Using the CODIT model to explain secondary metabolites of xylem in defence systems of temperate trees against decay fungi.

BACKGROUND: In trees, secondary metabolites (SMs) are essential for determining the effectiveness of defence systems against fungi and why defences are sometimes breached. Using the CODIT model (Compartmentalization of Damage/Dysfunction in Trees), we explain defence processes at the cellular level. CODIT is a highly compartmented defence system that relies on the signalling, synthesis and transport of defence compounds through a three-dimensional lattice of parenchyma against the spread of decay fungi in xylem. SCOPE: The model conceptualizes 'walls' that are pre-formed, formed during and formed after wounding events. For sapwood, SMs range in molecular size, which directly affects performance and the response times in which they can be produced. When triggered, high-molecular weight SMs such as suberin and lignin are synthesized slowly (phytoalexins), but can also be in place at the time of wounding (phytoanticipins). In contrast, low-molecular weight phenolic compounds such as flavonoids can be manufactured de novo (phytoalexins) rapidly in response to fungal colonization. De novo production of SMs can be regulated in response to fungal pathogenicity levels. The protective nature of heartwood is partly based on the level of accumulated antimicrobial SMs (phytoanticipins) during the transitionary stage into a normally dead substance. Effectiveness against fungal colonization in heartwood is largely determined by the genetics of the host. CONCLUSION: Here we review recent advances in our understanding of the role of SMs in trees in the context of CODIT, with emphasis on the relationship between defence, carbohydrate availability and the hydraulic system.We also raise the limitations of the CODIT model and suggest its modification, encompassing other defence theory concepts. We envisage the development of a new defence system that is modular based and incorporates all components (and organs) of the tree from micro- to macro-scales.

Mechanical properties and structure-function trade-offs in secondary xylem of young roots and stems.

Bending and torsional properties of young roots and stems were measured in nine woody angiosperms. The variation in mechanical parameters was correlated to wood anatomical traits and analysed with respect to the other two competing functions of xylem (namely storage and hydraulics). Compared with stems, roots exhibited five times greater flexibility in bending and two times greater flexibility in torsion. Lower values of structural bending and structural torsional moduli (Estr and Gstr, respectively) of roots compared with stems were associated with the presence of thicker bark and a greater size of xylem cells. Across species, Estr and Gstr were correlated with wood density, which was mainly driven by the wall thickness to lumen area ratio of fibres. Higher fractions of parenchyma did not translate directly into a lower wood density and reduced mechanical stiffness in spite of parenchyma cells having thinner, and in some cases less lignified, cell walls than fibres. The presence of wide, partially non-lignified rays contributed to low values of Estr and Gstr in Clematis vitalba. Overall, our results demonstrate that higher demands for mechanical stability in self-supporting stems put a major constraint on xylem structure, whereas root xylem can be designed with a greater emphasis on both storage and hydraulic functions.

Vessel diameter is related to amount and spatial arrangement of axial parenchyma in woody angiosperms.

Parenchyma represents a critically important living tissue in the sapwood of the secondary xylem of woody angiosperms. Considering various interactions between parenchyma and water transporting vessels, we hypothesize a structure-function relationship between both cell types. Through a generalized additive mixed model approach based on 2,332 woody angiosperm species derived from the literature, we explored the relationship between the proportion and spatial distribution of ray and axial parenchyma and vessel size, while controlling for maximum plant height and a range of climatic factors. When factoring in maximum plant height, we found that with increasing mean annual temperatures, mean vessel diameter showed a positive correlation with axial parenchyma proportion and arrangement, but not for ray parenchyma. Species with a high axial parenchyma tissue fraction tend to have wide vessels, with most of the parenchyma packed around vessels, whereas species with small diameter vessels show a reduced amount of axial parenchyma that is not directly connected to vessels. This finding provides evidence for independent functions of axial parenchyma and ray parenchyma in large vesselled species and further supports a strong role for axial parenchyma in long-distance xylem water transport.

Vessel-associated cells in angiosperm xylem: Highly specialized living cells at the symplast-apoplast boundary.

BACKGROUND: Vessel-associated cells (VACs) are highly specialized, living parenchyma cells that are in direct contact with water-conducting, dead vessels. The contact may be sparse or in large tight groups of parenchyma that completely surrounds vessels. VACs differ from vessel distant parenchyma in physiology, anatomy, and function and have half-bordered pits at the vessel-parenchyma juncture. The distinct anatomy of VACs is related to the exchange of substances to and from the water-transport system, with the cells long thought to be involved in water transport in woody angiosperms, but where direct experimental evidence is lacking. SCOPE: This review focuses on our current knowledge of VACs regarding anatomy and function, including hydraulic capacitance, storage of nonstructural carbohydrates, symplastic and apoplastic interactions, defense against pathogens and frost, osmoregulation, and the novel hypothesis of surfactant production. Based on microscopy, we visually represent how VACs vary in dimensions and general appearance between species, with special attention to the protoplast, amorphous layer, and the vessel-parenchyma pit membrane. CONCLUSIONS: An understanding of the relationship between VACs and vessels is crucial to tackling questions related to how water is transported over long distances in xylem, as well as defense against pathogens. New avenues of research show how parenchyma-vessel contact is related to vessel diameter and a new hypothesis may explain how surfactants arising from VAC can allow water to travel under negative pressure. We also reinforce the message of connectivity between VAC and other cells between xylem and phloem.

Corneoscleral stiffening increases IOP spike magnitudes during rapid microvolumetric change in the eye.

Factors governing the steady-state IOP have been extensively studied; however, the dynamic aspects of IOP are less understood. Clinical studies have suggested that intraocular pressure (IOP) fluctuation may be associated with glaucoma risk. This study aims to investigate how stiffening of corneoscleral biomechanical properties affects IOP spikes induced by rapid microvolumetric change. Porcine eyes (n = 25 in total) were subjected to volumetric infusions before and after external treatment of a circular area (11 mm diameter) in either the central cornea or posterior sclera. The treated area in the control group was immersed in phosphate-buffered saline (PBS) for 40 min, while the treated area of the chemical crosslinking group was immersed in 4% glutaraldehyde/PBS for 40 min. A subset of the sham-treated eyes was also subjected to volumetric infusions at a raised steady-state IOP. The magnitude of IOP spikes increased after localized chemical crosslinking of either the cornea (27.5% increase, p < 0.001) or the sclera (14.3% increase, p < 0.001) with corneal crosslinking having a stronger effect than scleral crosslinking (p = 0.018). We also observed that raising the steady-state IOP from 15 to 25 mmHg resulted in marked increase in IOP spike magnitudes by 63.9% (p < 0.001). These results suggested that an increased corneoscleral stiffness could significantly increase IOP spike magnitudes at the same volumetric change. Corneal stiffness appeared to have a strong impact on the IOP spike magnitude and may play a major role in regulating rapid volume-pressure dynamics. An increase in steady-state IOP also resulted in larger IOP fluctuations due to the increased "apparent" stiffness of the ocular shell, suggesting a potential interaction between the magnitude of IOP and its fluctuations. Corneoscleral properties may represent additional pathways for understanding and managing glaucoma risk and warrant future investigation.

A global analysis of parenchyma tissue fractions in secondary xylem of seed plants.

Parenchyma is an important tissue in secondary xylem of seed plants, with functions ranging from storage to defence and with effects on the physical and mechanical properties of wood. Currently, we lack a large-scale quantitative analysis of ray parenchyma (RP) and axial parenchyma (AP) tissue fractions. Here, we use data from the literature on AP and RP fractions to investigate the potential relationships of climate and growth form with total ray and axial parenchyma fractions (RAP). We found a 29-fold variation in RAP fraction, which was more strongly related to temperature than with precipitation. Stem succulents had the highest RAP values (mean ± SD: 70.2 ± 22.0%), followed by lianas (50.1 ± 16.3%), angiosperm trees and shrubs (26.3 ± 12.4%), and conifers (7.6 ± 2.6%). Differences in RAP fraction between temperate and tropical angiosperm trees (21.1 ± 7.9% vs 36.2 ± 13.4%, respectively) are due to differences in the AP fraction, which is typically three times higher in tropical than in temperate trees, but not in RP fraction. Our results illustrate that both temperature and growth form are important drivers of RAP fractions. These findings should help pave the way to better understand the various functions of RAP in plants.

Weak tradeoff between xylem safety and xylem-specific hydraulic efficiency across the world's woody plant species.

The evolution of lignified xylem allowed for the efficient transport of water under tension, but also exposed the vascular network to the risk of gas emboli and the spread of gas between xylem conduits, thus impeding sap transport to the leaves. A well-known hypothesis proposes that the safety of xylem (its ability to resist embolism formation and spread) should trade off against xylem efficiency (its capacity to transport water). We tested this safety-efficiency hypothesis in branch xylem across 335 angiosperm and 89 gymnosperm species. Safety was considered at three levels: the xylem water potentials where 12%, 50% and 88% of maximal conductivity are lost. Although correlations between safety and efficiency were weak (r(2)  < 0.086), no species had high efficiency and high safety, supporting the idea for a safety-efficiency tradeoff. However, many species had low efficiency and low safety. Species with low efficiency and low safety were weakly associated (r(2)  < 0.02 in most cases) with higher wood density, lower leaf- to sapwood-area and shorter stature. There appears to be no persuasive explanation for the considerable number of species with both low efficiency and low safety. These species represent a real challenge for understanding the evolution of xylem.

Are needles of Pinus pinaster more vulnerable to xylem embolism than branches? New insights from X-ray computed tomography.

Plants can be highly segmented organisms with an independently redundant design of organs. In the context of plant hydraulics, leaves may be less embolism resistant than stems, allowing hydraulic failure to be restricted to distal organs that can be readily replaced. We quantified drought-induced embolism in needles and stems of Pinus pinaster using high-resolution computed tomography (HRCT). HRCT observations of needles were compared with the rehydration kinetics method to estimate the contribution of extra-xylary pathways to declining hydraulic conductance. High-resolution computed tomography images indicated that the pressure inducing 50% of embolized tracheids was similar between needle and stem xylem (P50 needle xylem = -3.62 MPa, P50 stem xylem = -3.88 MPa). Tracheids in both organs showed no difference in torus overlap of bordered pits. However, estimations of the pressure inducing 50% loss of hydraulic conductance at the whole needle level by the rehydration kinetics method were significantly higher (P50 needle = -1.71 MPa) than P50 needle xylem derived from HRCT. The vulnerability segmentation hypothesis appears to be valid only when considering hydraulic failure at the entire needle level, including extra-xylary pathways. Our findings suggest that native embolism in needles is limited and highlight the importance of imaging techniques for vulnerability curves.

The amount of parenchyma and living fibers affects storage of nonstructural carbohydrates in young stems and roots of temperate trees.

PREMISE OF THE STUDY: Concentrations of nonstructural carbohydrates (NSCs) are used as proxies for the net carbon balance of trees and as indicators of carbon starvation resulting from environmental stress. Woody organs are the largest NSC-storing compartments in forest ecosystems; therefore, it is essential to understand the factors that affect the size of this important storage pool. In wood, NSC are predominantly deposited in ray and axial parenchyma (RAP); however, direct links between nutrient storage and RAP anatomy have not yet been established. Here, we tested whether the NSC storage capacity of wood is influenced by the amount of RAP. METHODS: We measured NSC concentrations and RAP fractions in root and stem sapwood of 12 temperate species sampled at the onset of winter dormancy and in stem sapwood of four tropical trees growing in an evergreen lowland rainforest. The patterns of starch distribution were visualized by staining with Lugol's solution. KEY RESULTS: The concentration of NSCs in sapwood of temperate trees scales tightly with the amount of RAP and living fibers (LFs), with almost all RAP and LFs being densely packed with starch grains. In contrast, the tropical species had lower NSC concentrations despite their higher RAP and LFs fraction and had considerable interspecific differences in starch distribution. CONCLUSIONS: The differences in RAP and LFs abundance affect the ability of sapwood to store NSC in temperate trees, whereas a more diverse set of functions of RAP might be pronounced in species growing in a tropical environment with little seasonality.

Three-Dimensional Strains in Human Posterior Sclera Using Ultrasound Speckle Tracking.

Intraocular pressure (IOP) induced strains in the peripapillary sclera may play a role in glaucoma progression. Using inflation testing and ultrasound speckle tracking, the 3D strains in the peripapillary sclera were measured in nine human donor globes. Our results showed that the peripapillary sclera experienced through-thickness compression and meridional stretch during inflation, while minimal circumferential dilation was observed when IOP was increased from 10 to 19 mmHg. The maximum shear was primarily oriented in the through-thickness, meridional cross sections and had a magnitude slightly larger than the first principal strain. The tissue volume had minimal overall change, confirming near-incompressibility of the sclera. Substantial strain heterogeneity was present in the peripapillary region, with local high strain areas likely corresponding to structural heterogeneity caused by traversing blood vessels. These 3D strain characteristics provide new insights into the biomechanical responses of the peripapillary sclera during physiological increases of IOP. Future studies are needed to confirm these findings and investigate the role of these biomechanical characteristics in ocular diseases.

Rapid measurement of individual cone photoreceptor pointing using focus diversity.

A novel method is presented to rapidly measure the pointing direction of individual human cone photoreceptors using adaptive-optics (AO) retinal imaging. For a fixed entrance pupil position, the focal plane is rapidly modulated to image the guided light in various axial planes. For cones with different pointing directions, this focus diversity will cause a shift in their apparent position, allowing for their relative pointing to be determined. For four normal human subjects, retinal images were acquired, registered, and the positions of individual cones tracked throughout the dataset. Variation in cone tilt was 0.02 radians, agreeing with other objective measurements on the same subjects at the same retinal locations.

Variation in Tracheid Dimensions of Conifer Xylem Reveals Evidence of Adaptation to Environmental Conditions.

Globally distributed extant conifer species must adapt to various environmental conditions, which would be reflected in their xylem structure, especially in the tracheid characteristics of earlywood and latewood. With an anatomical trait dataset of 78 conifer species growing throughout China, an interspecific study within a phylogenetic context was conducted to quantify variance of tracheid dimensions and their response to climatic and soil conditions. There was a significant difference in tracheid diameter between earlywood and latewood while no significant difference was detected in tracheid wall thickness through a phylogenetically paired t-test. Through a phylogenetic principle component analysis, Pinaceae species were found to be strongly divergent in their tracheid structure in contrast to a conservative tracheid structure in species of Cupressaceae, Taxaceae, and Podocarpaceae. Tracheid wall thickness decreased from high to low latitudes in both earlywood and latewood, with tracheid diameter decreasing for latewood only. According to the most parsimonious phylogenetic general least square models, environment and phylogeny together could explain about 21∼56% of tracheid structure variance. Our results provide insights into the effects of climate and soil on the xylem structure of conifer species thus furthering our understanding of the trees' response to global change.

Anatomical features associated with water transport in imperforate tracheary elements of vessel-bearing angiosperms.

BACKGROUND AND AIMS: Imperforate tracheary elements (ITEs) in wood of vessel-bearing angiosperms may or may not transport water. Despite the significance of hydraulic transport for defining ITE types, the combination of cell structure with water transport visualization in planta has received little attention. This study provides a quantitative analysis of structural features associated with the conductive vs. non-conductive nature of ITEs. METHODS: Visualization of water transport was studied in 15 angiosperm species by dye injection and cryo-scanning electron microscopy. Structural features of ITEs were examined using light and electron microscopy. KEY RESULTS: ITEs connected to each other by pit pairs with complete pit membranes contributed to water transport, while cells showing pit membranes with perforations up to 2 µm were hydraulically not functional. A close relationship was found between pit diameter and pit density, with both characters significantly higher in conductive than in non-conductive cells. In species with both conductive and non-conductive ITEs, a larger diameter was characteristic of the conductive cells. Water transport showed no apparent relationship with the length of ITEs and vessel grouping. CONCLUSIONS: The structure and density of pits between ITEs represent the main anatomical characters determining water transport. The pit membrane structure of ITEs provides a reliable, but practically challenging, criterion to determine their conductive status. It is suggested that the term tracheids should strictly be used for conductive ITEs, while fibre-tracheids and libriform fibres are non-conductive.

Phylogeny Best Explains Latitudinal Patterns of Xylem Tissue Fractions for Woody Angiosperm Species Across China.

Investigating space allocation patterns of plant secondary xylem along a latitudinal gradient is useful to evaluate structure-function tradeoffs in woody angiosperm xylem. An anatomical dataset including 700 woody angiosperm species across China was compiled together with latitudinal and climate data for each species. The relative tissue fractions of vessels, fibers, and parenchyma (including ray and axial parenchyma) in xylem were analyzed to determine the effect of latitudinal differences and phylogeny on anatomical variation. The analyses revealed a trade-off between vessel and non-vessel fraction across latitude, with tissue fraction trade-offs mainly occurring between vessels and fibers, and between fibers and total parenchyma. Among 13 climate variables, thermal indices generally had greater explanatory power than moisture indices in bi-variate models for all cell types, while mean annual temperature, mean temperature of the coldest month, and annual actual evapotranspiration were included in the top multi-variate models explaining variance of different tissue fractions. Phylogeny and climate together explained 57-73% of the total variation in xylem space occupancy, with phylogeny alone accounting for over 50% of the variation. These results contribute to our knowledge of wood structure-function and are relevant to better understand forest response to climate change.

Investigation of the viscous heating artefact arising from the use of thermocouples in a focused ultrasound field.

Accurate temperature measurements in therapeutic ultrasound fields are necessary for understanding damage mechanisms, verification of thermal modelling and calibration of non-invasive clinical thermometry. However, artefactual heating, primarily due to viscous forces which result from motion relative to the surrounding tissue, occurs when metal thermocouples are used in an ultrasound field. The magnitude and time dependence of this artefact has been characterized by comparison with novel thin-film thermocouples (TFTs) at 1-2 cm focal depths in fresh degassed ex vivo bovine liver. High-intensity focused ultrasound exposures (1.7 MHz; free-field spatial-peak temporal-average intensities 40-600 W cm(-2)) were used. Subtraction of the TFT data from that obtained for other thermocouples yielded the time dependence of the viscous heating artefact. This was found to be intensity independent up to 600 W cm(-2) (below the threshold for cavitation and lesion formation) and remained significant at radial distances out to the first side lobe in the focal plane. The contribution of viscous heating to cooling was also found to be significant for at least 5 s after the end of insonation. The ratio of viscous artefact to absorptive heating after 5 s was: 1.76 +/- 0.07 for a fine-wire, 0.45 +/- 0.07 and 1.93 +/- 0.07 for two different sheathed-wires and 0.24 +/- 0.07 for a needle thermocouple.

The Parenchyma of Secondary Xylem and Its Critical Role in Tree Defense against Fungal Decay in Relation to the CODIT Model.

This review examines the roles that ray and axial parenchyma (RAP) plays against fungal pathogens in the secondary xylem of wood within the context of the CODIT model (Compartmentalization of Decay in Trees), a defense concept first conceived in the early 1970s by Alex Shigo. This model, simplistic in its design, shows how a large woody perennial is highly compartmented. Anatomical divisions in place at the time of infection or damage, (physical defense) alongside the 'active' response by the RAP during and after wounding work together in forming boundaries that function to restrict air or decay spread. The living parenchyma cells play a critical role in all of the four walls (differing anatomical constructs) that the model comprises. To understand how living cells in each of the walls of CODIT cooperate, we must have a clear vision of their complex interconnectivity from a three-dimensional perspective, along with knowledge of the huge variation in ray parenchyma (RP) and axial parenchyma (AP) abundance and patterns. Crucial patterns for defense encompass the symplastic continuum between both RP and AP and the dead tissues, with the latter including the vessel elements, libriform fibers, and imperforate tracheary elements (i.e., vasicentric and vascular tracheids). Also, the heartwood, a chemically altered antimicrobial non-living substance that forms the core of many trees, provides an integral part of the defense system. In the heartwood, dead RAP can play an important role in defense, depending on the genetic constitution of the species. Considering the array of functions that RAP are associated with, from capacitance, through to storage, and long-distance water transport, deciding how their role in defense fits into this suite of functions is a challenge for plant scientists, and likely depends on a range of factors. Here, we explore the important role of RAP in defense against fungal pathogens and the trade-offs involved from a viewpoint for structure-function relations, while also examining how fungi can breach the defense system using an array of enzymes in conjunction with the physically intrusive hyphae.

Mapping 3D Strains with Ultrasound Speckle Tracking: Method Validation and Initial Results in Porcine Scleral Inflation.

This study aimed to develop and validate a high frequency ultrasound method for measuring distributive, 3D strains in the sclera during elevations of intraocular pressure. A 3D cross-correlation based speckle-tracking algorithm was implemented to compute the 3D displacement vector and strain tensor at each tracking point. Simulated ultrasound radiofrequency data from a sclera-like structure at undeformed and deformed states with known strains were used to evaluate the accuracy and signal-to-noise ratio (SNR) of strain estimation. An experimental high frequency ultrasound (55 MHz) system was built to acquire 3D scans of porcine eyes inflated from 15 to 17 and then 19 mmHg. Simulations confirmed good strain estimation accuracy and SNR (e.g., the axial strains had less than 4.5% error with SNRs greater than 16.5 for strains from 0.005 to 0.05). Experimental data in porcine eyes showed increasing tensile, compressive, and shear strains in the posterior sclera during inflation, with a volume ratio close to one suggesting near-incompressibility. This study established the feasibility of using high frequency ultrasound speckle tracking for measuring 3D tissue strains and its potential to characterize physiological deformations in the posterior eye.

Influence of Age on Ocular Biomechanical Properties in a Canine Glaucoma Model with ADAMTS10 Mutation.

Soft tissue often displays marked age-associated stiffening. This study aims to investigate how age affects scleral biomechanical properties in a canine glaucoma model with ADAMTS10 mutation, whose extracellular matrix is concomitantly influenced by the mutation and an increased mechanical load from an early age. Biomechanical data was acquired from ADAMTS10-mutant dogs (n = 10, 21 to 131 months) and normal dogs (n = 5, 69 to 113 months). Infusion testing was first performed in the whole globes to measure ocular rigidity. After infusion experiments, the corneas were immediately trephined to prepare scleral shells that were mounted on a pressurization chamber to measure strains in the posterior sclera using an inflation testing protocol. Dynamic viscoelastic mechanical testing was then performed on dissected posterior scleral strips and the data were combined with those reported earlier by our group from the same animal model (Palko et al, IOVS 2013). The association between age and scleral biomechanical properties was evaluated using multivariate linear regression. The relationships between scleral properties and the mean and last measured intraocular pressure (IOP) were also evaluated. Our results showed that age was positively associated with complex modulus (p<0.001) and negatively associated with loss tangent (p<0.001) in both the affected and the normal groups, suggesting an increased stiffness and decreased mechanical damping with age. The regression slopes were not different between the groups, although the complex modulus was significantly lower in the affected group (p = 0.041). The posterior circumferential tangential strain was negatively correlated with complex modulus (R = -0.744, p = 0.006) showing consistent mechanical evaluation between the testing methods. Normalized ocular rigidity was negatively correlated with the last IOP in the affected group (p = 0.003). Despite a mutation that affects the extracellular matrix and a chronic IOP elevation in the affected dogs, age-associated scleral stiffening and loss of mechanical damping were still prominent and had a similar rate of change as in the normal dogs.