Vegetation type, not the legacy of warming, modifies the response of microbial functional genes and greenhouse gas fluxes to drought in Oro-Arctic and alpine regions.

Climate warming and summer droughts alter soil microbial activity, affecting greenhouse gas (GHG) emissions in Arctic and alpine regions. However, the long-term effects of warming, and implications for future microbial resilience, are poorly understood. Using one alpine and three Arctic soils subjected to in situ long-term experimental warming, we simulated drought in laboratory incubations to test how microbial functional-gene abundance affects fluxes in three GHGs: carbon dioxide, methane, and nitrous oxide. We found that responses of functional gene abundances to drought and warming are strongly associated with vegetation type and soil carbon. Our sites ranged from a wet, forb dominated, soil carbon-rich systems to a drier, soil carbon-poor alpine site. Resilience of functional gene abundances, and in turn methane and carbon dioxide fluxes, was lower in the wetter, carbon-rich systems. However, we did not detect an effect of drought or warming on nitrous oxide fluxes. All gene-GHG relationships were modified by vegetation type, with stronger effects being observed in wetter, forb-rich soils. These results suggest that impacts of warming and drought on GHG emissions are linked to a complex set of microbial gene abundances and may be habitat-specific.

How does the pattern of root metabolites regulating beneficial microorganisms change with different grazing pressures?

Grazing disturbance can change the structure of plant rhizosphere microbial communities and thereby alter the feedback to promote plant growth or induce plant defenses. However, little is known about how such changes occur and vary under different grazing pressures or the roles of root metabolites in altering the composition of rhizosphere microbial communities. In this study, the effects of different grazing pressures on the composition of microbial communities were investigated, and the mechanisms by which different grazing pressures changed rhizosphere microbiomes were explored with metabolomics. Grazing changed composition, functions, and co-expression networks of microbial communities. Under light grazing (LG), some saprophytic fungi, such as Lentinus sp., Ramichloridium sp., Ascobolus sp. and Hyphoderma sp., were significantly enriched, whereas under heavy grazing (HG), potentially beneficial rhizobacteria, such as Stenotrophomonas sp., Microbacterium sp., and Lysobacter sp., were significantly enriched. The beneficial mycorrhizal fungus Schizothecium sp. was significantly enriched in both LG and HG. Moreover, all enriched beneficial microorganisms were positively correlated with root metabolites, including amino acids (AAs), short-chain organic acids (SCOAs), and alkaloids. This suggests that these significantly enriched rhizosphere microbial changes may be caused by these differential root metabolites. Under LG, it is inferred that root metabolites, especially AAs such as L-Histidine, may regulate specific saprophytic fungi to participate in material transformations and the energy cycle and promote plant growth. Furthermore, to help alleviate the stress of HG and improve plant defenses, it is inferred that the root system actively regulates the synthesis of these root metabolites such as AAs, SCOAs, and alkaloids under grazing interference, and then secretes them to promote the growth of some specific plant growth-promoting rhizobacteria and fungi. To summarize, grasses can regulate beneficial microorganisms by changing root metabolites composition, and the response strategies vary under different grazing pressure in typical grassland ecosystems.

DNA methylation mediates overgrazing-induced clonal transgenerational plasticity.

Overgrazing generally induces dwarfism in grassland plants, and these phenotypic traits could be transmitted to clonal offspring even when overgrazing is excluded. However, the dwarfism-transmitted mechanism remains largely unknown, despite generally thought to be enabled by epigenetic modification. To clarify the potential role of DNA methylation on clonal transgenerational effects, we conducted a greenhouse experiment with Leymus chinensis clonal offspring from different cattle/sheep overgrazing histories via the demethylating agent 5-azacytidine. The results showed that clonal offspring from overgrazed (by cattle or sheep) parents were dwarfed and the auxin content of leaves significantly decreased compared to offspring from no-grazed parents'. The 5-azaC application generally increased the auxin content and promoted the growth of overgrazed offspring while inhibited no-grazed offspring growth. Meanwhile, there were similar trends in the expression level of genes related to auxin-responsive target genes (ARF7, ARF19), and signal transduction gene (AZF2). These results suggest that DNA methylation leads to overgrazing-induced plant transgenerational dwarfism via inhibiting auxin signal pathway.

Grazing reduces plant sexual reproduction but increases asexual reproduction: A global meta-analysis.

Grazing affects grasslands worldwide. However, the global patterns and general mechanisms of how grazing affects plant reproductive traits are poorly understood, especially in the context of different climates and grazing duration. We conducted a meta-analysis of 114 independent grazing studies worldwide that measured plant reproductive traits in grasslands. The results showed that the number of tillers of plant increased under grazing. Grazing did not affect the number of reproductive branches of forbs, but significantly reduced the number of reproductive branches of grasses. Grazing increased the number of vegetative branches of all plants and reduced the proportion of reproductive branches. Grazing significantly reduced the number of flowers in forbs. Seed yield in the two plant functional groups was reduced compared with no-grazing. Under grazing, the sexual reproduction of grasses decreased much more substantially than that of forbs. This may be due to biomass allocation pattern of grasses under grazing (i.e., belowground versus aboveground). Under grazing, plants tended to adopt rapid, low-input asexual reproduction rather than long-term, high-risk sexual reproduction. This study represents the first large-scale evaluation of plant reproductive trait responses under grazing and demonstrates that grazing inhibits sexual reproduction and promotes asexual reproduction. The effect of grazing on plant sexual reproduction was influenced by grazing intensity, mean annual precipitation, and grazing duration. These results will assist in the development of sustainable grazing management strategies to improve the balance between human welfare and grassland ecosystem health.

Plant roots send metabolic signals to microbes in response to long-term overgrazing.

Overgrazing directly and indirectly affects soil microorganisms, which can have feedback effects on plant growth. Little is known about the root metabolites plants produce and whether they recruit beneficial microbes in response to overgrazing. Here, we used the dominant grassland species Leymus chinensis to explore correlations between root metabolites and the rhizosphere microbiome shaped by long-term overgrazing, which was determined by using LC-MS technology and high-throughput sequencing. In total, 839 metabolites were detected, with 41 significantly higher and 3 significantly lower in overgrazing versus grazing exclusion plots. The rhizosphere bacterial community was changed, but the fungal community was not altered. Moreover, 11 bacterial orders were found only in the overgrazed samples, and these showed close relationships to root metabolites and certain soil properties. Of these, Latescibacterales, B10-SB3A, and Nitrosococcales are known to be involved in growth promotion, C and N metabolism, respectively. In addition, root metabolites play an important role in mediating root-fungi interactions. The beneficial fungal orders Agaricales and Sordariales have a tread to be higher maybe due to root metabolites, mainly facilitate nutrient absorption and protect organic carbon in the soil, respectively. Our results indicate that grassland plants send metabolic signals to recruit key beneficial bacteria and stabilize fungal communities to alleviate grazing-induced stress in typical grassland ecosystems.

Growth-Defense Trade-Offs Induced by Long-term Overgrazing Could Act as a Stress Memory.

Long-term overgrazing (OG) is one of the key drivers of global grassland degradation with severe loss of productivity and ecosystem functions, which may result in stress memory such as smaller stature of grassland plants. However, how the OG-induced stress memory could be regulated by phytohormones is unknown. In this study, we investigated the changes of four phytohormones of cloned offspring of Leymus chinensis that were developed from no-grazing (NG) plants and OG plants with a grazing history of 30 years. The concentrations of auxin (IAA) and gibberellic acid (GA) in OG plant leaves were 45% and 20% lower than control, respectively. Meanwhile, the level of abscisic acid (ABA) in OG leaves nearly doubled compared with that in NG leaves. The situation was quite similar in roots. Unexpectedly, no significant changes in the jasmonic acid (JA) level were observed between OG and NG plants. The changes in gene expression patterns between OG and NG plants were also investigated by transcriptomic analysis. In total, 302 differentially expressed genes (DEGs) were identified between OG and NG plants, which were mainly classified into the functions of synthesis, receptor, and signal transduction processes of phytohormones. The expression of 24 key genes related to the biosynthesis and signal transduction of IAA and GA was downregulated in OG plants. Among them, OASA1 and AO1 (regulating the biosynthesis of IAA and ABA, respectively) were reduced significantly by 88 and 92%, respectively. In addition, the content of secondary metabolites related to plant defense such as flavonoids and phenols was also increased in leaves. Taken together, the decrease of positive plant growth-related hormones (IAA and GA) together with the increase of plant stress-related hormones or factors (ABA, flavonoids, and phenols) induced the growth-defense trade-offs for L. chinensis adaptation to long-term OG stress. The findings reported in this study shed new light on the mechanism of plant-animal interaction in the grassland ecosystem and provide a deeper insight into optimizing grazing management and sustainable utilization of grassland.

Do soil depth and plant community composition interact to modify the resistance and resilience of grassland ecosystem functioning to drought?

While the effect of drought on plant communities and their associated ecosystem functions is well studied, little research has considered how responses are modified by soil depth and depth heterogeneity. We conducted a mesocosm study comprising shallow and deep soils, and variable and uniform soil depths, and two levels of plant community composition, and exposed them to a simulated drought to test for interactive effects of these treatments on the resilience of carbon dioxide fluxes, plant functional traits, and soil chemical properties. We tested the hypotheses that: (a) shallow and variable depth soils lead to increased resistance and resilience of ecosystem functions to drought due to more exploitative plant trait strategies; (b) plant communities associated with intensively managed high fertility soils, will have more exploitative root traits than extensively managed, lower fertility plant communities. These traits will be associated with higher resistance and resilience to drought and may interact with soil depth and depth heterogeneity to amplify the effects on ecosystem functions. Our results showed that while there were strong soil depth/heterogeneity effects on plant-driven carbon fluxes, it did not affect resistance or resilience to drought, and there were no treatment effects on plant-available carbon or nitrogen. We did observe a significant increase in exploitative root traits in shallow and variable soils relative to deep and uniform, which may have resulted in a compensation effect which led to the similar drought responses. Plant community compositions representative of intensive management were more drought resilient than more diverse "extensive" communities irrespective of soil depth or soil depth heterogeneity. In intensively managed plant communities, root traits were more representative of exploitative strategies. Taken together, our results suggest that reorganization of root traits in response to soil depth could buffer drought effects on ecosystem functions.

Drought soil legacy overrides maternal effects on plant growth.

Maternal effects (i.e. trans-generational plasticity) and soil legacies generated by drought and plant diversity can affect plant performance and alter nutrient cycling and plant community dynamics. However, the relative importance and combined effects of these factors on plant growth dynamics remain poorly understood.We used soil and seeds from an existing plant diversity and drought manipulation field experiment in temperate grassland to test maternal, soil drought and diversity legacy effects, and their interactions, on offspring plant performance of two grassland species (Alopecurus pratensis and Holcus lanatus) under contrasting glasshouse conditions.Our results showed that drought soil legacy effects eclipsed maternal effects on plant biomass. Drought soil legacy effects were attributed to changes in both abiotic (i.e. nutrient availability) and biotic soil properties (i.e. microbial carbon and enzyme activity), as well as plant root and shoot atom 15N excess. Further, plant tissue nutrient concentrations and soil microbial C:N responses to drought legacies varied between the two plant species and soils from high and low plant diversity treatments. However, these diversity effects did not affect plant root or shoot biomass.These findings demonstrate that while maternal effects resulting from drought occur in grasslands, their impacts on plant performance are likely minor relative to drought legacy effects on soil abiotic and biotic properties. This suggests that soil drought legacy effects could become increasingly important drivers of plant community dynamics and ecosystem functioning as extreme weather events become more frequent and intense with climate change. A plain language summary is available for this article.

Soil multifunctionality and drought resistance are determined by plant structural traits in restoring grassland.

It is increasingly recognized that belowground responses to vegetation change are closely linked to plant functional traits. However, our understanding is limited concerning the relative importance of different plant traits for soil functions and of the mechanisms by which traits influence soil properties in the real world. Here we test the hypothesis that taller species, or those with complex rooting structures, are associated with high rates of nutrient and carbon (C) cycling in grassland. We further hypothesized that communities dominated by species with deeper roots may be more resilient to drought. These hypotheses were tested in a 3-yr grassland restoration experiment on degraded ex-arable land in southern England. We sowed three trait-based plant functional groups, assembled using database derived values of plant traits, and their combinations into bare soil. This formed a range of plant trait syndromes onto which we superimposed a simulated drought 2 yr after initial establishment. We found strong evidence that community weighted mean (CWM) of plant height is negatively associated with soil nitrogen cycling and availability and soil multifunctionality. We propose that this was due to an exploitative resource capture strategy that was inappropriate in shallow chalk soils. Further, complexity of root architecture was positively related to soil multifunctionality throughout the season, with fine fibrous roots being associated with greater rates of nutrient cycling. Drought resistance of soil functions including ecosystem respiration, mineralization, and nitrification were positively related to functional divergence of rooting depth, indicating that, in shallow chalk soils, a range of water capture strategies is necessary to maintain functions. Finally, after 3 yr of the experiment, we did not detect any links between the plant traits and microbial communities, supporting the finding that traits based on plant structure and resource foraging capacity are the main variables driving soil function in the early years of grassland conversion. We suggest that screening recently restored grassland communities for potential soil multifunctionality and drought resilience may be possible based on rooting architecture and plant height. These results indicate that informed assembly of plant communities based on plant traits could aid in the restoration of functioning in degraded soil.

Drought reduces floral resources for pollinators.

Climate change is predicted to result in increased occurrence and intensity of drought in many regions worldwide. By increasing plant physiological stress, drought is likely to affect the floral resources (flowers, nectar and pollen) that are available to pollinators. However, little is known about impacts of drought at the community level, nor whether plant community functional composition influences these impacts. To address these knowledge gaps, we investigated the impacts of drought on floral resources in calcareous grassland. Drought was simulated using rain shelters and the impacts were explored at multiple scales and on four different experimental plant communities varying in functional trait composition. First, we investigated the effects of drought on nectar production of three common wildflower species (Lathyrus pratensis, Onobrychis viciifolia and Prunella vulgaris). In the drought treatment, L. pratensis and P. vulgaris had a lower proportion of flowers containing nectar and O. viciifolia had fewer flowers per raceme. Second, we measured the effects of drought on the diversity and abundance of floral resources across plant communities. Drought reduced the abundance of floral units for all plant communities, irrespective of functional composition, and reduced floral species richness for two of the communities. Functional diversity did not confer greater resistance to drought in terms of maintaining floral resources, probably because the effects of drought were ubiquitous across component plant communities. The findings indicate that drought has a substantial impact on the availability of floral resources in calcareous grassland, which will have consequences for pollinator behaviour and populations.

Plant attributes explain the distribution of soil microbial communities in two contrasting regions of the globe.

We lack strong empirical evidence for links between plant attributes (plant community attributes and functional traits) and the distribution of soil microbial communities at large spatial scales. Using datasets from two contrasting regions and ecosystem types in Australia and England, we report that aboveground plant community attributes, such as diversity (species richness) and cover, and functional traits can predict a unique portion of the variation in the diversity (number of phylotypes) and community composition of soil bacteria and fungi that cannot be explained by soil abiotic properties and climate. We further identify the relative importance and evaluate the potential direct and indirect effects of climate, soil properties and plant attributes in regulating the diversity and community composition of soil microbial communities. Finally, we deliver a list of examples of common taxa from Australia and England that are strongly related to specific plant traits, such as specific leaf area index, leaf nitrogen and nitrogen fixation. Together, our work provides new evidence that plant attributes, especially plant functional traits, can predict the distribution of soil microbial communities at the regional scale and across two hemispheres.

Drought neutralises plant-soil feedback of two mesic grassland forbs.

Plant-soil feedbacks (PSFs) describe the effect of a plant species on soil properties, which affect the performance of future generations. Here we test the hypothesis that drought alters PSFs by reducing plant-microbe associations and nutrient uptake. We chose two grassland forb species, previously shown to respond differently to soil conditioning and drought, to test our hypothesis. We conditioned unsterilised grassland soil with one generation of each species, and left a third soil unconditioned. We grew a second generation consisting of each combination of plant species, soil, and drought in a full factorial design, and measured soil microbial community and nutrient availability. Scabiosa columbaria displayed negative PSF (smaller plants) under non-droughted conditions, but neutral under drought, suggesting that drought disrupts plant-soil interactions and can advantage the plant. Photosynthetic efficiency of S. columbaria was reduced under drought, but recovered on rewetting regardless of soil conditioning, indicating that PSFs do not impede resilience of this species. Sanguisorba minor showed positive PSFs (larger plants), probably due to an increase in soil N in conspecific soil, but neutral PSF under drought. PSF neutralisation appeared to occur through drought-induced change in the soil microbial community for this species. When S. minor was planted in conspecific soil, photosynthetic efficiency declined to almost zero, with no recovery following rewetting. We attributed this to increased demand for water through higher demand for nutrients with positive PSF. Here we show that drought neutralises PSFs of two grassland forbs, which could have implications for plant communities under climate change.

Root architecture governs plasticity in response to drought.

AIMS: Root characteristics are important for predicting plant and ecosystem responses to resource scarcity. Simple, categorical traits for roots could be broadly applied to ecosystem function and restoration experiments, but they need to be evaluated for their role and behaviour under various stresses, including water limitation. We hypothesised that more complex root architectures allow more plastic responses to limited water than do tap roots. METHODS: We carried out two greenhouse experiments: one with a range of grassland plant species; the other with only species of Asteraceae to test the responsiveness of root architectural classes to location of limited water in the soil column. Using trait screening techniques and X-ray tomography, we measured the plasticity of the roots in response to water location. RESULTS: Plasticity of root biomass was lowest in tap rooted species, while fibrous and rhizomatous roots allocated biomass preferentially to where the soil was wettest. X-ray tomography indicated that root morphology was least plastic in rhizomatous species. CONCLUSIONS: Our results provide a starting point to effective categorisation of plants in terms of rooting architecture that could aid in understanding drought tolerance of grassland species. They also demonstrate the utility of X-ray tomography in root analyses.

Fifty important research questions in microbial ecology.

Microbial ecology provides insights into the ecological and evolutionary dynamics of microbial communities underpinning every ecosystem on Earth. Microbial communities can now be investigated in unprecedented detail, although there is still a wealth of open questions to be tackled. Here we identify 50 research questions of fundamental importance to the science or application of microbial ecology, with the intention of summarising the field and bringing focus to new research avenues. Questions are categorised into seven themes: host-microbiome interactions; health and infectious diseases; human health and food security; microbial ecology in a changing world; environmental processes; functional diversity; and evolutionary processes. Many questions recognise that microbes provide an extraordinary array of functional diversity that can be harnessed to solve real-world problems. Our limited knowledge of spatial and temporal variation in microbial diversity and function is also reflected, as is the need to integrate micro- and macro-ecological concepts, and knowledge derived from studies with humans and other diverse organisms. Although not exhaustive, the questions presented are intended to stimulate discussion and provide focus for researchers, funders and policy makers, informing the future research agenda in microbial ecology.

Plant functional group composition modifies the effects of precipitation change on grassland ecosystem function.

Temperate grassland ecosystems face a future of precipitation change, which can alter community composition and ecosystem functions through reduced soil moisture and waterlogging. There is evidence that functionally diverse plant communities contain a wider range of water use and resource capture strategies, resulting in greater resistance of ecosystem function to precipitation change. To investigate this interaction between composition and precipitation change we performed a field experiment for three years in successional grassland in southern England. This consisted of two treatments. The first, precipitation change, simulated end of century predictions, and consisted of a summer drought phase alongside winter rainfall addition. The second, functional group identity, divided the plant community into three groups based on their functional traits- broadly described as perennials, caespitose grasses and annuals- and removed these groups in a factorial design. Ecosystem functions related to C, N and water cycling were measured regularly. Effects of functional groupidentity were apparent, with the dominant trend being that process rates were higher under control conditions where a range of perennial species were present. E.g. litter decomposition rates were significantly higher in plots containing several perennial species, the group with the highest average leaf N content. Process rates were also very strongly affected by the precipitation change treatmentwhen perennial plant species were dominant, but not where the community contained a high abundance of annual species and caespitose grasses. This contrasting response could be attributable to differing rooting patterns (shallower structures under annual plants, and deeper roots under perennials) and faster nutrient uptake in annuals compared to perennials. Our results indicate that precipitation change will have a smaller effect on key process rates in grasslandscontaining a range of perennial and annual species, and that maintaining the presence of key functional groups should be a crucial consideration in future grassland management.