Genetic variation in fruitfulness in a hinoki (Chamaecyparis obtusa Endl.) seed orchard and its impact on the maintenance of genetic diversity in seedlots

Genetic variation in seed/cone production among clones was studied in a hinoki (Chamaecyparis obtusa Endl.) seed orchard containing 25 plus-trees by analyzing the number of cones, the yield of cones and seeds of individual ramets for 5 successive years (1982 to 1986). There was significant variation among clones each year and parental contribution in the seed orchard. Specifically, in the years 1982, 1983, 1984, 1985, and 1986, 20% of the clones produced 37.2, 60.6, 36.0, 44.3, and 44.8% of the total cones, respectively. The size of the crop greatly influenced the parental balance in the resulting seed/cone crops. The product moment correlation coefficients and Spearman’s coefficients of rank correlation were small and insignificant between consecutive years, but large and highly significant between alternate years, suggesting the presence of carry-over effects in seed/cone production. The broad-sense heritability on a clone mean basis was 0.74 ± 0.15 for the number of cones, 0.72 ± 0.14 for the yield of cones, and 0.68 ± 0.13 for the yield of seeds. The corresponding heritabilities from analyses combined over all years were 0.24, 0.558, and 0.724, respectively. These results indicate that seed/cone production in hinoki is under strong genetic control. Several managerial measures are discussed that maintain the genetic diversity in seedlots used for reforestation, by reducing the variation in seed/cone production among clones and producing seed crops with equal contributions from all parents.     

Individual variation in low boiling point monoterpenes emitted from hinoki (Chamaecyparis obtusa) needles

Variations in the compositions of low-boilingpoint (LBP) monoterpenes in needle samples of 50 hinoki (Chamaecyparis obtusa) trees were investigated using the headspace technique. Considerable compositional variations were revealed, especially in sabinene composition. The sabinene composition varied from 4.9% to 78.0% of the total LBP monoterpenes.-Pinene, myrcene, and limonene also showed considerable variations (9.0%–32.7%, 5.5%–22.6%, 3.6%–29.0% respectively). Analysis of the monoterpene composition allowed definition of four chemotypes based on the contingency table test. No correlation was observed between tree size and LBP monoterpene composition, indicating that the compositional variation in LBP monoterpene exists genetically in this population of hinoki.Part of this paper was presented at the 11th annual meeting of the Chugoku Shikoku branch of the Japan Wood Research Society, Matsue, September 1999     

Liquid culture and transformation of hinoki cypress (Chamaecyparis obtusa Sieb. et Zucc.)

Hinoki cypress (Chamaecyparis obtusa) is one of the most important timber resource forest trees in Japan. Because seed production from a seed orchard of hinoki cypress is not constant every year, micropropagation from a limited amount of material is useful. Up to now, the conventional tissue culture method using solid medium has been used. Here a new method using liquid culture in tubes rotated vertically is described. Shoot primordium of hinoki cypress was inoculated in Campbell and Durzan’s (CD) liquid medium containing different cytokinins (6-benzylaminopurine (BAP), Zeatin, thidiazurone (TDZ)), and the container tubes were rotated vertically around the axis at 2 times / min. Culture room temperature was 25°C and light condition was 16 h photoperiod per day of fluorescent lamps. Zeatin at 1μM concentration was the best for maintaining the shoot primordium production and TDZ induced callus on the surface of the shoot primordia. After shoot primordium multiplication in the liquid culture, they were transplanted to agar medium for shoot elongation. A high concentration of agar (up to 16 g/L) or AVF (anti vitrification factor from Dr. Nairn, 1995) was effective to prevent vitrification of the shoots. Transformation of shoot primordium was done using particle bombardment with vectors containingβ-glucuronidase (GUS) gene or herbicide resistance gene (bar). Positive result for transient transformation was observed with the histo-chemical study for transformation with GUS. Integration of a useful herbicidebar gene into the shoot primordium culture system was also tried and stably transformed plants were obtained. This is the first report of stable transformation of Japanese conifer using practically useful gene. The generous supply of AVF-B from Dr. B.J. Nairn, Tasman Forestry, NZ is also appreciated.     

Seasonal change in the temperature coefficientQ 10 for respiration of field-grown hinoki cypress (Chamaecyparis obtusa) trees

The effect of temperature upon nighttime respiration was examined on four different sized sample trees in a 17-year-old hinoki cypress (Chamaecyparis obtusa (Sieb. et Zucc.) Endl.) stand over two years. Seasonal changes inQ ₁₀ values and their responses to mean temperature were investigated. On the basis of the monthly relationships between nighttime respiration (r) and temperature inside a chamber (θ),r=r ₀exp (kθ), theQ ₁₀ value (=exp(10k)) was calculated. TheQ ₁₀ values were high (Q ₁₀≥3.0) in winter when mean air temperature was low, and gradually decreased toward summer (Q ₁₀≤1.5) through spring with increasing temperature. TheQ ₁₀ values were negatively correlated with mean air temperature. The response ofQ ₁₀ values to mean air temperature was described by a single equation, regardless of tree size. This result, which might be characteristic of this species, shows that respiration ofC. obtusa trees is promoted by slight increases of air temperature in winter season. On the other hand, temperature sensitivity of total respiration reduced during growing season when ambient temperature was high. These chaning temperature sensitivity according to seasons may depend on the seasonal change of the ratio of growth respiration to total respiration. It is concluded that changes in temperature due to changing seasons not only change respiration rate, but also change the response of respiration rate to temperature by shiftingQ ₁₀ values.     

Progeny differences of hinoki (Chamaecyparis obtusa) and sawara (C. pisifera) against resinous stem canker disease and spatial distribution of damage (disease severity) in a progeny test

The incidence and severity of resinous stem canker disease were investigated in hinoki (Chamaecyparis obtusa) and sawara (C. pisifera) at a progeny test located in Yamatsuri Town, Fukushima Prefecture, Japan. Symptoms of the disease were observed in 307 trees out of 933 investigated trees (32.9%). The damage was more severe on lower slopes than on upper slopes, indicating that micro-environmental factors are causally associated with the occurrence of the disease. The severity of the disease varied both among nine open-pollinated progenies derived from hinoki plus-tree clones and among 13 progenies derived from pollination between hinoki plus-tree clones and the mixed-pollen of hinoki plus-tree clones. The severity of the disease also varied with height above the ground. This tendency was most obvious where the disease was most severe. An estimation of the narrow sense heritability,h ², of 0.214, was obtained from analysis of the 13 progenies derived from pollination with mixed-pollen. The heritability derived from material that had not been subjected to selection against the disease, indicated that breeding to enhance resistance would be possible to control the disease. The interaction between environments and progenies in relation to severity of the disease is also discussed. Ms. Setsuko Chiba and Mr. Mamoru Ueta gave us vigorous supports, and Mr. Haruki Orita, Dr. Tomiyasu Miyaura, and Dr. Shinichiro Ito gave us many helpful comments. We deeply appreciated their cooperation. The study was supported by a grant from the Ministry of Agriculture, Forestry. and Fisheries of Japan.     

Water status of hinoki cypress, Chamaecyparis obtusa, attacked by secondary woodboring insects after typhoon strike

We estimated the water status of six 31-year-old hinoki cypress (Chamaecyparis obtusa) trees that were struck by a large typhoon. Measurements were made of the trunk water-conduction area of xylem, leaf water potentials, and diurnal changes in trunk diameters. The secondary woodboring insects, Callidiellum rufipenne and Ovalisia vivata, on the trees were captured by the sticky-trap banding method. In trees on which these wood borers were captured, water conduction areas were reduced, leaf water potentials were lower, and trunk diameter changes (R) fluctuated during the day, suggesting that the trees had been damaged by the typhoon and were under water stress. The secondary woodboring insects attacked the water-stressed trees.     

Effect of Ca on Al-induced activation of antioxidant enzymes in the needles of hinoki cypress (Chamaecyparis obtusa)

The effect of CA(NO₃)₂ on the active oxygen scavenging system in hinoki cypress (Chamaecyparis obtusa) seedlings cultured in a nutrient solution containing aluminum was examined. The hinoki cypress seedlings were transferred to nutrient solutions containing 5 mM AlCl₃ together with various concentrations of Ca(NO₃)₂ in pots containing glass beads and Teflon tips. The growth in height and dry matter allocation to each organ was little influenced over a period of 12 weeks by either Al or the concentration of Ca(NO₃)₂. The activity of superoxide dismutase (SOD) in the needles was stimulated by Al, and the effect of Al was lowered significantly by simultaneous application of 25 mM Ca(NO₃)₂. At week 1, the activity of catalase (CAT) in the needles was increased by Al, but the effect was no longer observed at week 12. The Al concentration in the roots was increased by treatment with Al, whereas the Al concentration in needles was not. These results indicate that rhizospheric Al stress stimulates antioxidative enzyme activities in hinoki cypress needles and the activation of the enzymes is suppressed by addition of Ca. The transmission of Al stress to the needles, which induced a change in the enzyme activity, is not caused by the transfer of the Al ion itself from roots to needles. This work was supported in part by funding from the Japan Science and Technology Corporation, the CREST program 1996–2001, and the Center for Forest Decline Studies.     

Effects of elevated CO2 concentration on the respiratory behavior of the branches of field-grown hinoki cypress (Chamaecyparis obtusa)

The effects of elevated atmospheric CO₂ concentrations on the nighttime respiration were examined for two sample branches of a hinoki cypress tree (Chamaecyparis obtusa) growing in the field with an open gas exchange system for a one-year period from July 1994 to June 1995. The branches were of a similar size and located at a similar position within the crown. One branch was subjected to an elevated CO₂ concentration of 800 μmol mol⁻¹ and the other was subjected to ambient air which had a CO₂ concentration of about 370 μmol mol⁻¹. Nighttime respiration rate was higher in elevated CO₂ level than in ambient CO₂ level. The relationship between nighttime respiration and the corresponding nighttime air temperature was fitted by the exponential function in every month of the year. The segregation of regression lines between the two CO₂ treatments increased gradually as the seasons progressed during the treatment period. TheQ ₁₀ values for nighttime respiration were lower in elevated CO₂ (1.9 ≤Q ₁₀ ≤ 3.7) than in ambient CO₂ (2.4 ≤Q ₁₀ ≤ 4.5) in every month of the year. TheQ ₁₀ was inversely related to the monthly mean nighttime air temperature in both elevated and ambient CO₂. The estimated daily nighttime respiration rate under both CO₂ treatments had a similar seasonal pattern, which almost synchronized with the temperature change. The respiration ratio of elevated CO₂ to ambient CO₂ increased gradually from 1.1 to 1.6 until the end of the experiment. Our results indicate that the CO₂ level and the temperature have a strong interactive effect on respiration and suggest that a potential increase in respiration of branches will occur when ambient CO₂ increases.     

Efficient plant regeneration of Hinoki cypress (Chamaecyparis obtusa) via somatic embryogenesis

This report describes the efficient plant regeneration of Chamaecyparis obtusa Sieb et Zucc. via somatic embryogenesis. Embryogenic cultures were initiated from megagametophytes containing immature zygotic embryos. Embryogenic cultures were maintained and proliferated by 2–3-week interval subcultures in medium supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzylaminopurine. High maturation frequencies of cotyledonary embryos were obtained on maturation medium containing maltose, polyethylene glycol, activated charcoal, and abscisic acid. Somatic embryos germinated readily after transfer to plant growth regulator-free medium. Growth of regenerated emblings has been monitored in a greenhouse.     

Responses of antioxidant enzymes in the needles of hinoki cypress (Chamaecyparis obtusa) seedlings to nutrient solutions containing various calcium/aluminum ratios

An attempt was made to examine the possible connection between the various ratios of calcium/aluminum (Ca/Al) in the nutrient solution of plant cultures and the active oxygen scavenging system of hinoki cypress (Chamaecyparis obtusa) seedlings. The hinoki cypress seedlings were transferred to nutrient solutions containing 5 mM AlCl₃ together with various concentrations of Ca(NO₃)₂ in pots containing glass beads and Teflon tips. The activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) in the needles were estimated at several stages during the 7-day treatment. The samples treated with the lower Ca/Al solutions show the highest SOD activities. The activities of APX and GR, both of them involved in the ascorbate-glutathione cycle, showed the same tendency with decrease to higher Ca/Al ratio. These results indicate that rhizospheric Ca might compete with Al and ameliorate Al toxicity on and in the roots, the Al stress is not transformed to the needles after a few days, and the ascorbate-glutathione cycle in the hinoki cypress needles might fluctuate and be suppressed by the rhizospheric Al stress during the 7 days. This work was supported in part by funding from the Japan Science & Technology Corporation, the CREST program 1996–2001, and the Center for Forest Decline Studies.     

The effect of bark decortication for hiwada production on xylem and phloem formation in Chamaecyparis obtusa

Of all plant materials used to cover the roofs of traditional Japanese buildings, Japanese cypress (Chamaecyparis obtusa) bark, hiwada, has the longest service life and has been used from ancient times. However, wood and bark properties after hiwada harvest have not been evaluated in detail. We studied whether decortication for hiwada production in winter affected xylem and phloem formation. Decorticated trees still preserved all inner bark and part of the outer bark, and both decorticated and control trees had similar annual ring structures at all stem heights in the xylem and phloem. In both xylem and inner bark, no significant difference in ring width at any stem height was found between annual rings before and after decortication. Thus, this study revealed that the decortication of bark for hiwada production does not affect the formation of xylem and the inner and outer bark if decortication is carried out by highly skilled workers in winter.     

Detection of genetic variation among seed and seedlings ofChamaecyparis obtusa using allozyme markers

The effects of nursery practices on genetic composition and structure were studied inChamaecyparis obtusa by analyzing seeds and one-, two-, and three-year-old seedlings. Enzyme polymorphisms of the6Pgd-2, G6pd, Got, Gk, Shd-2, Pod, Dia-1, andPgm loci were used. OnlyDia-1 indicated significantly different allelic frequencies between seeds and three-year-old seedlings. Most of the genetic diversity of the parental clones in this seed orchard could be transferred to the offsprings. Genetic diversity on the basis of an average expected heterozygosity did not differ between seeds and seedlings. A slight excess of homozygotes in seeds and a significant excess of heterozygotes in seedlings of different ages to the expected panmictic proportions were found. Most of the differences in genotypic distributions among the seed and seedling stages, probably resulted from viability selection favoring outcrosses, and selective removal of inbred offsprings including self-fertilized ones prior to their transplantation in the nursery. A part of this paper was orally presented at the 107th Annual Meeting of the Japanese Forestry Society (1996).     

The composition of volatiles from tatami mats containing hinoki (<Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis>) wood-wool and its decline over the long term

Volatiles inside tatami mats containing hinoki (Chamaecyparis obtusa) wood-wool as padding were analyzed. Volatiles were collected with solid-phase microextraction (SPME) fibers in a small chamber prepared in tatami mats and assayed by gas chromatography. Most of the detected compounds are typically found in hinoki extractives. Monoterpenes rapidly decreased at the beginning of the experiment, while the dispersion of sesquiterpenes that contain hydroxyl groups was observed to last more than 1 year. These sesquiterpenes, T-cadinols, and α-cadinols may play a role in the suppression of house dust mites. Part of this report was presented at the 55th Annual Meeting of the Japan Wood Research Society in Kyoto, March 2005     

The penetration rate of laser pulses transmitted from a small-footprint airborne LiDAR: a case study in closed canopy, middle-aged pure sugi (Cryptomeria japonica D. Don) and hinoki cypress (Chamaecyparis obtusa Sieb. et Zucc.) stands in Japan

Penetration rates of laser pulses transmitted from a small-footprint airborne light detection and ranging (LiDAR) system in closed-canopy, middle-aged (40–50 years old) hinoki cypress (Chamaecyparis obtusa Sieb. et Zucc.) and sugi (Cryptomeria japonica D. Don) plantations that had similar levels of canopy openness were studied. The number of transmitted pulses was 107 427 points/ha for the hinoki cypress stand and 122 883 points/ha for the sugi stand. The penetration rates of the first (P_f) and second pulses (P_s) that reached the ground were 0.1% and 1.0% in the hinoki cypress stand, and 2.6% and 5.5% in the sugi stand, respectively. Penetration rates for the total number of pulses (P_f+s) that reached the ground within each stand were 1.1% and 8.1%, respectively, and values for P_f+s, P_f, and P_s for sugi and hinoki cypress stands were significantly different (P < 0.001). The results of this study indicated that the generation of accurate digital terrain models in dense hinoki cypress stands with complex topographies using only the data from laser pulses with poor laser penetration rates is likely to be difficult.     

Phenological observations onLarix principis-rupprechtii Mayr. in primary seed orchard

Through 5 years of phenological observations onLarix principis-rupprechtii Mayr. in primary seed orchard and studies on population and individuals of clones, the annual periodic phenological laws were revealed and the annual phenological periodic table was drawn up. The correlation between various phenophases, the air temperature and active accumulated temperature were analyzed and expounded. The authors also analyzed the similarities and differences of phenophases among clonal individuals as well as the blooming properties of male and female flowers at the same time. This study could provide theoretical reference for working out the production plan of improved varieties and other management measures in seed orchard ofLarix principis-rupprechtii. Biography: ZHANG Xin-bo (1956-), male, senior engineer in Shanxi Academy of Forestry Science and Northern Research Institute of Forestry of China, Taiyuan 030012, P. R. China. Responsible editor: Zhu Hong     

Identification of chloroplast DNA haplotypes ofAbies firma andA. homolepis using a polymerase chain reaction with species-specific primers

DNA sequences of the chloroplast spacer region between thetrnP andtrnW genes (234 bps) were determined for two Japanese fir species,Abies firma Sieb. et Zucc. andA. homolepis Sieb. et Zucc., using four individuals from each species. No intraspecific variation was found in either species, but interspecific sequence polymorphism was detected between the two species. The interspecific variation was three nucleotide changes, from the 148th to 150th nucleotide position. These three nucleotides were TAC inA. firma and GTA inA. homolepis, which were inverted. In order to identify the cpDNA haplotypes between the two species, this inversion polymorphism was utilized to develop a new marker. Species-specific primers were designed so that the 3′ ends of the primers would anneal to the mutation site, in order for the two haplotypes to be easily identified by a polymerase chain reaction (PCR).     

Changes in soil chemical and physical characteristics in Japanese cypress (Chamaecyparis obtusa Endl.) stands by mixture of deciduous broad-leaved trees in the northern Kanto region of Japan

In order to clarify the effects of a mixture of deciduous broad-leaved trees on soil fertility, we investigated litter biomass accumulation, mineral soil chemical and physical characteristics, characteristics of nitrogen mineralization, and the mutual relationships between them in Japanese cypress (Chamaecyparis obtusa) stands mixed with deciduous broad-leaved trees at different ratios (mixture ratio; MR = 0, 16, 33, 43, 100% by basal area) in the northern Kanto region of Japan. Litter biomass in the forest floor and mineral soil was 19.1 Mg ha⁻¹ in MR 0% and decreased approximately 60 % in MR 33%, MR 43% and MR100%. The permeability at 0–5 cm soil depth in MR100% was twice as much as that in MR 0%. Increases in soil permeability were likely due to larger soil pores in the higher MR with much accumulated deciduous broad-leaves. At 0–5 cm soil depth, the differences in carbon concentration among the plots were not clear. On the other hand, carbon concentrations at 5–10 cm depth increased from 90 g kg⁻¹ to 147 g kg⁻¹ with increases in MR from 0% to 100%. Concentrations of exchangeable bases increased two to four times with increases in MR from 0 to 100% at 0–10 cm depth. Soil pH (H₂O) generally increased with increases in MR at each depth. The rates of net nitrogen mineralization at 0–5 cm depthin vitro increased from 25 to 87 mg kg⁻¹ 2 weeks⁻¹ with increases in MR from 0 to 100%. However, increases in nitrification with increases in MR were not clear compared with nitrogen mineralization. These results indicated that a mixture of deciduous broad-leaved trees in a Japanese cypress stand was effective in preventing soil fertility decline. This study was supported by a grant from the Showa Shell Sekiyu Fundation for Promotion of Environmental Research. A part of this study was presented at the 7th International Congress of Ecology (1998).     

Traumatic resin-canal formation caused by inoculation withCistella japonica in secondary phloem ofChamaecyparis obtusa

In the resinous lesion development caused by the inoculation withCistella japonica onChamaecyparis obtusa stem, numerous traumatic resin-canals were formed at certain radial intervals in the secondary phloem. These formation started in the ring grown in the current or the previous year of inoculation and the resin-canals were subsequently formed in the newly grown rings. The resin-canals were arranged in tangential series in one to two lines in an annual-ring. Artificially induced resin-canal formation was similar to that in naturally infected resinous stem canker. The results from the present study reconfirmed thatCi. japonica is the causal agent of the disease.     

Butt rot ofChamaecyparis obtusa (Sieb. et Zucc.) Endlicher trees caused byPerenniporia subacida in Shikoku district, Japan: Pathogen, distribution of damaged trees in the stand, and soil investigation

The rot ofChamaecyparis obtusa (hinoki) trees was studied in the northern part of Kagawa Prefecture, Japan. Among 158 hinoki examined, butt rot was found in 28.6, 58.3, and 100% of trees in the 29, 30, and 34-year-old stands, respectively. All trees with butt rot, and 11 trees without it had rotted roots. Thirteen hinoki were peeled byCervus nippon (sika deer) and all of them were infected with butt rot. The maximum height of rot in deer-damaged trees was 2.6 m from the ground level, but was 1.9 m in undamaged trees. About eighty-two percent of rotted trees showed rotted areas of less than 40 cm² on the cross section of stems at the ground level. White mycelia and black flecks sometimes appeared in the rotted wood. Basidiocarps of polyporaceous fungus were often found on felled logs and rotted stumps of hinoki and identified asPerenniporia subacida. Basidiomycetous fungus was isolated frequently from rotted wood of roots and stems, and determined to beP. subacida by comparative study on cultural characteristics. An inoculation experiment and wood-decay test proved that the fungus was the cause of the rot of hinoki. Few absorbing roots of living trees were found in the clayey subsurface soil of the high dry bulk density and the less soil aeration. Some absorbing roots had root rot and the rot spread from the base of the absorbing root to the central part of the woody root. This is the first report on the rot of hinoki caused byP. subacida in Japan.     

Seedling growth and establishment in natural stands of yellow-cedar (Chamaecyparis nootkatensis) seedlings derived from the use of modified seed dormancy-breaking treatments

Following dispersal from the parent tree, seeds of yellow-cedar (Chamaecyparis nootkatensis[D. Don] Spach) exhibit low germination, primarily as a result of coat-imposed dormancy. Dormancy of the mature (intact) seed is effectively terminated by traditional warm/cold treatments. A chemical treatment using the anaesthetic 1-propanol combined with a three day warm water soak (30 °C), a two day GA₃ treatment and 60 d of moist chilling not only promotes high germinability of yellow-cedar seeds, but also elicits vigorous post-germinative growth following seedling emergence under nursery greenhouse conditions. Here we compare the effectiveness of the more traditional warm/cold treatments with the chemical treatment in terms of their capacity to elicit vigorous growth and establishment in natural stands following transplant of seedlings from a nursery greenhouse environment. Two seed lots (42313 and 43697) and open-pollinated seed from parent trees 13-6 and 19-8 showed equivalent seedling growth in natural stands following the chemical treatment and two traditional warm/cold treatments typically used for dormancy breakage by the forest industry and by the Ministry of Forests in British Columbia. The chemical protocol offers the advantage of reducing the time required to break seed dormancy. We have now demonstrated that it yields seedlings that exhibit vigorous growth and are capable of withstanding the vagaries of the environment.