues pour être responsables de cas de toxicité sur d’autres continents. / The most studied symbiotic associations Epichloë- cool seasons grasses concern perennial ryegrass (Lolium perenne) with Epichloë festucae var. lolii and tall fescue (L. arundinaceum) with E. coenophiala. These fungi can synthesize alkaloids that could be toxic to livestock such as ergovaline and lolitrem B. My thesis work was to explore the factors related to the production of mycotoxins in grasses. Different studies have been conducted on tall fescue and perennial ryegrass to reveal i) the presence of endophyte-infected toxigenic strains in the South West of France and ii) the influence of environmental conditions of Saint-Affrique (Aveyron) on the synthesis of ergovaline and lolitrem B in forage varieties known to be responsible for toxicity on other continents.

Ergovaline is produced by the endophyte Neotyphodium coenophialum (Morgan-Jones and Gams) in tall fescue (Schedonorus arundinacea (Schreb.) Dumort. = Festuca arundinacea Schreb.) and is blamed for a multitude of costly livestock disorders. Testing of pastures is common in both research and on farm situations. Since ergovaline is known to be unstable and affected by many variables, the objective of this study was to determine the effect of sample handling and storage on the stability of this compound. Homogeneous milled tall fescue sub-samples were analyzed for ergovaline concentration using HPLC after a range of sample handling procedures or storage. Ergovaline was unstable in milled material after 24 hours in storage, regardless of temperature. The decrease in ergovaline after 24 hours ranged from 17 to 60%. These results show that tall fescue sample handling and storage have a significant effect on ergovaline concentrations. In conclusion, accurate laboratory analysis of ergovaline content may require that samples be transported immediately to the laboratory on ice for immediate analysis. Most laboratories are not equipped for same day analysis, therefore researchers and producers should acknowledge that laboratory ergovaline results may be lower than the actual content in the field.

Perennial ryegrass (PRG, Lolium perenne) is a hardy cool-season grass that is infected with the endophytic fungus Neotyphodium lolii, which enables the plant to be insect repellant and drought resistant, lowering the use of insecticides and fertilizers. However, this fungus produces the compound lolitrem B (LB, m/z 686.4) which causes the tremorgenic neurotoxicity syndrome 'ryegrass staggers' in livestock consuming forage which contains <2000 ppb LB. Ergovaline (EV, m/z 534) is a vasoconstrictor normally associated with tall fescue (Festuca arudinacea), but has also been found in endophyte-infected PRG. Past research has shown a strong linear correlation between levels of LB and EV in PRG. The purpose of this study was to examine the linear relationship between EV and LB in feces and determine common metabolites. To accomplish this, four groups of steers (n=6/group) consumed endophyte- infected PRG over 70 days consumed the following averages of LB and EV: group I 2254ppb LB/633 ppb EV; group II 1554ppb LB/ 373ppb EV, group III 1011ppb LB/259ppb EV, and group IV 246ppb LB/<100ppb EV. Group I in week 4 was inadvertently given a washout period at which time the steers consumed the amount of LB and EV given to group IV (control). Both feed and feces samples were extracted using difference solid phase extraction methods and quantified by HPLC-fluorescence for LB and EV. Concentrations of EV and LB obtained through HPLC-fluorescence in both PRG and feces showed a linear relationship. Additional screening for metabolites was conducted LC-MS/MS and showed possible oxidation and reduction metabolites for both toxins. / Graduation date: 2013