Fungal loline alkaloids in grass–endophyte associations confer resistance to the rice leaf bug, Trigonotylus caelestialium

Plant symbiotic fungi (endophytes) of the genus Neotyphodium [anamorphs, asexual derivatives of Epichloë (Ascomycota: Clavicipitaceae)] often associate with grass species of the subfamily Pooideae, which includes important forage and turf species. These endophytes are known to produce a range of alkaloids that enhance their host's resistance to insects or are toxic to grazing animals. Among the alkaloids, loline alkaloids (saturated 1‐aminopyrrolizidines) are generally observed in the highest concentrations in many Neotyphodium–grass symbiotic associations, and are known to be toxic to insects but not to mammals. Some Neotyphodium‐infected grasses have enhanced resistance to rice leaf bug, Trigonotylus caelestialium (Kirkaldy) (Heteroptera: Miridae), one of the major pests for rice production in Japan. Our laboratory experiments quantified the effects of purified loline (N‐formylloline) and in planta synthesis of loline alkaloids by meadow fescue [Lolium pratense (Huds.) S.J. Darbyshire (syn. Festuca pratensis Huds.)]–Neotyphodium uncinatum (Gams, Petrini & Schmidt) Glenn, Bacon & Hanlin and Italian ryegrass (Lolium multiflorum Lam.)–Neotyphodium occultans Moon, Scott & Christensen associations on the development and survival of T. caelestialium. No‐choice feeding assays with laboratory populations of the insect revealed that their growth was significantly decreased by the infected grasses, and the effect was greater for N. uncinatum than for N. occultans, in keeping with differences in N‐formylloline concentrations in the plants. Artificial feeding of N‐formylloline through feeding sachets indicated that the chemical has an adverse effect on survival of larvae, even at the lowest concentration tested (50 µg/g), which was considerably lower than the typical concentrations in many Neotyphodium–grass associations. The results confirmed the ability of Neotyphodium‐infected forage grasses to control T. caelestialium propagation in meadows, which may cause damages to nearby rice paddies.     

Mollusc herbivory influenced by endophytic clavicipitaceous fungal infections in grasses

Clavicipitaceous fungi of the genus Neotyphodium occur widely as mutualistic, systemic, seed‐borne infections in festucoid grasses. Grass infection by these fungi is associated with the presence of a range of secondary metabolites (SM), several of which have been demonstrated to confer to the plant resistance against herbivorous vertebrates and insects. An initial experiment demonstrated that endophytic infection by Neotyphodium can influence the utilisation of grasses by Deroceras, with feeding preferences and impact on plant yields affected differentially by endophytes with different SM profiles. The role of Neotyphodium SM in feeding preferences of Deroceras slugs were then evaluated in artificial diets. Among the indole diterpenoids tested, lolitrem B was demonstrated to reduce feeding, while diets containing paxilline, lolitriol, α‐paxitriol and β‐paxitriol tended to be preferred over that of untreated diet. The pyrrolopyrazine alkaloid peramine had no effect. Among the ergopeptine alkaloids tested in the diets, ergotamine and ergovaline were demonstrated to be phagostimulatory. These results with artificial diets were generally consistent with Deroceras reticulatum preferences among plants of known Neotyphodium endophyte strain and SM profile. Deroceras slugs obtained from sites containing contrasting frequencies of Neotyphodium‐infected grasses, exhibited differential responses to Neotyphodium SM incorporated into artificial diet. This study demonstrates that infection of grasses by different isolates of Neotyphodium endophytes differentially influence herbivory by molluscs, reflecting their SM profile. These results offer an explanation for variable acceptability of grasses to molluscs and their importance in the diet of molluscs in the field reported in previous studies in both natural and agricultural systems. Neotyphodium endophytes potentially offer novel approaches to management of mollusc pests in agricultural gramineous crops.     

Contribution of ergot alkaloids to suppression of a grass-feeding caterpillar assessed with gene knockout endophytes in perennial ryegrass

Neotyphodium and Epichloë species (Ascomycota: Clavicipitaceae) are fungal symbionts (endophytes) of grasses. Many of these endophytes produce alkaloids that enhance their hosts’ resistance to insects or are toxic to grazing mammals. The goals of eliminating from forage grasses factors such as ergot alkaloids that are responsible for livestock disorders, while retaining pasture sustainability, and of developing resistant turf grasses, require better understanding of how particular alkaloids affect insect herbivores. We used perennial ryegrass Lolium perenne L. (Poaceae) symbiotic with Neotyphodium lolii × Epichloë typhina isolate Lp1 (a natural interspecific hybrid), as well as with genetically modified strains of Lp1 with altered ergot alkaloid profiles, to test effects of ergot alkaloids on feeding, growth, and survival of the black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae), a generalist grass‐feeding caterpillar. Neonates or late instars were provided clippings from glasshouse‐grown plants in choice and rearing trials. Wild‐type endophytic grass showed strong antixenosis and antibiosis, especially to neonates. Plant‐endophyte symbiota from which complex ergot alkaloids (ergovaline and lysergic acid amides such as ergine) or all ergot alkaloids were eliminated by endophyte gene knockout retained significant resistance against neonates. However, this activity was reduced compared to that of wild‐type Lp1, providing the first direct genetic evidence that ergot alkaloids contribute to insect resistance of endophytic grasses. Similarity of larval response to the two mutants suggested that ergovaline and/or ergine account for the somewhat greater potency of wild‐type Lp1 compared to the knockouts, whereas simpler ergot alkaloids contribute little to that added resistance. All of the endophyte strains also produced peramine, which was probably their primary resistance component. This study suggests that ergot alkaloids can be eliminated from an endophyte of perennial ryegrass while retaining significant insect resistance.     

Detrimental and neutral effects of wild barley–Neotyphodium fungal endophyte associations on insect survival

Neotyphodium (Clavicipitaceae: Balansieae) fungal endophyte infection does not always confer temperate grass resistance to insect herbivores, although reports indicate that over 40 species are adversely affected by its infection. Laboratory and glasshouse experiments were conducted to improve our knowledge of the anti‐insect properties of Neotyphodium‐infected (E+) non‐commercial grasses, and E+ wild barley (Hordeum) specifically. Neotyphodium infection of four plant inventory (PI) lines of wild barley conferred resistance to Mayetiola destructor (Say) (Diptera: Cecidomyiidae), whereas none of the E+ wild barley accessions reduced the survival of Rhopalosiphum padi (L.) (Homoptera: Aphididae). Metopolophium dirhodum (Walker) (Homoptera: Aphididae) densities were significantly lower on the E+ clones of Hordeum brevisubulatum ssp. violaceum (Boissier and Hohenacker) (PI 440420), compared with densities on endophyte‐free (E–) plants of this species in population growth experiments. Neotyphodium infection of three Hordeum bogdanii (Wilensky) PI lines did not confer resistance to M. dirhodum; however, one of these E+ lines (PI 314696) was resistant to this aphid in a second population growth experiment. Our results provide additional evidence that the outcome of a grass–endophyte–insect interaction is influenced by the host grass species or genotype, Neotyphodium species or genotype, and the insect species involved. They also reinforce this phenomenon for non‐commercial grass–endophyte–insect interactions and underscore the potential role of endophytes in mediating wild barley–insect interactions and their potential to act as defensive mutualists.     

Controlling Myzus persicae with recombinant endophytic fungi Chaetomium globosum expressing Pinellia ternata agglutinin: using recombinant endophytic fungi to control aphids

Aims: Sap‐sucking insect pests have become the major threats to many crops in recent years; however, only a few biopesticides have been developed for controlling those pests. Here, we developed a novel pest management strategy, which uses endophytes to express anti‐pest plant lectins. Methods and Results: The fungal endophyte of Chaetomium globosum YY‐11 with anti‐fungal activities was isolated from rape seedlings. Pinellia ternata agglutinin (pta) gene was cloned into YY‐11 mediated by Agrobacterium tumefaciens. The positive transformants, as selected by antibiotic resistance, were evaluated using PCR and Western blot assay. We found that the recombinant endophytes colonized most of the crops, and the resistance of rape inoculated with recombinant endophytic fungi significantly inhibited the growth and reproduction of Myzus persicae. Conclusions: Our results showed that the recombinant endophytes expressing Pinellia ernata agglutinin (PTA) may endow hosts with resistance against sap‐sucking pests. Significance and Impact of the Study: This research may have important implications for using endophytes to deliver insecticidal plant lectin proteins to control sap‐sucking pests for crop protection.     

Evidence for leaf endophyte regulation of root symbionts: effect of <Emphasis Type="Italic">Neotyphodium</Emphasis> endophytes on the pre-infective state of mycorrhizal fungi

Neotyphodium endophytes and arbuscular mycorrhizal (AM) fungi are common constituents of natural grasslands. The plant–endophyte symbiosis can introduce changes in soil conditions that affect the density and activity of different functional groups of soil organisms. In the present work we performed in vitro assays to evaluate the effect of root and endophyte exudates on the pre-infective state of mycorrhizal fungi (Gigaspora margarita and G. rosea). Plant roots of Bromus setifolius from populations of Patagonia, and four strains of Neotyphodium were used to obtain the exudates. Root exudates of infected plants, at a high concentration, significantly increased AMF hyphal branches and length relative to exudates from naturally endophyte free plants. The effect of Neotyphodium endophyte exudates on AMF mycelial length varied depending on strain and the concentration used, suggesting a differential interaction between endophyte and AMF species. AMF hyphal branches were increased by Neotyphodium fungal exudates in both mycorrhizal species. A few previous studies have suggested that Neotyphodium endophytes can reduce mycorrhizal sporulation and colonization of host roots in commonly-cultivated agronomic hosts. In this study we report the opposite effect in B. setifolius. This study reports the direct and positive effect of root exudates from plants in symbiosis with Neotyphodium, on AMF pre-infective state. Further, identical effects were detected using exudates from Neotyphodium endophytes.     

Endophyte–grass complexes and the relationship between feeding preference and performance in a grass herbivore

Overseeding non‐endophytic turfgrass lawns with endophytic turfgrass is considered promising for the development of integrated pest management programs in urban landscapes. In this context, a better understanding of the variability in the preference–performance response of insect herbivores exposed to endophytic grasses could be useful to develop efficient practices. Specifically, while endophytic varieties that are strongly toxic and avoided could provide good control of mobile pests, varieties that are toxic but not avoided could be more suitable for the control of sedentary pests. In this laboratory study, we investigated how the infection of 10 grass varieties by Neotyphodium endophytes affected the feeding performance and preference of newly hatched nymphs of the hairy chinch bug, Blissus leucopterus hirtus Montandon (Heteroptera: Lygaeidae), a common turfgrass pest in north‐eastern USA. We found that endophytes generally induced a strong relationship between feeding performance and preference in this herbivore. However, two endophytic varieties did not conform to this relationship, with one variety being highly toxic but not avoided and the other less toxic but highly avoided. These results provide a solid basis to further explore the impacts of endophytic grasses on the dispersal and survival of insect pests in mixed stands of endophytic and non‐endophytic plants.     

Genetic diversity in epichloid endophytes of Hordelymus europaeus suggests repeated host jumps and interspecific hybridizations

Epichloid fungal endophytes (Epichloë and Neotyphodium spp.) are excellent model systems for studying speciation processes because of their variable life history traits that are linked to host grass fitness. Presumed jumps to new hosts and subsequent somatic hybridizations appear to be common among epichloid endophytes resulting in increased genetic variation upon which selection can act and speciation be initiated. In this study, we explored the endophyte diversity of a rare European native woodland grass species, Hordelymus europaeus, along a latitudinal transect covering the entire distribution range of H. europaeus. From 28 populations in six countries, isolates were sampled and molecularly characterized. Based on the sequences of tubB and tefA, six distinct epichloid taxa (interspecific hybrid or cryptic haploid species) were found, of which four were novel and two have been previously reported from this host. Of the novel endophytes, two were presumed to be interspecific hybrids and two of nonhybrid origin. While previously known endophytes of H. europaeus are seed‐born and strictly asexual, one of the novel nonhybrid endophytes found in the glacial refugium of the Apennine peninsula reproduced sexually in cultured plants. This is the first case of a seed‐borne, but sexually reproducing endophyte of this host. We discuss the origin, and possible ancestral species, of the six epichloid taxa using phylogenetic analyses. Repeated host jumps and somatic hybridizations characterize the diversity of the endophytes. To date, no other grass species is known to host a larger diversity of endophytes than H. europaeus.     

Neotyphodium Endophyte Changes Phytoextraction of Zinc in Festuca arundinacea and Lolium perenne

The effect of Neotyphodium endophytes on growth parameters and zinc (Zn) tolerance and uptake was studied in two grass species of Festuca arundinacea and Lolium perenne. Plants were grown under different Zn concentrations (control, 200, 400, 800, and 1800 mg kg^?1) in potted soil for 5 months. The results showed that the number of plant tillers was 85 and 51% greater in endophyte infected Festuca (FaEI) and Lolium (LpEI), respectively, compared to their endophyte free (EF) plants. Roots and shoots dry weights in infected Festuca were 87 and 9% greater than non-infected counterparts but in opposite, EF Lolium had 47 and 8% greater root and shoot dry weights than LpEI. Endophyte infected Festuca and Lolium improved chlorophyll fluorescence as Fv/Fm at high concentrations of Zn, showing their better chlorophyll functions and significant reduction of Zn stress in endophyte infected plants. Shoots of endophyte infectedFestuca had 82% greater concentration of Zn than EF Festuca when grown in soil containing 1800 mg kg^?1 Zn. Festuca and Lolium may tolerate high Zn concentration in soil without reduction in shoot and root growth. Endophyte infection in Festuca may help the grass accumulate and transport more Zn in aboveground parts under Zn-stress, thereby aiding phytoremediation of contaminated soils.     

Positive association between mycorrhiza and foliar endophytes in Poa bonariensis, a native grass

The interaction between mycorrhiza and leaf endophytes (Neotyphodium sp.) was studied in three Poa bonariensis populations, a native grass, differing significantly in endophyte infection. The association between endophytes and mycorrhizal fungi colonisation was assessed by analysing plant roots collected from the field. We found that roots from endophyte-infected populations showed a significantly higher frequency of colonisation by mycorrhizal fungi and that soil parameters were not related to endophyte infection or mycorrhiza colonization. In addition, we did not observe significant differences in the number of AM propagules in soils of the three populations sites. We also report the simultaneous development of Paris-type and Arum-type mycorrhiza morphology within the same root systems of P. bonariensis. The co-occurrence of both colonisation types in one and the same root system found in the three populations, which differed in Neotyphodium infection, suggests that foliar endophytes do not determine AM morphology. The percentage of root length colonised by different types of fungal structures (coils, arbuscules, longitudinal hyphae and vesicles) showed significant and positive differences in arbuscular frequency associated with endophyte infection, whereas the much smaller amounts of vesicles and hyphal coils did not differ significantly.     

Mealybug, Phenococcus solani, and barley aphid, Sipha maydis, response to endophyte-infected tall and meadow fescues

Mealybugs and aphids are insects which damage grass species. The effects of fungal endophytes on the feeding of the mealybug, Phenococcus solani Ferris (Homoptera: Pseudococcidae), and barley aphid, Sipha maydis Passerini (Homoptera: Aphididae), on tall fescue, Festuca arundinacea Schreb. and meadow fescue, Festuca pratensis Huds., were studied under greenhouse conditions. Mealybugs preferred endophyte‐free (E–) clones over their endophyte‐infected (E+) counterparts. E+ plants had a significantly lower number of mealybugs than E– plants. A mixture of E+ and E– plants supported intermediate mealybug numbers, between pure plantings of E+ and E– grasses. Barley aphids released on to plant materials were deterred from feeding and could not persist on E+ plants. E– plants did not survive because of aphid damage, while E+ plants generally re‐grew, but were damaged to some degree. The results showed that the use of pure stands of endophyte‐infected grasses or a mixed stand of infected and non‐infected plants may increase the persistence and durability of turf and forage grass species in the presence of foliar damaging insects.     

Population studies of native grass-endophyte symbioses provide clues for the roles of host jumps and hybridization in driving their evolution

Fungal endophytes in the genera Epichloë and Neotyphodium, collectively termed the epichloae, have fascinated biologists for decades. These intriguing fungi, also referred to as ‘class 1 or clavicipitaceous endophytes’, spend the large majority, or even their entire life cycle, within the tissues of their cool‐season grass hosts without eliciting any symptoms of infection. While all epichloae reside within the intercellular spaces of aboveground vegetative grass tissues, the species at the symbiotic extreme are known as Neotyphodium, and the intimacy of their interaction extends to the reproductive (flowering) stage. At this point, fungal filaments (hyphae) nondestructively invade the developing ovaries of their host and are incorporated into perfectly viable, healthy seeds. Thus, these endophytes live solely within the tissues of their host plants and are transmitted maternally from generation to generation. A second life history characteristic of interest is that while all Epichloë and some Neotyphodium species are haploid, a great many of the strictly seed‐transmitted Neotyphodium spp. are interspecific hybrids. This phenomenon may be critical for the success of these symbioses over longer spans of evolutionary time and will be discussed in greater detail below. A third characteristic, and one of the primary reasons these grass endophytes have received so much attention over the last three decades, is the strong mutualistic nature these relationships often exhibit. In exchange for photosynthetically derived carbon, the endophytes protect their cool‐season grass hosts from grazing herbivores and a variety of abiotic stresses. It has been hypothesized that these three biological phenomena are related ( Schardl & Craven 2003 ), perhaps with the former two driving the third, and it is here that the recent article in Molecular Ecology entitled ‘Genetic diversity in epichloid endophytes of Hordelymus europaeus suggests repeated host jumps and interspecific hybridizations’, by Oberhofer & Leuchtmann (2012) , provides critical clues to linking these traits together. While the large majority of studies have focused on documenting the ever‐increasing list of mutualistic qualities attributed to these fungi, very few have taken an exhaustive population‐level approach to document plant and endophyte genotypes within a naturally occurring system ( Faeth et al. 2010 ; Jani et al. 2010 ; Tintjer & Rudgers 2006 ). Such information is crucial to more fully elucidate the factors shaping grass‐endophyte symbioses and those often driving these relationships to mutualistic extremes.     

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.     

Endophyte-grass-herbivore interactions: the case of Neotyphodium endophytes in Arizona fescue populations

Neotyphodium endophytes in introduced agronomic grasses are well known to increase resistance to herbivores, but little is known of interactions between Neotyphodium endophytes and herbivores in native grass populations. We investigated whether endophytes mediate plant-herbivore interactions in a native grass species, Festuca arizonica in the southwestern United States, in two ways. First, to test the prediction that the presence and frequency of endophyte-infected (E+) plants should increase with increasing herbivory, we determined endophyte frequencies over a 4-year period in six natural Arizona fescue populations. We compared Neotyphodium frequency among plants growing inside and outside long-term vertebrate grazing exclosures. Second, we experimentally tested the effects of Neotyphodium infection, plant clone, and soil nutrients on plant resistance to the native grasshopper Xanthippus corallipes. Contrary to predictions based upon the hypothesis that endophytes increase herbivore resistance, levels of infection did not increase in plants subjected to grazing outside of exclosures relative to ungrazed plants within exclosures. Instead, endophyte frequencies tended to be greater inside the exclosures, where long-term vertebrate grazing was reduced. The grasshopper bioassay experiment corroborated these long-term patterns. Survival of grasshoppers did not differ between infected (E+) and uninfected (E–) plants. Instead, mean relative growth rate of grasshoppers was higher on E+ grasses than on E– ones. Growth performance of newly hatched grasshopper nymphs varied among host plant clones, although two of six clones accounted for most of this variation. Our results suggest that Neotyphodium-grass-herbivore interactions may be much more variable in natural communities than predicted by studies of agronomically important Neotyphodium-grass associations, and herbivory is not always the driving selective force in endophyte-grass ecology and evolution. Thus, alternative hypotheses are necessary to explain the wide distribution and variable frequencies of endophytes in natural plant populations. Received: 15 February 1999 / Accepted: 19 July 1999     

Arthropod abundance in tall fescue,Lolium arundinaceum,pastures containing novel ‘safe’ endophytes

Poor livestock performance on tall fescue is linked to infection by a fungal endophyte that enhances grass resistance to stress, including herbivory, while producing ergot alkaloids toxic to vertebrate grazers. Novel ‘safe’ endophyte/grass associations produce no ergot alkaloids yet retain stand persistence, but they could be more susceptible to insect outbreaks. We tested the hypothesis that grass‐feeding insects are more abundant in novel endophyte pastures compared with those containing common endophyte. Above‐ and below‐ground herbivores were sampled across two growing seasons in pastures containing common strain endophyte (KY31), novel endophytes (MaxQ or AR584) or endophyte free. We also sampled natural enemies as an indicator of possible tritrophic effects. With a few exceptions, numbers of predatory, chewing (grasshoppers, crickets, caterpillars) or sucking insects (leafhoppers, planthoppers) were similar in common and novel endophyte pastures. Contrary to our hypothesis, Aphrodes spp. leafhoppers were more abundant in KY31 than other pasture types in 2008; their nymphs also were more abundant in KY31 than in MaxQ in 2009. Adults (but not total numbers or nymphs) of another leafhopper, Psammotettix striatus, were less abundant in KY31 than other pasture types in 2009. Popillia japonica and Cyclocephala spp. grubs did not differ in density, weight or instar among the endophyte‐containing associations. In feeding assays, armyworm (Mythimna unipuncta) survival was higher on MaxQ and NE9301, and weight gain was higher on NE9301, suggesting armyworm outbreaks could be more severe on such grasses, but that result may not be attributable solely to alkaloids because common strain and endophyte‐free grasses did not differ in either parameter. Caterpillar abundance did not differ among pasture types in the field. Our results suggest that re‐seeding common strain endophytic pastures with livestock‐safe novel endophyte/grass associations to alleviate fescue toxicosis is unlikely to promote markedly higher populations of plant‐feeding insects.     

A new group of endophytes in European grasses

Poaceae of subfamily Festucoideae frequently harbour endophytic, symbiotic fungi in their aerial parts. Isolations were made from 21 different species of Poaceae, belonging mainly to the genera Lolium and Festuca. Pure cultures of 45 fungal isolates were obtained. Morphology, isozyme and molecular analysis showed the presence of three groups of endophytes. Most of the isolates could be accommodated with the previously described species of Neotyphodium and Epichloë (the e endophytes). Two new endophytes isolated from Koeleria cristata and Melica ciliata were shown to belong to this group. Gliocladium like and Phialophora like isolates were obtained from Lolium perenne and Festuca gigantea respectively (the p endophytes). Nine isolates especially those from annual Lolium produced rod like conidia. These isolates belong to Acremonium sensu stricto and are morphologically similar to Acremonium chilense.     

Insect oviposition preference between Epichloë‐symbiotic and Epichloë‐free grasses does not necessarily reflect larval performance

Variation in plant communities is likely to modulate the feeding and oviposition behavior of herbivorous insects, and plant‐associated microbes are largely ignored in this context. Here, we take into account that insects feeding on grasses commonly encounter systemic and vertically transmitted (via seeds) fungal Epichloë endophytes, which are regarded as defensive grass mutualists. Defensive mutualism is primarily attributable to alkaloids of fungal origin. To study the effects of Epichloë on insect behavior and performance, we selected wild tall fescue (Festuca arundinacea) and red fescue (Festuca rubra) as grass–endophyte models. The plants used either harbored the systemic endophyte (E+) or were endophyte‐free (E?). As a model herbivore, we selected the Coenonympha hero butterfly feeding on grasses as larvae. We examined both oviposition and feeding preferences of the herbivore as well as larval performance in relation to the presence of Epichloë endophytes in the plants. Our findings did not clearly support the female's oviposition preference to reflect the performance of her offspring. First, the preference responses depended greatly on the grass–endophyte symbiotum. In F. arundinacea, C. hero females preferred E+ individuals in oviposition‐choice tests, whereas in F. rubra, the endophytes may decrease exploitation, as both C. hero adults and larvae preferred E? grasses. Second, the endophytes had no effect on larval performance. Overall, F. arundinacea was an inferior host for C. hero larvae. However, the attraction of C. hero females to E+ may not be maladaptive if these plants constitute a favorable oviposition substrate for reasons other than the plants' nutritional quality. For example, rougher surface of E+ plant may physically facilitate the attachment of eggs, or the plants offer greater protection from natural enemies. Our results highlight the importance of considering the preference of herbivorous insects in studies involving the endophyte‐symbiotic grasses as host plants.     

Fungal endophyte infection changes growth attributes in Lolium multiflorum Lam

Abstract Lolium multiflorum is a successful invader of postagricultural succession in the Inland Pampa grasslands in Argentina, becoming a dominant species in the plant community. Individual plants of this annual species are naturally highly infected with fungal endophytes (Neotyphodium sp.) from early successional stages. We assessed the effect of Neotyphodium infection on the biology of L. multiflorum. We evaluated growth attributes between endophyte infected (E+) and uninfected (E–) plants under non‐competitive conditions during the normal growing season. E+ plants produced significantly more vegetative tillers and allocated more biomass to roots and seeds. Although seed germination rates were greater in endophyte free plants, the rate of emergence and the final proportion of emerged seedlings were similar between the biotypes. The greater production of vegetative tillers, and the greater resource allocation to roots and seeds are likely to confer an ecological advantage to E+ plants, thus enabling their dominance over the E– individuals in natural grasslands.     

Presence of Neotyphodium-like endophytes in European grasses

In order to improve knowledge of Neotyphodium‐likc fungi in European grasses, microscopic detection of endophytic mycelium was achieved in living grasses and in seeds. Samples of grasses were gathered near permanent pastures or along paths in France (93 samples, 13 genera, 22 species). Typical convoluted mycelium was found inside leaf sheaths of four genera, especially in Festuca and Lolium spp. The results suggest that endophytes were present most often in the southern regions where plants may suffer from summer drought. Endophytes were looked for in 489 seed samples (63 genera, 237 species) received from 24 European botanical gardens. Neotyphodium was found in seeds of only six genera (22 species), especially in Festuca (15 species) and Lolium (three species). No endophytic mycelium was found in the other 215 species, although other work had demonstrated the presence of the fungus in 39 of the species analysed. Neotyphodium‐likc mycelium was detected for the first time in Festuca juncifolia, F. trachyphylla, F. vaginata, F. pseudodalmatica, F. rupicaprina, F. arenaria, Vulpia ciliata and Micropyrum tenellum. Further studies are needed for the correct identification of the Neotyphodium species observed.     

Microsite conditions influence nutritive value characteristics of a tall fescue cultivar devoid of,or infected with a native,or a novel non-ergogenic endophyte

Tall fescue [Lolium arundinaceum, Schreb., S.J. Darbysh.] productivity and persistence often benefits from association with Neotyphodium coenophialum [Morgan-Jones and Gams], Glenn, Bacon, and Hanlin) endophyte. The influence of novel, non-ergogenic endophytes on nutritive value is unclear, especially when simultaneous stresses (e.g., defoliation and shading) are imposed on the association. We conducted a field experiment using Jesup tall fescue that had either a native or novel non-ergogenic fungal endophyte (AR542; referred to as MaxQ?), or that was endophyte free. Dry matter production and nutritive value including crude protein (CP), non-structural carbohydrates (TNC), ergo- and loline alkaloids, and phenolics were determined for plants stockpiled or clipped repeatedly in sites differing in the amount of light. Productivity varied less among sites when plants were infected with a native endophyte compared to novel or no endophyte. The trend suggests that native endophyte contributed to resilience of the host in this experiment. Leaf dry matter content was affected by host–endophyte association interacting with light availability suggesting differences in leaf composition could occur. Herbage CP increased, whereas TNC decreased with increasing shade. The concentration of loline alkaloids, irrespective of host–endophyte association, tended to increase in leaves with decreasing light availability and could be related to the relatively greater N concentrations in shade-grown leaves. Phenolics decreased in leaves, but increased in stembases as light availability decreased. The combination of increased loline alkaloids in leaves and phenolics in stembases, suggests that shade-grown tall fescue might have some competitive advantage based on the known anti-herbivory attributes of loline alkaloids and phenolic compounds.