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Many pathways of invasion have been posited, but ecologists lack an experimental framework to identify which mechanisms are dominant in a given invasion scenario. Plant-soil feedbacks (PSFs) are one such mechanism that tend to initially facilitate, but over time attenuate, invasive species' impacts on plant diversity and ecosystem function. PSFs are typically measured under greenhouse conditions and are often assumed to have significant effects under field conditions that change over time. However, direct tests of PSFs effects in natural settings and their change over time are rare. Here we compare the role of PSFs with the effects of biomass in limiting the dominance of an invasive species and impacts on resident species diversity. We characterized the effects of the invader Bromus inermis (Leyss.) on native plant communities over time and measured changes in its conspecific PSFs and vegetative growth to understand their integrated effects on community diversity. To do so, we combined data from a 6-year field study documenting the rate and impacts of invasion with a short-term greenhouse experiment quantifying PSF as a function of time since invasion in the field. We found that the nature and strength of B. inermis PSFs did not change over time and were not mediated by soil microbial communities. Though PSFs impacted B. inermis reproduction, they did not sufficiently limit vegetative growth to diminish the negative impacts of B. inermis biomass on native species. B. inermis experienced the full strength of its negative PSFs immediately upon invasion, but they were ineffective at reducing B. inermis vigor to facilitate the recovery of the native plant community. We recommend that conservation efforts focus on limiting B. inermis vegetative growth to facilitate community recovery., Competing Interests: None., (© 2024 The Author(s). Ecology and Evolution published by John Wiley & Sons Ltd.)

Fungi play vital regulatory roles in terrestrial ecosystems. Local community assembly mechanisms, including deterministic and stochastic processes, as well as the size of regional species pools (gamma diversity), typically influence overall soil microbial community beta diversity patterns. However, there is limited evidence supporting their direct and indirect effects on beta diversity of different soil fungal functional groups in forest ecosystems. To address this gap, we collected 1606 soil samples from a 25-ha subtropical forest plot in southern China. Our goal was to determine the direct effects and indirect effects of regional species pools on the beta diversity of soil fungi, specifically arbuscular mycorrhizal (AM), ectomycorrhizal (EcM), plant-pathogenic, and saprotrophic fungi. We quantified the effects of soil properties, mycorrhizal tree abundances, and topographical factors on soil fungal diversity. The beta diversity of plant-pathogenic fungi was predominantly influenced by the size of the species pool. In contrast, the beta diversity of EcM fungi was primarily driven indirectly through community assembly processes. Neither of them had significant effects on the beta diversity of AM and saprotrophic fungi. Our results highlight that the direct and indirect effects of species pools on the beta diversity of soil functional groups of fungi can significantly differ even within a relatively small area. They also demonstrate the independent and combined effects of various factors in regulating the diversities of soil functional groups of fungi. Consequently, it is crucial to study the fungal community not only as a whole but also by considering different functional groups within the community., (© 2024. The Author(s).)

Rationale: The sampling throughput of immediate drop-on-demand technology (I.DOT) coupled with an open port sampling interface (OPSI) is limited by software communication. To enable much-needed high-throughput mass spectrometry (MS) analysis capabilities, a novel software was developed that allows for flexible sample selection from a 96-well plate and for maximized analysis throughput using I.DOT/OPSI-MS coupling., Methods: Wells of a 96-well I.DOT plate were filled with propranolol solution and were used to test maximum sampling throughput strategies to minimize analysis time. Demonstration of chemical reaction monitoring was done using acid-catalyzed ring closure of 2,3-diaminonaphthalene (DAN) with nitrite to form 2,3-naphthotriazole (NAT). Analytes were detected in positive electrospray ionization mode using selected reaction monitoring., Results: A maximum throughput of 1.54 s/sample (7.41 min/96-well plate with three technical replicates) was achieved, and it was limited by the peak width of the MS signal resulting in an occasional slight overlap between the peaks. Relative standard deviation was 10 ± 1% with all tested sampling strategies. Chemical reaction monitoring of DAN to NAT using nitrite was successfully accomplished with 2 s/sample throughout showing almost complete transformation in 10 min with no signal overlap., Conclusions: This work illustrates the development of a noncontact, automated I.DOT/OPSI-MS system with improved throughput achieved through an optimized software interface. Its achievable analysis time and precision make it a viable approach for drug discovery and in situ reaction monitoring studies., (© 2023 John Wiley & Sons Ltd.)

Climate change is increasing the frequency and severity of short-term (~1 y) drought events-the most common duration of drought-globally. Yet the impact of this intensification of drought on ecosystem functioning remains poorly resolved. This is due in part to the widely disparate approaches ecologists have employed to study drought, variation in the severity and duration of drought studied, and differences among ecosystems in vegetation, edaphic and climatic attributes that can mediate drought impacts. To overcome these problems and better identify the factors that modulate drought responses, we used a coordinated distributed experiment to quantify the impact of short-term drought on grassland and shrubland ecosystems. With a standardized approach, we imposed ~a single year of drought at 100 sites on six continents. Here we show that loss of a foundational ecosystem function-aboveground net primary production (ANPP)-was 60% greater at sites that experienced statistically extreme drought (1-in-100-y event) vs. those sites where drought was nominal (historically more common) in magnitude (35% vs. 21%, respectively). This reduction in a key carbon cycle process with a single year of extreme drought greatly exceeds previously reported losses for grasslands and shrublands. Our global experiment also revealed high variability in drought response but that relative reductions in ANPP were greater in drier ecosystems and those with fewer plant species. Overall, our results demonstrate with unprecedented rigor that the global impacts of projected increases in drought severity have been significantly underestimated and that drier and less diverse sites are likely to be most vulnerable to extreme drought., Competing Interests: Competing interests statement:The authors declare no competing interest.

Root-centric studies have revealed fast taxonomic turnover across root neighborhoods, but how such turnover is accompanied by changes in species functions and phylogeny (i.e., β diversity) remains largely unknown. As β diversity can reflect the degree of community-wide biotic homogenization, such information is crucial for better inference of below-ground assembly rules, community structuring, and ecosystem processes. We collected 2480 root segments from 625 0-30 cm soil profiles in a subtropical forest in China. Root segments were identified into 138 species with DNA-barcoding with six root morphological and architectural traits measured per species. By using the mean pairwise (D pw ) and mean nearest neighbor distance (D nn ) to quantify species ecological differences, we first tested the non-random functional and phylogenetic turnover of root neighborhoods that would lend more support to deterministic over stochastic community assembly processes. Additionally, we examined the distance-decay pattern of β diversity, and finally partitioned β diversity into geographical and environmental components to infer their potential drivers of environmental filtering, dispersal limitation, and biotic interactions. We found that functional turnover was often lower than expected given the taxonomic turnover, whereas phylogenetic turnover was often higher than expected. Phylogenetic D pw (e.g., interfamily species) turnover exhibited a distance-decay pattern, likely reflecting limited dispersal or abiotic filtering that leads to the spatial aggregation of specific plant lineages. Conversely, both functional and phylogenetic D nn (e.g., intrageneric species) exhibited an inverted distance-decay pattern, likely reflecting strong biotic interactions among spatially and phylogenetically close species leading to phylogenetic and functional divergence. While the spatial distance was generally a better predictor of β diversity than environmental distance, the joint effect of environmental and spatial distance usually overrode their respective pure effects. These findings suggest that root neighborhood functional homogeneity may somewhat increase forest resilience after disturbance by exhibiting an insurance effect. Likewise, root neighborhood phylogenetic heterogeneity may enhance plant fitness by hindering the transmission of host-specific pathogens through root networks or by promoting interspecific niche complementarity not captured by species functions. Our study highlights the potential role of root-centric β diversity in mediating community structures and functions largely ignored in previous studies., (© 2023 The Ecological Society of America.)

Here, we present a rapid droplet sampling interface (RDSI) electrospray ionization mass spectrometry (ESI-MS) system as a high-throughput, low-volume, noncontact, and minimal-carryover approach for characterization of liquids. Liquid characterization was achieved by combining droplet ejection with an open-face microflow capillary with a 2.5 μL/min continuous flow of carrier solvent. Through this implementation, single 0.3 nL droplets containing the analyte effectively mix with 4-8 nL of carrier solvent and create a combined electrospray plume. The carrier solvent continuously cleaned the system, eliminating carryover. A sampling rate of 5 Hz was achieved for droplets containing 1 μM propranolol or 5 μM leu-enkephalin with each droplet fully baseline-resolved (138 ± 32 ms baseline-to-baseline). Using a SCIEX API4000 mass spectrometer, a lower limit of quantification (LLOQ) of propranolol was 15 nM, corresponding to 1.16 fg of propranolol in the droplet, and was linear across 3 orders of magnitude. Quantitation could be achieved by adding an isotopically labeled internal standard, as done in conventional ESI. Signal transients were faster than the acquisition speed of the mass spectrometer, resulting in artificially high reproducibility of 15-30% RSD droplet-to-droplet. Analyte-solvent mixing ratios could be controlled by adjusting droplet positioning along the open-face capillary with an optimal position about 0.4 mm from the tip end. The range of analyte coverage was exemplified by measures of peptides and drugs in methanol, water, and buffer solutions. In a comparison to the Open Port Sampling Interface (OPSI) implemented on the same system, the RDSI had 78× greater sensitivity, 6× greater throughput and used significantly less carrier solvent.

The rhizosphere is the narrow region of soil surrounding the roots of plants that is influenced by root exudates, root secretions, and associated microbial communities. This region is crucial to plant growth and development and plays a critical role in nutrient uptake, disease resistance, and soil transformation. Understanding the function of exogenous compounds in the rhizosphere starts with determining the spatiotemporal distribution of these molecular components. Using liquid microjunction surface-sampling probe mass spectrometry (LMJ-SSP-MS) and microfluidic devices with attached microporous membranes enables in situ, nondisruptive, and nondestructive spatiotemporal measurement of exogenous compounds from plant roots. However, long imaging times (>2 h) can negatively affect plant heath and limit temporal studies. Here, we present a novel strategy to optimize the number and location of sampling sites on these microporous membrane-covered microfluidic devices. This novel, "structure-driven" sampling workflow takes into consideration the channel structure of the microfluidic device to maximize sampling from the channels and minimize acquisition time (∼4× less time in some cases while providing similar chemical image accuracy), thus reducing stress on plants during in situ LMJ-SSP-MS analysis.

Plant organ growth results from the combined activity of cell division and cell expansion. The co-ordination of these two processes depends on the interplay between multiple hormones that determine the final organ size. Using the semidominant Hairy Sheath Frayed1 ( Hsf1 ) maize mutant that hypersignals the perception of cytokinin (CK), we show that CK can reduce leaf size and growth rate by decreasing cell division. Linked to CK hypersignaling, the Hsf1 mutant has an increased jasmonic acid (JA) content, a hormone that can inhibit cell division. The treatment of wild-type seedlings with exogenous JA reduces maize leaf size and growth rate, while JA-deficient maize mutants have increased leaf size and growth rate. Expression analysis revealed the increased transcript accumulation of several JA pathway genes in the Hsf1 leaf growth zone. A transient treatment of growing wild-type maize shoots with exogenous CK also induced the expression of JA biosynthetic genes, although this effect was blocked by the co-treatment with cycloheximide. Together, our results suggest that CK can promote JA accumulation, possibly through the increased expression of specific JA pathway genes.

Enzymatic biodegradation of polymers, such as polyamides (PA), has the potential to cost-effectively reduce plastic waste, but enhancements in degradation efficiency are needed. Engineering enzymes through directed evolution is one pathway toward identification of critical domains needed for improving activity. However, screening such enzymatic libraries (100s-to-1000s of samples) is time-consuming. Here we demonstrate the use of robotic autosampler (PAL) and immediate drop on demand technology (I.DOT) liquid handling systems coupled with open-port sampling interface-mass spectrometry (OPSI-MS) to screen for PA6 and PA66 hydrolysis by 6-aminohexanoate-oligomer endo -hydrolase (nylon hydrolase, NylC) in a high-throughput (8-20 s/sample) manner. The OPSI-MS technique required minimal sample preparation and was amenable to 96-well plate formats for automated processing. Enzymatic hydrolysis of PA characteristically produced soluble linear oligomer products that could be identified by OPSI-MS. Incubation temperatures and times were optimized for PA6 (65 °C, 24 h) and PA66 (75 °C, 24 h) over 108 experiments. In addition, the I.DOT/OPSI-MS quantified production of PA6 linear dimer (8.3 ± 1.6 μg/mL) and PA66 linear monomer (13.5 ± 1.5 μg/mL) by NylC with a lower limit of detection of 0.029 and 0.032 μg/mL, respectively. For PA6 and PA66, linear oligomer production corresponded to 0.096 ± 0.018% and 0.204 ± 0.028% conversion of dry pellet mass, respectively. The developed methodology is expected to be utilized to assess enzymatic hydrolysis of engineered enzyme libraries, comprising hundreds to thousands of individual samples.

Plants exhibit differential behaviours through changes in biomass development and distribution in response to environmental cues, which may impact crops uniquely. We conducted a mesocosm experiment in pots to determine the root and shoot behavioural responses of wheat, T. aestivum. Plants were grown in homogeneous or heterogeneous and heavily or lightly fertilized soil, and alone or with a neighbour of the same or different genetic identity (cultivars: CDC Titanium, Carberry, Glenn, Go Early, and Lillian). Contrary to predictions, wheat did not alter relative reproductive effort in the presence of neighbours, more nutrients, or homogenous soil. Above and below ground, the plants' tendency to use potentially shared space exhibited high levels of plasticity. Above ground, they generally avoided shared, central aerial space when grown with neighbours. Unexpectedly, nutrient amount and distribution also impacted shoots; plants that grew in fertile or homogenous environments increased shared space use. Below ground, plants grown with related neighbours indicated no difference in neighbour avoidance. Those in homogenous soil produced relatively even roots, and plants in heterogeneous treatments produced more roots in nutrient patches. Additionally, less fertile soil resulted in pot-level decreases in root foraging precision. Our findings illustrate that explicit coordination between above- and belowground biomass in wheat may not exist.

Successful host plant colonization by tree-killing bark beetle-symbiotic fungal complexes depends on host suitability, which is largely determined by host defense metabolites such as monoterpenes. Studies have shown the ability of specific blends of host monoterpenes to influence bark beetles or their fungal symbionts, but how biologically relevant blends of host monoterpenes influence bark beetle-symbiotic fungal interaction is unknown. We tested how interactions between two host species (lodgepole pine or jack pine) and two fungal symbionts of mountain pine beetle (Grosmannia clavigera or Ophiostoma montium) affect the performance of adult female beetles in vitro. Beetles treated with the propagules of G. clavigera or O. montium or not treated (natural fungal load) were introduced into media amended with a blend of the entire monoterpene profile of either host species and beetle performance was compared. Overall, host blends altered beetle performance depending on the fungal species used in the beetle amendment. When beetles were amended with G. clavigera, their performance was superior over beetles amended with O. montium in either host blend. Furthermore, G. clavigera-amended beetles performed better in media amended with host blends than without a host blend; in contrast, O. montium-amended beetles performed better in media without a host blend than with a host blend. Overall, this study showed that host defense metabolites affect host suitability to bark beetles through influencing their fungal symbionts and that different species of fungal symbionts respond differentlly to host defense metabolites., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The plant rhizosphere is a complex and dynamic chemical environment where the exchange of molecular signals between plants, microbes, and fungi drives the development of the entire biological system. Exogenous compounds in the rhizosphere are known to affect plant-microbe organization, interactions between organisms, and ultimately, growth and survivability. The function of exogenous compounds in the rhizosphere is still under much investigation, specifically with respect to their roles in plant growth and development, the assembly of the associated microbial community, and the spatiotemporal distribution of molecular components. A major challenge for spatiotemporal measurements is developing a nondisruptive and nondestructive technique capable of analyzing the exogenous compounds contained within the environment. A methodology using liquid microjunction-surface sampling probe-mass spectrometry (LMJ-SSP-MS) and microfluidic devices with attached microporous membranes was developed for in situ , spatiotemporal measurement of amino acids (AAs) from bacterial biofilms and plant roots. Exuded arginine was measured from a living Pantoea YR343 biofilm, which resulted in a chemical image indicative of biofilm growth within the device. Spot sampling along the roots of Populus trichocarpa with the LMJ-SSP-MS resulted in the detection of 15 AAs. Variation in AA concentrations across the root system was observed, indicating that exudation is not homogeneous and may be linked to local rhizosphere architecture and different biological processes along the root.

Neurological symptoms of post-acute sequelae of COVID-19 (PASC), also known as Long COVID, are recognized. Four neurological syndromes (transverse myelitis, ischemic stroke, headache, and Guillain-Barré syndrome) associated with PASC are reviewed here, with a particular focus on issues related to rehabilitation.

Competition is often highlighted as a major force influencing plant species diversity. However, there are multiple facets of competition (e.g., strength, intransitivity, and size asymmetry) that may have independent and differential impacts on diversity, making understanding the degree to which competition structures communities difficult. Unfortunately, field-based experiments that decouple multiple facets of competition are lacking, limiting our ability to test theoretical frameworks and reducing understanding of the actual linkages among competition and coexistence. Here, we experimentally manipulate the size structure of local grassland communities to examine the relative impacts of competitive size asymmetry (i.e., competitive advantage based on relative size) and intensity (i.e., mean effect of neighbors on plant growth) on species loss and gain. Increased competitive size asymmetry was associated with increased species loss and decreased species gain, while no relationship was found between competitive intensity and species loss and gain. Furthermore, the probability of loss was not dependent on a species initial size, suggesting that small species may not always be the losers of size-asymmetric interactions. Instead, loss was dependent on species rarity, where loss was higher for rare species. Overall, these results suggest that competitive size asymmetry may be more important for species loss than intensity in some plant communities and demonstrates the importance of decoupling different aspects of competition to better understand their drivers and ecological consequences., (© 2022 The Ecological Society of America.)

Grazing by wild and domesticated grazers occurs within many terrestrial ecosystems worldwide, with positive and negative impacts on biodiversity. Management of grazed lands in support of biological conservation could benefit from a compiled dataset of animal biodiversity within and adjacent to grazed sites. In this database, we have assembled data from the peer-reviewed literature that included all forms of grazing, co-occurring species, and site information. We reviewed 3,489 published articles and found 245 studies in 41 countries that surveyed animal biodiversity co-occurring with grazers. We extracted 16,105 observations of animal surveys for over 1,200 species in all terrestrial ecosystems and on all continents except Antarctica. We then compiled 28 different grazing variables that focus on management systems, assemblages of grazer species, ecosystem characteristics, and survey type. Our database provides the most comprehensive summary of animal biodiversity patterns that co-occur with wild and domesticated grazers. This database could be used in future conservation initiatives and grazing management to enhance the prolonged maintenance of ecosystems and ecosystem services., (© 2022. The Author(s).)

Background: Localized disturbances within grasslands alter biological properties and may shift species composition. For example, rare species in established communities may become dominant in successional communities if they exhibit traits well-suited to disturbance conditions. Although the idea that plant species exhibit different trait 'strategies' is well established, it is unclear how ecological selection for specific traits may change as a function of disturbance. Further, there is little data available testing whether disturbances select for single trait-characters within communities (homogenization), or allow multiple trait-types to persist (diversification). We investigated how (a) traits and (b) functional diversity of post-disturbance gap communities compared to those in adjacent undisturbed grasslands, and (c) if altered functional diversity resulted in the homogenization or diversification of functional traits., Methods: Here we emulate the impacts of an extreme disturbance in a native grassland site. We measured plant community composition of twelve paired 50 × 50 cm plots (24 total) in Alberta, Canada. Each pair consisted of one undisturbed plot and one which had all plants terminated 2 years prior. We used species abundances and a local trait database to calculate community weighted means for maximum height, specific leaf area, specific root length, leaf nitrogen percent, and root nitrogen percent. To test the impacts of disturbance on community functional traits, we calculated functional diversity measures and compared them between disturbed and undisturbed communities., Results: Within 2 years, species richness and evenness in disturbed communities had recovered and was equivalent to undisturbed communities. However, disturbed and undisturbed communities had distinct community compositions, resulting in lower functional divergence in disturbed plots. Further, disturbance was linked to increases in community-weighted mean trait values for resource-acquisitive traits, such as specific leaf area, and leaf and root nitrogen., Discussion: Disturbance had lasting effects on the functional traits and diversity of communities, despite traditional biodiversity measures such as richness and evenness recovering within 2 years. The trait space of gap communities shifted compared to undisturbed communities such that gap communities were dominated by traits enhancing resource uptake and growth rates. Overall, these results show that short-term disturbance fundamentally changes the functional character of early-successional communities, even if they superficially appear recovered., Competing Interests: The authors declare that they have no competing interests., (© 2022 Smith et al.)

Urbanization transforms environments in ways that alter biological evolution. We examined whether urban environmental change drives parallel evolution by sampling 110,019 white clover plants from 6169 populations in 160 cities globally. Plants were assayed for a Mendelian antiherbivore defense that also affects tolerance to abiotic stressors. Urban-rural gradients were associated with the evolution of clines in defense in 47% of cities throughout the world. Variation in the strength of clines was explained by environmental changes in drought stress and vegetation cover that varied among cities. Sequencing 2074 genomes from 26 cities revealed that the evolution of urban-rural clines was best explained by adaptive evolution, but the degree of parallel adaptation varied among cities. Our results demonstrate that urbanization leads to adaptation at a global scale.

The rhizosphere is a challenging ecosystem to study from a systems biology perspective due to its diverse chemical, physical, and biological characteristics. In the past decade, microfluidic platforms ( e.g. plant-on-a-chip) have created an alternative way to study whole rhizosphere organisms, like plants and microorganisms, under reduced-complexity conditions. However, in reducing the complexity of the environment, it is possible to inadvertently alter organism phenotype, which biases laboratory data compared to in situ experiments. To build back some of the complexity of the rhizosphere in a fully-defined, parameterized approach we have developed a rhizosphere-on-a-chip platform that mimics the physical structure of soil. We demonstrate, through computational simulation, how this synthetic soil structure can influence the emergence of molecular "hotspots" and "hotmoments" that arise naturally from the plant's exudation of labile carbon compounds. We establish the amenability of the rhizosphere-on-a-chip for long-term culture of Brachypodium distachyon , and experimentally validate the presence of exudate hotspots within the rhizosphere-on-a-chip pore spaces using liquid microjunction surface sampling probe mass spectrometry.

Quantitative measure of a drug and its associated metabolite(s) with single-cell resolution is often limited by sampling throughput or other compromises that limit broad use. Here, we demonstrate the use of single-cell printing-liquid vortex capture-mass spectrometry (SCP-LVC-MS) to quantitatively measure the intracellular concentrations of amiodarone (AMIO) and its metabolite, N-desethylamiodarone (NDEA), from thousands of single cells across several AMIO incubation concentrations ranging from 0 to 10 μM. Concentrations obtained by SCP-LVC-MS were validated through comparison with average assays and traditional measurement of cells in bulk. Average of SCP-LVC-MS measurements and aggregate vial collection assay the concentrations differed by < 5%. Both AMIO and NDEA had clear log-normal distributions with similar standard deviation of concentrations in the cell population. The mean of both AMIO and NDEA intracellular concentrations were positively correlated with AMIO incubation concentration, increasing from 0.026 to 0.520 and 0.0055 to 0.048 mM for AMIO and NDEA, respectively. The standard deviation of AMIO and NDEA log-normal distribution fits were relatively similar in value across incubation concentrations, 0.15-0.19 log 10 (mM), and exhibited a linear trend with respect to each other. The single cell-resolved conversion ratio of AMIO to NDEA increased with decreasing incubation concentration, 7 ± 2%, 18 ± 3%, and 20 ± 7% for 10.0, 1.0, and 0.1 μM AMIO incubation concentrations, respectively. Association with simultaneously measured lipids had several ions with statistically significant difference in intensity but no clear correlations with AMIO intracellular content was observed., (© 2021. UT- Battelle, LLC, under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Vertical root segregation and the resulting niche partitioning can be a key underpinning of species coexistence. This could result from substantial interspecific variations in root profiles and rooting plasticity in response to soil heterogeneity and neighbours, but they remain largely untested in forest communities. In a diverse forest in subtropical China, we randomly sampled > 4000 root samples from 625 0-30 cm soil profiles. Using morphological and DNA-based methods, we identified 109 woody plant species, determined the degree of vertical fine-root segregation, and examined rooting plasticity in response to soil heterogeneity and neighbour structure. We found no evidence of vertical fine-root segregation among cooccurring species. By contrast, root abundance of different species tended to be positively correlated within soil zones. Underlying these findings was a lack of interspecific variation in fine-root profiles with over 90% of species concentrated in the 0-10 cm soil zone with only one species dominating in the 10-20 cm soil zone. Root profiles exhibited low responsiveness to root neighbours but tended to be shallow in soils with low phosphorus and copper content. These findings suggest that if there is niche differentiation leading to coexistence in this diverse forest, it would be occurring by mechanisms other than vertical fine-root segregation., (© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.)

Desert steppe, a unique ecotone between steppe and desert in Eurasia, is considered highly vulnerable to global change. However, the long-term impact of warming and nitrogen deposition on plant biomass production and ecosystem carbon exchange in a desert steppe remains unknown. A 12-year field experiment was conducted in a Stipa breviflora desert steppe in northern China. A split-design was used, with warming simulated by infrared radiators as the primary factor and N addition as the secondary factor. Our long-term experiment shows that warming did not change net ecosystem exchange (NEE) or total aboveground biomass (TAB) due to contrasting effects on C 4 (23.4% increase) and C 3 (11.4% decrease) plant biomass. However, nitrogen addition increased TAB by 9.3% and NEE by 26.0% by increasing soil available N content. Thus, the studied desert steppe did not switch from a carbon sink to a carbon source in response to global change and positively responded to nitrogen deposition. Our study indicates that the desert steppe may be resilient to long-term warming by regulating plant species with contrasting photosynthetic types and that nitrogen deposition could increase plant growth and carbon sequestration, providing negative feedback on climate change.

Mass spectrometry (MS) has become the de facto tool for routine quantitative analysis of biomolecules. MS is increasingly being used to reveal the spatial distribution of proteins, metabolites, and pharmaceuticals in tissue and interest in this area has led to a number of novel spatially resolved MS technologies. Most spatially resolved MS measurements are qualitative in nature due to a myriad of potential biases, such as sample heterogeneity, sampling artifacts, and ionization effects. As applications of spatially resolved MS in the pharmacological and clinical fields increase, demand has become high for quantitative MS imaging and profiling data. As a result, several varied technologies now exist that provide differing levels of spatial and quantitative information. This review provides an overview of MS profiling and imaging technologies that have demonstrated quantitative analysis from tissue. Focus is given on the fundamental processes affecting quantitative analysis in an array of MS imaging and profiling technologies and methods to address these biases.Graphical abstract.

Rationale: The ability to quantify drugs and metabolites in tissue with sub-mm resolution is a challenging but much needed capability in pharmaceutical research. To fill this void, a novel surface sampling approach combining laser ablation with the commercial dropletProbe automated liquid surface sampling system (LA-dropletProbe) was developed and is presented here., Methods: Parylene C-coated 200 × 200 μm tissue regions of mouse brain and kidney thin tissue sections were analyzed for propranolol by laser ablation of tissue directly into a preformed liquid junction. Propranolol was detected by high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) in positive electrospray ionization mode. Quantitation was achieved via application of a stable-isotope-labeled internal standard and an external calibration curve., Results: The absolute concentrations of propranolol determined from 200 × 200 μm tissue regions were compared with the propranolol concentrations obtained from 2.3-mm-diameter tissue punches of adjacent, non-coated sections using standard bulk tissue extraction protocols followed by regular HPLC/MS/MS analysis. The average concentration of propranolol in both organs determined by the two employed methods agreed to within ±12%. Furthermore, the relative abundances of phase II hydroxypropranolol glucuronide metabolites were recorded and found to be consistent with previous results., Conclusions: This work illustrates that depositing a thin layer of parylene C onto thin tissue prior to analysis, which seals the surface and prevents direct liquid extraction of the drug from the tissue, coupled to the novel LA-dropletProbe surface sampling system is a viable approach for sub-mm resolution quantitative drug distribution analysis., (© 2020 John Wiley & Sons Ltd.)

Ecosystems are spatially heterogenous in plant community composition and function. Shrub occurrence in grasslands is a visually striking example of this, and much research has been conducted to understand the functional implications of this pattern. Within savannah ecosystems, the presence of tree and shrub overstories can have significant impacts on the understory herbaceous community. The exact outcomes, however, are likely a function of the spatial arrangement and traits of the overstory species. Here we test whether there are functional linkages between the spatial patterning of a native shrub and the standing biomass, community composition, and overall nutrient cycling of a neighbouring grassland understory communities within the Aspen Parkland of central Alberta, Canada. In a paired grassland-shrub stand study, we found the native shrub, Elaeagnus commutata , has relatively few stand-level impacts on the composition and standing biomass of the ecosystem. One factor contributing to these limited effects may be the overdispersion of shrub stems at fine spatial scales, preventing areas of deep shade. When we looked across a shrub density gradient and incorporated shrub architecture into our analyses, we found these shrub traits had significant associations with species abundance and root biomass in the understory community. These results suggest that stem dispersion patterns, as well as local stand architecture, are influential in determining how shrubs may affect their herbaceous plant understory. Thus, it is important to incorporate shrub spatial and architectural traits when assessing shrub-understory interactions., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Annals of Botany Company.)

Presented is a tethered, liquid-extraction-sampling interface designed for the mass spectrometric surface sampling/analysis of 3D objects. The tethered, open port sampling interface (TOPSI) incorporates a vacuum line between the sampling probe and ionization source, which enables the ability for an extended, tethered sample transfer line. Herein, several designs of the hand-held TOPSI are presented and evaluated on the basis of the analytical metrics of analyte transport time, peak width, and analyte sensitivity. The best analytical metrics were obtained with capillary flow resistances arranged in a particular order and the vacuum region set at 6.2 kPa. This TOPSI design incorporated a transfer capillary 1 m in length, while retaining a fast analyte transport time (12 s), short signal peak width (5 s baseline-to-baseline), and high analyte signal at 90% of that obtained with a regular open port sampling interface (OPSI). The hand-held TOPSI was demonstrated for the characterization of extracted small molecules and metabolites from the surface of mint and rosemary leaves.

Rationale: Spatially resolved and accurate quantitation of drug-related compounds in tissue is a much-needed capability in drug discovery research. Here, application of an integrated laser ablation-dropletProbe-mass spectrometry surface sampling system (LADP-MS) is reported, which achieved absolute quantitation of propranolol measured from <500 × 500 μm thin tissue samples., Methods: Mouse liver and kidney thin tissue sections were coated with parylene C and analyzed for propranolol by a laser ablation/liquid extraction workflow. Non-coated adjacent sections were microdissected for validation and processed using standard bulk tissue extraction protocols. High-performance liquid chromatography with positive ion mode electrospray ionization tandem mass spectrometry was applied to detect the drug and its metabolites., Results: Absolute propranolol concentration in ~500 × 500 μm tissue regions measured by the two methods agreed within ±8% and had a relative standard deviation within ±17%. Quantitation down to ~400 × 400 μm tissue regions was shown, and this resolution was also used for automated mapping of propranolol and phase II hydroxypropranolol glucuronide metabolites in kidney tissue., Conclusions: This study exemplifies the capabilities of integrated laser ablation-dropletProbe-mass spectrometry (LADP-MS) for high resolution absolute drug quantitation analysis of thin tissue sections. This capability will be valuable for applications needing to quantitatively understand the spatial distribution of small molecules in tissue., (© 2021 John Wiley & Sons Ltd.)

Disturbances have altered community dynamics in boreal forests with unknown consequences for belowground ecological processes. Soil fungi are particularly sensitive to such disturbances; however, the individual response of fungal guilds to different disturbance types is poorly understood. Here, we profiled soil fungal communities in lodgepole pine forests following a bark beetle outbreak, wildfire, clear-cut logging, and salvage-logging. Using Illumina MiSeq to sequence ITS1 and SSU rDNA, we characterized communities of ectomycorrhizal, arbuscular mycorrhizal, saprotrophic, and pathogenic fungi in sites representing each disturbance type paired with intact forests. We also quantified soil fungal biomass by measuring ergosterol. Abiotic disturbances changed the community composition of ectomycorrhizal fungi and shifted the dominance from ectomycorrhizal to saprotrophic fungi compared to intact forests. The disruption of the soil organic layer with disturbances correlated with the decline of ectomycorrhizal and the increase of arbuscular mycorrhizal fungi. Wildfire changed the community composition of pathogenic fungi but did not affect their proportion and diversity. Fungal biomass declined with disturbances that disrupted the forest floor. Our results suggest that the disruption of the forest floor with disturbances, and the changes in C and nutrient dynamics it may promote, structure the fungal community with implications for fungal biomass-C., (© 2020 The Authors New Phytologist © 2020 New Phytologist Foundation.)

Anthropogenic disturbance has generated a significant loss of biodiversity worldwide and grazing by domestic herbivores is a contributing disturbance. Although the effects of grazing on plants are commonly explored, here we address the potential multi-trophic effects on animal biodiversity (e.g. herbivores, pollinators and predators). We conducted a meta-analysis on 109 independent studies that tested the response of animals or plants to livestock grazing relative to livestock excluded. Across all animals, livestock exclusion increased abundance and diversity, but these effects were greatest for trophic levels directly dependent on plants, such as herbivores and pollinators. Detritivores were the only trophic level whose abundance decreased with livestock exclusion. We also found that the number of years since livestock was excluded influenced the community and that the effects of grazer exclusion on animal diversity were strongest in temperate climates. These findings synthesise the effects of livestock grazing beyond plants and demonstrate the indirect impacts of livestock grazing on multiple trophic levels in the animal community. We identified the potentially long-term impacts that livestock grazing can have on lower trophic levels and consequences for biological conservation. We also highlight the potentially inevitable cost to global biodiversity from livestock grazing that must be balanced against socio-economic benefits., (© 2020 John Wiley & Sons Ltd/CNRS.)

Wide-field coherent anti-Stokes Raman scattering (CARS) microscopy offers an attractive means for the rapid and simultaneous acquisition of vibrationally resolved images across a large field of view. A major challenge in the implementation lies in how to achieve sufficiently strong excitation fields necessary to drive the third-order optical responses over the large focal region. Here, we report a new wide-field CARS microscope enabled by a total internal reflection excitation scheme using a femtosecond Ti:Sapphire oscillator to generate pump and broadband near-infrared Stokes pulses. The spectrally broad Stokes pulse, in combination with its inherent chirp, offers not only access to a wide range of Raman modes spanning ∼1000 to ∼3500 c m -1 but also a straightforward means to select vibrational transitions within this range by simply varying the time delay between the pulses. The unique capabilities of this wide-field CARS microscope were validated by acquiring high-quality CARS images from the model and complex biological samples on conventional microscope coverslips.

For tree seedlings in boreal forests, ectomycorrhizal (EM) fungal networks may promote, while root competition may impede establishment. Thus, disruption to EM fungal networks may decrease seedling establishment owing to the loss of positive interactions among neighbors. Widespread tree mortality can disrupt EM networks, but it is not clear whether seedling establishment will be limited by the loss of positive interactions or increased by the loss of negative interactions with surrounding roots. Depending upon the relative influence of these mechanisms, widespread tree mortality may have complicated consequences on seedling establishment, and in turn, the composition of future forests. To discern between these possible outcomes and the drivers of seedling establishment, we determined the relative importance of EM fungal networks, root presence, and the bulk soil on the establishment of lodgepole pine and white spruce seedlings along a gradient of beetle-induced tree mortality. We manipulated seedling contact with EM fungal networks and roots through the use of mesh-fabric cylinders installed in soils of lodgepole pine forests experiencing a range of overstorey tree mortality caused by mountain pine beetle. Lodgepole pine seedling survival was higher with access to EM fungal networks in undisturbed pine forests in comparison with that in beetle-killed stands. That is, overstorey tree mortality shifted fungal networks from being a benefit to a cost on seedling survival. In contrast, overstorey tree mortality did not change the relative strength of EM fungal networks, root presence and the bulk soil on survival and biomass of white spruce seedlings. Furthermore, the relative influence of EM fungal networks, root presence, and bulk soils on foliar N and P concentrations was highly contingent on seedling species and overstorey tree mortality. Our results highlight that following large-scale insect outbreak, soil-mediated processes can enable differential population growth of two common conifer species, which may result in species replacement in the future.

The ability to observe dynamic chemical processes (e.g., signaling, transport, etc.) in vivo or in situ using nondestructive chemical imaging opens a new door to understanding the complex dynamics of developing biological systems. With the advent of "biology-on-a-chip" devices has come the ability to monitor dynamic chemical processes in a controlled environment, using these engineered habitats to capture key features of natural systems while allowing visual observation of system development. Having the capability to spatially and temporally map the chemical signals within these devices may yield new insights into the forces that drive biosystem development. Here, a porous membrane sealed microfluidic device was designed to allow normal microfluidic operation while enabling continuous, location specific sampling and chemical characterization by liquid microjunction surface sampling probe mass spectrometry (LMJ-SSP MS). LMJ-SSP was used to extract fluids with nL-to-μL/min flow rates directly from selected areas of the microfluidic device without negatively impacting the device function. These extracts were subsequently characterized using MS. This technique was used to acquire MS images of the entirety of several multi-input microfluidic devices having different degrees of fluid mixing. LMJ-SSP MS imaging visualized the spatial distribution of chemical components within the microfluidic channels and could visualize chemical reactions occurring in the device. These microfluidic devices with a porous membrane wall are wholly compatible with the construction of biology-on-a-chip devices. This ultimately would enable correlation of biosystem physical structure with an evolving chemical environment.

Nutrient distribution and neighbours can impact plant growth, but how neighbours shape root-foraging strategy for nutrients is unclear. Here, we explore new patterns of plant foraging for nutrients as affected by neighbours to improve nutrient acquisition. Maize (Zea mays) was grown alone (maize), or with maize (maize/maize) or faba bean (Vicia faba) (maize/faba bean) as a neighbour on one side and with or without a phosphorus (P)-rich zone on the other in a rhizo-box experiment. Maize demonstrated root avoidance in maize/maize, with reduced root growth in 'shared' soil, and increased growth away from its neighbours. Conversely, maize proliferated roots in the proximity of neighbouring faba bean roots that had greater P availability in the rhizosphere (as a result of citrate and acid phosphatase exudation) compared with maize roots. Maize proliferated more roots, but spent less time to reach, and grow out of, the P patches away from neighbours in the maize/maize than in the maize/faba bean experiment. Maize shoot biomass and P uptake were greater in the heterogeneous P treatment with maize/faba bean than with maize/maize system. The foraging strategy of maize roots is an integrated function of heterogeneous distribution of nutrients and neighbouring plants, thus improving nutrient acquisition and maize growth. Understanding the foraging patterns is critical for optimizing nutrient management in crops., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

Laser capture microdissection is a valuable technique in individually isolating single cells whether in tissue networks or deposited from a cell suspension. New developments have enabled coupling of laser capture microdissection with mass spectrometry via liquid vortex capture sampling probe. This enables online metabolic profiling of sectioned cells. Here, we describe the protocol used to deposit, isolate, and individually chemically characterize single Allium cepa and Chlamydomonas reinhardtii cells by laser capture microdissection-liquid vortex capture mass spectrometry.

Plants regularly encounter patchily distributed soil nutrients. A common foraging response is to proliferate roots within high-quality patches. The influence of the social environment on this behaviour has been given limited attention, despite important fitness consequences of competition for soil resources among plants. Using the common sunflower ( Helianthus annuus L.), we compared localized root proliferation in a high-quality patch by plants grown alone to that of plants in two different social environments: with a neighbouring plant sharing equal access to the high-quality patch, and with a neighbouring plant present but farther from the high-quality patch such that the focal individual was in closer proximity to the high-quality patch. Sunflowers grown alone proliferated more roots within high-nutrient patches than lower-nutrient soil. Plants decreased root proliferation within a high-nutrient patch when it was equidistant to a neighbour. Conversely, plants increased root proliferation when they were in closer proximity to the patch relative to a nearby neighbour. Such contingent responses may allow sunflowers to avoid competition in highly contested patches, but to also pre-empt soil resources from neighbours when they have better access to a high-quality patch. We also compared patch occupancy by sunflowers grown alone with two equidistant high-quality patches to occupancy by sunflowers grown with two high-quality patches and a neighbour. Plants grown with a neighbour decreased root length within shared patches but did not increase root length within high-quality patches they were in closer proximity to, perhaps because resource pre-emption may be less important for individuals when resources are more abundant. These results show that nutrient foraging responses in plants can be socially contingent, and that plants may account for the possibility of pre-empting limited resources in their foraging decisions.

Global change drivers (GCDs) are expected to alter community structure and consequently, the services that ecosystems provide. Yet, few experimental investigations have examined effects of GCDs on plant community structure across multiple ecosystem types, and those that do exist present conflicting patterns. In an unprecedented global synthesis of over 100 experiments that manipulated factors linked to GCDs, we show that herbaceous plant community responses depend on experimental manipulation length and number of factors manipulated. We found that plant communities are fairly resistant to experimentally manipulated GCDs in the short term (<10 y). In contrast, long-term (≥10 y) experiments show increasing community divergence of treatments from control conditions. Surprisingly, these community responses occurred with similar frequency across the GCD types manipulated in our database. However, community responses were more common when 3 or more GCDs were simultaneously manipulated, suggesting the emergence of additive or synergistic effects of multiple drivers, particularly over long time periods. In half of the cases, GCD manipulations caused a difference in community composition without a corresponding species richness difference, indicating that species reordering or replacement is an important mechanism of community responses to GCDs and should be given greater consideration when examining consequences of GCDs for the biodiversity-ecosystem function relationship. Human activities are currently driving unparalleled global changes worldwide. Our analyses provide the most comprehensive evidence to date that these human activities may have widespread impacts on plant community composition globally, which will increase in frequency over time and be greater in areas where communities face multiple GCDs simultaneously., Competing Interests: The authors declare no conflict of interest.

Premise: Plants generally increase root growth in areas with high nutrients in heterogeneous soils, a phenomenon called foraging precision. The physiology of this process is not well understood, but it may involve shoot-root signaling via leaf veins. If this is true, then damage to leaf veins, but not to nearby mesophyll, would reduce plant foraging precision., Methods: To test this hypothesis, we imposed two leaf damage treatments on Plantago asiatica and Prunus jamasakura, removing either the tip of each main vein or mesophyll tissue between the veins with a 3-mm-diameter hole punch. After 30 days or 20 weeks of plant growth, we measured root biomass in the soil in response to soil nutrient concentration., Results: When leaf mesophyll was damaged, root biomass of both species was greater in nutrient-rich patches than in nutrient-poor patches. However, when leaf veins were damaged, root biomass was similar between patches., Conclusions: These results suggest the importance of shoot-root signaling in plants, emphasizing that physiological processes are not necessarily restricted to single organs. The idea that herbivores that damage leaf veins may affect a plant's ability to selectively forage in high-nutrient patches is novel, with implications for natural and managed systems., (© 2019 Botanical Society of America.)

Invasive species can alter the structure and function of the communities they invade, as well as lead to biotic homogenization across their invasive range, thus affecting large-scale diversity patterns. The mechanisms by which invasive species can lead to biotic homogenization are poorly understood. We argue that invasive species acting as strong, deterministic, and consistent filters within and across invaded communities are likely to cause biotic homogenization at multiple spatial scales. We studied Bromus inermis Leyss. invasion into eight grasslands covering most of the grassland and parkland natural regions of Alberta (western Canada). Specifically, we tested whether B. inermis (1) has a strong impact on species richness and composition, (2) consistently alters resources (nutrients, light, and soil moisture), imposing the same ecological filter to species establishment and persistence across sites, and hence (3) whether it leads to biotic homogenization within and across sites. We recorded plant cover and resources across native-to-invaded transition areas combining space-for-time substitutions with time-series data analyses. Bromus inermis invasion was associated with rapid biotic homogenization of communities, within and among the eight grasslands. The sharp changes in species relative abundances following invasion was the initial driver of biotic homogenization, and species loss was delayed. Supporting the idea that biotic homogenization can occur when an invasive species presents a broad and consistent filter, resources modified by B. inermis invasion (particularly light and certain nutrients) were altered rather consistently within and across sites. The 50% reduction in light was likely the initial driver of biotic homogenization, and the increase in nutrient availability probably facilitates the displacement of species from the invaded areas and could lead to the establishment of self-reinforcing dynamics. Overall, our results support the idea that invaders acting as strong, deterministic, and consistent ecological filters are likely to cause biotic homogenization of the communities they invade., (© 2019 by the Ecological Society of America.)

Ectomycorrhizal fungi (plant symbionts) are diverse and exist within spatially variable communities that play fundamental roles in the functioning of terrestrial ecosystems. However, the underlying ecological mechanisms that maintain and regulate the spatial structuring of ectomycorrhizal fungal communities are both complex and remain poorly understood. Here, we use a gradient of mountain pine beetle ( Dendroctonus ponderosae ) induced tree mortality across eleven stands in lodgepole pine ( Pinus contorta ) forests of western Canada to investigate: (i) the degree to which spatial structure varies within this fungal group, and (ii) how these patterns may be driven by the relative importance of tree mortality from changes in understory plant diversity, productivity and fine root biomass following tree death. We found that the homogeneity of the ectomycorrhizal fungal community increased with increasing tree death, aboveground understory productivity and diversity. Whereas, the independent effect of fine root biomass, which declined along the same gradient of tree mortality, increased the heterogeneity of the ectomycorrhizal fungal community. Together, our results demonstrate that large-scale biotic disturbance homogenizes the spatial patterns of ectomycorrhizal fungal communities., Competing Interests: The authors declare there are no competing interests.

We report a method that enables untargeted, high throughput, and quantitative mass spectrometric analysis of single cells from cell suspension without needing additional sample preparation procedures (e.g., molecular tagging) through the combination of single-cell printer technology and liquid vortex capture-mass spectrometry (SCP-LVC-MS). The operating principle behind the SCP-LVC-MS technology is single cell isolation via small droplet piezoelectric ejection followed by capture of the droplet into an LVC-MS sampling probe. Once exposed to an appropriate solvent, the cell is lysed, extracted, and analyzed by MS. The SCP-LVC-MS approach was validated by measuring the lipid composition of microalgae, Chlamydomonas reinhardtii (ChRe) and Euglena gracilis (EuGr), and HeLa cells in their native growth media. Numerous diacylglyceryltrimethylhomo-Ser (DGTS), phosphatidylcholine (PC), monogalactosyldiacylglycerol (MGDG), and digalactosyldiacylglycerol (DGDG) lipids were observed in single cells. Continuous solvent flow ensures that cells are analyzed rapidly, and no signal carryover between cells is observed. ChRe and EuGr microalgae mixed together in the same solution were differentiated cell-by-cell in real-time based on differences between levels of diacylglyceryltrimethylhomo-Ser (DGTS) and phosphatidylcholine (PC) lipids measured in each cell. Several DGTS lipids present in ChRe were quantified with single-cell resolution by normalizing to a DGTS(32:0) internal standard added to the LVC probe solvent during analysis. Quantitative peak areas were validated by comparing to bulk lipid extracts. Lastly, peak area distributions comprised of hundreds of cells were compared for ChRe after 5 days of nitrogen-limited and normal growth conditions, which show clear differences and the ability to resolve cellular population differences with single-cell resolution.

Hypothesis: Controlling nanomaterial interfaces for emerging technologies has driven the need to understand the molecular species located there; however, challenges arise using traditional analytical techniques to directly characterize the molecular structure and local environments of these interfacial species due to their low relative populations. We hypothesized that vibrational sum frequency generation (vSFG) spectroscopy would be uniquely sensitive to the chemical modification of nanoparticle surfaces that is obscured using traditional bulk sensitive methods., Experiments: Octadecylamine ligands were removed from model CdSe quantum dot surfaces using a common precipitation-resuspension procedure with polar protic and aprotic nonsolvents. Vibrational spectra of the ligands at the surface were collected with vSFG to directly probe the ligand ordering and coverage. Photoluminescence (PL), optical absorption, NMR, and mass spectrometry measurements were conducted for comparison., Findings: vSFG was found to be sensitive to subtle changes in ligand disorder over multiple precipitation-resuspension washes, and a limit to the number of ligand molecules removed from the surface and subsequent amount of disorder introduced to their packing was clearly observed. We also find that nonsolvents do not remain associated with the surface after washing., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Multiple sclerosis is increasingly being recognized in association with cardiovascular dysfunction, which can manifest with cardiomyopathy, pulmonary edema (Kaplan et al., 2015), orthostasis (Kaplan et al., 2015) and other complications. Takotsubo cardiomyopathy is a syndrome of acute-onset ventricular dysfunction in the absence of obstructive coronary artery disease, which on rare occasion has been observed in the setting of demyelinated plaques in the brainstem. We report on two cases of Takotsubo cardiomyopathy with medullary demyelination secondary to multiple sclerosis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Premise of the Study: The rapid leaf movement of Mimosa pudica is expected to be costly because of energetic trade-offs with other processes such as growth and reproduction. Here, we assess the photosynthetic opportunity cost and energetic cost of the unique leaf closing behavior of M. pudica., Methods: In the greenhouse, we employed novel touch-stimulation machines to expose plants to one of three treatments: (1) untouched control plants; (2) plants touch-stimulated to close their leaves during the day to incur energetic costs associated with leaf movement and reduced photosynthesis; (3) plants touched at night to assess the effects of touch alone. M. pudica is nyctinastic and closes its leaves at night; thus, touching at night does not impart additional costs. We directly assessed costs by comparing physical traits. Leaf re-opening response was measured to assess the potential for plant behavioral plasticity to impact photosynthetic opportunity costs., Key Results: The cost of rapid leaf closure behavior was expressed as a 47% reduction in reproductive biomass accounting for the effect of touch. Touch itself changed physical traits such as biomass, with touched plants being generally bigger. Plants touched at night re-opened their leaflets 26% quicker than plants touched during the day., Conclusions: We reason that the reproductive allocation costs incurred by M. pudica can be attributed to a combination of photosynthetic opportunity cost and the energetic cost associated with increased stimulation of leaf movement and that behavioral plasticity has the potential to alter photosynthetic opportunity costs., (© 2018 Botanical Society of America.)

Current analytical methods are not capable of providing rapid, sensitive, and comprehensive chemical analysis of a wide range of cellular constitutes of single cells (e.g., lipids, metabolites, proteins, etc.) from dispersed cell suspensions and thin tissues. This capability is important for a number of critical applications, including discovery of cellular mechanisms for coping with chemical or environmental stress and cellular response to drug treatment, to name a few. Here we introduce an optically guided platform and methodology for rapid, automated recognition, sampling, and chemical analysis of surface confined individual cells utilizing a novel hybrid laser capture microdissection/liquid vortex capture/mass spectrometry system. The system enabled automated analysis of single cells by reliably detecting and sampling them either through laser ablation from a glass microscope slide or by cutting the entire cell out of a poly(ethylene naphthalate)-coated membrane substrate that the cellular sample is deposited on. Proof of principle experiments were performed using thin tissues of Allium cepa and cultured Euglena gracilis and Phacus cell suspensions as model systems for single cell analysis using the developed method. Reliable, hands-off laser ablation sampling coupled to liquid vortex capture/mass spectrometry analysis was conducted for hundreds of individual Allium cepa cells in connected tissue. In addition, more than 300 individual Euglena gracilis and Phacus cells were analyzed automatically and sampled using laser microdissection sampling with the same liquid vortex capture/mass spectrometry analysis system. Principal component analysis-linear discriminant analysis, applied to each mass spectral dataset, was used to determine the accuracy of differentiation of the different algae cell lines.

Although the negative effects of root herbivores on plant fitness are expected to be similar to those of above-ground herbivores, the study of below-ground plant defences is limited compared to the rich literature on above-ground defences. Current theory predicts that concentrations of defensive chemicals above- and below-ground should be correlated, as the evolutionary drivers that shape plant defence are similar across the whole plant. We conducted a field study to measure root condensed tannin concentrations in Populus tremuloides , and determine how they related to leaf condensed tannin concentrations, tree position within the stand (edge vs. interior), tree size, and time of year. Overall, root tannin concentrations were substantially lower than leaf tannin concentrations. At individual sampling periods, root and leaf tannin concentrations were uncorrelated with each other, and did not vary with stand position or size. Across the growing season both root and leaf tannin concentrations did show similar trends, with both highest in the early summer, and declining through mid-summer and fall. These results suggest that the mechanisms that influence leaf and root tannin levels in aspen are independent within individual stems, possibly due to different evolutionary pressures experienced by the different tissue types or in response to localized (roots vs. foliage) stressors. However, across individual stems, the similar patterns in chemical defence over time, independent of plant size or stand position indicate that larger scale processes can have consistent effects across individuals within a population, such as the relative investment in defence of tissues in the spring versus the fall. Overall, we conclude that using theories based on above-ground defence to predict below-ground defences may not be possible without further studies examining below-ground defence.

Pollination is critical for plant fitness and population dynamics, yet little attention is paid to the role of flowering and plant-pollinator interactions in structuring plant communities, including community responses to environmental change. Changes in arbuscular mycorrhizal fungi (AMF), nutrient abundances, and plant litter all affect plant access to different resources, and are known regulators of community structure. Each factor can also affect flowering and plant-pollinator interactions, potentially contributing to changes in community structure. To test whether AMF, nutrients, and litter influenced the relationship between pollination and community structure, we conducted a 5-yr field experiment applying fungicide, adding fertilizer, and removing plant litter in native grassland. We measured the distribution of flowers and floral visits among species in year three and linked these measures to changes in plant composition and species richness between years three and five. We hypothesized that an uneven distribution of flowers and visits among species would lead to greater community change, but that the treatments would disrupt this relationship by altering sexual allocation and recruitment. Consistent with our hypothesis, communities with uneven flower distributions exhibited greater changes in community composition and richness under ambient conditions. However, AMF suppression neutralized this relationship and regulated the other treatment effects, highlighting the potential importance of AMF for stabilizing recruitment dynamics. Combined, AMF suppression and nutrient addition caused species losses when few species flowered, likely by compounding stresses for those species. The treatment effects on the relationship between flowering and community composition were more nuanced, but were likely driven by increased competition and altered flowering among species. By contrast, community composition was more stable when visitation rates were uneven among species, irrespective of any treatments. This suggests that some species require high visitation rates to maintain their populations due to greater dependence on sexual reproduction. Combined, these results highlight the importance of flowering and floral visitation to the dynamics of grassland communities. They also suggest that altered recruitment dynamics is a major, yet understudied, mechanism by which environmental change affects communities. Consequently, understanding the effects of environmental change on plant communities will require study of both plant growth and sexual reproduction., (© 2018 by the Ecological Society of America.)

Rapidly determining root growth patterns is biologically important and technically challenging. Current methods focus on direct observation of roots and require destructive excavations or time-consuming root tracing. We developed a novel methodology based on analyzing soil particle displacement, rather than direct observation of roots. This inferred root growth method uses digital image correlation (DIC) analysis, an established and high-throughput method used in many engineering and science disciplines. By applying DIC analyses to repeated images of plants grown in clear window boxes, we produced visually intuitive and quantifiable strain maps, indicating the magnitude and direction of soil movement. From this, we could infer root growth and rapidly quantify root system metrics. Strain measures were closely associated with the spatial distribution of roots and correlated with root length measured using conventional approaches. The method also allowed for the detection of root proliferation in nutrient-enriched soil patches, indicating its suitability for quantifying biological patterns. This novel application of DIC in root biology is effective, scalable, low cost, flexible and complementary to existing technologies. This method offers a new tool for answering questions in plant biology and will be particularly useful in studies involving temporal dynamics of root processes., (© 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.)

Rationale: Laser microdissection-liquid vortex capture/electrospray ionization mass spectrometry (LMD-LVC/ESI-MS) has potential for on-line classification of tissue but an investigation into what analytical conditions provide best spectral differentiation has not been conducted. The effects of solvent, ionization polarity, and spectral acquisition parameters on differentiation of mouse brain tissue regions are described., Methods: Individual 40 × 40 μm microdissections from cortex, white, grey, granular, and nucleus regions of mouse brain tissue were analyzed using different capture/ESI solvents, in positive and negative ion mode ESI, using time-of-flight (TOF)-MS and sequential window acquisitions of all theoretical spectra (SWATH)-MS (a permutation of tandem-MS), and combinations thereof. Principal component analysis-linear discriminant analysis (PCA-LDA), applied to each mass spectral dataset, was used to determine the accuracy of differentiation of mouse brain tissue regions., Results: Mass spectral differences associated with capture/ESI solvent composition manifested as altered relative distributions of ions rather than the presence or absence of unique ions. In negative ion mode ESI, 80/20 (v/v) methanol/water yielded spectra with low signal/noise ratios relative to other solvents. PCA-LDA models acquired using 90/10 (v/v) methanol/chloroform differentiated tissue regions with 100% accuracy while data collected using methanol misclassified some samples. The combination of SWATH-MS and TOF-MS data improved differentiation accuracy., Conclusions: Combined TOF-MS and SWATH-MS data differentiated white, grey, granular, and nucleus mouse tissue regions with greater accuracy than when solely using TOF-MS data. Using 90/10 (v/v) methanol/chloroform, tissue regions were perfectly differentiated. These results will guide future studies looking to utilize the potential of LMD-LVC/ESI-MS for tissue and disease differentiation., (Copyright © 2018 John Wiley & Sons, Ltd.)