Forces on and in the cell walls of living plants.

Environmental influences and differential growth subject plants to mechanical forces. Forces on the whole plant resolve into tensile forces on its primary cell walls and both tensile and compression forces on the secondary cell wall layers of woody tissues. Forces on cell walls are further resolved into forces on cellulose microfibrils and the noncellulosic polymers between them. Many external forces on plants oscillate, with time constants that vary from seconds to milliseconds. Sound waves are a high-frequency example. Forces on the cell wall lead to responses that direct the oriented deposition of cellulose microfibrils and the patterned expansion of the cell wall, leading to complex cell and tissue morphology. Recent experiments have established many of the details of which cell wall polymers associate with one another in both primary and secondary cell walls, but questions remain about which of the interconnections are load bearing, especially in primary cell walls. Direct cellulose-cellulose interactions appear to have a more important mechanical role than was previously thought, and some of the noncellulosic polymers may have a role in keeping microfibrils apart rather than cross-linking them as formerly envisaged.

Drying of virus-containing particles: modelling effects of droplet origin and composition.

BACKGROUND AND PURPOSE: Virus-containing aerosol droplets emitted by breathing, speech or coughing dry rapidly to equilibrium with ambient relative humidity (RH), increasing in solute concentration with effects on virus survival and decreasing in diameter with effects on sedimentation and respiratory uptake. The aim of this paper is to model the effect of ionic and macromolecular solutes on droplet drying and solute concentration. METHODS: Deliquescence-efflorescence concepts and Kohler theory were used to simulate the evolution of solute concentrations and water activity in respiratory droplets, starting from efflorescence data on mixed NaCl/KCl aerosols and osmotic pressure data on respiratory macromolecules. RESULTS: In NaCl/KCl solutions total salt concentrations were shown to reach 10-13 M at the efflorescence RH of 40-55%, depending on the K:Na ratio. Dependence on K:Na ratio implies that the evaporation curves differ between aerosols derived from saliva and from airway surfaces. The direct effect of liquid droplet size through the Kelvin term was shown to be smaller and restricted to the evolution of breath emissions. Modelling the effect of proteins and glycoproteins showed that salts determine drying equilibria down to the efflorescence RH, and macromolecules at lower RH. CONCLUSION: Differences in solute composition between airway surfaces and saliva are predicted to lead to different drying behaviour of droplets emitted by breathing, speech and coughing. These differences may influence the inactivation of viruses.

Nanostructural deformation of high-stiffness spruce wood under tension.

Conifer wood is an exceptionally stiff and strong material when its cellulose microfibrils are well aligned. However, it is not well understood how the polymer components cellulose, hemicelluloses and lignin co-operate to resist tensile stress in wood. From X-ray scattering, neutron scattering and spectroscopic data, collected under tension and processed by novel methods, the ordered, disordered and hemicellulose-coated cellulose components comprising each microfibril were shown to stretch together and demonstrated concerted, viscous stress relaxation facilitated by water. Different cellulose microfibrils did not all stretch to the same degree. Attempts were made to distinguish between microfibrils showing large and small elongation but these domains were shown to be similar with respect to orientation, crystalline disorder, hydration and the presence of bound xylan. These observations are consistent with a major stress transfer process between microfibrils being shear at interfaces in direct, hydrogen-bonded contact, as demonstrated by small-angle neutron scattering. If stress were transmitted between microfibrils by bridging hemicelluloses these might have been expected to show divergent stretching and relaxation behaviour, which was not observed. However lignin and hemicellulosic glucomannans may contribute to stress transfer on a larger length scale between microfibril bundles (macrofibrils).

Aerosol Transmission of SARS-CoV-2: Physical Principles and Implications.

Evidence has emerged that SARS-CoV-2, the coronavirus that causes COVID-19, can be transmitted airborne in aerosol particles as well as in larger droplets or by surface deposits. This minireview outlines the underlying aerosol science, making links to aerosol research in other disciplines. SARS-CoV-2 is emitted in aerosol form during normal breathing by both asymptomatic and symptomatic people, remaining viable with a half-life of up to about an hour during which air movement can carry it considerable distances, although it simultaneously disperses. The proportion of the droplet size distribution within the aerosol range depends on the sites of origin within the respiratory tract and on whether the distribution is presented on a number or volume basis. Evaporation and fragmentation reduce the size of the droplets, whereas coalescence increases the mean droplet size. Aerosol particles containing SARS-CoV-2 can also coalesce with pollution particulates, and infection rates correlate with pollution. The operation of ventilation systems in public buildings and transportation can create infection hazards via aerosols, but provides opportunities for reducing the risk of transmission in ways as simple as switching from recirculated to outside air. There are also opportunities to inactivate SARS-CoV-2 in aerosol form with sunlight or UV lamps. The efficiency of masks for blocking aerosol transmission depends strongly on how well they fit. Research areas that urgently need further experimentation include the basis for variation in droplet size distribution and viral load, including droplets emitted by "superspreader" individuals; the evolution of droplet sizes after emission, their interaction with pollutant aerosols and their dispersal by turbulence, which gives a different basis for social distancing.

Structure of native cellulose microfibrils, the starting point for nanocellulose manufacture.

There is an emerging consensus that higher plants synthesize cellulose microfibrils that initially comprise 18 chains. However, the mean number of chains per microfibril in situ is usually greater than 18, sometimes much greater. Microfibrils from woody tissues of conifers, grasses and dicotyledonous plants, and from organs like cotton hairs, all differ in detailed structure and mean diameter. Diameters increase further when aggregated microfibrils are isolated. Because surface chains differ, the tensile properties of the cellulose may be augmented by increasing microfibril diameter. Association of microfibrils with anionic polysaccharides in primary cell walls and mucilages leads to in vivo mechanisms of disaggregation that may be relevant to the preparation of nanofibrillar cellulose products. For the preparation of nanocrystalline celluloses, the key issue is the nature and axial spacing of disordered domains at which axial scission can be initiated. These disordered domains do not, as has often been suggested, take the form of large blocks occupying much of the length of the microfibril. They are more likely to be located at chain ends or at places where the microfibril has been mechanically damaged, but their structure and the reasons for their sensitivity to acid hydrolysis need better characterization.This article is part of a discussion meeting issue 'New horizons for cellulose nanotechnology'.

Hydrogen-Bonding Network and OH Stretch Vibration of Cellulose: Comparison of Computational Modeling with Polarized IR and SFG Spectra.

Hydrogen bonds play critical roles in noncovalent directional interactions determining the crystal structure of cellulose. Although diffraction studies accurately determined the coordinates of carbon and oxygen atoms in crystalline cellulose, the structural information on hydrogen atoms involved in hydrogen-bonding is still elusive. This could be complemented by vibrational spectroscopy; but the assignment of the OH stretch peaks has been controversial. In this study, we performed calculations using density functional theory with dispersion corrections (DFT-D2) for the cellulose Iß crystal lattices with the experimentally determined carbon and oxygen coordinates. DFT-D2 calculations revealed that the OH stretch vibrations of cellulose are highly coupled and delocalized through intra- and interchain hydrogen bonds involving all OH groups in the crystal. Additionally, molecular dynamics (MD) simulations of a single cellulose microfibril showed that the conformations of OH groups exposed at the microfibril surface are not well-defined. Comparison of the computation results with the experimentally determined IR dichroism of uniaxially aligned cellulose microfibrils and the peak positions of various cellulose crystals allowed unambiguous identification of OH stretch modes observed in the vibrational spectra of cellulose.

Diffraction evidence for the structure of cellulose microfibrils in bamboo, a model for grass and cereal celluloses.

BACKGROUND: Cellulose from grasses and cereals makes up much of the potential raw material for biofuel production. It is not clear if cellulose microfibrils from grasses and cereals differ in structure from those of other plants. The structures of the highly oriented cellulose microfibrils in the cell walls of the internodes of the bamboo Pseudosasa amabilis are reported. Strong orientation facilitated the use of a range of scattering techniques. RESULTS: Small-angle neutron scattering provided evidence of extensive aggregation by hydrogen bonding through the hydrophilic edges of the sheets of chains. The microfibrils had a mean centre-to-centre distance of 3.0 nm in the dry state, expanding on hydration. The expansion on hydration suggests that this distance between centres was through the hydrophilic faces of adjacent microfibrils. However in the other direction, perpendicular to the sheets of chains, the mean, disorder-corrected Scherrer dimension from wide-angle X-ray scattering was 3.8 nm. It is possible that this dimension is increased by twinning (crystallographic coalescence) of thinner microfibrils over part of their length, through the hydrophobic faces. The wide-angle scattering data also showed that the microfibrils had a relatively large intersheet d-spacing and small monoclinic angle, features normally considered characteristic of primary-wall cellulose. CONCLUSIONS: Bamboo microfibrils have features found in both primary-wall and secondary-wall cellulose, but are crystallographically coalescent to a greater extent than is common in celluloses from other plants. The extensive aggregation and local coalescence of the microfibrils are likely to have parallels in other grass and cereal species and to influence the accessibility of cellulose to degradative enzymes during conversion to liquid biofuels.

Isolation of high quality lignin as a by-product from ammonia percolation pretreatment of poplar wood.

A two-step process combining percolation-mode ammonia pretreatment of poplar sawdust with mild organosolv purification of the extracted lignin produced high quality, high purity lignin in up to 31% yield and 50% recovery. The uncondensed fraction of the isolated lignin was up to 34%, close to that the native lignin (40%). Less lignin was recovered after pretreatment in batch mode, apparently due to condensation during the longer residence time of the solubilised lignin at elevated temperature. The lignin recovery was directly correlated with its molecular weight and its nitrogen content. Low nitrogen incorporation, observed at high ammonia concentration, may be explained by limited homolytic cleavage of ß-O-4 bonds. Ammonia concentrations from 15% to 25% (w/w) gave similar results in terms of lignin structure, yield and recovery.

How cellulose stretches: synergism between covalent and hydrogen bonding.

Cellulose is the most familiar and most abundant strong biopolymer, but the reasons for its outstanding mechanical performance are not well understood. Each glucose unit in a cellulose chain is joined to the next by a covalent C-O-C linkage flanked by two hydrogen bonds. This geometry suggests some form of cooperativity between covalent and hydrogen bonding. Using infrared spectroscopy and X-ray diffraction, we show that mechanical tension straightens out the zigzag conformation of the cellulose chain, with each glucose unit pivoting around a fulcrum at either end. Straightening the chain leads to a small increase in its length and is resisted by one of the flanking hydrogen bonds. This constitutes a simple form of molecular leverage with the covalent structure providing the fulcrum and gives the hydrogen bond an unexpectedly amplified effect on the tensile stiffness of the chain. The principle of molecular leverage can be directly applied to certain other carbohydrate polymers, including the animal polysaccharide chitin. Related but more complex effects are possible in some proteins and nucleic acids. The stiffening of cellulose by this mechanism is, however, in complete contrast to the way in which hydrogen bonding provides toughness combined with extensibility in protein materials like spider silk.

Organosolv pretreatment of Sitka spruce wood: conversion of hemicelluloses to ethyl glycosides.

A range of Organosolv pretreatments, using ethanol:water mixtures with dilute sulphuric acid, were applied to Sitka spruce sawdust with the aim of generating useful co-products as well as improving saccharification yield. The most efficient of the pretreatment conditions, resulting in subsequent saccharification yields of up to 86%, converted a large part of the hemicellulose sugars to their ethyl glycosides as identified by GC/MS. These conditions also reduced conversion of pentoses to furfural, the ethyl glycosides being more stable to dehydration than the parent pentoses. Through comparison with the behaviour of model compounds under the same reaction conditions it was shown that the anomeric composition of the products was consistent with a predominant transglycosylation reaction mechanism, rather than hydrolysis followed by glycosylation. The ethyl glycosides have potential as intermediates in the sustainable production of high-value chemicals.

Structure of cellulose microfibrils in primary cell walls from collenchyma.

In the primary walls of growing plant cells, the glucose polymer cellulose is assembled into long microfibrils a few nanometers in diameter. The rigidity and orientation of these microfibrils control cell expansion; therefore, cellulose synthesis is a key factor in the growth and morphogenesis of plants. Celery (Apium graveolens) collenchyma is a useful model system for the study of primary wall microfibril structure because its microfibrils are oriented with unusual uniformity, facilitating spectroscopic and diffraction experiments. Using a combination of x-ray and neutron scattering methods with vibrational and nuclear magnetic resonance spectroscopy, we show that celery collenchyma microfibrils were 2.9 to 3.0 nm in mean diameter, with a most probable structure containing 24 chains in cross section, arranged in eight hydrogen-bonded sheets of three chains, with extensive disorder in lateral packing, conformation, and hydrogen bonding. A similar 18-chain structure, and 24-chain structures of different shape, fitted the data less well. Conformational disorder was largely restricted to the surface chains, but disorder in chain packing was not. That is, in position and orientation, the surface chains conformed to the disordered lattice constituting the core of each microfibril. There was evidence that adjacent microfibrils were noncovalently aggregated together over part of their length, suggesting that the need to disrupt these aggregates might be a constraining factor in growth and in the hydrolysis of cellulose for biofuel production.

Comparative structure and biomechanics of plant primary and secondary cell walls.

Recent insights into the physical biology of plant cell walls are reviewed, summarizing the essential differences between primary and secondary cell walls and identifying crucial gaps in our knowledge of their structure and biomechanics. Unexpected parallels are identified between the mechanism of expansion of primary cell walls during growth and the mechanisms by which hydrated wood deforms under external tension. There is a particular need to revise current "cartoons" of plant cell walls to be more consistent with data from diverse approaches and to go beyond summarizing limited aspects of cell walls, serving instead as guides for future experiments and for the application of new techniques.

Nanostructure of cellulose microfibrils in spruce wood.

The structure of cellulose microfibrils in wood is not known in detail, despite the abundance of cellulose in woody biomass and its importance for biology, energy, and engineering. The structure of the microfibrils of spruce wood cellulose was investigated using a range of spectroscopic methods coupled to small-angle neutron and wide-angle X-ray scattering. The scattering data were consistent with 24-chain microfibrils and favored a "rectangular" model with both hydrophobic and hydrophilic surfaces exposed. Disorder in chain packing and hydrogen bonding was shown to increase outwards from the microfibril center. The extent of disorder blurred the distinction between the I alpha and I beta allomorphs. Chains at the surface were distinct in conformation, with high levels of conformational disorder at C-6, less intramolecular hydrogen bonding and more outward-directed hydrogen bonding. Axial disorder could be explained in terms of twisting of the microfibrils, with implications for their biosynthesis.

Distribution of (1->4)-beta-galactans, arabinogalactan proteins, xylans and (1->3)-beta-glucans in tracheid cell walls of softwoods.

Polysaccharides were located in the walls of normal and compression wood tracheids of Pinus radiata (radiata pine), Picea sitchensis (Sitka spruce) and Picea abies (Norway spruce) by transmission electron microscopy using immunogold labelling with monoclonal antibodies to (1-->4)-beta-galactan (LM5), (1-->3)-beta-glucan, arabinogalactan proteins (AGPs) (MAC207) and heteroxylans (LM10 and LM11). In fully differentiated compression wood tracheids, (1-->4)-beta-galactan was found in the S2((L)) layer and, to a smaller extent, at the interface between the compound middle lamella and the S1 layer. (1-->4)-beta-Galactan appeared to be displaced from, or modified in, the S1 layer during cell wall formation. (1-->3)-beta-Glucan (callose) was confined to the helical cavities in the inner S2 layer of severe compression wood. MAC207 AGP glycan epitope was found exclusively in the S1 and S3 layers of normal wood tracheids and in the S1 and inner S2 layers of compression wood tracheids. Binding of LM10, which specifically recognizes unsubstituted or low-substituted xylans, occurred at similar locations to the MAC207 epitope, whereas binding of LM11, which recognizes more highly substituted as well as unsubstituted xylans, occurred throughout the tracheid walls with the exception of the primary wall. Immunogold labelling showed that the different wall layers of softwood tracheids have different polysaccharide compositions which change abruptly during cell wall formation.

Vibrational spectroscopy of biopolymers under mechanical stress: processing cellulose spectra using bandshift difference integrals.

Mechanical stretching of covalent bonds, for example when a fibrous polymer is loaded in tension, results in their stretching vibrational bands in the infrared or Raman spectrum being shifted to lower frequency. Conversely stretching a hydrogen bond shifts the stretching vibrational mode of the donor covalent X-H bond to higher frequency. These band shifts are small and difficult to detect in complex regions of the spectrum where differently affected bands overlap. This paper describes a method of integrating the difference spectra (spectrum under tensile strain minus spectrum at zero tensile strain) to recover the shape of the bands that are shifted and the spectral variation in bandshift. The application of this method to two sets of vibrational spectra of cellulose under tension is described. In one example, C-O-C stretching bands of highly crystalline tunicate cellulose were observed to shift to lower frequency under axial strain. In the other example, a group of overlapping O-D stretching bands in partially deuterated cellulose showed varied bandshifts under axial strain, some bandshifts being positive as expected due to extension of axially oriented hydrogen bonds while others were negative. The possibility of constructing spectral plots of bandshift has the potential to clarify the interpretation of overlapped, shifting bands in the vibrational spectra of polymers under tension.

Cell-wall structure and anisotropy in procuste, a cellulose synthase mutant of Arabidopsis thaliana.

In dark-grown hypocotyls of the Arabidopsis procuste mutant, a mutation in the CesA6 gene encoding a cellulose synthase reduces cellulose synthesis and severely inhibits elongation growth. Previous studies had left it uncertain why growth was inhibited, because cellulose synthesis was affected before, not during, the main phase of elongation. We characterised the quantity, structure and orientation of the cellulose remaining in the walls of affected cells. Solid-state NMR spectroscopy and infrared microscopy showed that the residual cellulose did not differ in structure from that of the wild type, but the cellulose content of the prc-1 cell walls was reduced by 28%. The total mass of cell-wall polymers per hypocotyl was reduced in prc-1 by about 20%. Therefore, the fourfold inhibition of elongation growth in prc-1 does not result from aberrant cellulose structure, nor from uniform reduction in the dimensions of the cell-wall network due to reduced cellulose or cell-wall mass. Cellulose orientation was quantified by two quantitative methods. First, the orientation of newly synthesised microfibrils was measured in field-emission scanning electron micrographs of the cytoplasmic face of the inner epidermal cell wall. The ordered transverse orientation of microfibrils at the inner face of the cell wall was severely disrupted in prc-1 hypocotyls, particularly in the early growth phase. Second, cellulose orientation distributions across the whole cell-wall thickness, measured by polarised infrared microscopy, were much broader. Analysis of the microfibril orientations according to the theory of composite materials showed that during the initial growth phase, their anisotropy at the plasma membrane was sufficient to explain the anisotropy of subsequent growth.

Conformation and mobility of the arabinan and galactan side-chains of pectin.

The function of the arabinan and galactan side-chains of pectin remains unknown. We describe 13C NMR experiments designed to yield spectra from the most mobile polymer components of hydrated cell walls isolated from a range of plant species. In pectin-rich cell walls, these corresponded to the pectic side-chains. The arabinan side-chains were in general more mobile than the galactans, but the long galactan side-chains of potato pectin showed high mobility. Due to motional line-narrowing effects these arabinan and galactan chains gave 13C NMR spectra of higher resolution than has previously been observed from 'solid' biopolymers. These spectra were similar to those reported for the arabinan and galactan polymers in the solution state, implying time-averaged conformations resembling those found in solution. The mobility of the highly esterified galacturonan in citrus cell walls overlapped with the lower end of the mobility range characteristic of the pectic side-chains. The cellulose-rich cell walls of flax phloem fibres gave spectra of low intensity corresponding to mobile type II arabinogalactans. Cell walls from oat coleoptiles appeared to contain no polymers as mobile as the pectic arabinans and galactans in primary cell walls of the other species examined. These properties of the pectic side-chains suggest a role in interacting with water.

Structural details of crystalline cellulose from higher plants.

It is commonly assumed that cellulose from higher plants contains the Ialpha and Ibeta crystalline allomorphs together with surface and disordered chains. For cellulose Ialpha, the evidence for its presence in higher plants is restricted to the C-4 signals in the solid-state (13)C NMR spectrum, which match those of crystalline cellulose Ialpha from algal sources. Algal cellulose Ialpha can be converted to the Ibeta form by high-temperature annealing. We used this approach to generate cellulose samples differing in Ibeta content from flax fibers and celery collenchyma, which respectively are representative of textile (secondary-wall) and primary-wall cellulose. It was then possible to isolate the detailed spectral contributions of the surface, Ibeta and Ialpha-like phases from linear combinations of the observed (13)C NMR and FTIR spectra. The (13)C NMR spectra resembled those of highly crystalline tunicate or algal cellulose Ibeta and Ialpha, with slight differences implying increased disorder and minor conformational discrepancies. The FTIR spectrum of the Ibeta form was closely similar to its more crystalline counterparts, but the FTIR spectrum of the Ialpha form was not. In addition to increased bandwith indicative of lower order, it showed substantial differences in the profile of hydroxyl stretching bands. These results confirm that higher plants synthesize cellulose Ibeta but show that the Ialpha-like chains, although conformationally quite similar to crystalline algal cellulose Ialpha, sit in a different hydrogen-bonding environment in higher plants. The differences are presumably occasioned by the small diameter of the crystallites and the influence of the crystallite surface on chain packing.

Polarized vibrational spectroscopy of fiber polymers: hydrogen bonding in cellulose II.

Vibrational spectroscopy using polarized incident radiation can be used to determine the orientation of X-H bonds with respect to coordinates such as crystallographic axes. The adaptation of this approach to polymer fibers is described here. It requires spectral intensity to be quantified around a 180 degrees range of polarization angles and not just recorded transversely and longitudinally as is normal in fiber spectroscopy. Mercerized cellulose II is used as an example. The unit cell of the cellulose II lattice contains six distinct hydroxyl groups engaged in a complex network of hydrogen bonds that hold the cellulose chains laterally together. A formalism is described to relate the variation in intensity of each O-H stretching mode to the angle between its transition moment and the chain axis as the polarization axis is rotated with respect to the fiber axis. It was necessary to include the effect of dispersion in chain orientation around the mean and the averaging of all rotational positions of the chains round their axis. The two crystallographically distinct O(2)-H groups, which are each hydrogen-bonded to only one acceptor oxygen, show a close match in orientation between the transition moments of their stretching bands and the O-H bond axis. The two O(3)-H groups each have a three-centered hydrogen bond to O-5 and O-6 of the next residue in the same chain. The transition moments of their stretching modes lay between the acceptor oxygens. Hydrogen bonding from the O(6)-H groups is still more complex but again the transition moment of each O-H bond lay within the cone of orientations described by the acceptor oxygens, provided that one additional acceptor oxygen excluded from the published crystal structure was considered. The transition moments for the O-H stretching modes were approximately aligned with the O-H bond axes, but the alignment was not necessarily exact. This approach is not restricted to hydroxyl groups, but it is particularly useful for the elucidation of hydrogen bonding in fibrous polymers for which crystallographic data on proton positions are not available.