Fungal endophyte infection increases tall fescue's survival,growth, and flowering in a reconstructed prairie

Grasslands in North America are increasingly threatened by land conversion and ecological degradation, prompting restoration efforts to increase native plant species diversity and improve wildlife habitat. A major challenge is the removal and management of nonnative invasive species such as tall fescue (Schedonorus arundinaceus), which has a symbiotic association with a fungal endophyte (Epichloë coenophiala) that modifies its ecological interactions. Using transplanted clumps of the cultivar Kentucky‐31, we tested the effects of endophyte infection on tall fescue's survival and performance (tiller production, flowering, and basal area) for 5 years in a central Kentucky reconstructed prairie. We predicted that endophyte infected (E+) clumps would have increased performance compared to endophyte‐free (E?) clumps. Overall, E+ clumps had greater survival, tiller production, flowering tiller production, and basal area, but not reproductive effort (proportion of tillers flowering) as compared to E? clumps. However, survival and trends in tiller number and basal area over the 5‐year period suggested experimental tall fescue populations were in decline in the reconstructed prairie, although the E? population declined more rapidly. Our study provides evidence that endophyte infection improved tall fescue's growth and survival in a postreconstruction plant community, at least in the early years following reconstruction, and may increase the invasive potential of this nonnative species in prairie restorations.     

Effects of the endophyte Epichloë coenophiala on the root microbial community and growth performance of tall fescue in different saline-alkali soils

Soil salinization is detrimental to plant growth and yield in agroecosystems worldwide. Epichloë endophytes, a class of clavicipitaceous fungi, enhance the resistance of host plants to saline-alkali stress. This study explored the effects of the systemic fungal endophyte Epichloë coenophiala on the root microbial community and growth performance of tall fescue (Lolium arundinaceum) growing under different saline-alkali stress conditions. Structural equation modeling (SEM) was conducted to analyze the direct and indirect effects (mediated by root microbial community diversity and soil properties) of the endophyte on the growth of tall fescue under saline-alkali stress. The endophyte-infected plants produced higher shoot and root biomass compared to endophyte-free plants under saline-alkali stress (200 and 400 mM). Endophyte infection increased the fungal community diversity and altered its composition in the roots, decreasing the relative abundance of Ascomycota and increasing that of Glomeromycota. Furthermore, endophyte infection decreased the bacterial community diversity and the relative abundance of dominant Proteobacteria. SEM showed that endophyte infection increased the shoot and root biomass under saline-alkali stress (200 and 400 mM) by increasing the arbuscular mycorrhizal fungal diversity in the roots, and soil total nitrogen and phosphorus concentrations. Therefore, it is important to examine aboveground microbes as factors influencing plant growth in saline-alkali stress by affecting belowground microbes and soil chemical properties.     

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.     

Transfer of endophyte-origin defensive alkaloids from a grass to a hemiparasitic plant

Plants growing in natural environments experience myriad interactions with a diverse assemblage of pathogens, parasites and mutualists. Many of these interactions involve symbiotic bacteria and fungi, but they also include macroparasitic plants. In this study, we investigated the interactions among a host grass (Lolium pratense, ex., Festuca pratensis), its symbiotic endophytic fungus (Neotyphodium uncinatum), a root hemiparasitic plant (Rhinanthus serotinus) of the host grass and a generalist herbivore (aphid Aulacorthum solani) of the hemiparasite. We demonstrate that the hemiparasitic plant acquires defending mycotoxins produced by the endophytic fungus living within their shared host grass. The uptake of defensive mycotoxins from the endophyte‐infected host grass enhances the resistance of the hemiparasitic plant to the generalist aphid herbivore. Endophyte infection increases the performance of the hemiparasitic plant, but reduces the growth of the host grass. In other words, the mutualistic endophytic fungus becomes parasitic in the presence of the hemiparasitic plant. Our results suggest that the outcomes of grass–endophyte interactions are conditional on the complexity of community‐level interactions; thus, the outcome of multispecies interactions may not be predictable from pair‐wise combinations of species.     

A fungal endophyte of a palatable grass affects preference of large herbivores

Temperate grasses frequently acquire resistance to herbivores through a symbiosis with epichloid fungi that produces alkaloids of variable deterrent effects. However, in those cases without apparent endophyte negative effects on domestic herbivores, it is not clear whether plant consumption or preference is affected or not. We performed three experiments with 1‐year‐old steers (Bos taurus, Aberdeen Angus) and the annual grass Lolium multiflorum, infected or not by Epichloë occultans to evaluate preference and to identify the underlying tolerance mechanisms. The first experiment evaluated steer preference for L. multiflorum cultivated in plots with three endophyte infection frequencies (low, medium and high), and investigated the role of canopy structure and plant nutritional traits on preference. The second experiment evaluated preference for chopped grass, offered in individual trays with contrasting infection frequencies (low and high), to discard possible effects associated with canopy structure and to focus on nutritional traits. The third experiment was performed with a tray + basket design that separated visual and olfactory stimuli from nutritional traits. High endophyte infection frequencies reduced consistently animal preference, even after short (~10 min) feeding events. However, we did not find significant evidence of plant structural, nutritional, visual or olfactory traits. Our results discarded several potential mechanisms; therefore, the dissuasive effect of fungal endophytes on animal consumption might be related to other mechanisms, including, likely, alkaloids and changes on grass metabolome.     

Endophyte species influence the biomass production of the native grass Achnatherum sibiricum (L.) Keng under high nitrogen availability

Research on the interaction of endophytes and native grasses normally takes infection status into account, but less often considers the species of endophyte involved in the interaction. Here, we examined the effect of endophyte infection, endophyte species, nitrogen availability, and plant maternal genotype on the performance of a wild grass, Achnatherum sibiricum. Six different Epichloë‐infected maternal lines of A. sibiricum were used in the study; three lines harbored Epichloë gansuensis (Eg), while three lines harbored Epichloë sibirica (Es). These endophytes are vertically transmitted, while Eg also occasionally produces stromata on host tillers. We experimentally removed the endophyte from some ramets of the six lines, with the infected (E+) and uninfected (E?) plants grown under varying levels of nitrogen availability. Eg hosts produced more aboveground biomass than Es hosts only under high nitrogen supply. Endophyte species did not show any influence on the maximum net photosynthetic rate (P_max), photosynthetic nitrogen use efficiency, or total phenolics of A. sibiricum under all nitrogen conditions. However, the plant maternal genotype did influence the P_max and shoot biomass of A. sibiricum. Our results show that endophyte species influenced the shoot biomass of A. sibiricum, and this effect was dependent on nitrogen supply. As with most coevolutionary interactions, A. sibiricum that harbored Eg and Es may show pronounced geographic variation in natural habitats with increased nitrogen deposition. In addition, stroma‐bearing endophyte (Eg) provides positive effects (e.g., higher biomass production) to A. sibiricum plants during the vegetative growth stage.     

Endophytes inconsistently affect plant communities across Schedonorus arundinaceus hosts

Fungal endophytes in cool-season grasses may affect communities at multiple trophic levels. However, it is unclear whether community-scale endophyte effects arise due to the endophyte itself or as a result of unique, endophyte–host interactions. We used a long-term field experiment to test whether common-toxic (CT) and non-ergot alkaloid-producing (novel) endophytes in Schedonorus arundinaceus (tall fescue) forage cultivars consistently affect communities across tall fescue hosts. Tilled plots (2 × 2 m; Guelph, ON) were seeded with Georgia 5 and Jesup cultivars containing either the CT or AR542 (novel) endophyte and allowed to be re-colonized by plant species from the local propagule pool. Non-seeded control plots were included to assess effects of seeding the non-native grass. We assessed plant, invertebrate, soil moisture, and soil nutrient responses to the endophyte–cultivar treatments after four growing seasons. Seeding tall fescue affected plant species abundances, but not richness, and did not consistently alter soil moisture and nutrient pools. Endophyte identity in the tall fescue cultivars affected the communities, but effects were not consistent between cultivars. Within Georgia 5, the AR542 endophyte reduced tall fescue abundance and altered the invertebrate community relative to CT plots. Within Jesup, the AR542 endophyte reduced species evenness and decreased soil moisture during dry periods relative to CT plots. Endophyte effects were not consistent between cultivars, and it is probable that the community-scale effects of endophyte infection in tall fescue cultivars arise due to unique interactions between cultivar and endophyte.     

Volatiles in perennial ryegrass infected with strains of endophytic fungus: impact on African black beetle host selection

Perennial ryegrass (Lolium perenne) is often infected with the fungal‐endophyte Neotyphodium lolii. In addition to the ‘wild‐type’ strain (E^WT), several ‘selected’ strains of N. lolii are now being marketed as AR1 (E^AR1) and AR37 (E^AR37). Each of these strains impact positively on L. perenne's resistance against many insects, including the African black beetles (Heteronychus arator). The impact of volatile oils produced specifically by each strain in the endophyte–grass association in enhancing the grass's resistance to insects is still largely unknown. Keeping these in view, we determined the volatile oil profiles produced by L. perenne infected with either E^WT or E^AR1 or E^AR37 and determined the impacts of these volatiles on the host‐selection behaviour of H. arator adults. In the absence of endophyte infection (E^–), L. perenne produced 18 different volatile oils. In L. perenne E^WT, quantities of 2‐ethyl‐1‐hexanol acetate (Rt = 14.5 min), (Z)‐2‐octen‐1‐ol (Rt = 22.2 min), and butylated hydroxyl toluene (Rt = 23.2 min) were 24, 16 and 26%, respectively, greater than L. perenne E^–. The strains E^AR1 and E^AR37 affected differently the quantities of the volatile compounds but not their identity. In the four‐choice bioassay, males and females of H. arator were equally attracted to each strain. In Y‐tube olfactometer, compared against E^–, H. arator adults were less attracted to L. perenne E^WT and E^AR1. The attractiveness of E^AR37 was similar in effect to E^– to H. arator. The results indicate that each strain of N. lolii alters the profile of volatile oils in L. perenne differently and that alteration can influence H. arator adult‐host selection.     

Climate change and Epichloë coenophiala association modify belowground fungal symbioses of tall fescue host

Human alteration of symbiont genetics among aboveground endophytic Epichloë coenophiala strains within tall fescue (Schedonorus arundinaceus) has led to widespread deployment of novel grass-endophyte combinations, yet little is known about their ecological consequences. In this study, clone pairs (endophyte-infected, endophyte-free) of two tall fescue genotypes received factorial combinations of increased temperature (+3 °C) and precipitation (+30% long-term annual mean) for 2 yr. We measured root arbuscular mycorrhizal fungi (AMF), dark septate endophyte (DSE) colonization, and soil AMF extraradical hyphae (ERH) length. We hypothesized that genetically distinct grass-E. coenophiala associations would differentially affect belowground fungi, and that these relationships would be climate-sensitive. Tall fescue genotype, endophyte presence, and climate treatment interactions affected AMF arbuscules, vesicles, and ERH. DSE decreased with E. coenophiala presence but increased with warming. Genetically distinct tall fescue-E. coenophiala associations may have divergent long-term impacts on other host-symbiont interactions, potentially impacting ecosystem function and response to climate change.     

The effects of carbohydrate supply and host genetic background on Epichloë endophyte and alkaloid concentrations in perennial ryegrass

The ecological effects of novel grass–endophyte associations used in agriculture have not been widely studied. Previous studies of asexual Epichloë-infected Lolium perenne suggest that endophyte concentration is altered in high sugar grasses (HSGs) selectively bred to produce higher concentrations of water-soluble carbohydrates relative to conventional cultivars. We investigated whether differences are due to the effects of altered carbohydrates, or genetic background, by growing multiple cultivars in both high-sugar trait expression and non-expression conditions (using light/temperature treatments). Endophyte and alkaloid concentrations were measured in three HSG and three NSG (normal-sugar grass) cultivars infected with Lp19 or AR37 endophyte strains. Low molecular weight (LMW) carbohydrates had a small effect, explaining <6% of the variation in endophyte concentration. Endophyte concentrations were strongly dependent on plant genotype and fungal strain, with the highest concentrations seen in Lp19, suggesting that the interaction is highly dependent on genetic compatibility. Changes in endophyte concentration due to altered environmental variables and genetic compatibility may have consequences for persistence, toxicity, and invasive potential of endophyte-infected plants.     

Role of foliar fungal endophytes in litter decomposition among species and population origins

Litter decomposition is a key ecosystem process which returns nutrients from dead plant material to mineral forms in the soil. We examined whether systemic fungal endophytes modulate recycling of nutrients directly by altering litter decomposition. We studied litter decomposition mediated by Epichloë endophytes in litter-bag experiments. We examined direct endophyte effects on litter decomposition in wild populations and cultivars of Schedonorus phoenix and Schedonorus pratensis. In the first experiment, endophyte presence tended to increase litter decomposition rate in cultivars of the two grass species (S. phoenix and S. pratensis). However, in the second experiment plant origin had a stronger influence than endophyte symbiosis in S. phoenix. Interestingly, the initial level of alkaloids was associated positively with decomposition in S. phoenix populations. Characteristics associated with litter quality were not clearly related to either endophytes or decomposition rate. Our results suggest that endophytes can enhance litter breakdown but their role in nutrient cycling is far more complex depending on plant population origin.     

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.     

Restoration of Native Warm Season Grassland Species in a Tall Fescue Pasture Using Prescribed Fire and Herbicides

Pastures dominated by tall fescue (Schedonorus phoenix (Scop.) Holub) cover much of the eastern United States, and there are increasing efforts to restore native grassland plant species to some of these areas. Prescribed fire and herbicide are frequently used to limit the growth of tall fescue and other non‐natives, while encouraging native grasses and forbs. A fungal endophyte, commonly present in tall fescue, can confer competitive advantages to the host plant, and may play a role in determining the ability of tall fescue plants to persist in pastures following restoration practices. We compared vegetation composition among four actively restored subunits of a tall fescue pasture (each receiving different combinations of prescribed fire and/or herbicide) and a control. We also measured the rate of endophyte infection in tall fescue present within each restoration treatment and control to determine if restoration resulted in lower tall fescue cover but higher endophyte infection rates (i.e. selected for endophyte‐infected individuals). Tall fescue cover was low in all restoration treatments and the control (1.1–17.9%). The control (unmanaged) had higher species richness than restoration treatments and plant community composition was indicative of succession to forest. Restoration practices resulted in higher cover of native warm season grasses, but in some cases also promoted a different undesirable species. We found no evidence of higher fungal endophyte presence in tall fescue following restoration, as all subunits had low endophyte infection rates (2.2–9.3%). Restoration of tall fescue systems using prescribed fire and herbicide may be used to promote native grassland species.     

Postglacial colonization history reflects in the genetic structure of natural populations of Festuca rubra in Europe

We conducted a large‐scale population genetic survey of genetic diversity of the host grass Festuca rubra s.l., which fitness can be highly dependent on its symbiotic fungus Epichloë festucae, to evaluate genetic variation and population structure across the European range. The 27 studied populations have previously been found to differ in frequencies of occurrence of the symbiotic fungus E. festucae and ploidy levels. As predicted, we found decreased genetic diversity in previously glaciated areas in comparison with nonglaciated regions and discovered three major maternal genetic groups: southern, northeastern, and northwestern Europe. Interestingly, host populations from Greenland were genetically similar to those from the Faroe Islands and Iceland, suggesting gene flow also between those areas. The level of variation among populations within regions is evidently highly dependent on the postglacial colonization history, in particular on the number of independent long‐distance seed colonization events. Yet, also anthropogenic effects may have affected the population structure in F. rubra. We did not observe higher fungal infection rates in grass populations with lower levels of genetic variability. In fact, the fungal infection rates of E. festucae in relation to genetic variability of the host populations varied widely among geographical areas, which indicate differences in population histories due to colonization events and possible costs of systemic fungi in harsh environmental conditions. We found that the plants of different ploidy levels are genetically closely related within geographic areas indicating independent formation of polyploids in different maternal lineages.     

Soil microbial and chemical responses to foliar Epichloë fungal infection in Lolium perenne,Hordeum brevisubulatum and Achnatherum inebrians

We tested whether the host species identity in grass-Epichloë symbioses affected soil chemical and microbial properties. We grew endophyte infected (E+) and endophyte free (E−) Lolium perenne, Hordeum brevisubulatum and Achnatherum inebrians for 18 months in field plots. In E+ soil of all three grasses, available phosphorus was lower whereas total soil nitrogen was higher. Endophyte effects on soil pH, microbial biomass nitrogen, total carbon and organic carbon as well as bacteria and fungi abundance were host species dependent. Ammonia oxidizing bacteria abundance was higher in E+ soils for all species. Bacterial community composition of E+ and E− soils were different only for Lolium perenne with soil pH being the key factor. Fungal community composition of E+ and E− soils was not different for the three grasses. This study confirmed that the effects of foliar Epichloë infection on belowground properties depended on host species identity.     

Epichloë endophyte affects the ability of powdery mildew (Blumeria graminis) to colonise drunken horse grass (Achnatherum inebrians)

The effect of the systemic seed-borne endophyte Epichloë gansuensis on the colonization by Blumeria graminis, the cause of powdery mildew disease, and the growth of the host grass Achnatherum inebrians, was studied under four soil water conditions. Infection incidence, disease lesion parameters, disease index, biomass production and growth parameters of the grass with and without the fungal endophyte were measured and counted after a period of disease. There was a significantly (P < 0.05) higher disease incidence and disease index for endophyte-free (E−) compared to endophyte-infected (E+) plants under different drought stresses. The presence of the endophyte significantly positively affected all of the host grass growth factors. The results of the present study demonstrate that the presence of the Epichloë endophyte reduced the ability of B. graminis to colonise A. inebrians and also conferred enhanced host plant growth at all soil water conditions tested.     

Plant–fungal symbiosis affects litter decomposition during primary succession

Microbial symbionts of plants can affect decomposition by altering the quality or quantity of host plant tissue (substrate) or the micro‐environment where decomposition occurs (conditioning). In C₃ grasses, foliar fungal endophytes (Clavicipitaceae) can increase plant resistance to drought and/or produce alkaloids that reduce herbivory – effects that may also influence host litter composition and subsequent litter decomposition. We studied the effect of the endophyte Epichloë sp. on litter decomposition in the Great Lakes dunes (USA) using a reciprocal design altering endophyte presence/absence in both American beachgrass Ammophila breviligulata substrate (litter bags) and its conditioning of the decomposition microenvironment. Symbiont treatments were crossed with rain‐out shelters that altered growing season precipitation. The first year of decomposition, senesced leaf substrate from A. breviligulata with Epichloë decomposed 21% faster than endophyte‐free substrate. By the third year, conditioning by live symbiotic plants reduced cumulative decomposition by 33% compared to plots planted with endophyte‐free plants. Of the traits we examined – litter quantity, C:N ratio, mineral composition, fungal colonization, and carbon chemistry – increased litter quantity via greater tiller production was the primary trait shift associated with endophyte symbiosis. Epichloë in A. breviligulata litter also altered litter nitrogen decomposition dynamics, as evidenced by lower nitrogen and protein content in decomposed tissue from plants that hosted the endophyte. Differences in initial litter quality and subsequent colonization by saprotrophic fungi were ruled out as key drivers. Altered precipitation had negligible effects on decomposing processes in the dunes. Grass–Epichloë symbiosis altered nutrient cycling through increasing the rate of litter decomposition when present in the litter and through reducing litter decomposition by conditioning the decomposition microenvironment. Epichloë are widespread symbionts of grasses. Thus, their effects on decomposition could be an important, but often overlooked, driver of nutrient cycling in grass‐dominated ecosystems.     

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.     

An ecological framework for understanding the roles of Epichloë endophytes on plant defenses against fungal diseases

Plants harbor a wide diversity of microorganisms in their tissues. Some of them have a long co-evolutionary history with their hosts, likely playing a pivotal role in regulating the plant interaction with other microbes such as pathogens. Some cool-season grasses are symbiotic with Epichloë fungal endophytes that grow symptomless and systemically in aboveground tissues. Among the many benefits that have been ascribed to endophytes, their role in mediating plant interactions with pathogens has been scarcely developed. Here, we explored the effects of Epichloë fungal endophytes on the interaction of host grasses with fungal pathogens. We made a meta-analysis that covered a total of 18 host grass species, 11 fungal endophyte species, and 22 fungal pathogen species. We observed endophyte-mediated negative effects on pathogens in vitro and in planta. Endophyte negative effects on pathogens were apparent not only in laboratory but also in greenhouse and field experiments. Epichloë fungal endophytes had negative effects on pathogen growth and spores' germination. On living plants, endophytes reduced both severity and incidence of the disease as well as colonization and subsequent infection of seeds. Symbiosis with endophytes showed an inhibitory effect on debilitator and killer pathogens, but not on castrators, and this effect did not differ among biotrophic or necrotrophic lifestyles. We found that this protection can be direct through the production of fungistatic compounds, the competition for a common resource, or the induction of plant defenses, and indirect associated with endophyte-generated changes in the abiotic or the biotic environment. Several mechanisms operate simultaneously and contribute differentially to the reduction of disease within grass populations.