Layers Of Dead Cells Research Articles

Fungi in ant-plant interactions: a key to enhancing plant nutrient-acquisition strategies.

This article is a Commentary on Gegenbauer et al. (2023), 238: 2210–2223.

Peridermal fruit skin formation in Actinidia sp. (kiwifruit) is associated with genetic loci controlling russeting and cuticle formation

BackgroundThe skin (exocarp) of fleshy fruit is hugely diverse across species. Most fruit types have a live epidermal skin covered by a layer of cuticle made up of cutin while a few create an outermost layer of dead cells (peridermal layer).ResultsIn this study we undertook crosses between epidermal and peridermal skinned kiwifruit, and showed that epidermal skin is a semi-dominant trait. Furthermore, backcrossing these epidermal skinned hybrids to a peridermal skinned fruit created a diverse range of phenotypes ranging from epidermal skinned fruit, through fruit with varying degrees of patches of periderm (russeting), to fruit with a complete periderm. Quantitative trait locus (QTL) analysis of this population suggested that periderm formation was associated with four loci. These QTLs were aligned either to ones associated with russet formation on chromosome 19 and 24, or cuticle integrity and coverage located on chromosomes 3, 11 and 24.ConclusionFrom the segregation of skin type and QTL analysis, it appears that skin development in kiwifruit is controlled by two competing factors, cuticle strength and propensity to russet. A strong cuticle will inhibit russeting while a strong propensity to russet can create a continuous dead skinned periderm.

Mangifera Indica mid-rib fibers as reinforcements for CNSL-epoxy composites

This research work explores the potential of Mangifera indica leaf mid-rib fibers as a reinforcement for polymer composites. The fibers were subjected to sodium bicarbonate treatment for 6 h, 12 h, 18 h, and 24 h duration and the effect of chemical treatment on the tensile strength of the fibers was evaluated. Maximum tensile strength of 104 MPa was obtained for 18 h treated fibers. Scanning electron microscopy of the fiber surface revealed surface impurities, layers of dead cells and degradation of fibrils, as some of the important features for variation in tensile strength. Fourier transform infrared spectra of all varieties of fibers supported function group modifications with chemical treatment. X-ray diffraction analysis of the fiber confirmed improvement in the crystallinity index of the fibers till duration of 18 h. Composites with untreated and 18 h treated fibers were considered for fabrication along with Cashew Nut Shell Liquid (CNSL)/epoxy matrix. Techniques such as hand layup and compression moulding were adopted for composite fabrication and mechanical properties of the cured composites were determined. Maximum values of tensile, flexural and impact strength obtained were 48 MPa, 70 MPa and 38 kJ/m2 respectively. Failure modes such as fiber pull-out, fiber shearing, fiber debonding and matrix shearing were observed through scanning electron micrographs of the Mangifera indica composites.

Reduction of ozone dosage by using ozone in ultrafine bubbles to reduce sludge volume

Sludge ozonation, which promotes sludge disintegration and solubilization, is a promising technology for reducing waste sludge volume from biological wastewater treatment process. However, if this technology is to be widely adopted, reducing the energy consumption associated with ozone generation will be necessary. We used ultra-fine bubbles (UFBs) as ozone carriers to determine if their use could improve the efficiency of ozone treatment and reduce the ozone dose required. We used a spiral, liquid-type UFB generator, which can introduce UFBs directly into a sludge suspension. The death ratio of bacteria in sludge was used as an indicator of sludge reduction. The ozone requirement was reduced by UFBs. The ozone consumption required to achieve a death ratio of 80% was 15 mg-O3/g-MLSS in the sludge treated with ozone supplied by UFBs versus 25 and 45 mg-O3/g-MLSS in sludges treated with ozone supplied as a spiral, liquid-type microbubbles and by a diffuser, respectively. When mixing water ozonated with UFBs with sludge, the depth of the dead cell layer from the surface to the interior of the sludge floc was larger than that of ozonated water lacking UFBs at the same rate of ozone consumption. Ozone in UFBs kills bacteria inside the flocs. However, the fragmentation of sludge flocs by shear forces in the UFB generator made a larger contribution to the acceleration of bacterial death in sludge treated with ozone supplied by UFBs.

A Layer of Dead Cells at the Periphery Protects Biofilms from Antimicrobial Peptides

Under stress or sub-optimal growth conditions, some bacterial species are able to aggregate into defensive structures known as biofilm. In biofilms, cells not only compete for resources, but also are able to benefit one another. To date, we do not have direct evidence on how cells in biofilm protect each other against antimicrobial peptides (AMPs). We have utilized a live microscopy platform to monitor the time-course of the attack of AMPs on Bacillus subtilis biofilms. Consistent with the recent discoveries of the absorption of AMPs into dead cells, we observe that cells on the periphery of a biofilm protect the interior cells by blocking the flux of AMP towards the interior of the biofilm. Our results demonstrate the dynamics of AMP activity on a dense microbial community where the growth and survival of the population is coordinated with various cell-to-cell communication processes. The experimental procedure and quantitative framework of our approach provides novel insight on how AMPs can be optimized for targeting bacterial populations rather than individual cells.

A review of current knowledge about the formation of native peridermal exocarp in fruit.

The outer skin layer in any plant is essential in offering a protective barrier against water loss and pathogen attack. Within fleshy fruit, the skin supports internal cell layers and can provide the initial cues in attracting seed-dispersing animals. The skin of a fruit, termed the exocarp, is a key element of consumer preference and a target for many breeding programs. Across fruiting species there is a huge diversity of exocarp types and these range from a simple single living cell layer (epidermis) often covered with a waxy layer, to complex multicellular suberised and dead cell layers (periderm), with various intermediate russet forms in between. Each exocarp can be interspersed with other structures such as hairs or spines. The epidermis has been well characterised and remains pluripotent with the help of the cells immediately under the epidermis. The periderm, in contrast, is the result of secondary meristematic activity, which replaces the epidermal layers, and is not well characterised in fruits. In this review we explore the structure, composition and mechanisms that control the development of a periderm type fruit exocarp. We draw upon literature from non-fleshy fruit species that form periderm tissue, from which a considerable amount of research has been undertaken.

Aggregation of Nanosized Hydroxyapatite Particles and Inhibition of Cell Adhesion on this Bio-Active Material as Key Factors that Limit its Biointegration

Abstract—Different synthetic calcium phosphate compounds are used as osteoplastic materials for filling of bone defects during surgical reconstruction of bone. Synthetic nanosized hydroxyapatite is of particular interest because of its similarity to the chemical composition of bone and its osteoinductive potential. However, it is also known that nanosized hydroxyapatite can inhibit the processes of biointegration of biomaterials, although we have no clear understanding of the mechanisms that are responsible for this effect. In our in vitro studies suppression of cell adhesion on nanosized hydroxyapatite that may lead to anoikis was shown. Studies in which rat subcutaneous heterotopic implantation methods were used showed that nanosized hydroxyapatite in vivo formed a slightly resorbable agglomerate that blocks cell migration into the material, as well as neocollagenesis processes in it, including the processes of encapsulation of the material. In this case, the processes of neocollagenesis along the periphery of the sample were observed only over a dense layer of dead cells (cellular debris) surrounding the agglomerated nanosized hydroxyapatite. Addition of demineralized bone collagen matrix to nanosized hydroxyapatite as a conductive component contributed to colonization of the material by the cells, resorption of nanosized hydroxyapatite, and active construction of the neocollagen matrix. These results show that modification of nanosized hydroxyapatite with a conductive component is required in order to prevent its agglomeration and thus increase its biointegration and osteoinductive potential in the body.

UV fluorescence excitation spectroscopy as a noninvasive predictor of epidermal proliferation and clinical performance of cosmetic formulations.

The epidermis is the outermost layer of skin and is composed of cells primarily containing keratin. It consists of about ten layers of living cells (keratinocytes) and ten layers of dead cells (corneocytes). Thinning of the epidermis and decreased proliferation of its cells are associated with aging related changes in skin, including wrinkling and laxity. Fluorescence excitation spectroscopy is a noninvasive method of monitoring characteristic excitation-emission peaks in skin that have been related to the epidermal and dermal composition. The magnitude of the peak that occurs at 295nm excitation (F295) has been linked to changes in epidermal thickness, proliferation, and skin aging. The goal of this study is to correlate changes in the F295 signal with proliferation of cells and thickening of the epidermis induced by cosmetic formulations. We hypothesize that two commonly used cosmetic ingredients, retinol and glycolic acid, will increase these markers that have been implicated in skin anti-aging. In a placebo-controlled study subjects' forearms were treated with formulations containing retinol or glycolic acid under occlusive patch for a period of 21 days. Skin fluorescence was measured at baseline and after treatment, and biopsies were taken following treatment for histological analysis of epidermal thickness and cell proliferation. After 21 days of treatment retinol and glycolic acid formulas significantly increased F295 (by 265.1±33.5% and 162.2±18.7% respectively), whereas the placebo control formula did not induce a change from baseline. Furthermore, retinol and glycolic acid treatments significantly increased epidermal thickness (by 63.1% and 7.8% respectively) and keratinocyte proliferation (by 236.9% and 62.8% respectively) versus placebo control. Increases in F295 were found to correlate with epidermal renewal, but more so with increased cell proliferation than epidermal thickness. We conclude that the F295 signal is a fast and reliable early indicator of epidermal remodeling in skin that can be used to distinguish between formulations with different cosmetic ingredients.

Epidermal maturation, vitamin C and hyaluronan metabolism

Epidermis, the surface layer of human skin, is formed by cells called keratinocytes. They divide (making more cells) in the deeper parts of the skin and gradually move towards the surface. Keratinocytes eventually form a layer of dead cells on the skin's surface, and this layer is the main protective barrier against things that might damage or irritate the skin, such as chemicals and germs. A viscous substance called hyaluronan surrounds keratinocytes in the lower, living cell layers. All kinds of injuries to the epidermis, like wounding, infection, and sunburn, trigger a strong inflammatory response by keratinocytes. This response includes increased division of keratinocytes, and their movement to help the injury site recover, as well as increased hyaluronan production and turnover. In many skin diseases the keratinocyte injury‐response is excessive, or even completely unfounded, and harmful. Therefore, factors and treatments that calm down this reaction are potential medicines for some skin diseases. This study from Kuopio, Finland, studied hyaluronan metabolism (i.e. breakdown) in a special model developed in the laboratory, in which keratinocytes separate to produce a normal‐looking epidermis. This model allowed the researchers to demonstrate that vitamin C suppresses both production and degradation of hyaluronan in the epidermis. Since rapid hyaluronan metabolism is an indicator of inflammation, vitamin C – which is an antioxidant ‐ might have a use for suppressing this reaction in keratinocytes. This study calls for more work on the vitamin's mechanism of action, and possible roles in clinical practice.

Influence of Atmospheric Pressure Non-thermal Plasma on Inactivation of Biofilm Cells

The objective of this study is to investigate the bactericidal efficiency of atmospheric pressure non-thermal (cold) dielectric barrier discharge plasma on biofilms of Staphylococcus aureus and Escherichia coli. In general, cold plasma is a mixture of electrons, ions, neutral atoms and molecules. The different particles in cold plasmas exhibit different energies, i.e. electrons are much more energetic than other particles. This feature of cold plasmas allows to produce chemically reactive species at near room temperature that can be used in biological applications. Bacteria inactivation was performed using the air direct plasma (with reactive species, UV light, excited species and electric fields). Discharge power density during the experiment was equal to 70 mW/cm2 and plasma dose was regulated by the treatment time. Bacterial biofilms were treated with the non-thermal plasma for 10–300 s. The most effective reduction in the number of S. aureus cells was found after 300 s of treatment and was 2.77 log10 that is 99.83%. When the biofilm of E. coli was used in the experiment, killing of bacteria was independent of the exposure time and the mortality of cells did not exceed 0.48 that is 66.7% kill. The lethal effect on E. coli and S. aureus cells were observed after 300 and 120 s of plasma treatment, respectively but it was necessary to remove the layers of dead cells. The proposed process of removing dead cell layers was carried out due to the probable shielding effect, i.e. dead cells prevent further penetration of active plasma species into the deeper layers of the biofilm. It was shown that the effectiveness of cell destruction by the non-thermal plasma depends on the thickness of biofilm.

Biofouling Mitigation in Forward Osmosis Using Graphene Oxide Functionalized Thin-Film Composite Membranes.

Forward osmosis (FO) is an emerging membrane process with potential applications in the treatment of highly fouling feedwaters. However, biofouling, the adhesion of microorganisms to the membrane and the subsequent formation of biofilms, remains a major limitation since antifouling membrane modifications offer limited protection against biofouling. In this study, we evaluated the use of graphene oxide (GO) for biofouling mitigation in FO. GO functionalization of thin-film composite membranes (GO-TFC) increased the surface hydrophilicity and imparted antimicrobial activity to the membrane without altering its transport properties. After 1 h of contact time, deposition and viability of Pseudomonas aeruginosa cells on GO-TFC were reduced by 36% and 30%, respectively, compared to pristine membranes. When GO-TFC membranes were tested for treatment of an artificial secondary wastewater supplemented with P. aeruginosa, membrane biofouling was reduced by 50% after 24 h of operation. This biofouling resistance is attributed to the reduced accumulation of microbial biomass on GO-TFC compared to pristine membranes. In addition, confocal microscopy demonstrated that cells deposited on the membrane surface are inactivated, resulting in a layer of dead cells on GO-TFC that limit biofilm formation. These findings highlight the potential of GO to be used for biofouling mitigation in FO.

Adaptive Evolution of Skin in Terrestrial Vertebrates and Possible Involvement of Endogenous Retroviruses.

The first terrestrial vertebrates emerged from water and adapted to living on land approximately 360 million years ago (late Devonian). In particular, amphibians are thought to have surface epithelia that changed from multilayered epithelia into keratinized stratified squamous epithelia by acquiring stratum corneum (SC), which is composed of several dead cell layers that serve as an air liquid interface barrier. Then, reptiles appeared and became a major terrestrial vertebrate group approximately 340 million years ago by forming hard SC. About 220 million years ago, mammals radiated by acquiring soft and moisturized SC, and endogenous retroviruses were thought to be actively integrated into mammalian genomes. Skin ASpartic Protease (SASPase)/ASPRV1 is the mammalian-specific endogenous retroviral-derived protease. SASPase-deficient mice had dry skin and aberrant accumulation of profilaggrin, which is another mammalian-specific gene that regulates SC barrier function and is a major predisposing factor for atopic dermatitis. These findings indicate that the retroviral element SASPase was integrated into the first mammalian species and was involved in the adaptive evolution of mammals, as it facilitates moisturization of skin SC. It is possible that other uncharacterized endogenous retroviruses were also involved in epidermal barrier function.

Evidence for Induction of a Tumor-Metastasis-Receptive Microenvironment in Bone Marrow and Other Organs As an Unwanted and Underestimated Side Effect of Chemotherapy/Radiotherapy

▪Background. There are well-known side effects of chemotherapy and radiotherapy that are mainly related to the toxicity and impaired function of vital organs; however, the induction by these therapies of expression of several pro-metastatic factors in various tissues and organs that in toto create a pro-metastatic microenvironment is still, surprisingly, not widely acknowledged. On the other hand, it is very well known that, after infusion into a host after myeloablative treatment by radio-chemotherapy, HSPCs home efficiently to BM in response to several chemotactic factors upregulated in the BM microenvironment.Hypothesis. To explain this phenomenon, we hypothesized that toxic damage in BM and other organs due to radio and/or chemotherapy administered for treatment of malignancies leads to upregulation in “bystander” tissues of several factors, such as chemokines, growth factors, bioactive phosphosphingolipids, and small- molecule alarmines, that attract normal circulating stem cells for regeneration but unfortunately also provide chemotactic signals to cancer cells that survived the initial treatment.Materials and Methods. The level of the most important chemotactic factors for cancer cells in organs exposed to radio/chemotherapy, such as stromal-derived factor 1 (SDF-1) and hepatocyte growth factor (HGF), were evaluated by RT-PCR and ELISA. The levels of sphingosine-1-phosphate, ceramide-1-phosphate, and selected alarmines (ATP, UTP) were evaluated by tandem mass spectrometry and/or ELISA. Human rhabdomyosarcoma and ovary cancer cell lines were employed as metastatic models in in vitro and in vivo assays.Results. We first observed that conditioned media from irradiated BM and other tissues chemoattract human cancer cells. Next, by employing a short in vivo tumor cell tissue-seeding assay we observed that increased numbers of human cancer cells seed (metastasize) to the BM and other organs in SCID/beige immunodeficient mice after irradiation or exposure to chemotherapy. Moreover, we observed that human cancer cells expanded more strongly in the presence of damaged/dying human BM cells, providing evidence that cells expand better on feeder layers of dead cells. Finally, we were able to ameliorate the in vitro migration of cancer cells to CM harvested from irradiated tissues or decrease seeding of in vivo-injected human cells into various organs in irradiated or chemotherapy-exposed mice by inhibiting particular chemoattractant–chemoattractant receptor axes (e.g., SDF-1–CXCR4 or S1P–S1P1R), and a very efficient protective effect was observed by reducing the pro-inflammatory response induced by radio/chemotherapy by administering non-steroid anti-inflammatory drugs or dexamethasone in vivo in experimental animals.Conclusions. We propose that a radio-chemotherapy-induced pro-metastatic microenvironment plays an important role in the metastasis of cancer cells to bones as well as other organs (lungs or liver), which are highly susceptible to chemotherapeutic agents or ionizing irradiation. This problem indicates the need to develop efficient anti-metastatic drugs that will work in combination with, or follow, standard therapies in order to prevent the possibility of therapy-induced spread of tumor cells that are resistant to treatment. Such cells possess characteristics of cancer stem cells and are highly migratory. Since many different chemoattractant-specific receptor axes are involved in this phenomenon, simple, intensive anti-inflammatory treatment by non-steroid agents to suppress induction of pro-metastatic factorsafter radio/chemotherapy seems to be an interesting alternative. DisclosuresNo relevant conflicts of interest to declare.

SLNs Based on Co-Processed Lipids for Topical Delivery of Terbinafine Hydrochloride

Onychomycosis is a fungal infection which is responsible for 50% of all nail disorders [1-3]. In 90% of the cases, onychomycosis is caused by dermatophytes (mainly Trichophyton rubrum), but can also be due to other fungi such as yeasts (mainly Candida albicans) or molds (mainly Scopulariopsis brevicaulis) [4]. Human nail, being composed of 80-90 layers of dead cells [5-7], prevents effective topical treatment of onychomycosis. Therefore, it is difficult to achieve therapeutic antimicrobial drug concentration in the nail. High incidences of onychomycosis and problems associated with its therapy have renewed the interest of formulation scientists to work towards alternative formulation approaches [8].

Deconstructing a plant macromolecular assembly: chemical architecture, molecular flexibility, and mechanical performance of natural and engineered potato suberins.

Periderms present in plant barks are essential protective barriers to water diffusion, mechanical breakdown, and pathogenic invasion. They consist of densely packed layers of dead cells with cell walls that are embedded with suberin. Understanding the interplay of molecular structure, dynamics, and biomechanics in these cell wall-associated insoluble amorphous polymeric assemblies presents substantial investigative challenges. We report solid-state NMR coordinated with FT-IR and tensile strength measurements for periderms from native and wound-healing potatoes and from potatoes with genetically modified suberins. The analyses include the intact suberin aromatic–aliphatic polymer and cell-wall polysaccharides, previously reported soluble depolymerized transmethylation products, and undegraded residues including suberan. Wound-healing suberized potato cell walls, which are 2 orders of magnitude more permeable to water than native periderms, display a strikingly enhanced hydrophilic–hydrophobic balance, a degradation-resistant aromatic domain, and flexibility suggestive of an altered supramolecular organization in the periderm. Suppression of ferulate ester formation in suberin and associated wax remodels the periderm with more flexible aliphatic chains and abundant aromatic constituents that can resist transesterification, attenuates cooperative hydroxyfatty acid motions, and produces a mechanically compromised and highly water-permeable periderm.

The Ecology of a Synthetic Biofilm: Synchronization without a Clock

Biofilms are the archetype for smart materials where microcolonies of live bacteria and layers of dead cells are encapsulated in a hydrated matrix of polysaccharides, proteins and exopolymeric substances. We engineered synthetic biofilms to study their ecology_i.e. the cells’ relationship to each other and their environment. In particular, we investigated the physical parameters governing prokayrotic cell-to-cell signaling in a vascularized model of a biofilm, comprised of bacteria that are genetically engineered to transmit and receive so-called quorum-sensing signals based on the lux operon functionally linked to fluorescent reporters. Numerical modeling of these experiments reveals that gene expression is vitally dependent on the location within the biofilm and proximity to microchannels in the array, elements easily accessible in the model. Moreover, we observe synchronization in fluorescence from the microcolonies comprising the biofilm in a repetitive, pulsatile environment as shown in the figure. We find that the concentration of luxR mRNA varies measurably with C6HSL concentration and time indicating autoregulation of luxR. Moreover, flow cytometry data identifies two phenotypes in the cell population: one with a high C6HSL threshold; another with a lower threshold, indicating bi-stability associated with positive autoregulation of luxR.View Large Image | View Hi-Res Image | Download PowerPoint Slide

The allergy gene: how a mutation in a skin protein revealed a link between eczema and asthma

Ichthyosis vulgaris is a common genetic skin disorder characterized by dry, scaly skin. About 1% of the European population have the full presentation of ichthyosis vulgaris; up to 10% have a milder, subclinical form. Atopic eczema is the most common, inflammatory skin condition, affecting 20% of children. It is often accompanied by a number of other allergies, including atopic asthma. Atopic eczema is a complex trait, where predisposing genes in combination with environmental stimuli produce the disease. Recently, we reported the first loss-of-function genetic mutations in the filaggrin gene as the cause of ichthyosis vulgaris. We noted people with ichthyosis vulgaris also have atopic eczema (and vice versa) and that the filaggrin gene sits in a known atopic eczema susceptibility locus. We went on to confirm that filaggrin mutations, carried by up to 10% of the population, are the major genetic predisposing factor for atopic eczema and the various allergies associated with atopic eczema. Filaggrin is a highly abundant protein expressed in the uppermost part of the epidermis that is critical to the formation and hydration of the stratum corneum—the outermost dead cell layers responsible for the barrier function of the skin. Filaggrin deficiency leads to a “leaky” skin barrier that allows higher than normal water loss (explaining the dry, scaly skin), as well as allowing entry of allergens through the epidermis where they trigger inflammatory and allergic immune responses (atopic eczema and allergies). This work has placed the skin barrier at the center stage of eczema and allergy research and has kick-started new therapy development programs aimed at repairing or enhancing skin-barrier function as a means of treating or preventing these very common diseases.

Novel Approaches to avoid Microbial Adhesion onto Biomaterials

Infections resulting from microbial adhesion to biomaterial surfaces have been observed on nearly all medical devices with severe economic and medical consequences [1]. Biofilm infections, mainly due to their antimicrobial resistance, pose a number of clinical challenges, including disease, chronic inflammation, and rapidly acquired antibiotic resistance. Independently of the superiority of the implant, virtually all medical devices are prone to microbial colonization and infection. Examples of such devices include prosthetic heart valves, orthopaedic implants, intravascular catheters, artificial hearts, left ventricular assist devices, cardiac pacemakers, vascular prostheses, cerebrospinal fluid shunts, urinary catheters, voice prostheses, ocular prostheses and contact lenses, and intrauterine contraceptive devices. A large amount of research to eliminate or reduce infections by developing anti-infective and anti-adhesive devices has been encouraged as a result of the significant resistance of biofilms to conventional antibiotic therapies. These improved devices may be produced by either mechanical design alternatives; physicochemical modification of the biomaterial surface (e.g. biosurfactants, plasma, atom transfer radical polymerization, brushes); anti-infective agents bound to the surface of the material (e.g. biosurfactants, silver, quaternary ammonium compounds, synthetic antibiotics); or release of toxic agents into the adjacent surroundings (e.g. chlorhexidine, antibiotics) [2,3]. The success of the mechanical design alternatives has been residual and with limited applicability [2]. Furthermore, the effectiveness of coatings designed to reduce adhesion by modification of the surface properties has also been reduced and greatly dependent on the bacterial species. Surfaces modified with poly(ethylene glycol) [4], poly(ethylene oxide) brushes [5], and hydrophilic polyurethanes [6], among many others, have been reported. Additionally, surface-bounded anti-infective agents are only toxic to the initial wave of incoming bacteria and provide little residual effects once layers of dead cells accumulate, which are also inflammatory.

Evidence that Cell Death is Associated with Zebra Chip Disease in Potato Tubers

Zebra chip (ZC) is an established and highly destructive disease of potato (Solanum tuberosum L.) that occurs in several southwestern states of the United States, Mexico, Central America, and New Zealand. The causal agent for this disease has not been identified. However, the bacterium “Candidatus Liberibacter solanacearum” and the potato psyllid, Bactericera cockerelli (Sulc), its insect vector, are associated with the disease. Tubers from ZC-affected potato plants exhibit dramatic browning of vascular tissue concomitant with “necrotic flecking” both of which can affect the entire tuber. Upon frying, these tubers develop a characteristic striped pattern of discoloration rendering them unmarketable. These characteristic ZC symptoms in the tubers have been suggested to be associated with general cell death, though no evidence to confirm this hypothesis has been shown. In order to determine if cell death is associated with ZC disease, a series of experiments were undertaken. Cell death was initially quantified by comparing cellular ion leakage from ZC-affected and ZC-free tubers. Levels of ion leakage were found to be significantly higher in ZC-affected tubers compared to ZC-free tubers. To examine further the association of cell death with ZC disease, ZC-affected and ZC-free tubers were compared using classical histochemical staining methods in conjunction with optical microscopy, which revealed layers of dead cells surrounding numerous, small, irregularly-shaped lesions throughout the parenchymatic medullary region, vascular ring and cortex of ZC-affected tubers. This cell death was confirmed using high-resolution, field-emission scanning electron microscopy (FE-SEM) of fresh-cut tuber tissue.

Strategic distribution of protective proteins within bran layers of wheat protects the nutrient-rich endosperm.

Bran from bread wheat (Triticum aestivum 'Babbler') grain is composed of many outer layers of dead maternal tissues that overlie living aleurone cells. The dead cell layers function as a barrier resistant to degradation, whereas the aleurone layer is involved in mobilizing organic substrates in the endosperm during germination. We microdissected three defined bran fractions, outer layers (epidermis and hypodermis), intermediate fraction (cross cells, tube cells, testa, and nucellar tissue), and inner layer (aleurone cells), and used proteomics to identify their individual protein complements. All proteins of the outer layers were enzymes, whose function is to provide direct protection against pathogens or improve tissue strength. The more complex proteome of the intermediate layers suggests a greater diversity of function, including the inhibition of enzymes secreted by pathogens. The inner layer contains proteins involved in metabolism, as would be expected from live aleurone cells, but this layer also includes defense enzymes and inhibitors as well as 7S globulin (specific to this layer). Using immunofluorescence microscopy, oxalate oxidase was localized predominantly to the outer layers, xylanase inhibitor protein I to the xylan-rich nucellar layer of the intermediate fraction and pathogenesis-related protein 4 mainly to the aleurone. Activities of the water-extractable enzymes oxalate oxidase, peroxidase, and polyphenol oxidase were highest in the outer layers, whereas chitinase activity was found only in assays of whole grains. We conclude that the differential protein complements of each bran layer in wheat provide distinct lines of defense in protecting the embryo and nutrient-rich endosperm.