Moving beyond the cambium necrosis hypothesis of post-fire tree mortality: cavitation and deformation of xylem in forest fires.

• It is widely assumed that post-fire tree mortality results from necrosis of phloem and vascular cambium in stems, despite strong evidence that reduced xylem conductivity also plays an important role. • In this study, experiments with Populus balsamifera were used to demonstrate two mechanisms by which heat reduces the hydraulic conductivity of xylem: air seed cavitation and conduit wall deformation. Heat effects on air seed cavitation were quantified using air injection experiments that isolate potential temperature-dependent changes in sap surface tension and pit membrane pore diameters. Heat effects on conduit wall structure were demonstrated using air conductivity measurements and light microscopy. • Heating increased vulnerability to cavitation because sap surface tension varies inversely with temperature. Heating did not affect cavitation via changes in pit membrane pore diameters, but did cause significant reductions in xylem air conductivity that were associated with deformation of conduit walls (probably resulting from thermal softening of viscoelastic cell wall polymers). • Additional work is required to understand the relative roles of cavitation and deformation in the reduction of xylem conductivity, and how reduced xylem conductivity in roots, stems, and branches correlates and interacts with foliage and root necroses to cause tree mortality. Future research should also examine how heat necrosis of ray parenchyma cells affects refilling of embolisms that occur during and after the fire event.

Mapping with RAD (restriction-site associated DNA) markers to rapidly identify QTL for stem rust resistance in Lolium perenne.

A mapping population was created to detect quantitative trait loci (QTL) for resistance to stem rust caused by Puccinia graminis subsp. graminicola in Lolium perenne. A susceptible and a resistant plant were crossed to produce a pseudo-testcross population of 193 F(1) individuals. Markers were produced by the restriction-site associated DNA (RAD) process, which uses massively parallel and multiplexed sequencing of reduced-representation libraries. Additional simple sequence repeat (SSR) and sequence-tagged site (STS) markers were combined with the RAD markers to produce maps for the female (738 cM) and male (721 cM) parents. Stem rust phenotypes (number of pustules per plant) were determined in replicated greenhouse trials by inoculation with a field-collected, genetically heterogeneous population of urediniospores. The F(1) progeny displayed continuous distribution of phenotypes and transgressive segregation. We detected three resistance QTL. The most prominent QTL (qLpPg1) is located near 41 cM on linkage group (LG) 7 with a 2-LOD interval of 8 cM, and accounts for 30-38% of the stem rust phenotypic variance. QTL were detected also on LG1 (qLpPg2) and LG6 (qLpPg3), each accounting for approximately 10% of phenotypic variance. Alleles of loci closely linked to these QTL originated from the resistant parent for qLpPg1 and from both parents for qLpPg2 and qLpPg3. Observed quantitative nature of the resistance may be due to partial-resistance effects against all pathogen genotypes, or qualitative effects completely preventing infection by only some genotypes in the genetically mixed inoculum. RAD markers facilitated rapid construction of new genetic maps in this outcrossing species and will enable development of sequence-based markers linked to stem rust resistance in L. perenne.

Enamel Etching for Universal Adhesives: Examination of Enamel Etching Protocols for Optimization of Bonding Effectiveness.

OBJECTIVE: The purpose of this study was to evaluate whether different enamel etching methods with reduced etching times would improve the bonding effectiveness of universal adhesives. METHODS AND MATERIALS: Three enamel etching methods, phosphoric acid ester monomer (PPM) etching, phosphoric acid (PPA) etching, and polyalkenoic acid (PLA) etching, and three universal adhesives, G-Premio Bond (GP), Prime&Bond elect (PE), and Scotchbond Universal Adhesive (SU), were evaluated. Initial bond strengths and fatigue strengths of universal adhesives to ground enamel and ground enamel etched for less than one, five, 10, and 15 seconds using different etching methods were determined. The bonded fatigue specimens were loaded using a sine wave at a frequency of 20 Hz for 50,000 cycles or until failure occurred with a staircase method. Atomic force micrograph (AFM) observations along with measurements of surface Ra roughness and modified surface area of enamel with different etching protocols were also conducted. RESULTS: The bond fatigue durability of universal adhesives to enamel with PPA etching from less than one to 15 seconds and PLA etching for 15 seconds was significantly higher than that to ground enamel. The bond fatigue durability to enamel with PPM etching was not increased compared with ground enamel. The surface Ra roughness and surface area obtained with AFM of enamel increased after PPA and PLA etching, and those values were significantly higher than those of ground enamel. Furthermore, surface Ra roughness and surface area with PPA etching were significantly higher than those with PLA etching. However, surface Ra roughness and surface area of enamel with PPM etching were similar to those of ground enamel regardless of etching time. CONCLUSION: PPA etching for less than one to 15 seconds and PLA etching for 15 seconds improve universal adhesive bonding, surface Ra roughness, and surface area of enamel. However, PPM etching is not effective, regardless of etching time, in improving bonds strengths, increasing surface roughness, and increasing surface area.

A strand of cardiac muscle. Its ultrastructure and the electrophysiological implications of its geometry.

The structure of a small strand of rabbit heart muscle fibers (trabecula carnea), 30-80 micro in diameter, has been examined with light and electron microscopy. By establishing a correlation between the appearance of regions of close fiber contact in light and electron microscopy, the extent and distribution of regions of close apposition of fibers has been evaluated in approximately 200 micro length of a strand. The distribution of possible regions of resistive coupling between fibers has been approximated by a model system of cables. The theoretical linear electrical properties of such a system have been analyzed and the implications of the results of this analysis are discussed. Since this preparation is to be used for correlated studies of the electrical, mechanical, and cytochemical properties of cardiac muscle, a comprehensive study of the morphology of this preparation has been made. The muscle fibers in it are distinguished from those of the rabbit papillary muscle, in that they have no triads and have a kind of mitochondrion not found in papillary muscle. No evidence of a transverse tubular system was found, but junctions of cisternae of the sarcoplasmic reticulum and the sarcolemma, peripheral couplings, were present. The electrophysiological implications of the absence of transverse tubules are discussed. The cisternae of the couplings showed periodic tubular extensions toward the sarcolemma. A regularly spaced array of Z line-like material was observed, suggesting a possible mechanism for sarcomere growth.

Cardiac muscle. A comparative study of Purkinje fibers and ventricular fibers.

With light and electron microscopy a comparison has been made of the morphology of ventricular (V) and Purkinje (P) fibers of the hearts of guinea pig, rabbit, cat, dog, goat, and sheep. The criteria, previously established for the rabbit heart, that V fibers are distinguished from P fibers by the respective presence and absence of transverse tubules is shown to be true for all animals studied. No evidence was found of a permanent connection between the sarcoplasmic reticulum and the extracellular space. The sarcoplasmic reticulum (SR) of V fibers formed couplings with the sarcolemma of a transverse tubule (interior coupling) and with the peripheral sarcolemma (peripheral coupling), whereas in P fibers the SR formed only peripheral couplings. The forms of the couplings were identical. The significance, with respect to excitation-contraction coupling, of the difference in the form of the couplings in cardiac versus skeletal muscle is discussed together with the electrophysiological implications of the differing geometries of bundles of P fibers from different animals.

Cardiac muscle. Its ultrastructure in the finch and hummingbird with special reference to the sarcoplasmic reticulum.

Cardiac muscle fibers of the hummingbird and finch have no transverse tubules and are smaller in diameter than those of mammalian hearts. The fibers are connected by intercalated discs which are composed of desmosomes and f. adherentes; small nexuses are often interspersed. As in cardiac muscle of several other animals, the junctional SR of the couplings is highly structured in these two birds but, in addition, and after having lost sarcolemmal contact, the junctional SR continues beyond the coupling to extend deep into the interior of the cells and to form belts around the Z-I regions of the sarcomeres. This portion of the sarcoplasmic reticulum, which we have named "extended junctional SR," and which is so prominent and invariant a feature of cardiac cells of hummingbirds and finches, has not been observed in chicken cardiac cells. The morphological differences between these species of birds may be related to respective differences in heart rates characteristic for these birds.

Synthetic strands of cardiac muscle: growth and physiological implication.

Cardiac muscle cells obtained fronm disaggregated embryonic chick hearts were cultured on difjerentially treated oriented substrata. Subsequent cell reaggregation, growth, and attachmwent produced linearly organized strands of cardiac muscle with dimensions suitable for electrophysiological analysis. Along the strand, areas that contained few muscle cells demonstrated reduced conduction velocity and were subject to propagation failure.

Egg yolk antibodies for detection and neutralization of Clostridium botulinum type A neurotoxin.

The objective of this research project was to determine the usefulness of an egg antibody platform for producing materials for the detection and neutralization of botulinum type A neurotoxin. Yield estimates for detection and neutralizing antibodies produced using methods described were calculated. Antibody specific to botulinum toxoid A (aToxoid) and toxin A (aBoNT/A) was produced by immunizing hens with botulinum toxoid A (toxoid) followed by increasing amounts of botulinum neurotoxin A (BoNT/A) in Freund incomplete adjuvant. Egg yolks were extracted with polyethylene glycol (PEG) for antibody detection and neutralization experiments. A model aToxoid/toxoid immunoassay using only egg yolk antibody was developed and had a detection limit of 1 pg/ml of toxoid. In an indirect enzyme-linked immunosorbent assay of BoNT/A-specific antibody, the aBoNT/A contained more BoNT/A-specific antibody than did the aToxoid, and aBoNT/A was as effective as commercial rabbit antibody. The aToxoid provided no protection against BoNT/A in a standard mouse neutralization assay; however, 1 mg of PEG-extracted aBoNT/A neutralized 4,000 lethal doses of BoNT/A injected intraperitoneally. Based on these results, we calculated that in 1 month one hen could produce more than 100 liters of antibody detection reagents or enough antibody to neutralize approximately 11.6 million mouse lethal doses of botulinum toxin. Utilization of an egg antibody platform is potentially rapid (28 to 70 days) and scalable to kilogram quantities using current egg production facilities with as few as 1,000 hens.

Study of the porosity of calcified chicken eggshell using atomic force microscopy and image processing.

In this work, the porosity of the layers of calcified chicken eggshell (vertical crystal layer VCL, palisade layer PL and mammillary layer ML) was evaluated using atomic force microscopy (AFM) and image processing (IP). AFM topographic images were obtained from different locations for each layer and along the cross-section of calcified eggshell. Roughness parameters, surface area values, pore size and shape, surface porosity, area occupied by pores and pore density were obtained from AFM and IP. It was observed that the thickest layer (PL) exhibited the highest degree of porosity (surface porosity = 2.75 ± 1.68%, pore density = 162 ± 60 pores/µm2) when compared to the other two layers. In general, the pores located in all layers ("bubble pores") had circular shape and similar sizes. Measurements revealed a porosity gradient along the cross-section which varied with position, i.e., increasing surface porosity from the VCL towards the region of the PL closer to the ML, and decreasing surface porosity from this location towards the ML innermost surface. This suggests that the calcified eggshell has a sandwich-like structure where porosity may influence gas exchange and mechanical properties. The combination of AFM with IP presented here provides a simple and precise method to study porosity in calcified chicken eggshell, and this methodology could be used to examine other types of porous biological materials.

Creating new landscapes and ecosystems: the Alberta Oil Sands.

Extraction of oil from the Alberta Oil Sands through surface mining involves the removal of the overburden and oil sand to a depth of up to 100 m and over extremely large areas. While the operation of the bitumen processing plants has serious environmental impacts on downstream habitats, this article focuses on the reclamation of areas from which the oil sands have been removed, processed, and returned. This reclamation following closure of the mines will entail the complete re-creation of landforms and ecosystems at a landscape scale, with the goal of producing suitable habitats for plants, animals, and people. Such projects will require a reasonable understanding of the geophysical and ecological processes that operate at a wide range of scales. Some information is provided on the climate, hydrology, vegetation, and land use (past and current) of the Oil Sands area, situated within the Boreal Plain ecozone, to provide a framework for discussion of issues to be addressed in, and proposed guidelines for, such large-scale reclamation. Although none of the mines has yet closed, numerous consultant reports have been produced with recommendations for various aspects of such reclamation projects (e.g., wetland hydrology, vegetation, wildlife habitat). The scientific basis of such reports is found to vary with respect to depth of understanding of the relevant processes.

Microstructure and mechanical properties of hard Acrocomia mexicana fruit shell.

Fruit and nut shells can exhibit high hardness and toughness. In the peninsula of Yucatan, Mexico, the fruit of the Cocoyol palm tree (Acrocomia mexicana) is well known to be very difficult to break. Its hardness has been documented since the 1500 s, and is even mentioned in the popular Maya legend The Dwarf of Uxmal. However, until now, no scientific studies quantifying the mechanical performance of the Cocoyol endocarp has been found in the literature to prove or disprove that this fruit shell is indeed "very hard". Here we report the mechanical properties, microstructure and hardness of this material. The mechanical measurements showed compressive strength values of up to ~150 and ~250 MPa under quasi-static and high strain rate loading conditions, respectively, and microhardness of up to ~0.36 GPa. Our findings reveal a complex hierarchical structure showing that the Cocoyol shell is a functionally graded material with distinctive layers along the radial directions. These findings demonstrate that structure-property relationships make this material hard and tough. The mechanical results and the microstructure presented herein encourage designing new types of bioinspired superior synthetic materials.

Using decomposition rates to infer how far back tree populations can be reconstructed.

To study forest dynamics without relying on the space-for-time substitution, one must be able to follow a population or stand of trees back or forward in time. The method of stand reconstruction looks back in time by aging all the live trees and aging and dating the time of death of dead standing and fallen trees. However, dead trees are lost by decomposition so the record becomes increasingly incomplete with passage of time. Here we present a model of the passage of trees from dead standing to dead decomposed but still datable to completely decomposed and thus undatable or lost. We then generalize a method for calculating the falling rate of dead trees originally proposed in 1985 by A. P. Gore, E. A. Johnson, and H. P. Lo. We do this by removing the assumption that no trees are lost by decomposition, i.e., by using the decomposition rate. Finally, in the most important result, the model allows estimation of how far back a good estimate of the numbers in the population can be made if the decomposition rates are known.

Mechanical properties in crumple-formed paper derived materials subjected to compression.

The crumpling of precursor materials to form dense three dimensional geometries offers an attractive route towards the utilisation of minor-value waste materials. Crumple-forming results in a mesostructured system in which mechanical properties of the material are governed by complex cross-scale deformation mechanisms. Here we investigate the physical and mechanical properties of dense compacted structures fabricated by the confined uniaxial compression of a cellulose tissue to yield crumpled mesostructuring. A total of 25 specimens of various densities were tested under compression. Crumple formed specimens exhibited densities in the range 0.8-1.3 g cm-3, and showed high strength to weight characteristics, achieving ultimate compressive strength values of up to 200 MPa under both quasi-static and high strain rate loading conditions and deformation energy that compares well to engineering materials of similar density. The materials fabricated in this work and their mechanical attributes demonstrate the potential of crumple-forming approaches in the fabrication of novel energy-absorbing materials from low-cost precursors such as recycled paper. Stiffness and toughness of the materials exhibit density dependence suggesting this forming technique further allows controllable impact energy dissipation rates in dynamic applications.

The importance of fires and floods on tree ages along mountainous gravel-bed streams.

This paper examines the commonly accepted assumption in the riparian literature that areas adjacent to streams do not burn. Using time-since-fire distributions, derived from stand-origin maps for a watershed in the front ranges of the Canadian Rocky Mountains, we found that the areas adjacent to streams and the whole study watershed have similar fire frequencies. In addition, the relative importance of fires and floods is regulated by a change in channel morphology associated with the creation of bars. The results demonstrate that fires solely control tree establishment along straight streams without bars, while the influence of floods is observed at the onset of lateral- and point-bar formation. This occurs because bars are formed in-channel and require smaller discharges in order to be flooded, compared to higher terraces. Consequently, bars are the only surfaces being flooded more frequently than they are being burned. Thus, overall the results indicate that, on this watershed, areas adjacent to streams are not less likely to burn than the uplands, except for lateral and point bars. The generality of these results to other systems should be tested as they have important implications for current forest ecological definition of "riparian zones," which typically include all fluvially derived landforms, from the channel banks to the terraces. Indeed, this study suggests that along smaller, headwater, gravel-bed mountain watersheds, the forests found on terraces are only influenced by fire and not fluvial processes and should therefore not be included in the riparian zone, while the forests on bars are the only surfaces currently being influenced by fluvial processes. Such a change in definition has implications for both ecologists and forest managers aiming to protect areas along streams as they now must take into account the effects of two disturbances on these small gravel-bed streams.

Morphogenesis of marek's disease virus in feather follicle epithelium.

Three evaluative systems, immunodiffusion, fluorescent antibody (FA), and electron microscopy (EM), were used to follow the morphogenesis of Marek's disease virus in inoculated chickens. Of the three, EM and FA were the most sensitive in detecting early stages of infection. Virus particles were found in skin biopsy specimens as early as 12 days post inoculation. Immature naked particles appeared first in the nucleus; later particles were enveloped in the cytoplasm and enclosed in cytoplasmic inclusion bodies. No evidence for continued virus replication was seen in feather follicles after an initial burst of heavy virus production, which lasted several weeks. Residual virus, however, was found occasionally in cytoplasmic inclusion bodies within keratinized material near the feathers. This was believed to contribute to the long-term shedding of infectious virus into the environment.

Distribution of radiolabeled alloantibodies in mice bearing 3-methylcholanthrene-induced sarcoma.

The distribution of purified 125I-labeled alloantibodies, prepared from the serum of DBA/2J mice obtained after immunization with C3H/HeJ spleen cells, was studied in immunosuppressed DBA/2J mice bearing either allogeneic C3H/HeJ 3-methylcholanthrene sarcomas growing s.c. or syngeneic SaD2 3-methylcholanthrene sarcomas. Once purified radiolabeled antibody was isolated from 125I-labeled immune gamma-globulin by a single adsorption onto C3H/HeJ RBC and elution from stroma prepared from these cells, by using 0.1 M glycine buffer (pH 3.0). Twice-purified alloantibody was then produced by Bio-Gel P-200 or Sephadex G-200 gel filtration chromatography or DEAE A-50 ion-exchange chromatography. In vitro, such purified antibodies bound specifically to C3H/HeJ RBC. In vivo, they localized to a significant extent following i.p. injection, preferentially in C3H/HeJ 3-methylcholanthrene sarcomas (4.4 to 8.9% of the injected dose per g of tumor equal to 1% of mouse weight), with mean tumor/blood ratios of 4.0 to 7.8, at 24 or 48 hr after injection. The percentage of injected dose localized in tumor and the tumor/blood ratio did not show significant differences with respect to time or method of antibody purification. Normal tissue/blood ratios in C3H/HeJ or SaD2 sarcoma-bearing mice were less than 0.9. The tumor/blood ratios in SaD2 sarcomas were approximately 0.6. Injection of 131I-labeled normal DBA/2J gamma-globulin resulted in normal tissue/blood and tumor/blood ratios of less than 0.9 in C3H/HeJ tumor-bearing mice.

Isolation and partial characterization of a lectin from Vicia faba.

A lectin present in the broad bean, Vicia faba, was isolated by affinity chromatography on Sephadex G-150. The active lectin had a molecular weight of 50 000. The lectin contained about 3% neutral sugar and it contained two different subunits. The subunits had molecular weights of 17 300 and 14 300. The lectin was mitogenic to human peripheral lymphocytes with some reduction in activity upon acetylation. The lectin competed with pea lectin for binding to murine tumor cells. Binding to tumor cells was inhibited by methyl-alpha-D-mannoside but not by D-galactose; binding was not influenced by the presence or absence of Ca2+, Mn2+ or ethylenediaminetetraacetate.

Studies on 6C3HED murine ascites tumor cell receptors for mannosyl-binding lectins.

We have investigated the receptor site activity present on 6C3HED tumor cells for concanavalin A, fava, lentil and pea lectins. The binding of the tritiated lectins to the tumor cells was inhibited by methyl-alpha-D-mannoside but not by D-galactose. The number of binding sites for the lectins was 3.5-10(6)/cell for concanavalin A, 3.3-10(6)/cell for fava, 3.6-10(6)/cell for lentil and 4.8-10(6)/cell for pea. The apparent association constants were 3.6 and 1.3 muM-1 for concanavalin A, 3.9 muM-1 for fava, 4.2 muM-1 for lentil and 4.6 and 0.6 muM-1 for pea. Competitive inhibition studies showed that lentil was a good inhibitor of pea binding; concanavalin A was a poor inhibitor of pea binding; and fava was a better inhibitor than concanavalin A but not as good as lentil. Reciprocal inhibition experiments indicated that concanavalin A and pea may bind to different receptors as well as to common receptors. This was also indicated by the observation that trypsin or protease treatment of the cells decreased the binding of pea lectin by 20-40 percent whereas concanavalin A binding was unaffected.

Chromatographic behavior of pea lectin receptors from the 6C3HED murine ascites tumor.

We have previously shown that pea lectin and concanavalin A bind to different receptors as well as to common receptors on the cell surface of 6C3HED murine ascites tumor cells (Allen, H.J. nd Johnson, E.A.Z. (1976) Biochim. Biophys. Acta 436, 557-566). We have characterized the metabolically labeled pea lectin receptor from trypsinates of viable tumor cells by gel filtration and affinity chromatography. The pea lectin receptor eluted as an apparent high molecular weight glycopeptice (Mr 200 000) from Sephadex G-200 columns. Other glycopeptides were present in the trypsinates but they did not bind to either concanavalin A or to pea lectin in significant amounts. The high molecular weight glycopeptide was resistant to digestion by a high concentration of trypsin but it was absent in non-specific protease digests of viable cells. Sequential lectin affinity chromatography of Nonidet P-40 extracts of cells permitted the probable identification of the native glycoprotein corresponding to the high molecular weight glycopeptide which binds to pea lectin.

Abnormal fatty acid composition in sarcolemma and sarcoplasmic reticulum from myotonic ADR mouse muscle.

The fatty acid composition of membrane lipids from sarcolemma and sarcoplasmic reticulum isolated from biceps and gastrocnemius muscle has been compared in normal (wildtype, +/adrmto or +/+) and affected (adrmto/adrmto) myotonic mice. The adrmto mouse exhibits an arrested development of the righting response, and arose spontaneously from the SWR/J strain. These mice exhibit classical myotonia similar to the human disease, Becker's myotonia [1]. Significant alterations, characterized by a decrease in the saturated fatty acid, palmitic acid (16:0), and the polyunsaturated fatty acid, arachidonic acid (20:4), and an increase in stearic (18:0) and linoleic (18:2) acids, were observed between sarcolemma and sarcoplasmic reticulum from normal and affected mice. These changes in fatty acid composition of muscle membrane from ADR mice may be adequate to cause an alteration in membrane fluidity and affect the function of ion channels. The fatty acid composition of erythrocytes ghosts was also examined, as a potential marker for alterations in muscle membranes. In erythrocyte ghosts isolated from affected mice, the only alteration observed was a decrease in the proportion of oleic acid (18:1), an effect completely different from those observed in muscle membranes. Therefore, erythrocyte ghosts do not serve as an adequate indicator of changes in fatty acid composition of muscle membranes in this model of myotonia.