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Growth suppression and defence signalling are simultaneous strategies that plants invoke to respond to abiotic stress. Here, we show that the drought stress response of poplar trees (Populus trichocarpa) is initiated by a suppression in cell wall derived methanol (MeOH) emissions and activation of acetic acid (AA) fermentation defences. Temperature sensitive emissions dominated by MeOH (AA/MeOH

Hyphae are microscopic filaments that elongate and branch to create networks of interconnected tubes. Understanding how they work remains a formidable challenge in experimental mycology. Important advances in hyphal research in the 20th century came from electron microscopy, which revealed clusters of cytoplasmic vesicles in the cell apex, and biochemical studies that identified the cell wall materials that are assembled at the tip. Early genetic experiments on hyphae based on mutant analysis were disappointing and provided little information on the relationship between genotype and phenotype. Progress has come more recently, in the first decades of this century, by combining the techniques of molecular genetics with modern imaging methods. Live-cell imaging has allowed us to study the dynamics of cell components in strains of fungi engineered with plasmids encoding proteins fused to fluorescent probes. This technology has provided significant insights on the growth process and yet the fundamentals of hyphal growth remain elusive.

Fungi show a variety of abilities in affecting metal speciation, toxicity, and mobility and mineral formation, dissolution or deterioration through several interacting biomechanical and biochemical mechanisms. A consequence of many metal-mineral interactions is the production of nanoparticles which may be in elemental, mineral or compound forms. Organisms may benefit from such nanomaterial formation through removal of metal toxicity, protection from environmental stress, and their redox properties since certain mycogenic nanoparticles can act as nanozymes mimicking enzymes such as peroxidase. With the development of nanotechnology, there is growing interest in the application of biological systems for nanomaterial production which may provide economic benefits and a lower damaging environmental effect than conventional chemical synthesis. Fungi offer some advantages since most are easily cultured under controlled conditions and well known for the secretion of metabolites and enzymes related to nanoparticle or nanomineral formation. Nanoparticles can be formed intracellularly or extracellularly, the latter being favourable for easy harvest, while the cell wall also provides abundant nucleation sites for their formation. In this article, we focus on the synthesis of nanoparticles and nanominerals by fungi, emphasizing the mechanisms involved, and highlight some possible applications of fungal nanomaterials in environmental biotechnology.

This technical focus article discusses the importance of concentration, cellular exposure and specificity for the application of organelle selective fluorescent dyes in fungi using DNA, membrane and cell wall stains as examples. Nonetheless, the presented considerations are generally applicable to all fluorescent dyes applied to living cells. The association of a fluorescent dye with its target molecule generally impairs molecule and consequently organelle function. Effective dye concentration, cellular exposure time and specificity to the target molecule are key factors that influence the biocompatibility of any fluorescent dye. Prominent molecules frequently used as fluorescent staining targets in fungal cell biology are: (i) DNA for nuclear labelling, (ii) α-/β-glucans and chitin for cell wall labelling, and (iii) phospholipids for plasma membrane and endomembrane labelling. In combination with live-cell imaging settings that reduce light stress, i.e. excitation intensities and exposure times set to the minimum that still achieves good signal-to-noise ratios, is the low dosage application of fluorescent markers as so called “vital dyes” essential for visualising cellular processes in an artefact-free fashion.

Biosynthesis of plant cell walls requires UDP-glucose as the substrate for cellulose biosynthesis, and as an intermediate for the synthesis of other matrix polysaccharides. The sucrose cleaving enzyme sucrose synthase (SUS) is thought to have a central role in UDP-glucose biosynthesis, and a long held and much debated hypothesis postulates that SUS is required to supply UDP-glucose to cellulose biosynthesis. To investigate the role of SUS in cellulose biosynthesis of Arabidopsis thaliana we characterized mutants in which four, or all six Arabidopsis SUS genes were disrupted. These sus mutants showed no growth phenotypes, vascular tissue cell wall defects or changes in cellulose content. Moreover, the UDP-glucose content of rosette leaves of the sextuple sus mutants was increased by approximately 20% compared to wild type. It can thus be concluded that cellulose biosynthesis is able to employ alternative UDP-glucose biosynthesis pathway(s), and thereby the model of SUS requirement for cellulose biosynthesis in Arabidopsis can be refuted.

Potato plants and their tubers in Egypt are affected by one of the most renowned soil-borne pathogen, Ralstonia solanacearum, that caused brown rot in potato tubers and wilt in plants. There is no efficient therapeutic bactericide so; control of bacterial wilt is very rough. The study investigated three different concentrations of seven essential plant oils under in vitro and in vivo conditions as a result of their effects on Ralstonia solanacearum growth and their possibility use as potato seed pieces dressing for controlling bacterial wilt disease incidence. In vitro, anise oil at the three tested different concentrations (0.04, 0.07, and 0.14% vol/vol) was the most effective one inhibiting the growth of T4 and W9 isolates of Ralstonia solanacearum then pursued by thyme, lemongrass, and clove oils. On the other hand, rocket oil at the tested concentration was the least effective one followed by fennel oil. However, wheat germ oil was not completely effective. In vivo, experiment revealed that anise oil at the three concentrations significantly reduced disease incidence and severity in sponta and hermes potato cultivars and their effect was associated with increase of peroxidase, polyphenoloxidase, phenols and the foliar fresh weight of treated plants as well as the weight of tubers/plant followed by thyme and lemongrass oils compared to the infected untreated control. Morphological differences in bacterial cell structure have been observed using a transmission electron microscope (TEM). Anise oil at higher concentration caused of cell wall rupture and degraded cellular components.

Boron (B) is an essential micronutrient for vascular plant growth. Both B deficiency and toxicity can impair tissue development in diverse plant species, but little is known about the effect of B on reproductive panicle development and grain yield. In this study, a mutant of Setaria italica exhibiting necrotic panicle apices was identified and designated as sibor1. Sequencing revealed a candidate gene, SiBOR1, with a G-to-A alteration at the seventh exon. Knockout transgenic lines generated by clustered regularly interspaced short palindromic repeats and their associated protein-9 also had necrotic panicles, verifying the function of SiBOR1. SiBOR1 encoded a membrane-localized B efflux transporter, co-orthologous to the rice BOR1 protein. SiBOR1 was dominantly expressed in panicles and displayed a distinct expression pattern from those of its orthologs in other species. The induced mutation in SiBOR1 caused a reduction in the B content of panicle primary branches, and B deficiency-associated phenotypes such as thicker cell walls and higher cell porosity compared with Yugu 1. Transcriptome analysis indicated that differentially expressed genes involved in cell wall biogenesis, jasmonic acid synthesis, and programmed cell death response pathways were enriched in sibor1. qPCR analysis identified several key genes, including phenylalanine ammonia-lyase (SiPAL) and jasmonate-ZIM-domain (SiJAZ) genes, responsive to B-deficient conditions. These results indicate that SiBOR1 helps to regulate panicle primary branch development to maintain grain yield in S. italica. Our findings shed light on molecular mechanisms underlying the relationship between B transport and plant development in S. italica.

Bacterial wilt is one of the main diseases of Solanum spp., which caused by Ralstonia solanacearum (RS), formerly known as Pseudomonas solanacearum. Different concentrations of chitosan nanoparticles have been evaluated as one of the alternative methods of disease management in vitro and in vivo to reduce the risks of pesticide residues. Results in vitro experiment indicated that RS5 isolate was the most virulence one compared to RS1 and RS3. Increasing concentration of nano-chitosan, lead to increase inhibition zone, and this was observed at higher concentrations (100 and 200 µg/ml). In vivo results showed the highest concentration of spraying chitosan nanoparticles increase percentage reduction of disease incidence and severity, in effected potato and tomato plants. Recorded data of disease incidence and severity in treated potato plants were 78.93 % and 71.85 %, while on tomato plants were 81.64 % and 77.63 %, respectively compared to untreated infected potato plants were recorded 15.38 %, 20.87 %, and tomato plants were 20.98 % and 28.64 %. Results also revealed that 100 µg/ml of chitosan nanoparticles the lowest treatments used as soil amended curative treatments led to incease percentage reduction of disease incidence and severity, respectively on potato and tomato plants, but less than preventive treatment. The results registered that on potato plant were 54.93 % and 52.65 %, whilst recorded on tomato plants were 59.93 % and 56.74 %. Transmission electron microscopy (TEM) micrpgraphs illustrated that morphological of healthy R. solanacearum cells were undesirably stained with uranyl. The electron-dense uranyl acetate dye was limited to the cell surface slightly than the cytoplasm, which designated the integrity of the cell film of healthy cells. While bacterial cells treated with nano-chitosan, showed modification in the external shape, such as lysis of the cell wall and loss of cell flagella. Also, the result of using Random amplified polymorphic DNA (RAPD)-PCR observed that differences in treated Ralstonia solanancearum genotype by nano-chitosan compared to the genotype of the same untreated isolate.

The cytosol of eukaryotic host cells is an intrinsically hostile environment for bacteria. Understanding how cytosolic pathogens adapt to and survive in the cytosol is critical to developing novel therapeutic interventions for these pathogens. The cytosolic pathogen Listeria monocytogenes requires glmR (previously known as yvcK), a gene of unknown function, for resistance to cell wall stress, cytosolic survival, inflammasome avoidance and ultimately virulence in vivo. A genetic suppressor screen revealed that blocking utilization of UDP-GlcNAc by a non-essential wall teichoic acid decoration pathway restored resistance to cell wall stress and partially restored virulence of ΔglmR mutants. In parallel, metabolomics revealed that ΔglmR mutants are impaired in the production of UDP-GlcNAc, an essential peptidoglycan and wall teichoic acid (WTA) precursor. We next demonstrated that purified GlmR can directly catalyze the synthesis of UDP-GlcNAc from GlcNAc-1P and UTP, suggesting that it is an accessory uridyltransferase. Biochemical analysis of GlmR orthologues suggest that uridyltransferase activity is conserved. Finally, mutational analysis resulting in a GlmR mutant with impaired catalytic activity demonstrated that uridyltransferase activity was essential to facilitate cell wall stress responses and virulence in vivo. Taken together these studies indicate that GlmR is an evolutionary conserved accessory uridyltransferase required for cytosolic survival and virulence of L. monocytogenes.ImportanceBacterial pathogens must adapt to their host environment in order to cause disease. The cytosolic bacterial pathogen Listeria monocytogenes requires a highly conserved protein of unknown function, GlmR (previously known as YvcK) to survive in the host cytosol. GlmR is important for resistance to some cell wall stresses and is essential for virulence. The ΔglmR mutant is deficient in production of an essential cell wall metabolite, UDP-GlcNAc, and suppressors which increase metabolite levels also restore virulence. Purified GlmR can directly catalyze the synthesis of UDP-GlcNAc and this enzymatic activity is conserved in pathogens from Firmicutes and Actinobacteria phyla. These results highlight the importance accessory cell wall metabolism enzymes in responding to cell wall stress in a variety of bacterial pathogens.

Fusarium moniliforme (F. moniliforme) and its secondary metabolite fumonisin pose a severe threat to food safety, and searching for effective antimicrobial agents is a focus of current research. In this study, the secondary structure of Sub3 was analyzed by circular dichroism, meanwhile, the inhibition rate of Sub3 against F. moniliforme infection of maize was studied. To explore the possible inhibition mechanisms, morphological and structural changes of spores treated with Sub3 at 0, 1/2MIC and MIC were observed by scanning electron microscopy and transmission electron microscopy; the cell wall integrity, membrane integrity, reactive oxygen species, mitochondrial membrane potential, ATP synthase activity, redox reactions, and the nuclear damage of F. moniliforme were also investigated. The results showed that Sub3 was mostly in the state of random in deionized water, while mainly showed the β-sheet structure in the hydrophobic environment of 50% TFE solution, indicating that Sub3 might generate partial structure deformation when acting on the cell membrane; and its minimum inhibitory concentration (MIC) on F. moniliforme spores was 0.2 g/L. Under the 1/2MIC and MIC, the inhibition rates of Sub3 against F. moniliforme infected maize were 34.3% and 75.6%, respectively. The results of inhibition mechanisms revealed that the defective pathogenicity of F. moniliforme caused by Sub3 was attributed to damages on both the cell wall and the cell membrane, which might upset balance of intracellular redox system and mitochondrial energy metabolism and trigger nucleus damage, ultimately leading to cell death. The results provided direct evidence for inhibition of F. moniliforme infection of maize by Sub3, and useful knowledge applicable for food preservation.

The conserved Tol-Pal trans-envelope complex is important for outer membrane (OM) stability and cell division in Gram-negative bacteria. It has been proposed to mediate OM constriction during cell division via tethering to the cell wall. Yet, recent studies suggest that the complex has additional roles in OM lipid homeostasis and septal cell wall separation. How the Tol-Pal complex functions to facilitate these many processes is unclear. To gain insights into its role(s), we applied transposon insertion sequencing, and report here a detailed network of genetic interactions with the tol-pal locus in Escherichia coli. We found one positive and >20 negative strong interactions based on fitness. Disruption of genes responsible for osmoregulated periplasmic glucan biosynthesis restores fitness and OM barrier function, but not cell division defects, in tol-pal mutants. In contrast, deletions of genes involved in OM homeostasis and cell wall remodelling give rise to synthetic growth defects in strains lacking Tol-Pal, especially exacerbating OM barrier and/or cell division defects. Notably, the ΔtolA mutant having additional defects in OM protein assembly (ΔbamB) exhibited severe division phenotypes, even under conditions where the single mutants divide normally; this highlights the possibility for OM phenotypes to indirectly influence the cell division process. Overall, our work provides insights into the intricate nature of Tol-Pal function, and reinforces the model that this complex plays crucial roles in cell wall-OM tethering, cell wall remodelling, and in particular, OM homeostasis.

4-Vinylguaiacol (4-VG) is one of the most common off-flavors found in sake. 4-VG is produced from its precursor, ferulic acid, which is a component of the cell wall of the rice endosperm. The release of ferulic acid in sake brewing is thought to be mediated by feruloyl esterase produced by either Aspergillus oryzae or Saccharomyces cerevisiae. To investigate the effect of FaeA, a feruloyl esterase produced by A. oryzae, its loss-of-function strain was produced by genome co-editing. The feruloyl esterase activity of the faeA-deficient strain was drastically reduced. Sake was fermented using koji with S. cerevisiae strain G046, which can convert ferulic acid to 4-VG. Fermented sake was analyzed by measuring the 4-VG content and sensory evaluation. 4-VG content was reduced to approximately 10% of that of sake fermented with control koji. Sensory evaluation revealed that 4-VG was almost undetectable. Our findings showed that disruption of faeA in A. oryzae is a promising strategy to reduce 4-VG off-flavors in sake.

Abstract: The bacterial cell wall peptidoglycan (PG) is a dynamic structure that is constantly synthesized, re-modeled and degraded during bacterial division and growth. Postsynthetic modifications modulate the action of endogenous autolysis during PG lysis and remodeling for growth and sporulation, but also they are a mechanism used by pathogenic bacteria to evade the host innate immune system. Modifications of the glycan backbone are limited to the C-2 amine and C-6 hydroxyl moieties of either GlcNAc or MurNAc residues. This paper reviews the functional roles and properties of peptidoglycan de-Nacetylases (distinct PG GlcNAc and MurNAc deacetylases) and recent progress through genetic studies and biochemical characterization to elucidate their mechanism of action, 3D structures, substrate specificities and biological functions. Since they are virulence factors in pathogenic bacteria, peptidoglycan deacetylases are potential targets for the design of novel antimicrobial agents.

The main defense response to Soybean mosaic virus (SMV) infection in soybean [Glycine max (L.) Merr.] is thought to be blockage of intercellular virus transport by callose deposition on plasmodesmata. But the specific regulatory mechanism remains largely unknown. In this study, we found that hydrogen peroxide (H2O2) signal downstream of NO was associated with the regulation of callose accumulation. Abundant H2O2 was produced on the cell membrane and cell wall in the incompatible combination of soybean cultivar Jidou 7 and SMV strain N3, whereas no obvious H2O2 was observed in the compatible combination of Jidou 7 and strain SC-8. When H2O2 production was inhibited, callose accumulation induced by SMV infection decreased to a level insufficient to restrict virus transport in the incompatible combination. The H2O2-associated transcriptome dynamics of soybean during SMV infection was investigated. Transcriptome and functional analysis using virus-induced gene silencing showed that GmSEOB and GmPAP27, two genes regulated by H2O2, functioned in resistance by positively regulating the accumulation of callose in response to SMV infection. These results lay a foundation for further research on the signal transduction and molecular regulation of callose deposition during soybean resistance to SMV infection.

The fungal cell wall and membrane are the principal targets of antifungals. Herein, we report that myricetin exerts antifungal activity against Candida albicans by damaging the cell wall integrity and notably enhancing the membrane permeability. In the presence of sorbitol, an osmotic protectant, the minimum inhibitory concentration (MIC) of myricetin against C. albicans increased from 20 to 40 and 80 ?g/ml in 24 and 72 h, respectively, demonstrating that myricetin disturbs the cell wall integrity of C. albicans. Fluorescence microscopic images showed the presence of propidium iodide-stained C. albicans cells, indicating the myricetin-induced initial damage of the cell membrane. The effects of myricetin on the membrane permeability of C. albicans cells were assessed using crystal violet-uptake and intracellular material-leakage assays. The percentage uptakes of crystal violet for myricetin-treated C. albicans cells at 1×, 2×, and 4× the MIC of myricetin were 36.5, 60.6, and 79.4%, respectively, while those for DMSO-treated C. albicans cells were 28.2, 28.9, and 29.7%, respectively. Additionally, myricetin-treated C. albicans cells showed notable DNA and protein leakage, compared with the DMSO-treated controls. Furthermore, treatment of C. albicans cells with 1× the MIC of myricetin showed a 17.2 and 28.0% reduction in the binding of the lipophilic probes diphenylhexatriene and Nile red, respectively, indicating that myricetin alters the lipid components or order in the C. albicans cell membrane, leading to increased membrane permeability. Therefore, these data will provide insights into the pharmacological worth of myricetin as a prospective antifungal for treating C. albicans infections.

Secondary cell walls (SCWs) in stem xylem vessel and fibre cells enable plants to withstand the enormous compressive forces associated with upright growth. It remains unclear if xylem vessel and fibre cells can directly sense mechanical stimuli and modify their SCW during development. We provide evidence that Arabidopsis SCW-specific Fasciclin-Like Arabinogalactan-proteins 11 (FLA11) and 12 (FLA12) are possible cell surface sensors regulating SCW development in response to mechanical stimuli. Plants overexpressing FLA11 (OE-FLA11) showed earlier SCW development compared to the wild-type (WT) and altered SCW properties that phenocopy WT plants under compression stress. By contrast, OE-FLA12 stems showed higher cellulose content compared to WT plants, similar to plants experiencing tensile stress. fla11, OE-FLA11, fla12, and OE-FLA12 plants showed altered SCW responses to mechanical stress compared to the WT. Quantitative polymerase chain reaction (qPCR) and RNA-seq analysis revealed the up-regulation of genes and pathways involved in stress responses and SCW synthesis and regulation. Analysis of OE-FLA11 nst1 nst3 plants suggests that FLA11 regulation of SCWs is reliant on classical transcriptional networks. Our data support the involvement of FLA11 and FLA12 in SCW sensing complexes to fine-tune both the initiation of SCW development and the balance of lignin and cellulose synthesis/deposition in SCWs during development and in response to mechanical stimuli.

This study aimed to examine the antifungal activity of wild bergamot (Monarda fistulosa) essential oil on Colletotrichum musae and Lasiodiplodia theobromae, the causative organisms of anthracnose and crown-rot diseases of banana fruits, respectively. Chitin synthase is a promising target for antifungal compounds, since it is involved in synthesizing chitin, which forms a major proportion of the fungal cell wall. Disc volatilisation method was employed to assess the in vitro antifungal activity of the oil. The vapours of this essential oil at 4 μL per Petriplate exhibited 100% growth inhibition of both the fungal pathogens, and at 66.66 μLL−1 , it significantly (P

Filamentous fungi are used as production hosts for various commercially valuable enzymes and chemicals including organic acids and secondary metabolites. We previously revealed that α-1,3-glucan and galactosaminogalactan (GAG) contribute to hyphal aggregation in the industrial fungus Aspergillus oryzae, and that production of recombinant protein in shake-flask culture is higher in a mutant lacking both α-1,3-glucan and GAG (AGΔ-GAGΔ) than in the parental strain. Here, we compared the productivity of the wild type, AGΔ-GAGΔ, and mutants lacking α-1,3-glucan (AGΔ) or GAG (GAGΔ) in batch culture with intermittent addition of glucose in a 5-L lab-scale bioreactor. The hyphae of the wild type and all mutants were dispersed by agitation, although the wild type and AGΔ formed small amounts of aggregates. Although mycelial weight was similar among the strains, the concentration of a secreted recombinant protein (CutL1) was the highest in AGΔ-GAGΔ. Evaluation of fluid properties revealed that the apparent viscosities of mycelial cultures of the wild type and AGΔ-GAGΔ decreased as the agitation speed was increased. The apparent viscosity of the AGΔ-GAGΔ culture tended to be lower than that of the wild-type strain at each agitation speed, and was significantly lower at 600 rpm. Overall, the lack of α-1,3-glucan and GAG in the hyphae improved culture rheology, resulting in an increase in recombinant protein production in AGΔ-GAGΔ. This is the first report of flow behavior improvement by a cell-surface component defect in a filamentous fungus.

In this study, major factors involved in regulating ethanol production from wood by the saprotrophic white-rot fungus Phanerochaete sordida YK-624 were investigated. P. sordida YK-624 produced ethanol from wood meal culture without the addition of any nutrients, and ethanol was produced from wood culture only when the oxygen concentration in headspace was reduced to ≤5%; thereafter, ethanol production ceased within a few days. Analyses of gene expression during aerobic incubation indicated that P. sordida simultaneously upregulates the glycolytic pathway from sugar uptake to pyruvate conversion during ethanol fermentation and suppresses pyruvate influx into the TCA cycle. Upon termination of ethanol fermentation, the expression of all tested genes was repressed, and the fungus ceased to grow. In contrast, the fungus could utilize ethanol for aerobic growth. These results suggest that ethanol fermentation by P. sordida functions as a short-term stress response system under anaerobic conditions during wood decay, enabling the fungus to rapidly resume growing when oxygen is supplied (e.g., following breakdown of plant cell walls or removal of the fungus from water immersion). This is the first report to describe the physiologic significance of ethanol fermentation in saprotrophic white-rot fungi.

Root associated bacteria were isolated from Suaeda nudiflora and two isolates were selected for this study: rhizospheric Bacillus megaterium and endophytic Pseudomonas aeruginosa. These isolates were inoculated into maize variety Narmada Moti during its germination. TTC (2, 3, 5-triphenyl tetrazolium chloride) staining was used to confirm the association of the isolates with the maize root. The effects of these root associated bacteria were tested alone and in combinations for cell wall reinforcement and the induction of defense enzymes such as phenylalanine ammonia lyase (PAL) and β-1,3-glucanase in the presence of fungal pathogen Aspergillus niger in maize. The results indicated that the rhizospheric bacteria had a greater fight response to fungal infection than the endophhytic bacteria due to cell wall lignification as well as the rapid induction of higher concentrations of defense related enzymes.

Yeast cells, fundamental in the brewery and bakery industry and present in human nutrition for thousands of years, from an attractive and novel encapsulation matrix. The structure of the yeast cells, which includes the thick and mechanically strong cell wall and the lipid plasma membrane, allows the encapsulation of both hydrophobic and hydrophilic active ingredients and provides high loading capacity. Yeast encapsulation mainly involves whole cells or cell walls of Saccharomyces cerevisiae, although Candida utilis, Kluyveromyces fragilis, Torulopsis lipofera, Endomyces vernalis, and oleaginous yeast (Cryptococcus curvatus) have also been used for encapsulation. The encapsulation process is simple and cost-effective and the yeast capsules prepared provide particular thermo-stability during heat processes, which is extremely beneficial for volatile compounds. Yeast encapsulation is also profitable for stabilization of the active compounds against light or oxygen damage, for along-lasting flavor release, and for targeted oral delivery.

The widespread availability of nano-enabled products in the global market may lead to the release of a substantial amount of engineered nanoparticles in the environment, which frequently display drastically different physiochemical properties than their bulk counterparts. The purpose of the study was to evaluate the impact of citrate-stabilised silver nanoparticles (AgNPs) on the plant Arabidopsis thaliana at three levels, physiological phytotoxicity, cellular accumulation and subcellular transport of AgNPs. The monodisperse AgNPs of three different sizes (20, 40 and 80 nm) aggregated into much larger sizes after mixing with quarter-strength Hoagland solution and became polydisperse. Immersion in AgNP suspension inhibited seedling root elongation and demonstrated a linear dose–response relationship within the tested concentration range. The phytotoxic effect of AgNPs could not be fully explained by the released silver ions. Plants exposed to AgNP suspensions bioaccumulated higher silver content than plants exposed to AgNO3 solutions (Ag+ representative), indicating AgNP uptake by plants. AgNP toxicity was size and concentration dependent. AgNPs accumulated progressively in this sequence: border cells, root cap, columella and columella initials. AgNPs were apoplastically transported in the cell wall and found aggregated at plasmodesmata. In all the three levels studied, AgNP impacts differed from equivalent dosages of AgNO3.

Mixed-linkage glucan (MLG), which is widely distributed in grasses, is a polysaccharide highly abundant in cell walls of grass endosperm and young vegetative tissues. Lichenases are enzymes that hydrolyze mixed-linkage glucan first identified in mixed-linkage glucan rich lichens. In this study, we identify a gene encoding a lichenase we name Brachypodium distachyon LICHENASE 1 (BdLCH1), which is highly expressed in the endosperm of germinating seeds and coleoptiles and at lower amounts in mature shoots. RNA in situ hybridization showed that BdLCH1 is primarily expressed in chlorenchyma cells of mature leaves and internodes. Disruption of BdLCH1 resulted in an eight-fold increase in mixed-linkage glucan content in senesced leaves. Consistent with the in situ hybridization data, immunolocalization results showed that mixed-linkage glucan was not removed in chlorenchyma cells of lch1 mutants as it was in wild type and implicate the BdLCH1 enzyme in removing mixed-linkage glucan in chlorenchyma cells in mature vegetative tissues. We also show that mixed-linkage glucan accumulation in lch1 mutants was resistant to dark induced degradation, and eight-week-old lch1 plants showed a faster rate of starch breakdown than wild type in darkness. Our results suggest a role for BdLCH1 in modifying the cell wall to support highly metabolically active cells.

The cell walls of plants are mainly made of cellulose and contain a large number of calories. However, the main component, cellulose, is an indigestible plant fiber that is thought to be difficult for humans to use as energy. Herbivores acquire energy through the degradation of cell wall-derived dietary fiber by microorganisms in the digestive tract. Herbivores, especially horses, have a highly developed cecum and large intestine, and plants are fermented for their efficient use with the help of microorganisms. Humans also have an intestinal tract with a wide lumen on the proximal side of the large intestine, in which fermentation occurs. The digestive process of horses is similar to that of humans, and many of the intestinal bacteria found in horses that degrade plants are also found in humans. Therefore, it is thought that humans also obtain a certain amount of energy from cell wall-derived dietary fiber. However, the intake of dietary fiber by modern humans is low; thus, the amount of calories derived from indigestible plant fiber is considered to be very low. Cellulose in the plant cell wall is often accompanied by hemicellulose, pectin, lignin, suberin, and other materials. These materials are hard to degrade, and cellulose is therefore difficult for animals to utilize. If the cell wall can be degraded to some extent by cooking, it is thought that humans can obtain calories from cell wall-derived dietary fiber. If humans can use the calories from the cell wall for their diet, it may compensate for human food shortages.

Background Phomopsis stem-end rot caused by Diaporthe citri, causes significant commercial postharvest losses of pummelo fruit during storage. Carvacrol (CVR) is a known generally recognized as safe (GRAS) and has the ability to prolong the preservation of harvested fruits. In the present study, the inhibitory effects of CVR treatment at the appropriate concentration on Phomopsis stem-end rot development of harvested pummelo fruit inoculated with D. citri were evaluated by the amounts of cell wall components, the activities and gene expressions of related enzymes involved in cell wall modification and lignin biosynthesis. Results Results indicated that CVR completely inhibited D. citri growth in vitro at 200 mg L-1 and significantly controlled Phomopsis stem-end rot development in harvested pummelo. The CVR treatment delayed peel softening and browning, and retarded electrolyte leakage, O2 •- production, and malondialdehyde content. The CVR-treated fruit maintained higher amounts of cell wall materials (CWM), protopectin, hemicelluloses, and cellulose, but exhibited lower water-soluble pectin (WSP) amount. Moreover, in D. citri-inoculated fruit, CVR treatment suppressed the activities and gene expressions of cell wall disassembling-enzymes, including pectin methylesterase, polygalacturonase, cellulase, and β-galactosidase, while the development of cell wall degradation was reduced. Meanwhile, the CVR treatment enhanced the lignin biosynthesis by increasing the activities and up-regulating the gene expressions of phenylalanine ammonialyase, cinnamic alcohol dehydrogenase, and peroxidase accompanied with elevated level of lignin in pummelo fruit. Conclusion The disease resistance to D. citri in pummelo fruit elicited by CVR treatment is related to delaying cell wall degradation and enhancing lignin biosynthesis. This article is protected by copyright. All rights reserved.

Lodging is a major problem limiting maize yield worldwide. However, the mechanisms of lodging resistance remain incompletely understood for maize. Here, we evaluated 443 maize accessions for lodging resistance in the field. Five lodging-resistant accessions and five lodging-sensitive accessions were selected for further research. The leaf number, plant height, stem diameter, and rind penetrometer resistance were similar between lodging-resistant and -sensitive inbred lines. The average thickness of sclerenchymatous hypodermis layer was thicker and the vascular area was larger in the lodging-resistant lines compared with lodging-sensitive lines. Although total lignin content in stem tissue did not significantly differ between lodging-resistant and -sensitive lines, phloroglucinol staining revealed that the lignin content of the cell wall in the stem cortex and in the stem vascular tissue near the cortex was higher in the lodging-resistant lines than in the lodging-sensitive lines. Analysis of strand-specific RNA-seq transcriptome showed that a total of 793 genes were up-regulated and 713 genes were down-regulated in lodging-resistant lines relative to lodging-sensitive lines. The up-regulated genes in lodging-resistant lines were enriched in cell wall biogenesis. These results indicated that modification of cell wall biosynthesis would contribute to lodging resistance of maize.

Fusarium crown rot (FCR), typically caused by Fusarium pseudograminearum, is a severe soil-borne disease that, in recent years, has become an emerging threat to Chinese wheat crops. For the first time in this study, we investigated and compared the proteomic characteristics of two Chinese wheat varieties (04 Zhong 36 and Xinmai 26) at 24, 48, and 72 h post-inoculation using label-free quantitative proteomic analysis. A total of 9 234 proteins were successfully quantified, of which 783 were differentially expressed after inoculation. These proteins were mainly involved in metabolic, single-organism, and cellular processes. Thirty-three proteins associated with defense, cell wall formation, photosynthesis, etc., showed consistently different expression between the two genotypes at multiple time points. In particular, chitinase, which degrades chitin in the fungal cell wall and limits fungal growth, was exclusively and consistently upregulated in 04 Zhong 36 across the three time points. Other proteins such as flavonoid O-methyltransferase, glycosyltransferase, and peroxidase were only upregulated in 04 Zhong 36, and proteins, including the berberine bridge enzyme and rubisco large subunit-binding protein, were specifically downregulated in Xinmai 26. The expression of transcripts encoding eight selected proteins through qRT-PCR analysis supported the proteomic profiles. Overall, the results of this study allow us to understand FCR resistance in wheat at the protein level. Some proteins and their corresponding genes may be useful resources for the genetic improvement of FCR resistance in wheat.

Protein glycosylation is one of the ubiquitous post-translational modifications in eukaryotic cells, which play important roles in plant growth and adverse response. In this study, we performed the first comprehensive wheat plasma membrane N-glycoproteome analysis under drought stress via glycopeptide HILIC enrichment and LC-MS/MS identification. In total, 414 glycosylated sites corresponding to 407 glycopeptides and 312 unique glycoproteins were identified, of which 173 plasma membrane glycoproteins with 215 N-glycosylation sites were significantly regulated by drought stress. Functional enrichment analysis reveals that the significantly regulated N-glycosylation proteins were particularly related to protein kinase activity involved in the reception and transduction of extracellular signal and plant cell wall remolding. The motifs and sequence structures analysis showed that the significantly regulated N-glycosylation sites were concentrated within [NxT] motif, and 79.5% of them were located on the random coil that is always on the protein surface and flexible regions, which could facilitate protein glycosylated modification and enhance protein structural stability via reducing protein flexibility. PNGase F enzyme digestion and glycosylation site mutation further indicated that N-glycosylated modification could increase protein stability. Therefore, N-glycosylated modification is involved in plant adaptation to drought stress by improving the stability of cell wall remodeling related plasma membrane proteins.

With active physiological and biochemical activities, tissue-specific protoplasts from cambial derivatives, could serve as a specific source for information on xylogenesis for softwood species resistant to stable genetic transformation and lacking available mutants. In this study, protoplasts were isolated from developing xylem of the Chinese red pine, Pinus massoniana, by enzymolysis. High-quality RNAs were extracted from developing xylem and their protoplasts for constructing transcriptome libraries. Using Illumina HiSeq 2500 PE150 platform, a total of 362,328,426 clean paired-end reads (54.35G) were generated from multiple cDNA libraries and assembled into 146,422 unigenes. The transcriptome data were further analysed to identify 1567 differentially expressed genes (DEGs) between the isolated protoplasts and developing xylem of P. massoniana (Masson pine), 1126 DEGs were upregulated in protoplasts relative to developing xylem cells and 441 were downregulated. Most of the differentially expressed genes in biological process terms are related to plant response, which may be due to the response to cell wall removal. Further, the expression pattern of 71 unigenes involved in lignin biosynthesis was verified by RNA-seq. This study is the first to report the transcriptome profiles of the developing xylem and its protoplasts of coniferous trees, which provide a new perspective and valuable resource for tracking transcriptional regulatory events in wood formation of Masson pine.

Cell wall modifications are of pivotal importance during plant development. Among cell wall components, xyloglucans are the major hemicellulose polysaccharide in primary cell walls of dicots and non-graminaceous monocots. They can connect the cellulose microfibril surface to affect cell wall mechanical properties. Changes in xyloglucan structure are known to play an important role in regulating cell growth. Therefore, the degradation of xyloglucan is an important modification that alters the cell wall. The α-XYLOSIDASE1 (XYL1) gene encodes the only α-xylosidase acting on xyloglucans in Arabidopsis thaliana. Here, we showed that mutation of XYL1 strongly influences seed size, seed germination, and fruit elongation. We found that the expression of XYL1 is directly regulated in developing seeds and fruit by the MADS-box transcription factor SEEDSTICK. We demonstrated that XYL1 complements the stk smaller seed phenotype. Finally, by atomic force microscopy, we investigated the role of XYL1 activity in maintaining cell stiffness and growth, confirming the importance of cell wall modulation in shaping organs.

Peptidoglycan—a mesh sac of glycans that are linked by peptides—is the main component of bacterial cell walls. Peptidoglycan provides structural strength, protects cells from osmotic pressure and contributes to shape. All bacterial glycans are repeating disaccharides of N-acetylglucosamine (GlcNAc) β-(1–4)-linked to N-acetylmuramic acid (MurNAc). Borrelia burgdorferi, the tick-borne Lyme disease pathogen, produces glycan chains in which MurNAc is occasionally replaced with an unknown sugar. Nuclear magnetic resonance, liquid chromatography–mass spectroscopy and genetic analyses show that B. burgdorferi produces glycans that contain GlcNAc–GlcNAc. This unusual disaccharide is chitobiose, a component of its chitinous tick vector. Mutant bacteria that are auxotrophic for chitobiose have altered morphology, reduced motility and cell envelope defects that probably result from producing peptidoglycan that is stiffer than that in wild-type bacteria. We propose that the peptidoglycan of B. burgdorferi probably evolved by adaptation to obligate parasitization of a tick vector, resulting in a biophysical cell-wall alteration to withstand the atypical torque associated with twisting motility., Tick chitobiose is co-opted to build the cell wall of Lyme disease pathogen Borrelia burgdorferi.

One strategy for improving the treatability of refractory wood species is biological incising, and its efficiency depends on how the microorganisms modify the porous structure of the wood. Evaluation of the bioincised wood treatability on a micro-scale can thus help to better understand the treatability enhancing mechanisms. In the present study, the biodegradation pattern and micro-scale treatability of Loblolly pine (Pinus taeda L.) heartwood were determined after bioincising with the white-rot fungus Physisporinus vitreus (Pers.: Fr.) P. Karsten isolate 136 and bacterium Bacillus subtilis UTB22. Oven-dried specimens with dimensions of 50 mm × 25 mm × 15 mm (L × T × R) were incubated with the microorganisms at (23±2) °C and (65±5) % relative humidity for six weeks. The control and exposed wood blocks were then pressure treated by 1 % fluorescent dye (fluorescein)-containing water to study the treatability pattern under a fluorescence microscope. The longitudinal and tangential air permeability and compression strength parallel to the grain of the specimens were also determined at the end of the incubation period. Scanning electron microscopic (SEM) studies showed that degradation by B. subtilis UTB22 was limited to the pit membranes, but the cell walls were also degraded to some extent by P. vitreus. The fungus caused a higher mass loss compared to the bacterium, whereas the permeability enhancing ability of the bacterium was more pronounced. The fluorescent dye tracer also showed that higher treatability with more uniformity was obtained by B. subtilis UTB22. The improvement in treatability by both microorganisms was mainly due to the degradation of the earlywood tracheids., Jedna od metoda poboljšanog impregniranja slabo poroznih vrsta drva jest tretiranje drva mikroorganizmima, a cilj tog postupka jest promjena porozne strukture drva. Evaluacija tretmana drva mikroorganizmima može pridonijeti boljem razumijevanju mehanizama poboljšanja prodora zaštitnog sredstva u drvo. U ovom je istraživanju promatrana biorazgradnja i impregnacija srži lobodijskog bora (Pinus taeda L.) na mikrorazini nakon izloženosti drva gljivi bijele truleži Physisporinus vitreus (Pers.: Fr.) P. Karsten izolat 136 i bakteriji Bacillus subtilis UTB22. Apsolutno suhi uzorci drva dimenzija 50 mm × 25 mm × 15 mm (L × T × R) zaraženi su mikroorganizmima pri 23±2 °C i 65±5 % relativne vlažnosti zraka tijekom šest tjedana. Nakon toga kontrolni i zaraženi blokovi drva tlačnim su postupkom impregnirani fluorescentnim bojilom otopljenim u vodi (1 %-tni fluorescein) kako bi se pod fluorescentnim mikroskopom promotrio učinak tretmana drva mikroorganizmima. Na kraju inkubacije utvrđena je propusnost zraka u uzdužnom i tangentnom smjeru te izmjerena čvrstoća na tlak paralelno s vlakancima. Istraživanje pretražnim elektronskim mikroskopom (SEM) pokazalo je da je razgradnja bakterijom B. subtilis UTB22 ograničena na membrane jažica, dok su stanične stijenke u određenoj mjeri razgrađene djelovanjem gljive P. vitreus. Gljiva je uzrokovala opsežniji gubitak mase nego bakterija, dok je sposobnost bakterije da poveća propusnost bila veća. Fluorescentno bojilo također je pokazalo da je bakterijom B. subtilis UTB22 postignuta bolja propusnost, s većom ujednačenošću. Poboljšanje propusnosti primjenom obaju mikroorganizama uglavnom je posljedica razgradnje traheida ranog drva.

Arabinogalactan-proteins (AGPs) are complex, hyperglycosylated plant cell wall proteins with little known about the biological roles of their glycan moieties in sexual reproduction. Here, we report that GLCAT14A, GLCAT14B and GLCAT14C, three enzymes responsible for the addition of glucuronic acid residues to AGPs, function in pollen development, polytubey block, and normal embryo development in Arabidopsis. Using biochemical and immunolabelling techniques, we demonstrated that the loss of function of the GLCAT14A, GLCAT14B and GLCAT14C genes resulted in disorganization of the reticulate structure of the exine wall, abnormal development of the intine layer, and collapse of pollen grains in glcat14a/b and glcat14a/b/c mutants. Synchronous development between locules within the same anther was also lost in some glcat14a/b/c stamens. In addition, we observed excessive attraction of pollen tubes targeting glcat14a/b/c ovules, indicating that the polytubey block mechanism was compromised. Monosaccharide composition analysis revealed significant reductions in all sugars in glcat14a/b and glcat14a/b/c mutants except for arabinose and galactose, while immunolabeling showed decreased amounts of AGP sugar epitopes recognized by glcat14a/b and glcat14a/b/c mutants compared to wild type. This work demonstrates the important roles that AG glucuronidation plays in Arabidopsis sexual reproduction and reproductive development. Supporting Information.

Inula viscosa (syn Dittrichia viscosa) (sticky fleabane) from the Asteraceae family, is well-known for its pharmacological activities. In this research, anatomical characterization of leaves and st...

Cell wall polysaccharide biosynthesis enzymes play important roles in plant growth, development and stress responses. The functions of cell wall polysaccharide synthesis enzymes in plant growth and development have been well studied. In contrast, their roles in plant responses to environmental stress are poorly understood. Previous studies have demonstrated that the rice cell wall cellulose synthase-like D4 protein (OsCSLD4) is involved in cell wall polysaccharide synthesis and is important for rice growth and development. This study demonstrated that the OsCSLD4 function-disrupted mutant nd1 was sensitive to salt stress, but insensitive to abscisic acid (ABA). The expression of some ABA synthesis and response genes was repressed in nd1 under both normal and salt stress conditions. Exogenous ABA can restore nd1-impaired salt stress tolerance. Moreover, overexpression of OsCSLD4 can enhance rice ABA synthesis gene expression, increase ABA content and improve rice salt tolerance, thus implying that OsCSLD4-regulated rice salt stress tolerance is mediated by ABA synthesis. Additionally, nd1 decreased rice tolerance to osmotic stress, but not ion toxic tolerance. The results from the transcriptome analysis showed that more osmotic stress-responsive genes were impaired in nd1 than salt stress-responsive genes, thus indicating that OsCSLD4 is involved in rice salt stress response through an ABA-induced osmotic response pathway. Intriguingly, the disruption of OsCSLD4 function decreased grain width and weight, while overexpression of OsCSLD4 increased grain width and weight. Taken together, this study demonstrates a novel plant salt stress adaptation mechanism by which crops can coordinate salt stress tolerance and yield.

The transformation of plants from juveniles to adults is a key process in plant growth and development, and the main regulatory factors are miR156 and SQUAMOSA promoter binding protein-like (SPL) transcription factors. Lilium is an ornamental bulb, but it has a long maturation time. In this experiment, Lilium bulbs were subjected to a temperature treatment of 15 °C for 4 weeks to initiate vegetative phase change. Transmission electron microscopy indicated the cell wall of bud core tissue undergoing vegetative phase change became thinner, the starch grains were reduced, and the growth of the juvenile stage was accelerated. The key transcription factors LbrSPL9 and LbrSPL15 were cloned, and the phylogenetic analysis showed they possessed high homology with other plant SPLs. Subcellular localization and transcription activation experiments confirmed LbrSPL9 and LbrSPL15 were mainly located in the nucleus and exhibited transcriptional activity. The results of in situ hybridization showed the expression levels of LbrSPL9 and LbrSPL15 were increased after temperature change treatment. The functional verification experiment of the transgenic plants confirmed that the overexpression of LbrSPL9 and LbrSPL15 could shorten maturation time. These findings help elucidate the regulatory mechanisms of phase transition in Lilium and provide a reference for breeding research in other bulbous flowers.

Two novel species of the genus Deinococcus, designated SYSU M49105T and SYSU M42101T, were isolated from freshwater samples of the Pearl River estuary in Guangdong, China. Phylogenetic analysis using 16S rRNA gene sequence indicated that strains SYSU M49105T and SYSU M42101T showed the highest sequence similarities to Deinococcus aetherius JCM 11751 T (93.6%) and Deinococcus multiflagellatus NBRC 112888 T (97.3%), respectively. Cells of both strains were Gram-staining positive, aerobic, coccus-shaped, oxidase-negative and non-motile. The cell wall contained meso-diaminopimelic acid as their diagnostic diamino acid. MK-8 was the predominant respiratory quinone for both strains. The polar lipid profile of SYSU M49105T contained two unidentified phosphoglycolipids, nine unidentified glycolipids, and five unidentified polar lipids. SYSU M42101T had one unidentified phosphoglycolipid, nine unidentified glycolipids, one unidentified aminophospholipid and four unidentified polar lipids. The major fatty acids of strains SYSU M49105T and SYSU M42101T were summed feature 3 (C16:1 ω7c and/ or C16:1 ω6c) and C16:0. The G + C contents of the novel isolates based on genomic DNAs were 69.6% and 67.4%, respectively. On the basis of phenotypic, genotypic and phylogenetic data, strains SYSU M49105T and SYSU M42101T should be considered to represent two novel species in the genus Deinococcus, for which the names Deinococcus aestuarii sp. nov. and Deinococcus aquaedulcis sp. nov. were proposed with the type strains SYSU M49105T (= KCTC 43258 T = CGMCC 1.18609 T) and SYSU M42101T (= KCTC 43257 T = CGMCC 1.18614 T), respectively.

Almost half of the world’s arable land has acidic pH. Aluminum salts present in acid soils dissociate to release Al3+ ions in the soil solution that inhibit root growth causing severe loss in crop yields. Aluminium toxicity accounts for the second highest loss in plant productivity after drought. Aluminium in high doses causes damage to the plant cell wall, cytoskeleton and DNA. One of the ways by which plants alleviate aluminium toxicity is by the exudation of citrate from the roots that chelates the free Al3+ and prevents its entry into the plant. In several crop plants Multidrug and Toxic Compound Extrusion (MATE) transporters regulate citrate exudation from the roots. The MATE proteins are ubiquitously present in bacteria, archaea, fungi, animals and plants. The origin and evolution of these membrane transporters in plants is not well known. Here, using protein sequence information we identify MATE transporters in major groups of land plants and their algal ancestors. Our study indicates that the MATE family members expanded in number and functionally diverse among the land plants. We also identify motifs present across the streptophyte clade and a conserved aspartate residue that might regulate citrate exudation. This study can provide leads to engineer MATE transporters to confer enhanced tolerance in acid soils.

ALUMINUM-ACTIVATED MALATE TRANSPORTER1 (ALMT1)-mediated malate exudation from roots is critical for aluminium (Al) resistance in Arabidopsis. Its upstream molecular signalling regulation is not yet well understood. The role of CALMODULIN-LIKE24 (CML24) in Al-inhibited root growth and downstream molecular regulation of ALMT1-meditaed Al resistance was investigated. CML24 confers Al resistance demonstrated by an increased root-growth inhibition of the cml24 loss-of-function mutant under Al stress. This occurs mainly through the regulation of the ALMT1-mediated malate exudation from roots. The mutation and overexpression of CML24 leads to an elevated and reduced Al accumulation in the cell wall of roots, respectively. Al stress induced both transcript and protein abundance of CML24 in root tips, especially in the transition zone. CML24 interacts with CALMODULIN BINDING TRANSCRIPTION ACTIVATOR2 (CAMTA2) and promotes its transcriptional activity in the regulation of ALMT1 expression. This results in an enhanced malate exudation from roots and less root-growth inhibition under Al stress. Both CML24 and CAMTA2 interacted with WRKY46 suppressing the transcriptional repression of ALMT1 by WRKY46. The study provides novel insights into understanding of the upstream molecular signalling of the ALMT1-depdendent Al resistance.

Cytochimera potato plants, which mixed with diploid and tetraploid cells, could cause the highest and significantly increased biomass yield than the polyploid and diploid potato plants. Polyploidization is an important approach in crop breeding for agronomic trait improvement, especially for biomass production. Cytochimera contains two or more mixed cells with different levels of ploidy, which is considered a failure in whole genome duplication. Using colchicine treatment with diploid (Dip) potato (Solanum chacoense) plantlets, this study generated tetraploid (Tet) and cytochimera (Cyt) lines, which, respectively, contained complete and partial cells with genome duplication. Compared to the Dip potato, we observed remarkably enhanced plant growth and biomass yields in Tet and Cyt lines. Notably, the Cyt potato straw, which was generated from incomplete genome doubling, was of significantly higher biomass yield than that of the Tet with a distinctively altered cell wall composition. Meanwhile, we observed that one layer of the tetraploid cells (about 30%) in Cyt plants was sufficient to trigger a gene expression pattern similar to that of Tet, suggesting that the biomass dominance of Cyt may be related to the proportion of different ploidy cells. Further genome-wide analyses of co-expression networks indicated that down-regulation (against Dip) of spliceosomal-related transcripts might lead to differential alternative splicing for specifically improved agronomic traits such as plant growth, biomass yield, and lignocellulose composition in Tet and Cyt plants. In addition, this work examined that the genome of Cyt line was relatively stable after years of asexual reproduction. Hence, this study has demonstrated that incomplete genome doubling is a promising strategy to maximize biomass production in potatoes and beyond.

To decarbonize the building sector, the use of durable wood materials must be increased. Inspiration for environmentally benign wood protection systems is sought in durable tree species depositing phenolic extractives in their heartwood. Based on the hypothesis that the micro-distribution of extractives influences durability, we compared the natural impregnation patterns of non-durable, but readily available Norway spruce to more durable Kurile larch by mapping the distribution of heartwood extractives with Confocal Raman Imaging and multivariate data decomposition. Phenolics of both species were associated with hydrophobic oleoresin, likely facilitating diffusion through the tissue. They accumulated preferentially in lignin-rich sub-compartments of the cell wall. Yet, the distribution of extractives was found not to be the same. The middle lamellae contained flavonoids in larch and aromatic waxes in spruce, which was also found in rays and epithelial cells. Spruce-lignans were tentatively identified in all cell types, while larch-flavonoids were not present in resin channels, hinting at a different origin of synthesis. Larch-oleoresin without flavonoids was only found in lumina, indicating that the presence of phenolics in the mixture influences the final destination. Together our findings suggest, that spruce heartwood-defense focuses on water regulation, while the more efficient larch strategy is based on antioxidants.

As an essential element, copper (Cu) is involved in various metabolic processes in plants. However, this metal becomes a potential stress factor when the concentration reaches the threshold level. A comparative analysis was conducted to investigate the potential role of two cytokinins [kinetin (KIN) and 6-benzylaminopurine (BAP)] in Cu stress alleviation in Ricinus communis seedlings by assessing the metal bioaccumulation, antioxidation mechanisms, anatomical changes as well as analysis of various essential elements and bioactive compounds in the cotyledonary leaves. Application of KIN and BAP regulated the Cu toxicity in castor seedlings via modulation of metal uptake and antioxidation mechanisms. The increase in antioxidant enzyme activities during Cu stress was further increased upon exposure to cytokinin treatments, and the enhancement reached up to 25 folds for CAT, 16 folds for POD, and 8 folds for SOD over the control, which helps the plant to alleviate the toxic effects of Cu stress. The FTIR analysis of the cotyledonary leaves revealed that the functional groups associated with cell wall materials contributed to the Cu sequestration in the cell wall, supported by the xylem wall thickening as observed in the SEM analysis. The GCMS analysis revealed that most of the secondary metabolites identified were phenolic and flavonoid compounds with antioxidant properties, which help the castor seedlings withstand the Cu stress, and the presence of these compounds was more prominent in cytokinin-treated seedlings. Findings revealed that BAP was more effective than KIN to withstand the Cu stress effects in castor seedlings.

Lipopolysaccharides (LPS) are cell wall components from Gram-negative bacteria and are composed of three covalently linked regions: the O-antigen, the core oligosaccharide, and the lipid A moiety, which carries most of their endotoxic activity. The objective of this study was to isolate and compare the lipid A structures from ruminal LPS derived from total mixed ration (TMR)- and pasture-fed cows, by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Ruminal bacteria were collected from two rumen-cannulated Holstein cows; one fed a TMR (60:40, forage–concentrate) and the other pasture fed. The representativeness of each sample was validated by comparing the rumen microbiome from the cows in our study to the core rumen microbiome from the previous literature. Lipopolysaccharides from each respective sample were extracted with a phenol–water extraction procedure and purified via ultracentrifugation. To isolate lipid A from the core and O-antigen, pure ruminal LPS samples were hydrolyzed with acetic acid. Lipid A derived from the TMR-fed cow potentially exhibited a tetra-acylated structure, whereas lipid A derived from the pasture-fed cow potentially exhibited a penta-acylated lipid A structure. Both samples were quantified using limulus amebocyte lysate (LAL) assay and exhibited low endotoxic activity, consistent with the MALDI-TOF MS observations. Results indicate that the lipid A acylation pattern differs between diets, and that ruminal bacteria express solely under-acylated lipid A structures contrary to hexa-acylated lipid A, typically expressed by bacteria such as E. coli.

Protein A (SpA) is one of the most importantStaphylococcus aureuscell wall proteins. It includes five immunoglobulin (Ig)-binding domains which can bind to immune complexes through the Fc region of immunoglobulins. The binding of SpA to the polymeric supports can be used to prepare affinity chromatography resins, which are useful for immunoprecipitation (IP) of antibodies. Protein A is also used to purify many anti-cancer antibodies. In this study, SpA was displayed on the surface ofBacillus subtiliscells using a sortase-mediated system to display the target protein to theB. subtiliscell wall. A series of plasmids consisting of cassettes for cell wall-directed protein A as well as negative controls were constructed and transformed intoB. subtilisWASD (wprA sigD) cells. SDS-PAGE, western blot, flow cytometry, functional IgG purification assay, and a modified ELISA assay were used to confirm the surface display of SpA and evaluate its function. Semi-quantitative ELISA results showed that the binding capacity of lyophilizedBs-SpA is 100 μg IgG from rabbit serum per 1 mg of cells under optimal experimental conditions. Low production costs, optimal performance, and the use of a harmless strain compared to a similar commercial product predict the possible use of SpA immobilization technology in the future.

BackgroundApplication of the wheat phytohormones abscisic acid (ABA) or gibberellic acid (GA) affect Fusarium head blight (FHB) disease severity; however, the molecular underpinnings of the elicited phenotypes remain unclear. Herein, the transcriptomic responses of an FHB-susceptible wheat cultivar ‘Fielder’ were characterized upon treatment with ABA, an ABA receptor antagonist (AS6), or GA in the presence or absence of Fusarium graminearum (Fg) challenge.ResultsA total of 30,876 differentially expressed genes (DEGs) where identified in ‘Fielder’ (26,004) and Fg (4,872). Fg challenge alone resulted in the most substantial wheat DEGs contributing to 57.2% of the total transcriptomic variation. Using a combination of topology overlap and correlation analyses, 9,689 Fg-related wheat DEGs were defined. Further enrichment analysis of the top 1% networked wheat DEGs identified critical expression changes within defense responses, cell structural metabolism, molecular transport, and membrane/lipid metabolism. Fg-challenged conditions also included the expression of a putative Fg ABA-biosynthetic cytochrome P450 and repression of wheat FUS3 for dysregulating ABA and GA crosstalk. ABA treatment alone elicited 4536 (32%) wheat DEGs common to those of the Fg-challenge, and Fg+ABA further enhanced 888 (12.5%) of them. These ABA elicited DEGs are involved in defense through both classical and non-classical phytohormone signaling and regulating cell wall structures including polyphenolic metabolism. Conversely, Fg+GA opposed 2239 (33%) Fg-elicited wheat DEGs, including modulating primary and secondary metabolism, defense responses, and flowering genes. ABA and jointly ABA⍰Fg⍰[Fg+ABA] treatments repressed, while Fg+GA induced an over-representation of wheat DEGs mapping to chromosome 6BL. Finally, compared to Fg+ABA, co-application of Fg+AS6 did not antagonize ABA biosynthesis or signal but rather elicited antagonistic Fg (557) and wheat (11) DEGs responses directly tied to stress responses, phytohormone transport, and FHB.ConclusionsComparative transcriptomics highlight the effects of wheat phytohormones on individual pathway and global metabolism simultaneously. Application of ABA may reduce FHB severity through misregulating defense mechanisms and cell wall fortification pathways. GA application may alter primary and secondary metabolism, creating a metabolic shift to ultimately reduce FHB severity. By comparing these findings to those previously reported for four additional plant genotypes, an additive model of the wheat-Fg interaction is proposed.

Sugar content is one of the most important quality traits of tomato. Cell wall invertase promotes sucrose unloading in the fruit by maintaining a gradient of sucrose concentration between source leaves and fruits, while invertase inhibitor (INVINH) regulates this process. In this study, knock-out of cell wall INVINH in tomato (SlINVINH1) was performed by genome editing using, CRISPR/Cas9 and Target-AID technologies. Most of the genome-edited lines set higher soluble solid content (SSC) fruit than the original cultivar ‘Suzukoma’, while fruit weight was different among the genome-edited lines. From these genome-edited lines, three lines (193–3, 199–2, and 247–2), whose SSC was significantly higher than ‘Suzukoma’ and fruit weight were almost the same as the original cultivar, were selected. The fruit weight and overall plant growth of the two lines were comparable to those of the original cultivar. In contrast, the fructose and glucose contents in the mature fruits of the two lines were significantly higher than those of the original cultivar. The mature fruits of genome edited line 193–3 showed the highest sugar content, and the fructose and glucose contents were 29% and 36% higher than that of the original cultivar, respectively. Whole genome sequence data showed no off-target mutations in the genome-edited lines. Non-target metabolome analysis of mature fruits revealed that fructose was the highest loading factor in principal component analysis (PCA) between the genome-edited line and the original cultivar, and no unexpected metabolites appeared in the genome-edited line. In this study, we succeeded in producing tomato lines with high sugar content without a decrease in fruit weight and deterioration of plant growth by knock-out of SlINVINH1 using genome editing technology. This study showed that functional disruption of SlINVINH1 is an effective approach to produce tomato cultivars with high sugar content.

Net blotch, caused by the ascomycete Drechslera teres, can compromise barley production. Beneficial bacteria strains are of substantial interest as biological agents for plant protection in agriculture. Belonging to the genus Paraburkholderia, a bacterium, referred to as strain B25, has been identified as protective for barley against net blotch. The strain Paraburkholderia phytofirmans (strain PsJN), which has no effect on the pathogen's growth, has been used as control. In this study, the expression of target genes involved in cell wall-related processes, defense responses, carbohydrate and phenylpropanoid pathways was studied under various conditions (with or without pathogen and/or with or without bacterial strains) at different time-points (0-6-12-48 h). The results show that specific genes were subjected to a circadian regulation and that the expression of most of them increased in barley infected with D. teres and/or bacterized with the strain PsJN. On the contrary, a decreased gene expression was observed in the presence of strain B25. To complement and enrich the gene expression analysis, untargeted metabolomics was carried out on the same samples. The data obtained show an increase in the production of lipid compounds in barley in the presence of the pathogen. In addition, the presence of strain B25 leads to a decrease in the production of defense compounds in this crop. The results contribute to advance the knowledge on the mechanisms occurring at the onset of D. teres infection and in the presence of a biocontrol agent limiting the severity of net blotch in barley.

Lectins are characterized of the carbohydrate-binding ability and play comprehensive roles in fungal physiology (e.g., defense response, development and host–pathogen interaction). Beauveria bassiana, a filamentous entomopathogenic fungus, has a lectin-like protein containing a Fruit Body_domain (BbLec1). BbLec1 could bind to chitobiose and chitin in fungal cell wall. BbLec1 proteins interacted with each other to form multimers, and translocated into eisosomes. Further, the interdependence between BbLec1 and the eisosome protein PliA was essential for stabilizing the eisosome architecture. To test the BbLec1 roles in B. bassiana, we constructed the gene disruption and complementation mutants. Notably, the BbLec1 loss resulted in the impaired cell wall in mycelia and conidia as well as conidial formation capacity. In addition, disruption of BbLec1 led to the reduced cytomembrane integrity and the enhanced sensitivity to osmotic stress. Finally, ΔBbLec1 mutant strain displayed the weakened virulence when compared with the wild-type strain. Taken together, BbLec1 traffics into eisosome and links the functionality of eisosome to development and virulence of B. bassiana.