Pathogenicity of the pinewood nematode, Bursaphelenchus xylophilus, to Japanese larch, Larix kaempferi, seedlings

Pathogenicity of the pine wood nematode, Bursaphelenchus xylophilus, to Japanese larch, Larix kaempferi, seedlings was tested with inoculation experiments under nursery conditions. Water suspensions of nematodes (mixed stages cultured on Botrytis cinerea or dispersal fourth-stage juveniles (DJ4) extracted from the adult Japanese pine sawyer, Monochamus alternatus) were injected into the stems of 2- and 3-year-old Japanese larch and Japanese black pine, Pinus thunbergii, seedlings growing in a nursery. In another treatment, Japanese pine sawyer adults holding DJ4 were released under a net that covered the upper half of the seedlings. Regardless of nematode inoculation method, Japanese larch seedlings were as susceptible as Japanese black pine seedlings to B. xylophilus under nursery conditions. The rate of disease development was similar on larch and pine seedlings. Nematode population densities were lower in the stems of dead larch seedlings than in the stems of dead pine seedlings. Histopathological observations revealed that the distribution of nematodes in the stems of dead larch seedlings was mostly limited to the cortex, phloem and cambial zone. Traumatic resin canal formation was one of the most characteristic symptoms in larch seedlings which was dissimilar to that in pine seedlings.     

The effects of Laccaria proxima and fibrous pulp waste on the growth of nine container-grown conifer seedling species

 Japanese larch (Larix kaempferi), white spruce (Picea glauca), black spruce (Picea mariana), red spruce (Picea rubens), jack pine (Pinus banksiana), mugo pine (Pinus mugo), red pine (Pinus resinosa), Japanese black pine (Pinus thunbergii) and Douglas-fir (Pseudotsuga menziesii var. menziesii), were inoculated to test the effective host range of the ectomycorrhizal fungus Laccaria proxima and the possibility of utilizing pulp waste as a potting medium for containerized seedling production. Laccaria proxima tended to improve the container growth of Japanese black pine and white spruce, and significantly improved that of jack pine, mugo pine, black spruce, red spruce and Douglasfir. The growth of red pine and Japanese larch were only slightly improved with L. proxima. Pulp waste (33% by volume) had negative effects on tree seedling growth, except for Douglasfir (no significant effect). The interactions of Laccaria proxima and pulp waste varied; the hosts were significantly positive (P<0.01) in the case of jack pine and black spruce, but there was no significant effect for the rest. Negative effects were found with Japanese black pine. Use of pulp waste in seedling production of jack pine, black spruce, mugo pine, red spruce and Douglasfir inoculated with L. proxima and of Japanese black pine both with and without L. proxima is feasible, but further research is necessary to determine the optimal percentage of pulp waste that can be utilized in seedling production of tree species and the field performance of these seedlings. Accepted: 30 August 1995     

Decline and Death of Pinus spp. in Delaware Caused by Bursaphelenchus xylophilus

Etiological studies to determine the cause of decline and death of Pinus spp. in Delaware were initiated in 1980. The pinewood nematode, Bursaphelenchus xylophilus, was found to be the major canse of mortality in Japanese black pine (Pinus thunbergii). When inoculated into healthy 5-yr-old Japanese black pines, B. xylophilus produced typical decline symptoms observed in the field. The xylophilous fungi most often associated with declining trees, Rhizosphaera pini, Fusarium spp., and Pestalotia funerea, were not pathogenic to Japanese black pine in greenhouse tests. Mineral analyses of soil and foliage showed no significant differences between healthy and infested trees. B. xylolyhilus was also found on loblolly pine (P. taeda), scrub pine (P. virginiana), Scots pine (P. sylvestris), red pine (P. resinosa), Eastern white pine (P. strobus), and pitch pine (P. rigida).     

Does ectomycorrhizal fungal community structure vary along a Japanese black pine (Pinus thunbergii) to black locust (Robinia pseudoacacia) gradient?

In this study we examined the role of the nitrogen-fixing tree, Robinia pseudoacacia (black locust), in ectomycorrhizal (ECM) formation and ECM community of Pinus thunbergii (Japanese black pine) seedlings. Two 200 m(2) experimental plots were established at the border between a Japanese black pine- and a black locust-dominated area in a coastal forest. The ECM fungal community of pine seedlings was examined by PCR-RFLP and sequence analysis. We analyzed the relationship between ECM formation, ECM community, growth, and nutrient status of pine seedlings and environmental conditions using the Mantel test and structural equation model. Percentages of ECM root tips, the number of ECM fungal species and ECM diversity on pine seedlings decreased in the black locust-dominated area. Cenococcum geophilum and Russula spp. were dominant in the Japanese black pine-dominated area, whereas Tomentella spp. were dominant in the black locust-dominated area. Nitrogen (N) concentration in soils or pine seedlings strongly influenced the percentage of ECM root tips, the number of ECM fungal species and ECM fungal similarity. These results imply the long-term eutrophication caused by N-fixing trees can change ECM formation and ECM community structure.     

Terminal settling velocity and physical properties of pollen grains in still air

Numerical simulation of wind pollination requires knowledge of pollen grain physical parameters such as size, shape factor, bulk density, and terminal settling velocity. The pollen grain parameters for Japanese cedar, Japanese cypress, short ragweed, Japanese black pine, and Japanese red pine were assessed for dry condition. Terminal settling velocities of dry pollen grains in still air were measured using image analysis of scattered light tracks in a dark settling tube. The measurement system was validated by comparing results to those obtained for standard microspheres of known size and density. Dry pollen grain shape factors indicate the resemblance of particles to spheres, except for pine pollen. Circularity factors of dry pine pollen grains were 0.90–0.86, suggesting more irregular shape than those of other pollen species. Aerodynamic diameters of dry pollen grains were calculated based on the terminal settling velocity. Aerodynamic diameters of Japanese cedar, Japanese cypress, and short ragweed closely resembled the projected area equivalent diameters, suggesting that aerodynamic behaviors of these pollen grains can be managed simply in numerical simulations. However, aerodynamic diameters of dry pine pollen grains were nearly 30 % smaller than projected area equivalent diameters. Sacci on dry pine pollen can reduce the terminal settling velocity through low density and shape effects attributed to their non-sphericity, engendering aerodynamic diameter smaller by more than 10 µm from area equivalent diameters.     

Comparative Studies between Portuguese and Japanese Isolates of the Pinewood Nematode, Bursaphelenchus xylophilus

Comparative studies between Portuguese (T and HF) and Japanese (S10, T4, C14-5 and OKD-1) isolates of the pinewood nematode Bursaphelenchus xylophilus have been made in order to provide information to better understand the possible origin of the Portuguese isolates, recently introduced in the European Union. The main comparative aspects investigated were pathogenicity (seedling mortality ratio), sexual compatibility, and DNA sequences of the rDNA region. Four-year-old Japanese black pine (Pinus thunbergii) seedlings were used as host plants for pathogenicity tests. The Portuguese isolates, and in particular isolate “T,” propagated in higher numbers than the Japanese isolates within pine seedlings. All combinations of crossings produced viable progeny, with higher numbers obtained when crossings were made between Japanese and Portuguese isolates, a possible situation of heterosis and/or inbreeding depression. Reciprocal crossings yielded different values, which may reflect a sex effect (maternal inheritance, mtDNA). Regarding DNA sequencing, both Portuguese isolates displayed nearly identical ITS 1, ITS2, and 5.8S rDNA base sequences as the Japanese isolates. Although biologically very similar, and possibly reflecting a common origin, the Portuguese isolates may present a serious threat to Japanese black pine, due to their higher virulence.     

Short-term effects of thinning and liming on forest soils of pitch pine and Japanese larch plantations in central Korea

The influences of thinning (50% of standing density) and liming (Ca+Mg, 2 Mg ha⁻¹) on soil chemical properties were investigated for 2 years (2001, 2002) in 40-year-old pitch pine (Pinus rigida Mill.) and 44-year-old Japanese larch (Larix leptolepis Gord.) plantations established on similar soils. In general, soil properties varied significantly among plantations and treatments. For both plantations, thinning significantly increased soil organic C (SOC) concentrations whereas there were no significant changes in soil pH and Ca and Mg concentrations. In addition, thinning increased total soil N and Na concentrations for the pitch pine plantation and available P concentration for the Japanese larch plantation in the second year after the treatment. Liming did not affect soil chemical characteristics for the pitch pine plantation except for Na concentration. However, for the Japanese larch plantation, liming significantly increased soil pH and K, Ca and Mg concentrations and decreased SOC and total soil N concentrations. For both plantations, soil Al concentration did not change after thinning and liming and decreased exponentially with increased pH values. The increases in SOC and total soil N concentrations after thinning were possibly due to increases in decomposition of organic matter and root death. Although differences were not statistically significant, soil available P concentration tended to increase at early stages of liming for both plantations. These results suggested that thinning and liming seemed to regulate soil chemical properties for pitch pine and Japanese larch plantations established on similar soils.     

On the Taxonomy and Morphology of the Pine Wood Nematode, Bursaphelenchus xylophilus (Steiner &Buhrer 1934) Nickle 1970

During the past 3 yr, nematologists in the United States have found specimens of Bursaphelenchus sp. in the wood of dead and dying pine trees. This nematode-host association resembles a similar interaction reported from Japan where pine trees are being killed by the pine wood nematode. This taxonomic research was conducted to determine if the Japanese pine wood nematode and similar populations in the United States are of the same species. Based upon typical morphological characters of original specimens of Bursaphelenchus xylophilus (Steiner and Buhrer 1934) Nickle 1970 that were rediscovered in the USDA Nematode Collection and genetic crosses among the Japanese and American nematode populations, it was concluded that they are all the same species, B. xylophilus.     

Detection of xylem cavitation in field-grown pine trees using the acoustic emission technique

The relationship between the ultrasonic acoustic emission (AE) and the needle water potential in field-grown Japanese black pine,Pinus thunbergii, and excised-pine shoots was investigated in summer. The AE technique was employed as a reliable indicator of cavitation events in the water columns of xylem tracheids even under field conditions. In excised shoots. AE events detected when the needle water potential fell below about-0.9 MPa. In the case of field-grown pine trees, however, relatively few AE event were detected when needle water potential was lower than-0.9 MPa. This suggests that the occurrence of cavitation events in Japanese black pine growing under field conditions is comparatively rare, even in summer.     

油松与赤松疱锈病病原及转主的研究

油松与赤松疱锈病是两针松类枝干上的重要病害,从油松和赤松主干病皮上取锈孢子分别向山芍药、芍药、返顾马先蒿、轮花马先蒿、长白茶藨子、东北茶藨子、香茶藨子和刺李进行室内和室外人工接种试验,终于在山芍药、芍药上接种成功,产生与自然状态下相同的典型症状。同样方法用夏孢子接种也都成功,在两种芍药上所产生的夏孢子堆与冬孢子柱以及夏孢子与冬孢子的形态也基本相同。由此可以肯定,油松疱锈病与赤松疱锈病是同一病原,病原菌为松芍柱锈菌[cronartium flaccidum(Alb·et Schw·)Wint]。取锈孢子用同样方法经多次在返顾马先蒿、轮花马先蒿、长白茶藨子、东北茶藨子、香茶藨子和刺李上接种,均未成功,因此认为这些植物与此两种疱锈病的发生无关,两种疱锈病在辽宁的转主寄主是山芍药(Paeonia obovata Haxim.)和芍药(P.lactiflora Pall.)。     

Phylogeography of the Korean pine (Pinus koraiensis) in northeast Asia: inferences from organelle gene sequences

Range-wide genetic variation of Korean pine (Pinus koraiensis) was assessed using maternally inherited mtDNA and paternally inherited cpDNA for 16 natural populations throughout northeast Asia in order to study its phylogeographical history during the Quaternary. The cpDNA variation indicated that there was no difference between populations on the Asian continent and those in the Japanese archipelago. In contrast, the mtDNA variation indicated that there was significant difference between the populations from the two regions, with each region having a different lineage. The continental populations exhibited no diversity in the mtDNA examined despite the species’ current extensive range and large populations. Conversely, while the Korean pine is rare in Japan, the Japanese populations exhibited greater levels of mtDNA diversity (H _T?=?0.502). The higher mtDNA diversity and evidence from numerous Korean pine macrofossil remains dated to the Pleistocene and recovered various sites in Japan suggest that the Japanese archipelago once served as a refugium to a much larger Korean pine population with a more extensive range than is the case today. The presence of the single mtDNA haplotype across the Asian continent suggests that the present widespread populations could have expanded from a single refugium population after the last glacial periods.     

Distribution of ectomycorrhizal and pathogenic fungi in soil along a vegetational change from Japanese black pine (Pinus thunbergii) to black locust (Robinia pseudoacacia)

The nitrogen-fixing tree black locust (Robinia pseudoacacia L.) seems to affect ectomycorrhizal (ECM) colonization and disease severity of Japanese black pine (Pinus thunbergii Parl.) seedlings. We examined the effect of black locust on the distribution of ECM and pathogenic fungi in soil. DNA was extracted from soil at depths of 0–5 and 5–10 cm, collected from the border between a Japanese black pine- and a black locust-dominated forest, and the distribution of these fungi was investigated by denaturing gradient gel electrophoresis. The effect of soil nutrition and pH on fungal distribution was also examined. Tomentella sp. 1 and Tomentella sp. 2 were not detected from some subplots in the Japanese black pine-dominated forest. Ectomycorrhizas formed by Tomentella spp. were dominant in black locust-dominated subplots and very little in the Japanese black pine-dominated forest. Therefore, the distribution may be influenced by the distribution of inoculum potential, although we could not detect significant relationships between the distribution of Tomentella spp. on pine seedlings and in soils. The other ECM fungi were detected in soils in subplots where the ECM fungi was not detected on pine seedlings, and there was no significant correlation between the distribution of the ECM fungi on pine seedlings and in soils. Therefore, inoculum potential seemed to not always influence the ECM community on roots. The distribution of Lactarius quieticolor and Tomentella sp. 2 in soil at a depth of 0–5 cm positively correlated with soil phosphate (soil P) and that of Tomentella sp. 2 also positively correlated with soil nitrogen (soil N). These results suggest the possibility that the distribution of inoculum potential of the ECM fungi was affected by soil N and soil P. Although the mortality of the pine seedlings was higher in the black locust-dominated area than in the Japanese black pine-dominated area, a pathogenic fungus of pine seedlings, Cylindrocladium pacificum, was detected in soil at depths of 0–5 and 5–10 cm from both these areas. This indicates that the disease severity of pine seedlings in this study was influenced by environmental conditions rather than the distribution of inoculum potential.     

Plant species effect on the decomposition and chemical changes of leaf litter in grassland and pine and oak forest soils

Aims

The aim of this study was to examine the effect of plant species differing in functional and phylogenetic traits on the decomposition processes of leaf litter in a grassland of Japanese pampas grass (Miscanthus sinensis) and adjacent forests of Japanese red pine (Pinus densiflora) and Japanese oak (Quercus crispula), representing sequential stages of secondary succession.

Methods

The litterbag experiments were carried out for 3 years in a temperate region of central Japan.

Results

The decomposition constant (Olson’s k) was 0.49, 0.39, and 0.56/year for grass, pine, and oak, respectively. Nitrogen mass decreased in grass leaf litter during decomposition, whereas the absolute amount of nitrogen increased in leaf litter of pine and oak during the first year. Holocellulose in grass leaf litter decomposed selectively over acid-unhydrolyzable residues more markedly than in leaf litter of pine and oak. ¹³C nuclear magnetic resonance analysis also revealed a decrease in the relative area of O-alkyl-C in grass.

Conclusions

The different decomposition among the three litter species implied that the secondary succession from grassland to pine forest and from pine to oak forests could decrease and increase, respectively, the rate of accumulation and turnover of organic materials and N in soils.     

Development of secondary pine forests after pine wilt disease in western Japan

Abstract. The development of secondary Pinus densiflora (Japanese red pine) forests after pine wilt disease was studied through phytosociological analysis, estimation of forest structure before disease and size-structure, tree ring and stem analyses. Following the end of the disease, the growth of previously suppressed small oak trees was accelerated. This is quite different from the development of forests following fire, which starts with the establishment of pine seedlings. Pine wilt disease shifted the dominance of secondary forests from Pinus densiflora to Quercus serrata oak forest. In pine forests, disturbance by fire is important for forest maintenance. In contrast, disturbance by pine wilt disease leads to an acceleration of succession from pine forest to oak forest.     

Improved AFLP protocol using dual-suppression PCR and its application to species with large genomes

To improve the amplified fragment length polymorphism assay, dual-suppression PCR was introduced into the preamplification step of the assay. The dual-suppression PCR blocked completely the amplification of fragments with the same sequence (Bsp1407I-Bsp1407I or NlaIII-NlaIII) at both ends and amplified selectively fragments with different adaptor sequences (Bsp1407I-NlaIII) at each end. Two protocols, referred to as A and B, were established for species with medium- and large-sized genomes, respectively. Both protocols incorporated the dual-suppression PCR. Protocol A resulted in high-quality electrophoretic profiles for black cottonwood and rice, which have medium-sized genomes. In protocol B, an intensely selective PCR step was added to protocol A. Protocol B yielded profiles for Japanese black pine and Japanese cedar that were improved significantly relative to protocol A: the number of strong peaks increased and that of low peaks decreased. Japanese black pine and Japanese cedar have large genomes. The optimal profiles were generated with a total of eight or nine selective nucleotides.     

Limonene: attractant kairomone for white pine cone beetles (Coleoptera: Scolytidae) in an eastern white pine seed orchard in western North Carolina

I report on the attraction of the white pine cone beetle, Conophthoru.s coniperda (Schwarz) (Coleoptera: Scolytidae), to traps baited with the host monoterpene limonene in western North Carolina. Both (+)- and (-)-limonene attracted male and female cone beetles to Japanese beetle traps in an eastern white pine, Pinus strobus L., seed orchard near Murphy, NC. Catches of cone beetles were directly proportional to the release rate of (-)-limonene; (+)-limonene was not tested for dose response. Attraction of cone beetles to the pheromone (+/-)-trans-pityol was increased significantly by both enantiomers of limonene. In all experiments, catches of C. coniperda were strongly male biased with no treatment effect on sex ratio. (- )-Limonene had no effect on trap catches of the predator Enoclerus nigripes (Say) to pityol, whereas (+)-limonene interrupted the attraction of E. nigripes to traps baited with pityol. Of six monoterpenes commonly found in white pine cones, only (-)-alpha-pinene elicited attraction of E. nigripes to Japanese beetle traps.     

Verticillium balanoides, a nematode endoparasite associated with pine needles of collapsing Japanese red pine trees in Tsukuba

Verticillium balanoides, isolated from nematodes associated with pine needles of collapsing Japanese red pine trees in Tsukuba, which is new to Japan, was redescribed with illustrations.     

The impact of host plant species on the larval development of the large pine weevil Hylobius abietis L.

• 1 The developmental performance of the large pine weevil Hylobius abietis was studied in the laboratory on four species of conifer, Corsican pine Pinus nigra var. maritima, Sitka spruce Picea sitchensis, Douglas‐fir Pseudotsuga menziesii and Japanese larch Larix kaempferi.
• 2 All species supported development, but, there was considerable variation in larval mortality, development time and weight of adults on emergence between host species.
• 3 Levels of mortality were highest in Japanese larch (77%) and lowest in Corsican pine (8.2%), and the heaviest adults emerged from Corsican pine (130 mg) and the smallest from Douglas‐fir (74 mg).
• 4 A constitutive plant defence chemical, lignin, found to vary within a northern provenance of Sitka spruce, also strongly affected larval development.
• 5 The significance of these findings is discussed in relation to the management of H. abietis.

     

A Taxonomie Note on Sclerodermas harmandi,Ectoparasite of Stem and Wood Boring Insect Larvae (Hymenoptera: Chrysidoidea'-Bethylidae) in South Korea

As a result of natural enemy survey against the Japanese pine sawyer (Monochamus alternates (Hope 1842)), the major vector of pine wood nematode (Bursaphelenchus xylophilus (Steiner & Buhrer 1934)), many samples of the ectoparasitic wasp, Sclerodermas harmandi (Buysson 1903) were collected nationwide in Korea from June to October in 2005. It is redescribed with the figures and biométrie data of apterous and alate females.     

Post-inoculation Population Dynamics of Bursaphelenchus xylophilus and Associated Bacteria in Pine Wilt Disease on Pinus thunbergii

Population dynamics of the pine wood nematode Bursaphelenchus xylophilus (PWN) and its accompanying bacteria in non-inoculated twigs along with the process of the disease was observed in Japanese black pine, Pinus thunbergii inoculated with PWN. In the non-inoculated twigs, bacteria could be detected when only a few pine needles became yellow. Once most needles had turned yellow or brown, the nematode began to appear and the bacterial populations increased. At the late stages of the disease when the inoculated pine was dying and the needles were totally wilted, the populations of both nematode and bacteria started to increase rapidly. Only a few bacterial species were found at the early stages. As the disease process advanced, the bacterial populations increased rapidly in both population size and variety of the species. However, Pseudomonas fluorescens , P. sp., Pantoea sp. and Sphimgomenas pancimobilis, remained dominant.