Isolation and characterisation of water soluble polysaccharides from Norway spruce and Scots pine

Water soluble polysaccharides from Norway spruce, Scots pine, and Siberian larch were compared. For all species the total amount of polysaccharides isolated from the heartwood was higher than that from the sapwood. The heartwood polysaccharides had a high content of galactose and arabinose units, and some glucuronic acid units, suggesting the presence of acidic arabinogalactans. The total amounts of recovered water-soluble arabinogalactans were 1.9 mg/g for spruce heartwood, 5.3 mg/g for pine heartwood, and as much as 106 mg/g for larch heartwood. The other water-soluble polysaccharides were mainly glucomannans. The average ratio of Gal:Ara:GlcA in the water-soluble arabinogalactans of spruce heartwood was about 4.3:1:1, pine heartwood about 4.5:1:0.2, and larch heartwood about 6.7:1:0.1. The corresponding molar ratios then being about 3.6:1:0.8 for spruce, 3.8:1:0.2 for pine, and 5.6:1:0.08 for larch. Thus, the content of glucuronic acid units was especially high in the spruce heartwood arabinogalactans. The content of arabinose was slightly higher in spruce and pine than in larch heartwood arabinogalactans.     

Polysaccharides in some industrially important softwood species

The content and composition of carbohydrates comprising polysaccharides in sapwood and heartwood of 12 industrially important pulpwood species were analysed. The polysaccharide content was between 60% and 80% (w/w) for all species, with cellulose as the predominant polysaccharide type. The carbohydrate composition suggested that the main non-cellulose polysaccharides were galactoglucomannans, except in Larix heartwood, where arabinogalactans were predominant, while the content of xylans were in the same range as the mannans in Pinus resinosa heartwood and Thuja occidentalis heartwood and sapwood. Pectins, i.e. polygalacturonic acids, were the main acidic polysaccharides in all species. The amount and composition of water-soluble carbohydrates from ground wood samples were also analysed, since these are important in mechanical pulping and as a possible source of bioactive polymers. The main polysaccharides released from the spruce species were mannans, together with starch from sapwood. Especially Abies balsamea stemwood, but also Abies sibirica heartwood, released considerable amounts of pectins, suggesting that fir species may release more troublesome anionic polysaccharides than spruce species. Heartwood of Larix lariciana, Larix decidua, Pinus banksiana, and Pinus resinosa released considerable amounts of acidic arabinogalactans. Thuja occidentalis released mainly arabinogalactans and pectins. Pseudotsuga menziesii heartwood released a large amount of arabinogalactans.     

Some factors influencing susceptibility to discoloring fungi and water uptake of Scots pine (Pinus sylvestris), Norway spruce (Picea abies) and Oriental spruce (Picea orientalis)

Abstract

The heartwood and sapwood from Scots pine (PS), Norway spruce (PA), and Oriental spruce (PO) were tested for susceptibility to discoloring fungi and water uptake. In addition, annual ring width and density were measured. The methods used were Mycologg for testing growth of fungi and a modified version of EN 927-5 to investigate water uptake. For pine, the heartwood showed a lower water uptake and no discoloring fungi growing in the tests. The heartwood had a significantly higher density and smaller annual ring width than the sapwood. In PA the heartwood had significantly lower discoloration than sapwood. The total water uptake in g/m² was significantly higher in sapwood, but not the calculated moisture content. As for wood properties, the density was significantly higher in sapwood compared to heartwood, although there were no differences in annual ring width. Regarding PO, differences in water uptake could be seen between sapwood and heartwood although the densities were similar. These results show that susceptibility to discoloring fungi and water uptake is hard to correlate to a single inherent property when looking at different wood species.     

Outdoor exposure of untreated Scots pine (Pinus sylvestris L.) and Norway spruce [Picea abies (L.) Karst.] wood samples

Abstract

Untreated Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) samples were exposed above ground in a durability test for 6 years. The samples consisted of three pieces of wood, 22×95×500 mm, screwed together; two pieces lengthwise with a third piece overlapping. Weight was measured, to calculate moisture content (MC), and samples checked regularly for cracks and fungal growth. Parameters investigated were heartwood/sapwood (pine), annual ring orientation (spruce), stand site, annual ring width and density. Stand site, annual ring width and density had no influence on MC or fungal growth for either pine or spruce. Spruce samples with vertical annual rings had fewer cracks than samples with horizontal annual rings. Pine sapwood samples had a high MC and a large amount of rot fungi, while heartwood had a lower MC and no rot. Most spruce samples were similar to pine heartwood, except from a few samples that had high MC and fungal growth. Those were all sawn from the outer part of the log. Therefore, it can be stated that spruce sawn from the inner part has almost the same properties as pine heartwood, while spruce from the outer part of the log has similar properties to pine sapwood.     

Decay resistance of sapwood and heartwood of untreated and thermally modified Scots pine and Norway spruce compared with some other wood species

Abstract

Thermal modification has been developed for an industrial method to increase the biological durability and dimensional stability of wood. In this study the effects of thermal modification on resistance against soft- and brown-rot fungi of sapwood and heartwood of Scots pine and Norway spruce were investigated using laboratory test methods. Natural durability against soft-rot microfungi was determined according to CEN/TS 15083-2 (2005) by measuring the mass loss and modulus of elasticity (MOE) loss after an incubation period of 32 weeks. An agar block test was used to determine the resistance to two brown-rot fungi using two exposure periods. In particular, the effect of the temperature of the thermal modification was studied, and the results were compared with results from untreated pine and spruce samples. The decay resistance of reference untreated wood species (Siberian larch, bangkirai, merbau and western red cedar) was also studied in the soft-rot test. On average, the soft-rot and brown-rot tests gave quite similar results. In general, the untreated heartwood of pine was more resistant to decay than the sapwood of pine and the sapwood and heartwood of spruce. Thermal modification increased the biological durability of all samples. The effect of thermal modification seemed to be most effective within pine heartwood. However, very high thermal modification temperature over 230°C was needed to reach resistance against decay comparable with the durability classes of “durable” or “very durable” in the soft-rot test. The brown-rot test gave slightly better durability classes than the soft-rot test. The most durable untreated wood species was merbau, the durability of which could be evaluated as equal to the durability class “moderately durable”.     

Chemical composition of earlywood and latewood in Norway spruce heartwood,sapwood and transition zone wood

This study focused on the distribution of wood components along a cross section of a spruce stem. Thin samples of earlywood and latewood were analysed by special micro-scale analytical techniques. Heartwood contained significantly more lignin and less cellulose than sapwood. The total content of hemicelluloses was the same along the radial direction, but the distribution of sugar units differed. The amounts of arabinoglucuronoxylan and pectins were larger in the heartwood. The transition zone between heartwood and sapwood had a specific composition, with less lignin and lipophilic extractives than heartwood and sapwood. For earlywood and latewood, significant differences were found in the distribution of sugar units in hemicelluloses. Latewood contained clearly more galactoglucomannan than earlywood, and conversely less pectins. The lipophilic extractives were also less concentrated in the latewood.Abbreviations EW or E earlywood - LW or L latewood - HW heartwood - SW sapwood - TZ transition zone wood - A.R. annual ring - AcBr Acetyl bromide - Ara arabinose - Xyl xylose - Gal galactose - Glc glucose - Man mannose - Rha rhamnose - GlcA glucuronic acid - MGlcA 4-O-methyl-glucuronic acid - GalA galacturonic acid - o.d. oven dry     

Growth and Wood Properties of Sitka Spruce (Picea sitchensis (Bong.) Carr.) in Nursing Mixtures Established on Nitrogen-deficient Mineral Soils

Rate of growth and wood properties of Sitka spruce (Picea sitchensis (Bong.) Carr.) were investigated in triplet mixtures with lodgepole pine (Pinus contorta Loud.) (Alaskan provenance) and Japanese larch (Larix kaempferi (Lamb.) Carr.), established on nitrogen-deficient, iron pan soils. These ?nursing? mixtures were compared with pure stands of Sitka spruce that had been either regularly or periodically fertilized with nitrogen. Japanese larch promoted a growth rate in Sitka spruce greater than that achieved in the lodgepole pine-nursed spruce and equivalent to the two pure spruce treatments over the duration of the experiment (current age 28 yrs). Growth of regularly fertilized pure Sitka spruce was not significantly greater than that of periodically fertilized pure spruce. Alaskan lodgepole pine controlled branch size on the lower part of the spruce stems more effectively than the other treatments, although this may have been a function of tree size. Branch characteristics of Japanese larch-nursed spruce, however, were similar to those of the pure spruce treatments. Japanese larch caused an imbalance in crown development in the spruce, although it is unclear from the present study whether this will have an influence on stem and wood quality by the end of the rotation. Overall, the evidence from this study suggests that Japanese larch is an effective nurse of Sitka spruce on nitrogen-deficient iron pans, maintaining a rate of growth similar to that of pure Sitka spruce periodically fertilized with nitrogen and higher than that observed in spruce nursed by Alaskan lodgepole pine.     

Polysaccharides in some industrially important hardwood species

The amount and composition of sugar units comprising polysaccharides in sapwood and heartwood, or stemwood, of 11 industrially important pulpwood species were analysed. The polysaccharide content was between 60 and 80% (w/w) for all species, with cellulose as the predominant polysaccharide type and glucuronoxylans as the main non-cellulosic polysaccharides. The second most abundant non-cellulosic polysaccharides were either pectins, i.e. polygalacturonic acids, or glucomannans. The amount of acidic sugar units were 15–23% of the total amount of non-cellulosic sugar units in all samples, with the Acacia species in the high end. The amount and composition of water-soluble carbohydrates from ground wood samples were also analysed, since these are important in mechanical and chemimechanical pulping, and as a possible source of bioactive polymers. Sapwood released more carbohydrates than heartwood for most species. It is to be noted that the relative amount of dissolved acidic sugar units was larger from the heartwood than from the sapwood for all species. Probably due to the mild treatment conditions, the main dissolved polysaccharides were xylans only for a few samples, while easily soluble galactans, arabinogalactans, or mannans dominated in most species. Pectins dominated in heartwood of Populus grandidentata. Generally, pectins and acidic xylans were the main acidic polysaccharides.     

Surface mould growth on wooden claddings – effects of transient wetting,relative humidity,temperature and material properties

ABSTRACT

Effects of climatic factors and material properties on the development of surface mould growth on wooden claddings were investigated in a laboratory experiment. Specimens of aspen (Populus tremula), Siberian larch (Larix Sibirica), American white oak (Querqus alba), Scots pine (Pinus sylvestris), Norway spruce (Picea abies) and thermally modified pine were incubated in eight climatic chambers at specified wetting periods (2 or 4?h per day), relative humidity (58–86%) and temperature conditions (10–27°C). Surface mould growth was assessed weekly for 13 weeks, and the results were evaluated statistically using Generalized Estimating Equations logistic regression models. All tested climatic factors had significant effects on the mould growth, and there were significant differences between the materials. The ranking of the materials varied with temperature and over time. Aspen, pine sapwood and oak were overall most susceptible to mould growth, and thermally modified pine least susceptible. There were significant differences between sapwood and heartwood for pine and spruce. The effect of density was tested on the spruce heartwood material, but was not found to be significant. The results can be used to further develop prediction models for mould growth on wooden claddings.     

Comparative susceptibility of pine,spruce and larch to sapstain

Summary This study compared the susceptibility of five UK‐grown conifer species to colonization by sapstain fungi in two trials carried out in consecutive years. The conifers consisted of Sitka spruce (Picea sitchensis), Japanese larch (Larix kaempferi), Norway spruce (Picea abies), Scots pine (Pinus sylvestris) and lodgepole pine (Pinus contorta). Freshly cut 1‐m logs were exposed to the available inoculum of sapstain fungi from April to August in a woodland environment in the south east of England. Logs of each species were removed after 1‐, 2‐ and 4‐month exposure and sampled destructively to assess the amount of sapstain. In the second trial, per cent moisture content and concentrations of nitrogen, carbohydrate and phenolic compounds in the sapwood were also measured at the start and end of the trial. After 2 months, only the sapwood of both pine species had significant levels of sapstain; mean values of 37% and 19% for lodgepole pine (year 1 and year 2 respectively) and 12% and 1% for Scots pine. After 4 months, the levels of sapstain in both pine species exceeded 60% in both years. By contrast, very little sapstain developed in the other conifer species with maximum mean values of 10% for Norway spruce, 3.5% for larch and less than 1% for Sitka spruce. Overall, the moisture content of the logs decreased progressively in all species over the length of the trial. However, pine logs tended to retain higher levels of moisture throughout the trial compared with spruce or larch. The relatively higher moisture content of pine sapwood may be closer to the optimal moisture content that sapstain fungi require for infection and colonization, thereby contributing to the increased susceptibility of pine compared with the other conifer species. The pine logs also suffered from some colonization by bark beetles (Ips sexdentatus), which increased the inoculum potential and the opportunity for colonization by sapstain fungi. In addition, particular phenolic compounds in conifer sapwood may play a role in determining the resistance of some species to sapstain. Notably the most resistant species, Sitka spruce, was the only softwood that still retained detectable levels of phenolics in the sapwood to the end of the trial.     

The Effects of Clear Polythene Cloches on Conifer Seedling Growth and Shoot Morphology

Nursery seedbeds sown with lodgepole pine, Scots pine, Douglasfir, Japanese larch and Sitka spruce were covered with clearpolythene cloches for 0, 8, 13 or 18 weeks from sowing. Sitkaspruce germinated poorly and was not studied further. In bothpine species longer durations of cloche cover increased thenumber and length of stem units resulting in taller 1 + 0 seedlings.The number of internodes held at the apex for elongation inthe second season was also increased but transplant shock preventedfull expression of this potential such that differences betweenthe treatments were reduced. Only when covered for 18 weeksdid Douglas fir and Japanese larch produce 1 + 0 seedlings whichwere taller than the uncovered treatments. Through the productionof ‘summer shoots’ in their second season Douglasfir and Japanese larch were able to compensate partially forthe reduction in ‘spring shoot’ length caused bytransplanting.     

Durability of thermally modified sapwood and heartwood of Scots pine and Norway spruce in the modified double layer test

In the present study, durability of untreated and thermally modified sapwood and heartwood of Scots pine and Norway spruce was examined using a modified double layer test. Base layer samples were partly on contact with ground where exposure conditions were harder than that in a double layer test above the ground. The base layer on ground contact gave results already after one year of exposure in Finnish climate, but the top layer of a double layer test element simulated more the situation of decking exposure.

Significant differences in durability and moisture content (MC) between the wood materials were detected after six years of exposure in the field. Thermally modified pine heartwood performed very well in all layers of the test element and only minor signs of decay were found in some of the base samples. Both sapwood and heartwood of thermally modified spruce were suffering only slight amounts of decay while thermally modified pine sapwood was slightly or moderately decayed. Untreated sapwood samples of pine and spruce were severely decayed or reached failure rating after six years in the field. Untreated heartwood samples performed clearly better. The highest MCs were measured from untreated and thermally modified pine samples. Thermal modification increased significantly the durability and decreased the MC values of all wood materials.     

Dissolution and dispersion of spruce wood components into hot water

Summary The dissolution and dispersion of components from Norway spruce (Picea abies) wood were examined in laboratory experiments to determine the factors influencing variations in dissolved and colloidal substances in mechanical pulp suspensions. Finely ground, fresh spruce wood was suspended in water at 90 °C and was agitated intensively for up to 12 h, after which the concentrations of dissolved and dispersed lipophilic extractives, lignans, carbohydrates and lignins were determined. Sapwood and heartwood were studied separately. Effects of pH and added electrolytes on the dissolution and dispersion of wood components were also investigated.Higher amounts of lipophilic extractives, and especially of triglycerides, were dispersed from sapwood than from heartwood. The release of lipophilic extractives continued for up to 3 h, after which the concentrations in the suspensions leveled off. At this stage the composition of the dissolved and dispersed lipophilic extractives equaled that of the wood. The amount of lipophilic extractives in the suspensions increased with increased pH, in the range of 4.5–6.7, but was lower in the presence of electrolytes. The dissolution of carbohydrates continued even beyond 3 h of agitation. The high water temperature induced hydrolytic reactions, thereby releasing especially arabinose. The release of arabinose through the hydrolytic cleavage from polysaccharides was more extensive at pH 4.5 than at pH 5.5 and 6.7. More polysaccharides containing galacturonic acid units (pectins) were dissolved at a higher pH. Much more polysaccharides containing glucose, most probably starch, were present in the sapwood suspensions. The dissolution of lignins also continued throughout the 12 h experiment. The measured UV-absorption, after extraction of lignans, was roughly the same for sapwood and heartwood suspensions. Slightly less lignins were released in the presence of electrolytes. Lignans were released only from heartwood.     

帽儿山主要森林类型凋落物层水文效应研究

对帽儿山地区不同林型的水文功能进行研究,通过分析不同林分凋落物层,得出人工林蓄水能力普遍比天然次生林强。在纯林中落叶松蓄水能力最强,水曲柳林最弱;混交林中红松水曲柳混交蓄水能力最强,云杉水曲柳林蓄水能力最弱;天然次生林蓄水能力较弱。蓄水能力总的排序：落叶松纯林〉红松纯林〉红松水曲柳混交林〉落叶松水曲柳混交林〉云杉纯林〉云杉水曲柳混交林〉天然次生林〉水曲柳纯林。蓄水能力与林分类型,枯落物分解状况及枯落物积累状况有关。     

The spreading of the S type of Heterobasidion annosum from Norway spruce stumps to the subsequent tree stand

The occurrence of Heterobasidion annosum in stumps and growing trees was investigated on 15 forest sites in southern Finland where the previous tree stand had been Norway spruce (Picea abies) infected by H. annosum, and the present stand was either Scots pine (Pinus sylvestris), lodgepole pine (Pinus contorta), Siberian larch (Larix siberica), silver birch (Betula pendula) or Norway spruce 8–53 years old. Out of 712 spruce stumps investigated of the previous tree stand, 26.3% were infected by the S group and 0.3% by the P group of H. annosum. The fungus was alive and the fruit bodies were active even in stumps cut 46 years ago. In the subsequent stand, the proportion of trees with root rot increased in spruce stands and decreased in stands of other tree species. On average, one S type genet spreading from an old spruce stump had infected 3.0 trees in the following spruce stand, 0.5 trees in lodgepole pine, 0.3 trees in Siberian larch, 0.05 trees in Scots pine and 0.03 trees in silver birch stand. Although silver birch generally was highly resistant to the S type of H. annosum, infected trees were found on one site that was planted with birch of a very northern provenance.     

The effect of Phlebiopsis gigantea and urea stump treatment against spore infection of Heterobasidion spp. on hybrid larch (Larix × eurolepis) in southern Sweden

The efficacy of stump treatment with 40% urea solution and spore suspension of Phlebiopsis gigantea against primary infection by Heterobasidion spp. on hybrid larch (Larix × eurolepis) was tested in two field studies in southern Sweden. In the first study, stumps treated with urea or P. gigantea were sampled by cutting a cross‐section disc after 3 months following exposure to natural conditions, and in the second study, stumps treated with only P. gigantea were sampled after 2 months. Spore traps made from hybrid larch, Norway spruce (Picea abies) and Scots pine (Pinus sylvestris) were used to estimate the abundance of ambient spores of Heterobasidion spp. in the second study. Urea significantly reduced infection frequency of Heterobasidion spp. compared to the control. Phlebiopsis gigantea was less effective at reducing infection frequency in the first study than in the second study. Infections were caused by both H. parviporum and H. annosum sensu stricto. The amount of H. annosum infection in proportion to Heterobasidion spp. was significantly lower on the stumps treated with P. gigantea compared to the control; however, the proportion of H. parviporum on the treated stumps was not higher than the control in the first study. Spore traps made of Scots pine had a significantly higher frequency of infection than Norway spruce and hybrid larch. Only the spore traps made of hybrid larch showed significant correlation with the control stumps in terms of relative infected area. Conclusively, it seems prudent to protect hybrid larch stumps from primary infection by Heterobasidion spp., and both urea and P. gigantea can be recommended as stump treatment agents on hybrid larch, even if urea seemed to present more stable results.     

Larval performances and life cycle completion of the Siberian moth, <Emphasis Type="Italic">Dendrolimus sibiricus</Emphasis> (Lepidoptera: Lasiocampidae), on potential host plants in Europe: a laboratory study on potted trees

The Siberian moth, Dendrolimus sibiricus, Tschtv. is the most harmful defoliator of coniferous forests in North Asia. The pest has already spread over the Urals and continues moving westwards. Recently, it has been recommended for quarantine in member countries by European and Mediterranean Plant Protection Organization (EPPO). The performances of the pest on coniferous species planted in Europe were assessed on a range of potted trees corresponding to the spectrum of economically important conifers in the EU: European larch Larix decidua, Norway spruce Picea abies, Scots pine Pinus sylvestris, European black pine Pinus nigra, and the North American species: Douglas fir Pseudotsuga menziesii and grand fir Abies grandis. Larvae showed a potential to survive and complete the development on all these host tree species. Favorable hosts were grand fir, European larch, and Douglas fir that allowed higher survival, better larval development, and as a result, yielded heavier pupae and adult moths with higher longevity. Black pine was a poor host but, however, could still support larval and pupal development. Norway spruce and Scots pine had an intermediate behavior. If accidentally introduced to Europe, the Siberian moth may become especially damaging in forest stands predominated by European larch and by the North American firs. Norway spruce and especially the two-needle pines will be less prone to intensive defoliation by this species. The fact that the pest may damage the range of economically important coniferous species should be taken into account in the pest risk assessment for Europe and also for North America where the Siberian moth occurrence is considered likely.     

In vitro testing for the resistance of conifers to the fungus Phaeolus schweinitzii (Fr.) Pat. on callus cultures

Defence reactions to the fungus Phaeolus schweinitzii were studied in spruce, pine, larch and Douglas-fir callus cultures. It was found that the defence reaction of the callus culture could indicate the degree of the resistance of the tree species to the wood-destroying fungus.     

H-NMR studies of water interactions in sitka spruce and western hemlock: moisture content determination and second moments

Summary The purpose of this study was to examine the influence of the moisture level on the cell-wall material in wood using pulsed proton nuclear magnetic resonance. The wood species used were western hemlock (Tsuga heterophylla (Raf.) Sarg.) and sitka spruce (Picea sitchensis (Bong.) Carr.), distinguishing between heartwood and sapwood regions. The moisture contents of the specimens were below the fibre saturation point and they were conditioned to equilibrium moisture contents based on initial desorption, adsorption and secondary desorption processes. From the FID experiments, the NMR-based moisture contents and the solid-wood lineshape second moments were determined. Average relative proton-spin densities, which were needed to calculate the NMR-based moisture contents, were determined from known moisture contents and they were: hemlock sapwood: 0.616; hemlock heartwood: 0.537; spruce sapwood: 0.679; and, spruce heartwood: 0.446. The average RSD value, considering both heartwood and sapwood, for western hemlock species was 0.577 and for sitka spruce was 0.563; these are close to published RSD values for other species. The condition as to how the equilibrium moisture content was attained did not influence the second moment for hemlock; however, for spruce sapwood, the second moments were sorption dependent. The hemlock M₂ decreased from about 5.1 × 10⁹ s^-2 at low M_NMR to 4.5 × 10⁹ s^-2 (heartwood) and 4.3 × 10⁹ s^-2 (sapwood) at higher m_nmr. The adsorption and secondary desorption M₂ for the spruce sapwood region decreased from about 5.0 × 10⁹ s^-2 at low m_nmr to about 4.1 × 10⁹ s^-2 near the M_F, whereas M₂ for the spruce heartwood decreased from about 4.3 × 10⁹ s^-2 at low M_NMR to about 3.5 × 10⁹ s^-2 near M_F. Extractives may have a key role in obtaining the RSD and second moments.This project was financially supported by the Science Council of British Columbia, MacMillan Bloedel Research. The Natural Sciences and Engineering Research Council of Canada is acknowledged for their support of the NMR spectrometer measurements     

CONIFEROUS LITTER AMENDMENTS AND THE GROWTH OF SITKA SPRUCE

The growth of semi-checked Sitka spruce (Picea sitcbensis Carr.)on heathland has been stimulated to different degrees by theannual application of litter of Scots pine (Pinus sylvestrisL.), Corsican pine (P. nigra var. calabrica Schneid), Lodgepolepine (P. contorta Dougl.), Japanese larch (Larix leptolepisGord.), and Sitka spruce, at rates corresponding to normal plantationconditions, following an initial heavier rate. Foliar analysissuggests that the response is at least partially attributableto the influence of the litters on the nitrogen nutrition ofthe spruce and differences between litters appear to be largelydetermined by their total nitrogen contents. In this respect,Japanese larch, with an annual needle fall generally greaterthan that of the pines and a higher nitrogen content, offersthe best promise as a nurse species for spruce.