BackgroundPine wilt disease is caused by the pine wood nematode, Bursaphelenchus xylophilus, which threatens pine forests and forest ecosystems worldwide and causes serious economic losses. In the 40 years since the pathogen was identified, the physiological changes occurring as the disease progresses have been characterized using anatomical and biochemical methods, and resistant trees have been selected via breeding programs. However, no studies have assessed the molecular genetics, e.g. transcriptional changes, associated with infection-induced physiological changes in resistant or susceptible trees.ResultsWe constructed seven subtractive suppression hybridization (SSH) cDNA libraries using time-course sampling of trees inoculated with pine wood nematode at 1, 3, or 7 days post-inoculation (dpi) in susceptible trees and at 1, 3, 7, or 14 dpi in resistant trees. A total of 3,299 sequences was obtained from these cDNA libraries, including from 138 to 315 non-redundant sequences in susceptible SSH libraries and from 351 to 435 in resistant SSH libraries. Using Gene Ontology hierarchy, those non-redundant sequences were classified into 15 subcategories of the biological process Gene Ontology category and 17 subcategories of the molecular function category. The transcriptional components revealed by the Gene Ontology classification clearly differed between resistant and susceptible libraries. Some transcripts were discriminative: expression of antimicrobial peptide and putative pathogenesis-related genes (e.g., PR-1b, 2, 3, 4, 5, 6) was much higher in susceptible trees than in resistant trees at every time point, whereas expression of PR-9, PR-10, and cell wall-related genes (e.g., for hydroxyproline-rich glycoprotein precursor and extensin) was higher in resistant trees than in susceptible trees at 7 and 14 dpi.ConclusionsFollowing inoculation with pine wood nematode, there were marked differences between resistant and susceptible trees in transcript diversity and the timing and level of transcripts expressed in common; in particular, expression of stress response and defense genes differed. This study provided new insight into the differences in the physiological changes between resistant and susceptible trees that have been observed in anatomical and biochemical studies.

Photosynthetic traits of two-year-old Japanese larch seedlings (Larix kaempferi Carr.) grown at elevated CO 2 concentrations were studied in relation to structural changes in the needles. Seedlings were grown at two CO 2 concentrations, 360 (AC) and 720 (EC) µmol mol -1 at high and low nutrient supply rates, high N (HN) and low N (LN). The photosynthetic capacity fell significantly in EC+LN, but increased significantly in EC+HN. Since the mesophyll surface area exposed to intercellular space per unit leaf area (A mes /A) is correlated with the photosynthetic rate, we measured A mes /A for larch needles growing in EC. Changes of A mes /A in both EC+HN and EC+LN were very similar to the changes in photosynthetic capacity. This suggests that the changes of A mes /A in EC probably caused the changes in the photosynthetic capacity. The changes of A mes /A in EC were attributed to changes in the mesophyll cell size and mesophyll cell number. The photosynthetic capacity in EC can be explained by taking morphological and structural adaptations into account as well as biochemical factors.

We evaluated the effects of elevated carbon dioxide concentration ([CO2]) and two nutrient regimes on stem growth rate, annual ring structure and temporal variations in photosynthetic characteristics of seedlings of Japanese larch (Larix kaempferi (Lamb.) Carr.). Seedlings were grown in phytotron chambers in an ambient (360 ppm) or an elevated (720 ppm) [CO2] in two nutrient regimes for one growing season. Elevated [CO2] reduced stem height and increased stem basal diameter compared with ambient [CO2]. The effect of elevated [CO2] on growth tended to be greater at high-nutrient supply than at low-nutrient supply. Elevated [CO2] had no significant effect on ring width or the number of tracheids per radial file. There was no obvious difference in cell wall thickness or the relative area of the cell wall between seedlings grown in ambient or elevated [CO2]. Although growth in elevated [CO2] resulted in a slight increase in cell diameter, the increase had a relatively minor effect on the relative area of the cell wall. Net assimilation rate increased in response to elevated [CO2]; however, the increase in whole-crown photosynthetic rate (Total Agrowth) in seedlings in the elevated [CO2] treatment was minimal because of the smaller specific needle area and acclimation of the photosynthetic characteristics of the needles to the growth [CO2]. In conclusion, we observed no obvious enhancement in the capacity for carbon fixation in Japanese larch seedlings grown in the presence of elevated [CO2] that might be attributable to changes in stem growth. However, elevated [CO2] caused changes in the temporal pattern of stem growth and in some anatomical features of the tracheids.

Knowledge of the genetic relationship between growth traits and wood properties is critical for their simultaneous genetic improvement. We measured the height and diameter at breast height (DBH) and wood quality traits, including stress wave velocity (SWV) as the selection criteria for wood stiffness, wood density, and Pilodyn penetration depth as selection criteria for wood density, at a progeny test site at stand age ca. 30, which comprised of full-sib families by a full diallel mating design with eight plus Larix kaempferi tree clones. We estimated the genetic parameters for each trait and phenotypic, genetic and residual correlation between traits. The contribution of specific combining ability and reciprocal effects were small for all traits. Growth traits showed high positive genetic correlation with average wood density of the outermost five rings (0.912 for height, 0.826 for DBH) and with SWV (0.738 for height, 0.762 for DBH), irrespective of small phenotypic correlations between them. Wood density and SWV also showed high genetic correlation. Pilodyn penetration depth showed high selection efficiency for average wood density of the outermost five rings (79.8 %) whereas SWV showed higher selection efficiency for wood density. Thus, simultaneous genetic improvement of growth traits and wood properties of L. kaempferi appears possible.

Heartwood in tree stems provides natural durability and ornamental values to wood products as well as antimicrobial properties to protect living stems from decay. Control of the amount and quality of heartwood is required because of its importance in wood utilization. Despite the importance, the mechanism of heartwood formation has been poorly understood. To obtain further knowledge for understanding this mechanism, we studied the seasonal variation of various components of heartwood formation in Larix kaempferi (Lamb.) Carrière. Dehydration of tracheids at the boundary between sapwood and intermediate wood occurred continuously during the study period. The death of ray parenchyma cells at intermediate wood occurred from spring to summer. The deposition of heartwood substance took place in autumn to winter. Thus, different components of heartwood formation were active in different seasons. Since heartwood formation is a system consisting of several components within a series of continuing processes, understanding the seasonal change of heartwood formation requires monitoring these components simultaneously.