Effect of feeding tall fescue seed infected by endophytic fungus (Acremonium coenophialum ) on reproductive performance in male rats

An experiment was conducted to determine the effect of feeding tall fescue seed infected by an endophytic fungus Acremonium coenophialum on the reproductive performance of male rats. Thirty 70-day-old rats were randomly allocated to four treatments: (I) fed 50% rat chow and 50% healthy fescue seed ad libitum (control; N=9; (II) same as I but restricted to the daily feed intake of III (N=7; (III) 50% fungal-infected fescue seed and 50% chow (N=7); and (IV) a mixture of 50% laboratory chow, 25% healthy fescue seed and 25% fungal-infected fescue seed (N=7). The rats were fed these diets for 42 days. During this time, body weights were taken weekly and feed intake was taken daily. At the end of the experiment, the rats were sacrificed and the testes and epididymides were excised and measured. Sperm parameters were assessed (concentration, percentage of motility and progressive motility) at the site of the cauda epididymis; the testes were homogenized and assessed for daily sperm production potential (DSP). Concentrations of spermatozoa (x 10(6)) among the various treatments were 645.6, 486.5, 387.4 and 457.1; motility and progressive motility measurements ranged from 48 to 50% and 2.3 to 2.4 (0-4), respectively. DSP values (per gram) and testicular weight were reduced (P<0.05) in Diet III. The data suggests that 50% fungal-infected fescue seed in the diet of rats does influence the DSP, testicular parechyma and epididymal weight in the rat.     

Evidence for chemical changes on the root surface of tall fescue in response to infection with the fungal endophyte Neotyphodium coenophialum

Endophyte-infected (E+) tall fescue (Festuca arundinacea Schreb.) plants grown in phosphorus (P) deficient soils accumulate more P in roots and shoots than noninfected isolines. In a growth chamber experiment, four tall fescue genotypes DN2, DN4, DN7, and DN11, infected with their naturally occurring strains of Neotyphodium coenophialum (Morgan-Jones & Gams) Glenn, Bacon & Hanlin, and their noninfected isolines (E-), were cultivated in nutrient solution at two P levels: 31 ppm (P+) and 0 ppm (P-) for 4 wk. The Fe³⁺ reducing activity of extracellular reductants and intact root tissues, and total phenolic concentration in roots and shoots were measured. Endophyte infection significantly increased Fe³⁺ reducing activity rate of extracellular reductants (9.6 × 10^-3 mol Fe³⁺ h-1 g-1 root FW) when compared to E- plants (3.9 × 10^-3) and Fe³⁺ reduction rate of intact root tissues (6.16 and 4.48 mol Fe³⁺ h-1 g-1 root FW, respectively for E+ and E- plants). In response to P deficiency, Fe³⁺ reduction rate of intact root tissues increased in E+ plants by 375% when compared to E- plants, whereas no significant differences were observed when P was provided. Total phenolic concentration was 20% greater in shoots of E+ plants than in E- plants. In response to P deficiency, total phenolic concentration significantly increased in roots of E+ plants by 7%, and decreased in roots of E- plants by 10%. The most active Fe³⁺ reducing zones were located along branching of secondary and tertiary roots. The Fe³⁺ reducing activity on the root surface and total phenolic concentration in roots and shoots increased dramatically in response to endophyte infection, especially under P limiting conditions.Visiting Scientist sponsored by the Fulbright Program No. 21133     

The effect of feeding tall fescue seed infected by Acremonium coenophialum on pregnancy and parturition in female rats

1. Female Sprague-Dawley rats (Rattus norvegicus) were randomly assigned to various dietary treatments containing: (1) 100% Purina rodent chow, ad libitum; (2) same as 1, but restricted to daily intake of 7; (3) 50% rodent chow (w/w) and 50% endophyte-free tall fescue (Festuca arundinacea) seed; (4) same as 3, but restricted to intake of 5; (5) 50% rodent chow, 25% endophyte-free tall fescue seed and 25% endophyte-infected (Acremonium coenophialum) tall fescue seed; (6) 50% rodent chow, 12.5% endophyte-free and 37.5% endophyte-infected tall fescue seed; and (7) 50% rodent chow and 50% endophyte infected tall fescue seed. 2. Average daily feed intakes and average daily weight gains decreased with higher levels of endophyte infected seed. 3. Frequency of litter production was affected by all endophyte-infected containing diets. 4. Conception was reduced only in dietary treatment (7). 5. Litter weights, number of pups per litter and weight per pup were proportionally reduced as higher levels of infected seed were incorporated in the ingested diets.     

Effects of consuming endophyte-infected tall fescue on growth, reproduction and lactation in mice selected for high fecundity

Effects of a diet containing endophyte-infected tall fescue seed (83% infected) were investigated using 2 lines of mice, one line selected for fecundity (L(+)) and the other a randomly selected control line (K). Treatments included a commercial stock diet (C), 50% stock plus 50% non-infected tall fescue seed (N), and 50% stock plus 50% infected tall fescue seed (I). The experiment was conducted using mice on respective treatments in 2 phases (successive generations), with 15 to 23 mated females per line and diet subgroups. Mated females of Phase 1 were assigned at random within line to experimental diets which were fed during gestation and through 21 d of lactation. Litters were standardized to 10 pups 1 d after birth. Stock diets were fed to all groups from Day 21 to weaning on Day 28. Weaned male and female pups were allotted to previous diets. Mated females in Phase 2 were managed as in Phase 1 through weaning at 28 d. Diets of males did not affect reproduction and data were pooled within female diets. Selected (L(+)) dams gave birth to more live pups than K dams (P<0.05) during both phases (+3.4 and +2.8 +/- 0.4 pups, respectively). Diet but not line affected littering rate of mated females in Phase 1 (71.3%, I; < 87.1%, C or 93.0%, N; P<0.05) and Phase 2 (82.1%, I < 93.8%, N or 97.1%, C; P<0.05). Diet had no effect on fecundity during Phase 1 but females on I diet had reduced (P<0.05) litter size by 1.9 and 3.2 +/- 0.5 pups compared with the females on N and C diets, respectively, in Phase 2. Feed consumption and weights of dams during lactation generally ranked C>N>I. Growth of pups during both phases also ranked C>N>I. Vaginal opening at 28 d differed by line (71.4%, K < 89.3%, L(+), P<0.05) and diet (56.8%, I < 92.0%, C or 92.2%, N, P<0.05). These results suggest both acute and chronic effects of consumption of endophyte-infected diets. Absence of line-by-diet interactions demonstrates that adverse effects were unrelated to genetic differences between lines.     

Endophytic fungi (Neotyphodium coenophialum) affect the growth and mineral uptake, transport and efficiency ratios in tall fescue (Festuca arundinacea)

Neotyphodium coenophialuminteracts mutualistically with its host grasses. Tall fescue (Festuca arundinacea Schreb.) plants infected by the fungal endophyte,Neotyphodium coenophialum(Morgan-Jones and Gams) Glenn, Bacon and Hanlin, often perform better than non-infected plants, especially in limited resource environments. However, there is a scarcity of information about endophyte-grass ecotypes interaction in Andisols of temperate regions. Clones of three tall fescue ecotypes (Fukaura, Koiwai and Showa) either infected with N. coenophialum (E+) or noninfected (E–) were grown in Andisols (Black Andisol: naturally low content of phosphorus, high in other nutrients; Red Andisol: naturally high content of phosphorus, low in other nutrients) for 133 days in a controlled environment. Cumulative shoot dry weight, daily regrowth rates (tiller number, plant height and shoot dry matter) after clippings and nutrient uptake, transport and efficiency ratios were measured. In Black Andisol, E+ plants had significantly higher cumulative shoot dry weight as well as daily regrowth rates than E– plants, while in Red Andisol the reverse was true. Among the ecotypes studied, Showa had the highest shoot growth. Significantly higher phosphorus (P), potassium (K), calcium (Ca) and magnesium (Mg) uptake as well as transport were identified in E+ vs. E– plants grown in Black Andisol. With few exceptions, values for nutrient efficiency ratios were not significantly different between E+ and E– plants grown in both soils. Significant three-way interaction (endophyte × ecotype × soil) for cumulative shoot dry weight and regrowth rate revealed that the ecotype specific regrowth responses to endophyte infection were depended on soil nutrient conditions. Vegetative growth and nutrient acquisition in tall fescue varied with ecotype and were modified by abiotic (soil fertility status) as well as biotic (endophyte infection) factors.     

The effect ofAcremonium coenophialum on the growth and nematode infestation of tall fescue

The presence of the endophytic fungusAcremonium coenophialum Morgan-Jones et Gams in tall fescue (Festuca arundinacea Schreb.) induces toxicity when this grass is grazed by cattle; however, there is evidence that removing the endophyte reduces the stand vigor and longevity of fescue. A field trial was conducted to determine the effects of water supply and the presence of the endophytic fungus on plant growth, drought tolerance, and soil nematode populations in Kentucky 31 tall fescue. The design included two factors, level of endophyte infection (0 and 75%) and irrigation regime (none, low, and high). Where water deficits occurred, herbage yield and leaf area were lower, and percentage dead tissue and canopy minus air temperature were greater in endophyte-free compared with endophyte-infected fescue. Soil populations ofPratylenchus scribneri andTylenchorhynchus acutus were substantially higher in the noninfected than in the endophyte-infected plots. The endophyte apparently confers drought tolerance to Kentucky 31 tall fescue, and this effect may be at least partially mediated through enhanced resistance to soil-borne nematodes.Published with the approval of the Director of the Ark. Agric. Exp. Stn.     

Metabolite analysis of the effects of elevated CO2 and nitrogen fertilization on the association between tall fescue (Schedonorus arundinaceus) and its fungal symbiont Neotyphodium coenophialum

Atmospheric CO₂ is expected to increase to between 550 ppm and 1000 ppm in the next century. CO₂‐induced changes in plant physiology can have ecosystem‐wide implications and may alter plant‐plant, plant‐herbivore and plant‐symbiont interactions. We examined the effects of three concentrations of CO₂ (390, 800 and 1000 ppm) and two concentrations of nitrogen fertilizer (0.004 g N/week versus 0.2 g N/week) on the physiological response of Neotyphodium fungal endophyte‐infected and uninfected tall fescue plants. We used quantitative PCR to estimate the concentration of endophyte under altered CO₂ and N conditions. We found that elevated CO₂ increased the concentration of water‐soluble carbohydrates and decreased the concentration of plant total amino acids in plants. Fungal‐derived alkaloids decreased in response to elevated CO₂ and increased in response to nitrogen fertilization. Endophyte concentration (expressed as the number of copies of an endophyte‐specific gene per total genomic DNA) increased under elevated CO₂ and nitrogen fertilization. The correlation between endophyte concentration and alkaloid production observed at ambient conditions was not observed under elevated CO₂. These results suggest that nutrient exchange dynamics important for maintaining the symbiotic relationship between fungal endophytes and their grass hosts may be altered by changes in environmental variables such as CO₂ and nitrogen fertilization.     

Effects of the Neotyphodium endophyte fungus on dormancy and germination rate of Lolium multiflorum seeds

Abstract Neotyphodium frequently occurs as an endophyte in grasses. Evidence shows enhanced fitness of endophyte infected grasses relative to non‐infected ones. Some studies of seed germination show endophyte enhancement of plant fitness in various environments, but inconsistent results indicate that further studies are needed. So far, experiments have failed to separate the confounded effects of population origin and seed management. For this reason, we evaluated the effects of endophyte infection on seed dormancy and germination in Lolium multiflorum using an experimental design controlling these factors. Depending on the year of seed production, endophyte infection modified seed response to light quality, affecting predominantly seed dormancy levels. Nevertheless, the endophyte did not affect base temperature or thermal time of germination. We concluded that endophytes were not a strong influence on germination behaviour. We speculate from our results that the presence of the endophyte changes germination by an indirect effect, in extending growth of the maternal plant during seed development and ripening. The direct effect of hyphae in the seed on seed behaviour was disregarded, because the difference between infected and non‐infected seed varied within the year of seed production. Future experiments should focus on effects of the endophyte on the canopy of parent plants during seed production and ripening, and, hence, on subsequent dormancy and germination of the seeds.     

A preliminary study of growth, serum prolactin and reproductive performance of beef heifers grazing Acremonium coenophialum -infected tall fescue

In an experiment with a 2x2 factorial arrangement of treatments, 32 virgin Angus heifers were used to study effects of tall fescue endophyte Acremonium coenophialum and energy supplementation on reproductive performance. Treatments were feeding of endophyte-free (EF) or endophyte-infected (EI) tall fescue without (0%) and with energy supplementation at one percent group mean body weight (1%). Blood samples were collected at 28-day intervals for prolactin (PRL). Starting 21 days before the breeding season, blood was also collected weekly for progesterone (P(4)) and PRL. After 99 days of grazing, heifers received Syncro-Mate-B (SMB) and were then artificially inseminated over a 30-day period. Heifers on EI-0% had a lower (P<0.05) average final body weight. Highest average daily gain was observed in heifers fed EF-1% (0.98 kg/day), whereas, heifers fed EI-0% had lowest average daily gain (0.53 kg/day; P<0.05). Endophyte-infected fescue depressed serum PRL concentrations (171 and 143 ng/ml vs 70 and 85 ng/ml; EF-0% and EF-1% vs EI-0% and EI-1%, respectively; P<0.05). No differences were observed among treatments in luteal activity before or after SMB, in pregnancy rates after a synchronized estrus or in overall pregnancy rates during the 30-day breeding season. Endophyte-infected fescue decreased average daily gain and depressed serum concentrations of prolactin, but had no effect on reproductive performance as measured in this study.     

Growth of endophyte, Neotyphodium, and its host plant, tall fescue (Festuca arundinacea), under 3D-clinorotation.

Growth of a filamentous fungus endophyte, Neotyphodium, and its host plant, tall fescue, Festuca arundinacea, was examined during the seed germination process under pseudo-microgravity [correction of micrgravity] generated by three dimensional (3D-) clinorotation. The shoot growth of tall fescue infected with the endophyte was remarkably suppressed on a 3D-clinostat compared with that of the ground control. Without being infected, shoot growth of tall fescue was not strongly affected by the 3D-clinorotation. Many aggregated hyphae were observed in the plant seed incubated for 1-day on the 3D-clinostat [correction of clinost] than in those kept on the ground. These results indicate that the clinorotation induces responses in the endophyte and its host plant different from those under normal gravity.     

Influence of phosphorus on the growth and ergot alkaloid content of Neotyphodium coenophialum-infected tall fescue (Festuca arundinacea Schreb.)

Tall fescue (Festuca arundinacea Schreb.) plants infected by the fungal endophyte Neotyphodium coenophialum (Morgan-Jones & Gams) (Glenn et al., 1996) often perform better than noninfected plants, especially in marginal resource environments. There is a lack of information about endophyte related effects on the rhizosphere of grasses. In a greenhouse experiment, four endophyte-infected (E+) tall fescue clones (DN2, DN4, DN7, DN11) and their endophyte-free (E–) forms were grown in limed (pH 6.3) Porter soil (low fertility, acidic, high aluminum and low phosphorus content, coarse-loamy mixed mesic Umbric Dystrochrept) at three soil P levels (17, 50, and 96 mg P kg^-1 soil) for five months. Excluding the genotype effect, endophyte infection significantly increased cumulative herbage DM yield by 8% at 17 mg P kg^-1 soil but reduced cumulative herbage DM yield by 12% at 96 mg P kg^-1 soil. With increased P availability in the soil, shoot and root DM, and root/shoot ratio in E+ plants were significantly less when compared to E– plants. Endophyte infection increased specific root length at 17 and 50 mg P kg^-1soil. At soil P level of 17 mg P kg^-1soil, E+ plants had significantly higher P concentrations both in roots and shoots. Similar relationships were found for Mg and Ca. E+ plants had significantly higher Zn, Fe, and Al concentration in roots, and lower Mn and Al concentration in shoots when compared to E– plants. Ergot alkaloid concentration and content in shoot of E+ plants increased with increasing P availability in the soil from 17 to 50 mg P kg^-1 but declined again at 96 mg P kg^-1 soil. Ergot alkaloid accumulation in roots increased linearly with P availability in the soil. Results suggest that endophyte infection affects uptake of phosphorus and other mineral nutrients and may benefit tall fescue grown on P-deficient soils. Phosphorus seems also to be involved in ergot alkaloid accumulation in endophyte-infected tall fescue.     

Effects of endophytic fungi on some drought tolerance mechanisms of tall fescue in a hydroponics culture

Neotyphodium, a seed-transmissible nonpathogenic fungal endophyte (symbiont) is considered beneficial because endophyte-infected grasses are more drought-tolerant, produce more dry matter, utilize soil nitrogen more efficiently, and deter insects. In this study, the effects of endophytes on physiological mechanisms of drought tolerance in tall fescue (Festuca arundinacea Schreb.) were studied in a greenhouse. Two clonally propagated genotypes of tall fescue (F. arundinacea Schreb.), naturally containing endophyte (EI), and their endophyte-free ramets (EF) were tested at three water stress treatments exerted by PEG 6000 in a hydroponics system. Relative water content (RWC), cell membrane stability (CMS), proline and chlorophyll contents in plant leaves were measured during water stress treatments. After harvest, K⁺, Ca²⁺, and Mg²⁺ contents were measured in plant roots and shoots. After 20 days under stress conditions, plants were transferred to basal hydroponics medium, and their survival after stress relief was evaluated. The results showed that endophyte considerably contributes to host grass water stress tolerance. Both genotypes of EI and EF plants did not differ in RWC, but, regardless of the infection status, genotype 75 had the higher RWC than genotype 83. EI clones of both genotypes maintained slightly higher chlorophyll content and membrane stability than EF clones, although these differences were not significant. The EI plants of genotype 83 concentrated significantly more proline than EF plants, but in the genotype 75, differences between EI and EF clones were not significant. Plant mineral absorption was also influenced by the endophyte presence. EI clones had the higher concentrations of K⁺ in the shoots of both genotypes. The Mg²⁺ and Ca²⁺ contents in EF plants of both genotypes were higher than EI plants in the roots, but in the shoots there were no differences between EI and EF clones. EI clones survived longer after stress removal. These results strongly suggest that Neotyphodium endophytes exert their effects on tall fescue drought tolerance through alteration of various physiological mechanisms involved. Published in Russian in Fiziologiya Rastenii, 2009, Vol. 56, No. 4, pp. 563–570. This test was submitted by the authors in English.     

Interaction between VA-mycorrhizal fungus and Azotobacter and their combined effects on growth of tall fescue

Amounts of N, P, K, Ca, Mg, Zn, Fe, and Mn absorbed by a nodulating and a non-nodulating (Non-nod) peanut genotype at two nitrogen fertilizer levels (nil and 200 kg N ha^–1) were determined in a field experiment. The amounts of nutrient elements in the plant parts were greatest for N, followed by K, Ca, Mg, P, Fe, Mn, and Zn in descending order. Although there were differences in the uptake of other nutrients, the major difference between Non-nod and nodulating genotypes was in nitrogen indicating the poor yield of the Non-nod line due to its inability to acquire N.Submitted as Journal Article No: 677 by International Crops Research Institute for the Semi-Arid Tropics (ICRISAT).     

Influence of temperature on alkaloid levels and fall armyworm performance in endophytic tall fescue and perennial ryegrass

The symbiotic relationships between Neotyphodium endophytes (Clavicipitacea) and certain cool‐season (C3) grasses result in the synthesis of several alkaloids that defend the plant against herbivory. Over a 3 month period we evaluated the effects of temperature on the expression of these alkaloids in tall fescue, Festuca arundinacea Schreb, and perennial ryegrass, Lolium perenne L. (Poaceae). Response surface regression analysis indicated that month, temperature, and their interaction had an impact on the alkaloid levels in both grasses. We aimed to identify the alkaloids most closely associated with enhanced resistance to the fall armyworm, Spodoptera frugiperda JE Smith (Lepidoptera: Noctuidae), and clarify the role of temperature in governing the expression of these alkaloids. The dry weights and survival of fall armyworms feeding on endophyte‐infected tall fescue or perennial ryegrass were significantly lower than for those feeding on uninfected grass, whereas endophyte infection had no significant influence on survival. For tall fescue, a four‐alkaloid model consisting of a plant alkaloid, perloline, and the fungal alkaloids ergonovine chanoclavine, and ergocryptine, explained 47% of the variation in fall armyworm dry weight, whereas a three‐alkaloid model consisting of the plant alkaloid perloline methyl ether and the fungal alkaloids ergonovine and ergocryptine explained 70% of the variation in fall armyworm dry weight on perennial ryegrass. Although temperature had a significant influence on overall alkaloid expression in both grasses, the influence of temperature on individual alkaloids varied over time. The levels of those alkaloids most closely linked to armyworm performance increased linearly or curvilinearly with increasing temperature during the last 2 months of the study. We conclude that the growth temperature of grasses can influence the performance of fall armyworm, and that this effect may be mediated through a set of plant‐ and endophyte‐related alkaloids.     

The effect of feeding endophyte infected tall fescue seed on reproductive performance in female rats

Female Sprague-Dawley rats (Rattus norvegicus) were randomly assigned to diets containing mixtures of rat chow and tall fescue (Festuca arundinacea) seed with 0, 5, 10, 20 and 40% infection levels of Acremonium coenophialum to assess the effect of the diets on the reproductive potential of rats. Rats fed 40% infected seed had decreased body weight, decreased mean percent body weight of uteri, failed to maintain normal estrous cycles and were unable to become pregnant. Animals fed a diet of 20% infected fescue seed had extended estrous cycles. There were no significant differences among the 0, 5 and 10% dietary treatments.     

Structural analysis of a peptide synthetase gene required for ergopeptine production in the endophytic fungus Neotyphodium lolii.

Lysergyl peptide synthetase 1 catalyzes the assembly of toxic ergopeptines from activated D-lysergic acid and three amino acids. The gene encoding this enzyme in the endophytic fungus Neotyphodium lolii was analyzed and compared to a homologous gene from the ergot fungus Claviceps purpurea. Each gene contained two introns, which were found in the same relative position within two modules of the gene. The 5' ends of the two genes were unusually divergent. Signature sequences determining substrate specificity were similar in adenylation domains that recognized identical amino acids but differed within the adenylation domain for the amino acid that varies between the major ergopeptines of the two fungi. Homologues were detected in several related endophytic fungi; the tall fescue endophyte Neotyphodium coenophialum contained a divergent, second copy of the gene. Our results provide new information on the structure and distribution of this important peptide synthetase involved in ergot alkaloid biosynthesis.     

Effect of feeding fungal endophyte (Acremonium coenophialum )-infected tall fescue seed on reproductive performance in CD-1 mice through continuous breeding

This study was designed to assess the effect of endophyte-infected (Acremonium coenophialum ) tall fescue (KY-31) seed (80% infected) on reproductive performance in CD-1 mice by continuous breeding. Twenty-four pairs of 70-d-old CD-1 mice were randomly allocated to four diets: 1) mouse chow ad libitum; 2) 40% infected fescue seed and 60% chow (w/w); 3) reduced intake (100% chow) similar to the intake, adjusted daily, in Diet 2; and 4) 60% infected fescue seed and 40% chow. Males and females were randomly paired (six pairs/treatment) and placed on the above diets. The mice were fed the corresponding diets for 80 d, although the pairs were separated on Day 60 (prior to the birth of the 3rd litter) and the females were monitored for one additional gestation period (20 d). The pregnancy data (litters produced) among the four treatments were 100.0 (18), 77.8 (14), 100.0 (18) and 80.0% (12) respectively. Similarly, the average number of pups born per litter among the four treatments was 11.8, 9.3, 10.1, and 9.8. When the chow treatment (1 and 3) and the fescue treatments (2 and 4) were pooled and compared, the percent pregnancy was 100.0 (n = 36) and 78.8 (n = 26), and the pups born per litter (means +/- SEM) were 11.0 +/- 0.5 and 9.5 +/- 0.6, respectively. Also the intervals between the three litters born during the 60-d cohabitation period were 21.6 +/- 1.1 and 24.5 +/- 0.9 d for the chow and fescue treatments, respectively. The results point out that 40 and 60% infected fescue seed in the diet of mice does influence (P < 0.05) their reproductive capacity as measured by percent pregnancy and litter size.     

Avian seed preference and weight loss experiments: the effect of fungal endophyte-infected tall fescue seeds

Summary The impact of endophytic fungus-infected seeds on seed predators is poorly understood. In this multiple trophic level investigation, seed preference experiments were conducted to determine whether five species of passerines (dark-eyed juncos, Junco hyemalis; American tree sparrows, Spizella arborea; song sparrows, Melospiza melodia; chipping sparrows, Spizella pusilla; and house sparrows, Passer domesticus) recognize and preferentially consume noninfected (NI) over infected (I) seeds of tall fescue (Festuca arundinacea). We predicted that the birds would refrain from eating I seeds because those seeds contain high concentrations of fungal alkaloids. When given a choice of NI fescue seeds and control seeds (millet), all bird species showed a significant preference for millet. However, individuals of all species consumed some NI seeds. When given a choice of NI and I fescue seeds, all species except the chippig sparrow ate significatly more NI than I fescue seed and the chipping sparrow showed the same trend. Thus, birds were able to distinguish between the two seed types and preferred NI seeds in choice tests. Additional experiments investigated weight changes in dark-eyed juncos fed diets containing different proportions of millet, NI, and I fescue seed. Significant differences in weight loss were observed for the various diets. Juncos showed greater weight loss when the proportion of fescue seed, especially the proportion of I seed, in their diet was greater. The potential significance of the finding that abundant grass seeds are made unavailable to predators by fungal infection is discussed in relation to foraging and competition in avian communities.     

Advanced data-mining strategies for the analysis of direct-infusion ion trap mass spectrometry data from the association of perennial ryegrass with its endophytic fungus, Neotyphodium lolii

Direct-infusion mass spectrometry (MS) was applied to study the metabolic effects of the symbiosis between the endophytic fungus Neotyphodium lolii and its host perennial ryegrass (Lolium perenne) in three different tissues (immature leaf, blade, and sheath). Unbiased direct-infusion MS using a linear ion trap mass spectrometer allowed metabolic effects to be determined free of any preconceptions and in a high-throughput fashion. Not only the full MS(1) mass spectra (range 150-1,000 mass-to-charge ratio) were obtained but also MS(2) and MS(3) product ion spectra were collected on the most intense MS(1) ions as described previously (Koulman et al., 2007b). We developed a novel computational methodology to take advantage of the MS(2) product ion spectra collected. Several heterogeneous MS(1) bins (different MS(2) spectra from the same nominal MS(1)) were identified with this method. Exploratory data analysis approaches were also developed to investigate how the metabolome differs in perennial ryegrass infected with N. lolii in comparison to uninfected perennial ryegrass. As well as some known fungal metabolites like peramine and mannitol, several novel metabolites involved in the symbiosis, including putative cyclic oligopeptides, were identified. Correlation network analysis revealed a group of structurally related oligosaccharides, which differed significantly in concentration in perennial ryegrass sheaths due to endophyte infection. This study demonstrates the potential of the combination of unbiased metabolite profiling using ion trap MS and advanced data-mining strategies for discovering unexpected perturbations of the metabolome, and generating new scientific questions for more detailed investigations in the future.     

Effect of organic fertilisers on the greening quality, shoot and root growth, and shoot nutrient and alkaloid contents of turf-type endophytic tall fescue, Festuca arundinacea

Increasing concern about the potential negative environmental impact of chemical fertilisers used in urban landscapes has provided impetus to develop organic fertilisers. However, little is known about the effect of organic fertilisers on turfgrass quality, growth and stress resistance. This study compared the effect of 11 organic fertilisers, applied at manufacturer's recommended rates, on greening quality, shoot and root growth, and shoot nutrient (an indication of nutrient uptake) and alkaloid content (an indication of insect resistance) in endophytic (infected with the fungus Neotyphodium coenophialum) tall fescue in the greenhouse. We measured turfgrass greening quality on a 1–9 scale weekly (9 being the highest), shoot and root growth monthly, and shoot contents of macro- and micronutrients and of various alkaloids at the end of 4 months. The results show that Corn Gluten and Cockadoodle Doo produce the highest turfgrass greening quality and shoot growth. Nature's Touch with enzymes enhanced root growth, and thus resulted in high root:shoot ratio, especially in third and fourth months after application. Compared with the most commonly used chemical fertiliser, Scott's Turf Builder, the organic fertilisers Cockadoodle Doo, Corn Gluten and Nature's Touch with enzymes generally resulted in better turf greening quality. Although Cockadoodle Doo, Vigoro and Scott's Turf Builder resulted in higher macronutrient contents in turfgrass shoots, there was no significant correlation between the nutrient contents in the fertilisers and in the shoots four months after application. Significant differences were found for all measured alkaloids in turfgrass shoots among the 13 treatments, and these differences varied with fertiliser. Overall, organic fertilisers produced higher turfgrass greening quality, root and shoot growth and insect resistance capacity (alkaloid content) compared with the chemical fertiliser, Scott's Turf Builder. On the basis of the high to excellent turfgrass greening quality ratings, root:shoot ratio, shoot nutrient and alkaloid contents in this study, we conclude that Cockadoodle Doo, Vigoro and Nature's Touch with enzymes are relatively superior organic fertilisers.