Pioneer Arabidopsis thaliana spans the succession gradient revealing a diverse root-associated microbiome.

BACKGROUND: Soil microbiomes are increasingly acknowledged to affect plant functioning. Research in molecular model species Arabidopsis thaliana has given detailed insights of such plant-microbiome interactions. However, the circumstances under which natural A. thaliana plants have been studied so far might represent only a subset of A. thaliana's full ecological context and potential biotic diversity of its root-associated microbiome. RESULTS: We collected A. thaliana root-associated soils from a secondary succession gradient covering 40 years of land abandonment. All field sites were situated on the same parent soil material and in the same climatic region. By sequencing the bacterial and fungal communities and soil abiotic analysis we discovered differences in both the biotic and abiotic composition of the root-associated soil of A. thaliana and these differences are in accordance with the successional class of the field sites. As the studied sites all have been under (former) agricultural use, and a climatic cline is absent, we were able to reveal a more complete variety of ecological contexts A. thaliana can appear and sustain in. CONCLUSIONS: Our findings lead to the conclusion that although A. thaliana is considered a pioneer plant species and previously almost exclusively studied in early succession and disturbed sites, plants can successfully establish in soils which have experienced years of ecological development. Thereby, A. thaliana can be exposed to a much wider variation in soil ecological context than is currently presumed. This knowledge opens up new opportunities to enhance our understanding of causal plant-microbiome interactions as A. thaliana cannot only grow in contrasting soil biotic and abiotic conditions along a latitudinal gradient, but also when those conditions vary along a secondary succession gradient. Future research could give insights in important plant factors to grow in more ecologically complex later-secondary succession soils, which is an impending direction of our current agricultural systems.

Methane-Derived Carbon as a Driver for Cyanobacterial Growth.

Methane, a potent greenhouse gas produced in freshwater ecosystems, can be used by methane-oxidizing bacteria (MOB) and can therefore subsidize the pelagic food web with energy and carbon. Consortia of MOB and photoautotrophs have been described in aquatic ecosystems and MOB can benefit from photoautotrophs which produce oxygen, thereby enhancing CH4 oxidation. Methane oxidation can account for accumulation of inorganic carbon (i.e., CO2) and the release of exometabolites that may both be important factors influencing the structure of phytoplankton communities. The consortium of MOB and phototroph has been mainly studied for methane-removing biotechnologies, but there is still little information on the role of these interactions in freshwater ecosystems especially in the context of cyanobacterial growth and bloom development. We hypothesized that MOB could be an alternative C source to support cyanobacterial growth in freshwater systems. We detected low δ13C values in cyanobacterial blooms (the lowest detected value -59.97‰ for Planktothrix rubescens) what could be the result of the use of methane-derived carbon by cyanobacteria and/or MOB attached to their cells. We further proved the presence of metabolically active MOB on cyanobacterial filaments using the fluorescein isothiocyanate (FITC) based activity assay. The PCR results also proved the presence of the pmoA gene in several non-axenic cultures of cyanobacteria. Finally, experiments comprising the co-culture of the cyanobacterium Aphanizomenon gracile with the methanotroph Methylosinus sporium proved that cyanobacterial growth was significantly improved in the presence of MOB, presumably through utilizing CO2 released by MOB. On the other hand, 13C-CH4 labeled incubations showed the uptake and assimilation of MOB-derived metabolites by the cyanobacterium. We also observed a higher growth of MOB in the presence of cyanobacteria under a higher irradiance regime, then when grown alone, underpinning the bidirectional influence with as of yet unknown environmental consequences.

Adapted dandelions trade dispersal for germination upon root herbivore attack.

A plant's offspring may escape unfavourable local conditions through seed dispersal. Whether plants use this strategy to escape insect herbivores is not well understood. Here, we explore how different dandelion (Taraxacum officinale agg.) populations, including diploid outcrossers and triploid apomicts, modify seed dispersal in response to root herbivore attack by their main root-feeding natural enemy, the larvae of the common cockchafer Melolontha melolontha. In a manipulative field experiment, root herbivore attack increased seed dispersal potential through a reduction in seed weight in populations that evolved under high root herbivore pressure, but not in populations that evolved under low pressure. This increase in dispersal potential was independent of plant cytotype, but associated with a reduction in germination rate, suggesting that adapted dandelions trade dispersal for establishment upon attack by root herbivores. Analysis of vegetative growth parameters suggested that the increased dispersal capacity was not the result of stress flowering. In summary, these results suggest that root herbivory selects for an induced increase in dispersal ability in response to herbivore attack. Induced seed dispersal may be a strategy that allows adapted plants to escape from herbivores.

Species-specific plant-soil feedbacks alter herbivore-induced gene expression and defense chemistry in Plantago lanceolata.

Plants actively interact with antagonists and beneficial organisms occurring in the above- and belowground domains of terrestrial ecosystems. In the past decade, studies have focused on the role of plant-soil feedbacks (PSF) in a broad range of ecological processes. However, PSF and its legacy effects on plant defense traits, such as induction of defense-related genes and production of defensive secondary metabolites, have not received much attention. Here, we study soil legacy effects created by twelve common grassland plant species on the induction of four defense-related genes, involved in jasmonic acid signaling, related to chewing herbivore defense (LOX2, PPO7), and in salicylic acid signaling, related to pathogen defense (PR1 and PR2) in Plantago lanceolata in response to aboveground herbivory by Mamestra brassicae. We also assessed soil legacy and herbivory effects on the production of terpenoid defense compounds (the iridoid glycosides aucubin and catalpol) in P. lanceolata. Our results show that both soil legacy and herbivory influence phenotypes of P. lanceolata in terms of induction of Pl PPO7 and Pl LOX2, whereas the expression of Pl PR1 and Pl PR2-1 is not affected by soil legacies, nor by herbivory. We also find species-specific soil legacy effects on the production of aucubin. Moreover, P. lanceolata accumulates more catalpol when they are grown in soils conditioned by grass species. Our study highlights that PSF can influence aboveground plant-insect interactions through the impacts on plant defense traits and suggests that aboveground plant defense responses can be determined, at least partly, by plant-specific legacy effects induced by belowground organisms.

Seasonal and herbivore-induced dynamics of foliar glucosinolates in wild cabbage (Brassica oleracea).

Levels of plant secondary metabolites are not static and often change in relation to plant ontogeny. They also respond to abiotic and biotic changes in the environment, e.g., they often increase in response to biotic stress, such as herbivory. In contrast with short-lived annual plant species, especially those with growing periods of less than 2-3 months, investment in defensive compounds of vegetative tissues in biennial and perennial species may also vary over the course of an entire growing season. In garden experiments, we investigated the dynamics of secondary metabolites, i.e. glucosinolates (GSLs) in the perennial wild cabbage (Brassica oleracea), which was grown from seeds originating from three populations that differ in GSL chemistry. We compared temporal long-term dynamics of GSLs over the course of two growing seasons and short-term dynamics in response to herbivory by Pieris rapae caterpillars in a more controlled greenhouse experiment. Long-term dynamics differed for aliphatic GSLs (gradual increase from May to December) and indole GSLs (rapid increase until mid-summer after which concentrations decreased or stabilized). In spring, GSL levels in new shoots were similar to those found in the previous year. Short-term dynamics in response to herbivory primarily affected indole GSLs, which increased during the 2-week feeding period by P. rapae. Herbivore-induced changes in the concentrations of aliphatic GSLs were population-specific and their concentrations were found to increase in primarily one population only. We discuss our results considering the biology and ecology of wild cabbage.

Changing soil legacies to direct restoration of plant communities.

It is increasingly acknowledged that soil biota may influence interactions among plant species; however, little is known about how to change historical influences of previous land management on soil biota, the so-called 'biotic soil legacy effect'. We used a two-phase plant community-soil feedback approach to study how plant species typical to original (i.e. undisturbed) and degraded fen meadows may influence effects of the soil community on Carex species that are dominant in fen meadows. In phase 1, soil from original, degraded, successfully and unsuccessfully restored fen meadows was conditioned by growing plants typical to original or to degraded fen meadows. In phase 2, interactions between Carex and neighbouring plant species were studied to quantify plant community-soil feedback effects in different neighbour plant mixtures. Soil conditioning with plants typical to original fen meadows resulted in significantly more Carex biomass than with plants typical to degraded fen meadows. These effects were strongest when the soil originated from unsuccessfully restored fen meadows. However, biomass of plants typical of degraded fen meadows was also higher in soil conditioned by typical fen meadow plants. We conclude that soil legacy effects of plants from degraded fen meadows can be altered by growing typical fen meadow plant species in that soil, as this enhances priority effects that favour growth of other typical fen meadow plants. As also plant species from degraded fen meadows benefitted from soil conditioning, further studies are needed to reveal if plant species can be chosen that change negative soil legacy effects for rare and endangered fen meadow plant species, but not for plant species that are typical to degraded fen meadows.

Plant-Soil Feedback Effects on Growth, Defense and Susceptibility to a Soil-Borne Disease in a Cut Flower Crop: Species and Functional Group Effects.

Plants can influence the soil they grow in, and via these changes in the soil they can positively or negatively influence other plants that grow later in this soil, a phenomenon called plant-soil feedback. A fascinating possibility is then to apply positive plant-soil feedback effects in sustainable agriculture to promote plant growth and resistance to pathogens. We grew the cut flower chrysanthemum (Dendranthema X grandiflora) in sterile soil inoculated with soil collected from a grassland that was subsequently conditioned by 37 plant species of three functional groups (grass, forb, legume), and compared it to growth in 100% sterile soil (control). We tested the performance of chrysanthemum by measuring plant growth, and defense (leaf chlorogenic acid concentration) and susceptibility to the oomycete pathogen Pythium ultimum. In presence of Pythium, belowground biomass of chrysanthemum declined but aboveground biomass was not affected compared to non-Pythium inoculated plants. We observed strong differences among species and among functional groups in their plant-soil feedback effects on chrysanthemum. Soil inocula that were conditioned by grasses produced higher chrysanthemum above- and belowground biomass and less leaf yellowness than inocula conditioned by legumes or forbs. Chrysanthemum had lower root/shoot ratios in response to Pythium in soil conditioned by forbs than by grasses. Leaf chlorogenic acid concentrations increased in presence of Pythium and correlated positively with chrysanthemum aboveground biomass. Although chlorogenic acid differed between soil inocula, it did not differ between functional groups. There was no relationship between the phylogenetic distance of the conditioning plant species to chrysanthemum and their plant-soil feedback effects on chrysanthemum. Our study provides novel evidence that plant-soil feedback effects can influence crop health, and shows that plant-soil feedbacks, plant disease susceptibility, and plant aboveground defense compounds are tightly linked. Moreover, we highlight the relevance of considering plant-soil feedbacks in sustainable horticulture, and the larger role of grasses compared to legumes or forbs in this.

Effects of population-related variation in plant primary and secondary metabolites on aboveground and belowground multitrophic interactions.

Insects feeding on aboveground and belowground tissues can influence each other through their shared plant and this is often mediated by changes in plant chemistry. We examined the effects of belowground root fly (Delia radicum) herbivory on the performance of an aboveground herbivore (Plutella xylostella) and its endoparasitoid wasp (Cotesia vestalis). Insects were reared on three populations of wild cabbage (Brassica oleracea) plants, exhibiting qualitative and quantitative differences in root and shoot defense chemistry, that had or had not been exposed to root herbivory. In addition, we measured primary (amino acids and sugars) and secondary [glucosinolate (GS)] chemistry in plants exposed to the various plant population-treatment combinations to determine to what extent plant chemistry could explain variation in insect performance variables using multivariate statistics. In general, insect performance was more strongly affected by plant population than by herbivory in the opposite compartment, suggesting that population-related differences in plant quality are larger than those induced by herbivory. Sugar profiles were similar in the three populations and concentrations only changed in damaged tissues. In addition to population-related differences, amino acid concentrations primarily changed locally in response to herbivory. Whether GS concentrations changed in response to herbivory (indole GS) or whether there were only population-related differences (aliphatic GS) depended on GS class. Poor correlations between performance and chemical attributes made biological interpretation of these results difficult. Moreover, trade-offs between life history traits suggest that factors other than food nutritional quality contribute to the expression of life history traits.

Root-Lesion Nematodes Suppress Cabbage Aphid Population Development by Reducing Aphid Daily Reproduction.

Empirical studies have shown that belowground feeding herbivores can affect the performance of aboveground herbivores in different ways. Often the critical life-history parameters underlying the observed performance effects remain unexplored. In order to better understand the cause for the observed effects on aboveground herbivores, these ecological mechanisms must be better understood. In this study we combined empirical experiments with a modeling approach to analyze the effect of two root feeding endoparasitic nematodes with different feeding strategies on the population growth of the aboveground feeding specialist aphid Brevicoryne brassicae on Brassica nigra. The aim was to test whether emerging differences in life history characteristics (days until reproduction, daily reproduction) would be sufficient to explain observed differences in aphid population development on plants with and without two species of nematodes. Aphid numbers were lower on plants with Pratylenchus penetrans in comparison to aphid numbers on plants with Meloidogyne spp. A dedicated experiment showed that aphid daily reproduction was lower on plants with P. penetrans (3.08 offspring female(-1) day(-1)) in comparison to both uninfested plants and plants with Meloidogyne spp. (3.50 offspring female(-1) day(-1)). The species-specific reduction of aphid reproduction appeared independent of changes in amino acids, soluble sugars or the glucosinolate sinigrin in the phloem. An individual-based model revealed that relatively small differences in reproduction rate per female were sufficient to yield a similar difference in aphid populations as was found in the empirical experiments.

Interactions Between a Belowground Herbivore and Primary and Secondary Root Metabolites in Wild Cabbage.

Plants are attacked by both above- and belowground herbivores. Toxic secondary compounds are part of the chemical defense arsenal of plants against a range of antagonists, and are subject to genetic variation. Plants also produce primary metabolites (amino acids, nutrients, sugars) that function as essential compounds for growth and survival. Wild cabbage populations growing on the Dorset coast of the UK exhibit genetically different chemical defense profiles, even though they are located within a few kilometers of each other. As in other Brassicaceae, the defensive chemicals in wild cabbages constitute, among others, secondary metabolites called glucosinolates. Here, we used five Dorset populations of wild cabbage to study the effect of belowground herbivory by the cabbage root fly on primary and secondary chemistry, and whether differences in chemistry affected the performance of the belowground herbivore. There were significant differences in total root concentrations and chemical profiles of glucosinolates, amino acids, and sugars among the five wild cabbage populations. Glucosinolate concentrations not only differed among the populations, but also were affected by root fly herbivory. Amino acid and sugar concentrations also differed among the populations, but were not affected by root fly herbivory. Overall, population-related differences in plant chemistry were more pronounced for the glucosinolates than for amino acids and sugars. The performance of the root herbivore did not differ among the populations tested. Survival of the root fly was low (<40%), suggesting that other belowground factors may override potential differences in effects related to primary and secondary chemistry.

Plants know where it hurts: root and shoot jasmonic acid induction elicit differential responses in Brassica oleracea.

Plants respond to herbivore attack by rapidly inducing defenses that are mainly regulated by jasmonic acid (JA). Due to the systemic nature of induced defenses, attack by root herbivores can also result in a shoot response and vice versa, causing interactions between above- and belowground herbivores. However, little is known about the molecular mechanisms underlying these interactions. We investigated whether plants respond differently when roots or shoots are induced. We mimicked herbivore attack by applying JA to the roots or shoots of Brassica oleracea and analyzed molecular and chemical responses in both organs. In shoots, an immediate and massive change in primary and secondary metabolism was observed. In roots, the JA-induced response was less extensive and qualitatively different from that in the shoots. Strikingly, in both roots and shoots we also observed differential responses in primary metabolism, development as well as defense specific traits depending on whether the JA induction had been below- or aboveground. We conclude that the JA response is not only tissue-specific but also dependent on the organ that was induced. Already very early in the JA signaling pathway the differential response was observed. This indicates that both organs have a different JA signaling cascade, and that the signal eliciting systemic responses contains information about the site of induction, thus providing plants with a mechanism to tailor their responses specifically to the organ that is damaged.

Broccoli and turnip plants display contrasting responses to belowground induction by Delia radicum infestation and phytohormone applications.

Induced responses to insect herbivory are a common phenomenon in the plant kingdom. So far, induced responses have mostly investigated in aerial plant parts. Recently it was found that root herbivore may also elicit both local and systemic responses affecting aboveground herbivores and their natural enemies. Using broccoli (Brassica oleracea subsp. italica L.) and turnip (Brassica rapa subsp. rapa L.), two cultivated brassicaceaous plants differing in their chemistry and morphology, we analysed the local and systemic induced responses triggered by Delia radicum L. damage, JA and SA application. We also assessed whether the root induction treatments affected D. radicum larval performance. Both D. radicum damage and JA induced changes in glucosinolate and sugar content as well as affected D. radicum performance, while SA application did not. Despite the uniform chemical responses, the effect on larval performance on broccoli and turnip plants was very different. On broccoli, JA root treatment reduced herbivore performance, whereas in turnips the same treatment enhanced it. JA- and D. radicum-induced responses followed similar patterns, which suggests that the JA signalling pathway is involved in root-induced responses to larval feeding. Glucosinolate induction cannot fully explain the differences found in the performance of D. radicum on the different species. Changes in other resistance factors might significantly contribute to the induced resistance in these brassicaceaeous species as well.

Tri-trophic effects of inter- and intra-population variation in defence chemistry of wild cabbage (Brassica oleracea).

The effect of direct chemical defences in plants on the performance of insect herbivores and their natural enemies has received increasing attention over the past 10 years. However, much less is known about the scale at which this variation is generated and maintained, both within and across populations of the same plant species. This study compares growth and development of the large cabbage butterfly, Pieris brassicae, and its gregarious pupal parasitoid, Pteromalus puparum, on three wild populations [Kimmeridge (KIM), Old Harry (OH) and Winspit (WIN)] and two cultivars [Stonehead (ST), and Cyrus (CYR)] of cabbage, Brassica oleracea. The wild populations originate from the coast of Dorset, UK, but grow in close proximity with one another. Insect performance and chemical profiles were made from every plant used in the experiment. Foliar glucosinolates (GS) concentrations were highest in the wild plants in rank order WIN > OH > KIM, with lower levels found in the cultivars. Caterpillar-damaged leaves in the wild cabbages also had higher GS levels than undamaged leaves. Pupal mass in P. brassicae varied significantly among populations of B. oleracea. Moreover, development time in the host and parasitoid were correlated, even though these stages are temporally separated. Parasitoid adult dry mass closely approximated the development of its host. Multivariate statistics revealed a correlation between pupal mass and development time of P. brassicae and foliar GS chemistry, of which levels of neoglucobrassicin appeared to be the most important. Our results show that there is considerable variation in quantitative aspects of defensive chemistry in wild cabbage plants that is maintained at very small spatial scales in nature. Moreover, the performance of the herbivore and its parasitoid were both affected by differences in plant quality.

A heritable glucosinolate polymorphism within natural populations of Barbarea vulgaris.

In natural populations of Barbarea vulgaris we found two distinctly different glucosinolate profiles. The most common glucosinolate profile is dominated (94%) by the hydroxylated form, (S)-2-hydroxy-2-phenylethyl-glucosinolate (glucobarbarin, BAR-type), whereas in the other type 2-phenylethyl-glucosinolate (gluconasturtiin, NAS-type) was most prominent (82%). NAS-type plants have a 108-fold increase of gluconasturtiin concentration in rosette leaves compared to BAR-type plants. The glucosinolate composition of both chemotypes is consistent throughout all plant organs and after induction with jasmonic acid. Although the glucosinolate profile of the roots has a more diverse composition than other plant organs, it still matches the chemotype. In 12 natural populations that we sampled in Germany, Belgium, France and Switzerland solely BAR-type plants were found. However, eight out of the 15 Dutch populations that were sampled contained 2-22% NAS-type plants. Controlled crosses showed that the chemotype was heritable and determined by a single gene with two alleles. The allele coding for the BAR-type was dominant and the allele for the NAS-type was recessive. The different glucosinolate profiles will yield different hydrolysis products upon damage, and therefore we expect them to differentially affect the multitrophic interactions associated with B. vulgaris in their natural environment.

Soil invertebrate fauna enhances grassland succession and diversity.

One of the most important areas in ecology is to elucidate the factors that drive succession in ecosystems and thus influence the diversity of species in natural vegetation. Significant mechanisms in this process are known to be resource limitation and the effects of aboveground vertebrate herbivores. More recently, symbiotic and pathogenic soil microbes have been shown to exert a profound effect on the composition of vegetation and changes therein. However, the influence of invertebrate soil fauna on succession has so far received little attention. Here we report that invertebrate soil fauna might enhance both secondary succession and local plant species diversity. Soil fauna from a series of secondary grassland succession stages selectively suppress early successional dominant plant species, thereby enhancing the relative abundance of subordinate species and also that of species from later succession stages. Soil fauna from the mid-succession stage had the strongest effect. Our results clearly show that soil fauna strongly affects the composition of natural vegetation and we suggest that this knowledge might improve the restoration and conservation of plant species diversity.