The growth promotion in endophyte symbiotic plants does not penalise the resistance to herbivores and bacterial microbiota.

A trade-off between growth and defence against biotic stresses is common in plants. Fungal endophytes of the genus Epichloë may relieve this trade-off in their host grasses since they can simultaneously induce plant growth and produce antiherbivore alkaloids that circumvent the need for host defence. The Epichloë ability to decouple the growth-defence trade-off was evaluated by subjecting ryegrass with and without Epichloë endophytes to an exogenous treatment with gibberellin (GA) followed by a challenge with Rhopalosiphum padi aphids. In agreement with the endophyte-mediated trade-off decoupling hypothesis, the GA-derived promotion of plant growth increased the susceptibility to aphids in endophyte-free plants but did not affect the insect resistance in endophyte-symbiotic plants. In line with the unaltered insect resistance, the GA treatment did not reduce the concentration of Epichloë-derived alkaloids. The Epichloë mycelial biomass was transiently increased by the GA treatment but at the expense of hyphal integrity. The response of the phyllosphere bacterial microbiota to both GA treatment and Epichloë was also evaluated. Only Epichloë, and not the GA treatment, altered the composition of the phyllosphere microbiota and the abundance of certain bacterial taxa. Our findings clearly demonstrate that Epichloë does indeed relieve the plant growth-defence trade-off.

Anti-herbivory defences delivered by Epichloë fungal endophytes: a quantitative review of alkaloid concentration variation among hosts and plant parts.

BACKGROUND AND AIMS: In the subfamily Poöideae (Poaceae), certain grass species possess anti-herbivore alkaloids synthesized by fungal endophytes that belong to the genus Epichloë (Clavicipitaceae). The protective role of these symbiotic endophytes can vary, depending on alkaloid concentrations within specific plant-endophyte associations and plant parts. METHODS: We conducted a literature review to identify articles containing alkaloid concentration data for various plant parts in six important pasture species, Lolium arundinaceum, Lolium perenne, Lolium pratense, Lolium multiflorum|Lolium rigidum and Festuca rubra, associated with their common endophytes. We considered the alkaloids lolines (1-aminopyrrolizidines), peramine (pyrrolopyrazines), ergovaline (ergot alkaloids) and lolitrem B (indole-diterpenes). While all these alkaloids have shown bioactivity against insect herbivores, ergovaline and lolitrem B are harmful for mammals. KEY RESULTS: Loline alkaloid levels were higher in the perennial grasses L. pratense and L. arundinaceum compared to the annual species L. multiflorum and L. rigidum, and higher in reproductive tissues than in vegetative structures. This is probably due to the greater biomass accumulation in perennial species that can result in higher endophyte mycelial biomass. Peramine concentrations were higher in L. perenne than in L. arundinaceum and not affected by plant part. This can be attributed to the high within-plant mobility of peramine. Ergovaline and lolitrem B, both hydrophobic compounds, were associated with plant parts where fungal mycelium is usually present, and their concentrations were higher in plant reproductive tissues. Only loline alkaloid data were sufficient for below-ground tissue analyses and concentrations were lower than in above-ground parts. CONCLUSIONS: Our study provides a comprehensive synthesis of fungal alkaloid variation across host grasses and plant parts, essential for understanding the endophyte-conferred defence extent. The patterns can be understood by considering endophyte growth within the plant and alkaloid mobility. Our study identifies research gaps, including the limited documentation of alkaloid presence in roots and the need to investigate the influence of different environmental conditions.

Genetic control underlying the flowering-drought tolerance trade-off in the Antarctic plant Colobanthus quitensis.

Plants inhabiting environments with stressful conditions often exhibit a low number of flowers, which can be attributed to the energetic cost associated with reproduction. One of the most stressful environments for plants is the Antarctic continent, characterized by limited soil water availability and low temperatures. Induction of dehydrins like those from the COR gene family and auxin transcriptional response repressor genes (IAAs), which are involved in floral repression, has been described in response to water stress. Here, we investigated the relationship between the water deficit-induced stress response and the number of flowers in Colobanthus quitensis plants collected from populations along a latitudinal gradient. The expression levels of COR47 and IAA12 genes in response to water deficit were found to be associated with the number of flowers. The relationship was observed both in the field and growth chambers. Watering the plants in the growth chambers alleviated the stress and stimualted flowering, thereby eliminating the trade-off observed in the field. Our study provides a mechanistic understanding of the ecological constraints on plant reproduction along a water availability gradient. However, further experiments are needed to elucidate the primary role of water availability in regulating resource allocation to reproduction in plants inhibiting extreme environments.

To tolerate drought or resist aphids? A new challenge to plant science is on the horizon.

Aphids are important herbivorous insects that can cause significant crop damage, leading to yield reduction and economic loss. One avenue being explored to reduce aphid impacts is the development of aphid-resistant plants. Under projected climate scenarios, it is expected that plants will be exposed to greater biotic and abiotic stress, including increased herbivorous insect infestation and exposure to prolonged periods of environmental stress, particularly drought. In response to these projections, plant-aphid interactions under drought conditions have been a subject of growing interest; however, few studies have looked at the impact of drought stress on plant resistance to aphids despite the potential importance for plant breeding. Here, we examine the latest scientific advances regarding variation in plant resistance to aphids under drought, emphasizing underlying mechanisms and functional trade-offs and propose a conceptual model relating plant tolerance to drought with plant resistance to aphids.

Environmental interference of plant-microbe interactions.

Environmental stresses can compromise the interactions of plants with beneficial microbes. In the present review, experimental results showing that stresses negatively affect the abundance and/or functionality of plant beneficial microbes are summarized. It is proposed that the environmental interference of these plant-microbe interactions is explained by the stress-mediated induction of plant signalling pathways associated with defence hormones and reactive oxygen species. These plant responses are recognized to regulate beneficial microbes within plants. The direct negative effect of stresses on microbes may also contribute to the environmental regulation of these plant mutualisms. It is also posited that, in stress situations, beneficial microbes harbour mechanisms that contribute to maintain the mutualistic associations. Beneficial microbes produce effector proteins and increase the antioxidant levels in plants that counteract the detrimental effects of plant stress responses on them. In addition, they deliver specific stress-protective mechanisms that assist to their plant hosts to mitigate the negative effects of stresses. Our study contributes to understanding how environmental stresses affect plant-microbe interactions and highlights why beneficial microbes can still deliver benefits to plants in stressful environments.

Can Aphid Herbivory Induce Intergenerational Effects of Endophyte-conferred Resistance in Grasses?

Plants have evolved mechanisms to survive herbivory. One such mechanism is the induction of defences upon attack that can operate intergenerationally. Cool-season grasses (sub-family Pooideae) obtain defences via symbiosis with vertically transmitted fungal endophytes (genus Epichloë) and can also show inducible responses. However, it is unknown whether these herbivore-induced responses can have intergenerational effects. We hypothesized that herbivory by aphids on maternal plants induces the intergenerational accumulation of endophyte-derived defensive alkaloids and resistance intensification in the progeny. We subjected mother plants symbiotic or not with Epichloë occultans, a species known for its production of anti-insect alkaloids known as lolines, to the aphid Rhopalosiphum padi. Then, we evaluated the progeny of these plants in terms of loline alkaloid concentration, resistance level (through herbivore performance), and shoot biomass. Herbivory on mother plants did not increase the concentration of lolines in seeds but it tended to affect loline concentration in progeny plants. There was an overall herbivore-induced intergenerational effect increasing the endophyte-conferred defence and resistance. Symbiotic plants were more resistant to aphids and had higher shoot biomass than their non-symbiotic counterparts. Since maternal herbivory did not affect the loline concentrations in seeds, the greater resistance of the progeny could have resulted from an inherited mechanism of epigenetic regulation. It would be interesting to elucidate the origin of this regulation since it could come from the host or the fungal symbiont. Thus, endophyte-driven differential fitness between symbiotic and non-symbiotic plants might be higher as generations pass on in presence of herbivores.

Fungal endophytes can eliminate the plant growth-defence trade-off.

A trade-off between growth and defence functions is commonly observed in plants. We propose that the association of plants with Epichloë fungal endophytes may eliminate this trade-off. This would be a consequence of the double role of these endophytes in host plants: the stimulation of plant growth hormones (e.g. gibberellins) and the fungal production of antiherbivore alkaloids. We put forward a model that integrates this dual effect of endophytes on plant growth and defence and test its predictions by means of meta-analysis of published literature. Our results support the notion that the enhanced plant resistance promoted by endophytes does not compromise plant growth. The limits and ecological benefits of this endophyte-mediated lack of plant growth-defence trade-off are discussed.

Getting ready for the ozone battle: Vertically transmitted fungal endophytes have transgenerational positive effects in plants.

Ground-level ozone is a global air pollutant with high toxicity and represents a threat to plants and microorganisms. Although beneficial microorganisms can improve host performance, their role in connecting environmentally induced maternal plant phenotypes to progeny (transgenerational effects [TGE]) is unknown. We evaluated fungal endophyte-mediated consequences of maternal plant exposure to ozone on performance of the progeny under contrasting scenarios of the same factor (high and low) at two stages: seedling and young plant. With no variation in biomass, maternal ozone-induced oxidative damage in the progeny that was lower in endophyte-symbiotic plants. This correlated with an endophyte-mediated higher concentration of proline, a defence compound associated with stress control. Interestingly, ozone-induced TGE was not associated with reductions in plant survival. On the contrary, there was an overall positive effect on seedling survival in the presence of endophytes. The positive effect of maternal ozone increasing young plant survival was irrespective of symbiosis and only expressed under high ozone condition. Our study shows that hereditary microorganisms can modulate the capacity of plants to transgenerationally adjust progeny phenotype to atmospheric change.

Seed-borne fungal endophytes constrain reproductive success of host plants under ozone pollution.

Tropospheric ozone is among the global change factors that pose a threat to plants and microorganisms. Symbiotic microorganisms can assist plants to cope with stress, but their role in the tolerance of plants to ozone is poorly understood. Here, we subjected endophyte-symbiotic and non-symbiotic plants of Lolium multiflorum, an annual species widely distributed in temperate grasslands, to high and low (i.e., charcoal-filtered air) ozone levels at vegetative and reproductive phases. Exposure to high ozone reduced leaf photochemical efficiency and greenness in both symbiotic and non-symbiotic plants. However, ozone-induced oxidative damage at biochemical level (i.e., lipid peroxidation) was mostly detected in symbiotic plants. Ozone exposure at the vegetative phase did not affect the reproductive investment in seeds, indicating full recovery from stress. Ozone exposure at the reproductive phase reduced biomass and seed production only in symbiotic plants indicating a symbiont-associated cost. At low ozone, endophyte-symbiotic plants showed a steeper slope in the relationship between seed number and seed weight (i.e., a number-weight trade-off) compared to non-symbiotic plants. However, when plants were treated at the reproductive phase, ozone increased the imbalance between seed number and seed weight in both endophyte-symbiotic and non-symbiotic plants. Plants with endophytes at the reproductive stage produced fewer seeds, which were not compensated by increased seed weight. Thus, fungal mycelium growing within ovaries or ozone-induced antioxidant systems may result in costs that finally depress the fitness of plants. Despite ozone pollution could destabilize plant-endophyte mutualisms and render them dysfunctional, other endophyte-mediated benefits (e.g., resistance to herbivory, tolerance to drought) could over-compensate these losses and explain the high incidence of the symbiosis in nature.

Functional roles of microbial symbionts in plant cold tolerance.

In this review, we examine the functional roles of microbial symbionts in plant tolerance to cold and freezing stresses. The impacts of symbionts on antioxidant activity, hormonal signaling and host osmotic balance are described, including the effects of the bacterial endosymbionts Burkholderia, Pseudomonas and Azospirillum on photosynthesis and the accumulation of carbohydrates such as trehalose and raffinose that improve cell osmotic regulation and plasma membrane integrity. The influence of root fungal endophytes and arbuscular mycorrhizal fungi on plant physiology at low temperatures, for example their effects on nutrient acquisition and the accumulation of indole-3-acetic acid and antioxidants in tissues, are also reviewed. Meta-analyses are presented showing that aspects of plant performance (shoot biomass, relative water content, sugar and proline concentrations and Fv /Fm ) are enhanced in symbiotic plants at low (-1 to 15 °C), but not at high (20-26 °C), temperatures. We discuss the implications of microbial symbionts for plant performance at low and sub-zero temperatures in the natural environment and propose future directions for research into the effects of symbionts on the cold and freezing tolerances of plants, concluding that further studies should routinely incorporate symbiotic microbes in their experimental designs.

Ontogenetic and trans-generational dynamics of a vertically transmitted fungal symbiont in an annual host plant in ozone-polluted settings.

Tropospheric ozone is an abiotic stress of increasing importance in the context of global climate change. This greenhouse gas is a potent phytotoxic molecule with demonstrated negative effects on crop yield and natural ecosystems. Recently, oxidative stress has been proposed as a mechanism that could regulate the interaction between cool-season grasses and Epichloë endophytes. We hypothesized that exposure of Lolium multiflorum plants, hosting endophytes to an ozone-polluted environment at different ontogenetic phases, would impact the trans-generational dynamics of the vertically transmitted fungal symbiont. Here, we found that the ozone-induced stress on the mother plants did not affect the endophyte vertical transmission but it impaired the persistence of the fungus in the seed exposed to artificial ageing. Endophyte longevity in seed was reduced by exposure of the mother plant to ozone. Although ozone exposure did not influence either the endophyte mycelial concentration or their compound defences (loline alkaloids), a positive correlation was observed between host fitness and the concentration of endophyte-derived defence compounds. This suggests that fungal defences in grass seeds were not all produced in situ but remobilized from the vegetative tissues. Our study reveals ozone trans-generational effects on the persistence of a beneficial symbiont in a host grass.

Simulated folivory increases vertical transmission of fungal endophytes that deter herbivores and alter tolerance to herbivory in Poa autumnalis.

BACKGROUND AND AIMS: The processes that maintain variation in the prevalence of symbioses within host populations are not well understood. While the fitness benefits of symbiosis have clearly been shown to drive changes in symbiont prevalence, the rate of transmission has been less well studied. Many grasses host symbiotic fungi (Epichloë spp.), which can be transmitted vertically to seeds or horizontally via spores. These symbionts may protect plants against herbivores by producing alkaloids or by increasing tolerance to damage. Therefore, herbivory may be a key ecological factor that alters symbiont prevalence within host populations by affecting either symbiont benefits to host fitness or the symbiont transmission rate. Here, we addressed the following questions: Does symbiont presence modulate plant tolerance to herbivory? Does folivory increase symbiont vertical transmission to seeds or hyphal density in seedlings? Do plants with symbiont horizontal transmission have lower rates of vertical transmission than plants lacking horizontal transmission? METHODS: We studied the grass Poa autumnalis and its symbiotic fungi in the genus Epichloë. We measured plant fitness (survival, growth, reproduction) and symbiont transmission to seeds following simulated folivory in a 3-year common garden experiment and surveyed natural populations that varied in mode of symbiont transmission. KEY RESULTS: Poa autumnalis hosted two Epichloë taxa, an undescribed vertically transmitted Epichloë sp. PauTG-1 and E. typhina subsp. poae with both vertical and horizontal transmission. Simulated folivory reduced plant survival, but endophyte presence increased tolerance to damage and boosted fitness. Folivory increased vertical transmission and hyphal density within seedlings, suggesting induced protection for progeny of damaged plants. Across natural populations, the prevalence of vertical transmission did not correlate with symbiont prevalence or differ with mode of transmission. CONCLUSIONS: Herbivory not only mediated the reproductive fitness benefits of symbiosis, but also promoted symbiosis prevalence by increasing vertical transmission of the fungus to the next generation. Our results reveal a new mechanism by which herbivores could influence the prevalence of microbial symbionts in host populations.

The plant hormone salicylic acid interacts with the mechanism of anti-herbivory conferred by fungal endophytes in grasses.

The plant hormone salicylic acid (SA) is recognized as an effective defence against biotrophic pathogens, but its role as regulator of beneficial plant symbionts has received little attention. We studied the relationship between the SA hormone and leaf fungal endophytes on herbivore defences in symbiotic grasses. We hypothesize that the SA exposure suppresses the endophyte reducing the fungal-produced alkaloids. Because of the role that alkaloids play in anti-herbivore defences, any reduction in their production should make host plants more susceptible to herbivores. Lolium multiflorum plants symbiotic and nonsymbiotic with the endophyte Epichloë occultans were exposed to SA followed by a challenge with the aphid Rhopalosiphum padi. We measured the level of plant resistance to aphids, and the defences conferred by endophytes and host plants. Symbiotic plants had lower concentrations of SA than did the nonsymbiotic counterparts. Consistent with our prediction, the hormonal treatment reduced the concentration of loline alkaloids (i.e., N-formyllolines and N-acetylnorlolines) and consequently decreased the endophyte-conferred resistance against aphids. Our study highlights the importance of the interaction between the plant immune system and endophytes for the stability of the defensive mutualism. Our results indicate that the SA plays a critical role in regulating the endophyte-conferred resistance against herbivores.

Vertically transmitted symbionts as mechanisms of transgenerational effects.

PREMISE OF THE STUDY: A transgenerational effect occurs when a biotic or abiotic environmental factor acts on a parental individual and thereby affects the phenotype of progeny. Due to the importance of transgenerational effects for understanding plant ecology and evolution, their underlying mechanisms are of general interest. Here, we introduce the concept that inherited symbiotic microorganisms could act as mechanisms of transgenerational effects in plants. METHODS: We define the criteria required to demonstrate that transgenerational effects are microbially mediated and review evidence from the well-studied, vertically transmitted plant-fungal symbiosis (grass-Epichloë spp.) in support of such effects. We also propose a basic experimental design to test for the presence of adaptive transgenerational effects mediated by plant symbionts. KEY RESULTS: An increasingly large body of literature shows that vertically transmitted microorganisms are common in plants, with potential to affect the phenotypes and fitness of progeny. Transgenerational effects could occur via parental modification of symbiont presence/absence, symbiont load, symbiont products, symbiont genotype or species composition, or symbiont priming. Several of these mechanisms appear likely in the grass-Epichloë endophytic symbiosis, as there is variation in the proportion of the progeny that carries the fungus, as well as variation in concentrations of mycelia and secondary compounds (alkaloids and osmolytes) in the seed. CONCLUSIONS: Symbiont-mediated transgenerational effects could be common in plants and could play large roles in plant adaptation to changing environments, but definitive tests are needed. We hope our contribution will spark new lines of research on the transgenerational effects of vertically transmitted symbionts in plants.

Metabolism or behavior: explaining the performance of aphids on alkaloid-producing fungal endophytes in annual ryegrass (Lolium multiflorum).

Plant-herbivore interactions are often mediated by plant microorganisms, and the "defensive mutualism" of epichloid fungal endophytes of grasses is an example. These endophytes synthesize bioactive alkaloids that generally have detrimental effects on the performance of insect herbivores, but the underlying mechanisms are not well understood. Our objective was to determine whether changes in the physiology and/or behavior of aphids explain the changes in performance of insects feeding on endophytic plants. We studied the interaction between the aphid Rhopalosiphum padi and the annual ryegrass Lolium multiflorum symbiotic (E+) or not symbiotic (E-) with the fungus Epichloë occultans that can synthesize loline alkaloids. We hypothesized that aphids feeding on E+ plants have higher energetic demands for detoxification of fungal alkaloids, thereby negatively impacting the individual performance, population growth, and structure. Aphids growing on E+ plants had lower values in morphometric and functional variables of individual performance, displayed lower birth rate, smaller population size, and dramatic structural changes. However, aphids exhibited lower values of standard metabolic rate (SMR) on E+ plants, which suggests no high costs of detoxification. Behavioral variables during the first 8 h of feeding showed that aphids did not change the phloem sap ingestion with the presence of fungal endophytes. We hypothesize that aphids may maintain phloem sap ingestion according to their fungal alkaloid tolerance capacity. In other words, when alkaloid concentrations overcome tolerance threshold, ingestion of phloem should decrease, which may explain the observed lower values of SMR in E+ feeding aphids.

Competing neighbors: light perception and root function.

Plant responses to competition have often been described as passive consequences of reduced resource availability. However, plants have mechanisms to forage for favorable conditions and anticipate competition scenarios. Despite the progresses made in understanding the role of light signaling in modulating plant-plant interactions, little is known about how plants use and integrate information gathered by their photoreceptors aboveground to regulate performance belowground. Given that the phytochrome family of photoreceptors plays a key role in the acquisition of information about the proximity of neighbors and canopy cover, it is tempting to speculate that changes in the red:far-red (R:FR) ratio perceived by aboveground plant parts have important implications shaping plant behavior belowground. Exploring data from published experiments, we assess the neglected role of light signaling in the control of root function. The available evidence indicates that plant exposure to low R:FR ratios affects root growth and morphology, root exudate profiles, and interactions with beneficial soil microorganisms. Although dependent on species identity, signals perceived aboveground are likely to affect root-to-root interactions. Root systems could also be guided to deploy new growth predominantly in open areas by light signals perceived by the shoots. Studying interactions between above- and belowground plant-plant signaling is expected to improve our understanding of the mechanisms of plant competition.

Chemical ecology mediated by fungal endophytes in grasses.

Defensive mutualism is widely accepted as providing the best framework for understanding how seed-transmitted, alkaloid producing fungal endophytes of grasses are maintained in many host populations. Here, we first briefly review current knowledge of bioactive alkaloids produced by systemic grass-endophytes. New findings suggest that chemotypic diversity of the endophyte-grass symbiotum is far more complex, involving multifaceted signaling and chemical cross-talk between endophyte and host cells (e.g., reactive oxygen species and antioxidants) or between plants, herbivores, and their natural enemies (e.g., volatile organic compounds, and salicylic acid and jasmonic acid pathways). Accumulating evidence also suggests that the tight relationship between the systemic endophyte and the host grass can lead to the loss of grass traits when the lost functions, such as plant defense to herbivores, are compensated for by an interactive endophytic fungal partner. Furthermore, chemotypic diversity of a symbiotum appears to depend on the endophyte and the host plant life histories, as well as on fungal and plant genotypes, abiotic and biotic environmental conditions, and their interactions. Thus, joint approaches of (bio)chemists, molecular biologists, plant physiologists, evolutionary biologists, and ecologists are urgently needed to fully understand the endophyte-grass symbiosis, its coevolutionary history, and ecological importance. We propose that endophyte-grass symbiosis provides an excellent model to study microbially mediated multirophic interactions from molecular mechanisms to ecology.

Global Change Factors Influence Plant-Epichloë Associations.

There is an increasing interest in determining the influence of global change on plant-microorganism interactions. We review the results of experiments that evaluated the effects of the global change factors carbon dioxide, ozone, temperature, drought, flooding, and salinity on plant symbioses with beneficial Epichloë endophytes. The factors affected the performance of both plants and endophytes as well as the frequency of plants symbiotic with the fungus. Elevated carbon dioxide levels and low temperatures differentially influenced the growth of plants and endophytes, which could compromise the symbioses. Furthermore, we summarise the plant stage in which the effects of the factors were quantified (vegetative, reproductive, or progeny). The factors ozone and drought were studied at all plant stages, but flooding and carbon dioxide were studied in just a few of them. While only studied in response to ozone and drought, evidence showed that the effects of these factors on symbiotic plants persisted trans-generationally. We also identified the putative mechanisms that would explain the effects of the factors on plant-endophyte associations. These mechanisms included the increased contents of reactive oxygen species and defence-related phytohormones, reduced photosynthesis, and altered levels of plant primary metabolites. Finally, we describe the counteracting mechanisms by which endophytes would mitigate the detrimental effects of the factors on plants. In presence of the factors, endophytes increased the contents of antioxidants, reduced the levels of defence-related phytohormones, and enhanced the plant uptake of nutrients and photosynthesis levels. Knowledge gaps regarding the effects of global change on plant-endophyte associations were identified and discussed.