Effect of Long-Term Forest Fertilization on Scots Pine Xylem Quality and Wood Borer Performance

We tested whether changes in long-term nutrient availability would affect the xylem quality and characteristics of Scots pine trees as a food source for the larvae of the xylophagous wood borer Hylotrupes bajulus L. (Cerambycidae). We looked for an effect of host plant growth and xylem structural traits on H. bajulus larval performance, and looked for delayed effects of long-term forest fertilization on xylem chemical quality. In general, larval performance was dependent on larval developmental stage. However, the growth of larvae also varied with host plant quality (increases in the concentration of nitrogen and carbon-based secondary compounds of xylem were correlated with a decrease in the larval growth rate). The greater annual growth of trees reduced tracheid length and correlated positively with second-instar H. bajulus growth rate. This is consistent with the hypothesis that intrinsic growth patterns of host plants influence the development of the xylophagous wood borer H. bajulus.     

Production of biocatalysts on the basis of recombinant heterologous xylanase producer strains in the Penicillium verruculosum fungus: Their application in the hydrolysis of timber and wood processing industry wastes

The aim of this work was to create biocatalysts with an increased heterologous expression of endo-β-1,4-xylanase of P. canescens using recombinant P. verruculosum strains, to analyze the properties of new enzyme preparations, and to study the saccharifying activity of these preparations in the hydrolysis of plant raw materials, such as hogged aspen and detarred pine wood wastes of the timber and wood processing industries. The xylanase activity of the existing enzymatic preparations is insufficiently high to hydrolyze a xylan-rich biomass. The creation of increasingly xylanolytically active P. verruculosum-based recombinant strains containing homologous or heterologous genes of xylanase and mannanase is therefore a problem of great interest. Using the methods of genetic engineering, we obtained enzymatic preparations that are biocatalysts for the hydrolysis of plant raw material wastes of the sawmilling and wood processing industries and, according to the data of chromatographic fractionation, have compositions of 45–60% cellulase and 20–50% xylanase (which is optimal for the saccharifying of bagasse, along with aspen and pine wood). The originality of our technique lies in the creation of biocatalysts with predetermined properties, thus reducing appreciably the cost of enzyme preparation by eliminating the need to mix components of the carbohydrase complex for the hydrolysis of plant raw materials, e.g., aspen and pine wood.     

Sterols of the phylum Zygomycota: Phylogenetic implications

The sterol composition of 42 fungal species representing six of the eight orders of the Zygomycota was determined using gas-liquid chromatography-mass spectrometry to assess whether the distribution of major sterols in this phylum has taxonomic or phylogenetic relevance. Ergosterol, 22-dihydroergosterol, 24-methyl cholesterol, cholesterol, and desmosterol were detected as the major sterols among the species studied. Ergosterol was the major sterol of the Dimargaritales, Zoopagales, and 13 of the 14 Mucorales families included in this study. Desmosterol appeared to be the characteristic sterol of the Mortierellaceae (Mucorales). 24-Methyl cholesterol was the major sterol of the Entomophthorales genera Entomophthora, Conidiobolus and Basidiobolus, but cholesterol was the sole sterol detected in Delacroixia coronatus. The Kickxellales species analyzed in this study were characterized by 22-dihydroergosterol as the major sterol. These results suggest that certain orders of the Zygomycota may be distinguished on the basis of major sterol. Also, if sterol structure has phylogenetic implications, then orders might be arranged in the order Kickxellales (C₂₈Δ^5,7) → Dimargaritales, Zoopagales and Mucorales (C₂₈Δ^5,7,22) on the basis of evolution of the predominant and presumably most competent sterol, ergosterol. Although the Entomophthorales would be expected to be more primitive than the above orders based on the predominance of C₂₈Δ^5,, it is not apparent from these data that members of the Zygomycota with ergosterol or its precursors as major sterols evolved from this taxon or the Chytridiomycota.     

Exogenous Brassinosteroid Facilitates Xylem Development in Pinus massoniana Seedlings

Brassinosteroids (BRs) are known to be essential regulators for wood formation in herbaceous plants and poplar, but their roles in secondary growth and xylem development are still not well-defined, especially in pines. Here, we treated Pinus massoniana seedlings with different concentrations of exogenous BRs, and assayed the effects on plant growth, xylem development, endogenous phytohormone contents and gene expression within stems. Application of exogenous BR resulted in improving development of xylem more than phloem, and promoting xylem development in a dosage-dependent manner in a certain concentration rage. Endogenous hormone determination showed that BR may interact with other phytohormones in regulating xylem development. RNA-seq analysis revealed that some conventional phenylpropanoid biosynthesis- or lignin synthesis-related genes were downregulated, but the lignin content was elevated, suggesting that new lignin synthesis pathways or other cell wall components should be activated by BR treatment in P. massoniana. The results presented here reveal the foundational role of BRs in regulating plant secondary growth, and provide the basis for understanding molecular mechanisms of xylem development in P. massoniana.     

Thermal Behavior of Deteriorated Wood Waste

Abstract

This paper investigates the thermal behavior of woody biomass waste—demolition wood of Japanese cedar (Cryptomeria japonica) and insect-attacked forest residue of Japanese red pine (Pinus densiflora)—using proximate analyses, thermogravimetry (TG), and differential thermal analysis (DTA), with comparison to virgin wood. For the pine samples, there was no significant difference in thermal behavior or elemental composition between the virgin pine and pine that had been damaged by insects, indicating that insect-damaged pine received here can be treated as virgin pine in terms of energy utilization. The cedar demolition wood used here was partly degraded by termites or fungi. Its degraded part had a lower weight loss rate under nitrogen and a broader exothermic peak in the char combustion stage under air than cedar virgin wood. The changes in the relative levels of the chemical components and the resultant chemical changes that occur upon fungal degradation might complicate char formation.     

Drywood termite feeding deterrents in sugar pine and antitermitic activity of related compounds

The feeding deterrent activity of sapwood extracts of sugar pine,Pinus lambertiana Dougl., and related compounds was determined against immatures of the western drywood termite,Incisitermes minor (Hagen). A bioassay was designed to quantify reductions in termite feeding caused by deterrent chemicals. Crude extracts and isolated fractions of sugar pine were deterrent and not preferred byI. minor at 0.5 mg/cm². Fatty acids occurring in sugar pine extracts had a broad range of deterrent activity. Long-chain saturated fatty acids were deterrent at 0.25 and 0.05 mg/cm². Unsaturated or intermediate length (C₈-C₁₄) acids, many not found in sugar pine wood, were less active than long-chain saturated acids. Related alpha-halogenated compounds were highly deterrent at 0.05 mg/cm² regardless of chain length or presence of a carboxylic acid moiety. Deposits of 2-iodooctadecanoic acid reduced termite feeding at 5 μg/cm², while 2-bromooctadecanoic acid had deterrent activity comparable to commercial wood preservatives. None of the halogenated compounds tested were termiticidal.     

Effects of fungal decay on properties of mechanical,chemical, and water absorption of wood plastic composites

This study investigated the associations between wood species and fungal resistance, as well as the effects of fungal decay on the properties of mechanical, chemical, and water absorption of wood polypropylene composites (WPCs) filled with white poplar, moso-bamboo, Chinese fir, Ramin, white pine, and rubber wood. Experimental results on weight losses and surface morphology both showed that fungal resistance of WPCs varied significantly with wood fiber species. Chinese fir and rubber wood filled composites separately presented the most and least durability against Coriolus versicolor (white rot) and Poria placenta (brown rot). In addition, fungal decay produced great differences in the properties of mechanical, chemical, and water absorptions between non-decayed and decayed composites. The decayed composites showed lower MOR, tensile strength, and impact strength, as well as higher MOE and water absorptions compared with non-decayed samples. These findings suggest that fungal decay could bring out dramatic influences on various properties of WPCs.     

Screening and Functional Analysis of the Peroxiredoxin Specifically Expressed in Bursaphelenchus xylophilus—The Causative Agent of Pine Wilt Disease

The pine wood nematode, Bursaphelenchus xylophilus, is the causal agent of pine wilt disease. Accurately differentiating B. xylophilus from other nematodes species, especially its related species B. mucronatus, is important for pine wood nematode detection. Thus, we attempted to identify a specific protein in the pine wood nematode using proteomics technology. Here, we compared the proteomes of B. xylophilus and B. mucronatus using Two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight (MALDI-TOF/TOF-MS) technologies. In total, 15 highly expressed proteins were identified in B. xylophilus compared with B. mucronatus. Subsequently, the specificity of the proteins identified was confirmed by PCR using the genomic DNA of other nematode species. Finally, a gene encoding a specific protein (Bx-Prx) was obtained. This gene was cloned and expressed in E. coli. The in situ hybridisation pattern of Bx-Prx showed that it was expressed strongly in the tail of B. xylophilus. RNAi was used to assess the function of Bx-Prx, the results indicated that the gene was associated with the reproduction and pathogenicity of B. xylophilus. This discovery provides fundamental information for identifying B. xylophilus via a molecular approach. Moreover, the purified recombinant protein has potential as a candidate diagnostic antigen of pine wilt disease, which may lead to a new immunological detection method for the pine wood nematode.     

Correlation of 13C‐NMR analysis with fungal decay tests of polymeric structural wood constituents. II. Ground contact tests

Heat treatment at relatively high temperatures (ranging from 150°C to 260°C) appear to be an effective method to improve the durability of wood. This study investigated the reasons for the decay resistance of heat‐treated and untreated wood as composed of polymeric structural constituents by solid‐state CP‐MAS ¹³C‐NMR analysis after fungal exposure in ground contact. An industrially used two‐stage heat treatment method under relatively mild conditions (<200°C) was used to treat the samples. Fungal exposure in ground contact resulted in strong degradation of the carbohydrates (cellulose and hemicellulose) of treated and untreated Scots pine, Radiata pine, and Simaruba. Fungal attack of the carbohydrates appeared to occur mainly at C4, resulting in cleavage and eventually depolymerization of cellulose and hemicellulose. The CP‐MAS ¹³C‐NMR spectra of the heat‐treated wood revealed similarities but also clear differences after fungal exposure in ground contact with the untreated wood. In ground contact fungi appeared to attack the carbohydrates of heat‐treated wood at C1 and possibly at C4 in order to cleave and eventually depolymerize cellulose and hemicellulose. An attack on the out‐of‐the‐ring alcoholic group, ? CH₂OH, of the carbohydrates of the heat‐treated wood was observed (particularly in treated Radiata pine). The fungus possibly tried to cleave the out‐of‐the‐ring CH2? OH group on the main H‐bond fixing sites of the crystalline cellulose structure in order to open the cellulose crystalline structure to an amorphous structure to decrease its water repellency and facilitate enzymatic cellulose degradation; this was also observed, but to a lesser extent, in untreated Radiata pine and untreated Scots pine. The opening of the glucose pyranose ring in heat‐treated Simaruba after fungal exposure, not observed in the untreated wood, was remarkable, and the thermal degradation of alpha‐arabinofuranose during heat treatment indicated more extensive decay. Demethoxylation and ring opening of the aromatic structure of lignin were observed, especially in the heat‐treated Radiata pine, Douglas fir, and Simaruba. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 102: 616–622, 2006     

Mevalocidin: A Novel, Phloem Mobile Phytotoxin from Fusarium DA056446 and Rosellinia DA092917

A multiyear effort to identify new natural products was built on a hypothesis that both phytotoxins from plant pathogens and antimicrobial compounds might demonstrate herbicidal activity. The discovery of one such compound, mevalocidin, is described in the current report. Mevalocidin was discovered from static cultures of two unrelated fungal isolates designated Rosellinia DA092917 and Fusarium DA056446. The chemical structure was confirmed by independent synthesis. Mevalocidin demonstrated broad spectrum post-emergence activity on grasses and broadleaves and produced a unique set of visual symptoms on treated plants suggesting a novel mode of action. Mevalocidin was rapidly absorbed in a representative grass and broadleaf plant. Translocation occurred from the treated leaf to other plant parts including roots confirming phloem as well as xylem mobility. By 24 hr after application, over 20 % had been redistributed through-out the plant. Mevalocidin is a unique phytotoxin based on its chemistry, with the uncommon attribute of demonstrating both xylem and phloem mobility in grass and broadleaf plants.     

Application of the “fusion” approach for the production of highly efficient biocatalysts based on recombinant strains of the fungus Penicillium verruculosum for the conversion of cellulose-containing biomass

Hydrolysis of cellulose-containing biomass mediated by biocatalysts (enzyme preparations, EP) is one of the most advanced and environmentally friendly methods of obtaining a range of useful substances. A new approach to creating recombinant EPs with predefined properties, which consists in applying fusion constructs for the cloning of genes encoding target enzymes, was used in the present study. A number of EPs with different properties was derived from a strain of the fungus Penicillium verruculosum using fusion constructs; these preparations are of interest primarily as additives enhancing the hydrolytic capacity of the basic cellulolytic complex from P. verruculosum. Use of the new EPs in combination with the basic EP from P. verruculosum resulted in an increase of the biocatalytic (hydrolytic) efficiency of the latter towards cellulose-containing raw materials of plant origin. Addition of 20% of the new EP to the basic EP without changing the total EP dose in the reaction mixture resulted in a significant (up to 70%) increase of the efficiency of hydrolysis of cellulose-containing substrates (ground aspen wood and shredded deresined pine wood).     

Novel Green Liquor Pretreatment of Loblolly Pine Chips to Facilitate Enzymatic Hydrolysis into Fermentable Sugars for Ethanol Production

Abstract

Softwood species generally have been found very recalcitrant to enzymatic hydrolysis of the carbohydrate fractions to monomeric sugars. To solve this problem, loblolly pine chips were pretreated with green liquor at 12–20% Total Titratable Alkali (TTA) (as Na₂O on wood) at 170°C for 800 H-factor. The yield of resulting pulp was 76.5–78.6% and the lignin content decreased from 29.2 to 20.2–22.4% and the total polysaccharide decreased from 62.6 to 53.8–55.0%, all based on the weight of original wood. When the pulp was subjected to enzymatic hydrolysis using 40 Filter Paper Unit (FPU)/g pulp, only 41% of the polysaccharides in wood were converted to monomeric sugars. This conversion figure is much lower than that of mixed southern hardwoods (80%) treated under similar conditions. If the green liquor treated pulp was further subjected to either oxygen delignification or mechanical refining prior to the enzymatic hydrolysis, the conversion rate increased to around 55% and 60%, respectively. Furthermore, combination of oxygen delignification and refining further increased the total sugar conversion to 78% of the total sugar in wood, approximately equal to that of the mixed southern hardwoods.     

Screening of Ophiostoma Species for Removal of Eucalyptus Extractives

Abstract

Ophiostoma species have been demonstrated to metabolize wood extractives and be useful to the pulp and paper industry. In order to have new isolates for the Asian market, Eucalyptus camaldulensis logs were harvested from forest sites in central Taiwan and 28 strains of the Ophiostoma genus were isolated from them. These strains were subsequently inoculated onto Eucalyptus wood chips to evaluate their effects on weight losses of wood and the amounts of acetone extractives degraded. At the same time, Gas Chromatography-Mass Spectroscopy (GC-MS) analysis was conducted and by using calibration curves and a database of GC-MS mass spectra, changes in lipophilic compounds were analyzed. Fatty acids, hydrocarbons, sterol compounds, sterol esters, and triglycerides were significantly reduced after two weeks’ inoculation by the fungal strains. The results show that six of the strains were capable of reducing the lipophilic fractions by more than 60% in a two-week treatment. DNA of the most effective strains were analyzed and found to be a variant of Ophiostoma querci.     

Identification of Autophagy in the Pine Wood Nematode Bursaphelenchus xylophilus and the Molecular Characterization and Functional Analysis of Two Novel Autophagy-Related Genes,BxATG1 and BxATG8

The pine wood nematode, Bursaphelenchus xylophilus, causes huge economic losses in pine forests, has a complex life cycle, and shows the remarkable ability to survive under unfavorable and changing environmental conditions. This ability may be related to autophagy, which is still poorly understood in B. xylophilus and no autophagy-related genes have been previously characterized. In this study, transmission electron microscopy was used to confirm that autophagy exists in B. xylophilus. The full-length cDNAs of BxATG1 and BxATG8 were first cloned from B. xylophilus, and BxATG1 and BxATG8 were characterized using bioinformatics methods. The expression pattern of the autophagy marker BxATG8 was investigated using in situ hybridization (ISH). BxATG8 was expressed in esophageal gland and hypodermal seam cells. We tested the effects of RNA interference (RNAi) on BxATG1 and BxATG8. The results revealed that BxATG1 and BxATG8 were likely associated with propagation of nematodes on fungal mats. This study confirmed the molecular characterization and functions of BxATG1 and BxATG8 in B. xylophilus and provided fundamental information between autophagy and B. xylophilus.     

Bacteria Associated with a Tree-Killing Insect Reduce Concentrations of Plant Defense Compounds

Bark beetles encounter a diverse array of constitutive and rapidly induced terpenes when attempting to colonize living conifers. Concentrations of these compounds at entry sites can rapidly reach levels toxic to beetles, their brood, and fungal symbionts. Large numbers of beetles can overwhelm tree defenses via pheromone-mediated mass attacks, but the mechanisms are poorly understood. We show that bacteria associated with mountain pine beetles can metabolize monoterpenes and diterpene acids. The abilities of different symbionts to reduce concentrations of different terpenes appear complementary. Serratia reduced concentrations of all monoterpenes applied to media by 55–75 %, except for α-pinene. Beetle-associated Rahnella reduced (?)- and (+)-α-pinene by 40 % and 45 %, respectively. Serratia and Brevundimonas reduced diterpene abietic acid levels by 100 % at low concentrations. However, high concentrations exhausted this ability, suggesting that opposing rates of bacterial metabolism and plant induction of terpenes are critical. The two major fungal symbionts of mountain pine beetle, Grosmannia clavigera and Ophiostoma montium were highly susceptible to abietic acid. Grosmannia clavigera did not reduce total monoterpene concentrations in lodgepole pine turpentine. We propose the ability of bark beetles to exert landscape-scale impacts may arise partly from micro-scale processes driven by bacterial symbionts.