Minor pollinosis fromCryptomeria japonica in Varese,northern Italy. Aerobiological monitoring and main clinical features

In the last 2 years we have conducted an aerobiological monitoring program ofCryptomeria japonica, a plant belonging to the family of Taxodiaceae that sometimes causes pollinosis in the period from February to April. Throughout 1994, we checked the incidence of its sensitization and the clinical effects in 85 subjects with correlated seasonal symptoms, who gave a positive skin prick test (SPT) for Betulaceae and/or Corylaceae. Twenty-five patients (29.4%; 19 M; 6 F; mean age, 38.8 years) all with oculorhinitis, were SPT positive to an allergenic extract ofCryptomeria. RAST confirmed this positivity in 44% of the cases. No patients showed monosensitization forCryptomeria to either SPT or RAST. Two subjects gave a positive result on specific nasal provocation. RAST inhibition showed no cross-reaction betweenCryptomeria and birch pollen. During the pollen season each patient made a list, scoring symptoms and specifying any drugs used, so we could correlate these elements with aerobiological observations. The pollen concentration probably exceeded the allergizing threshold forCryptomeria on 8 days during 3 months of recording. The intensity and duration of symptoms seemed to be mainly influenced by sensitization to Betulaceae and Corylaceae. It is thus possible that a combination of minor pollinosis may produce seasonal symptoms in allergic patients.     

Pollinosis in trentino (Northern Italy). Aerobiological and clinical research

Summary A study to evaluate and define the atmospheric pollen concentration in Trentino was carried out through the aerobiologic sampling in three localities chosen according to their different climatic conditions.1375 patients with pollinosis living in Trentino were studied retrospectively over the period ranging from 1986 to 1988 and selected according to the area they came from.Results have proved that the most allergenic pollen types are the following: Poaceae, Urticaceae (Parietaria), Compositae (Artemisia) and the tree pollen of Betulaceae and Corylaceae (Alnus, Betula, Corylus), and that pollinosis caused by such pollen, types has different features and frequencies according to the different localities.As far as symptoms are concerned, our data shows that rhinoconjuntivitis is more frequent in those patients who are allergic toParietaria while asthma results being more frequent in patients who are allergic to tree pollen.     

Formation and vertical distribution of sapwood and heartwood in Cryptomeria japonica D. Don

The formation and vertical distribution of sapwood and heartwood were studied with a 45-year-old Cryptomeria japonica D. Don. The tree was grown at a plantation with 1.5 m × 3.0 m spacing near Miao-Li, Taiwan and was felled on 27 February 1992. The thickness of sapwood and heartwood was expressed by a ring count and a linear measurement. The east-west (E-W) wood strips were collected from 0.3 m above ground upwards to the top of the tree at 2.5 m intervals. The sapwood thicknesses from the base to the 10.3 m tree level height are around 20–22 growth rings and 42±2 mm. At the top of the tree, the sapwood thickness is narrower. The heartwood, which decreases in thickness with increasing tree level heights is not found at the top of the tree. The heartwood appears as a conical shape in the tree trunk. There is no statistical difference in sapwood/heartwood thickness between E-W aspects. Tree level heights and the tree level age were found to be important parameters in determining the thickness of sapwood/heartwood.     

Aerobiological and clinical aspects ofParietaria officinalis in the area of Thessaloniki,Greece

Summary We collected the daily pollen samples during a 3-year period (Febr '87–Dec '89), using a Burkard volumetric trap, located on a high level area in the center of the city.Parietaria officinalis pollen was not differentiated under microscope from the other Urticaceae but through phenological criteria. The patients included in the detection of the sensitivity toP. officinalis pollen came from the Out-patient Clinic of Bronchial Asthma of the General Hospital «G. Papanikolaou». They had a seasonal pollinosis and they were submitted to Pricktest using a battery of 22 groups and aP. officinalis pollen extract. The Urticaceae pollen appears first in the atmosphere of Thessaloniki in the end of March, shows a peak in the beginning of May and continues to be present till the end of August. We detectedP. officinalis pollen sensitivity combined with other allergens in 24.1% of the patients and in 1.4% a monosensitivity toP. officinalis.     

Physical map of chloroplast DNA in sugi,Cryptomeria japonica

Summary To investigate the evolution of conifer species, we constructed a physical map of the chloroplast DNA of sugi, Cryptomeria japonica, with four restriction endonucleases, PstI, SalI, SacI and XhoI. The chloroplast genome of C. japonica was found to be a circular molecule with a total size of approximately 133 kb. This molecule lacked an inverted repeat. Twenty genes were localized on the physical map of C. japonica cpDNA by Southern hybridization. The chloroplast genome structure of C. japonica showed considerable rearrangements of the standard genome type found in vascular plants and differed markedly from that of tobacco. The difference was explicable by one deletion and five inversions. The chloroplast genome of C. japonica differed too from that of the genus Pinus which also lacks one of the inverted repeats. The results indicate that the conifer group originated monophyletically from an ancient lineage, and diverged independently after loss of an inverted repeat structure.     

A linkage map for sugi (Cryptomeria japonica) based on RFLP,RAPD, and isozyme loci

A linkage map for sugi was constructed on the basis of restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD), and isozyme loci using a three-generation pedigree prepared for genetic analysis of heartwood color. A total of 128 RFLP (123 cDNA and 5 genomic probes), 33 RAPD, 2 isozyme, and 1 morphological (dwarf) loci segregated in 73 progeny. Of the 164 segregating loci, 145 loci were distributed in 20 linkage groups. Of these loci, 91 with confirmed map positions were assigned to 13 linkage groups, covering a total of 887.3 cM. A clustering of markers with distorted segregation was observed in 6 linkage groups. In the four clusters, distortions with a reduction in the number of homozygotes from one parent only were found.Abbreviations MAS marker-assisted selection - PAGE polyacrylamide gel electrophoresis - QTL quantitative traits of loci - RAPD random amplified polymorphic DNA - RFLP restriction fragment length polymorphism This work was supported by a Grant-in-Aid from the Ministry of Agriculture, Forestry and Fisheries of Japan (Integrated Research Program for the Use of Biotechnological Procedures for Plant Breeding) and by a Grant-in-Aid from the Ministry of Education, Science and Culture of Japan (Cooperative Research, no. 04304017)     

Arthropod communities in a Japanese cedar (Cryptomeria japonica D. Don) plantation: Abundance,biomass and some properties

Arboreal arthropod communities were censused by insecticidal knockdown in a plantation of Japanese cedar,Cryptomeria japonica, in central Japan from April 1983 to February 1985 at intervals of two months. All arthropods sampled were allocated to higher taxonomic groups or guilds and the composition of the communities was analyzed with respect to abundance, biomass and their seasonal trends. The total densities fluctuated seasonally from 200 to 3500 m⁻²; corresponding biomass values ranged from 7 to 600 mg fresh weight m⁻². Maximum density and biomass generally occurred in summer and were minimum in winter. The consistently dominant guild in terms of abundance were the detritivores, mostly comprising Collembola and oribatid mites, accounting for 36–93% of all arthropods; phytophages, predators and detritivores were major guilds in terms of biomass, showing average proportions of 27%, 23% and 20%, respectively. Biomass ratios of predators to prey were generally high in some canopy communities, suggesting the importance of predation pressure in regulating the population levels of arthropods in forests. Average individual size of predators was approximately proportional to that of prey, irrespective of tree locality. Soil arthropod communities maintained densities approximately 10²–10³ times as large as the corresponding canopy communities throughout the year. The seasonal variations in abundance were much greater in canopy than in soil communities.     

Assignment of RFLP linkage groups to their respective chromosomes in aneuploids of sugi (Cryptomeria japonica)

Aneuploids of sugi (Cryptomeria japonica) were found in the open-pollinated progenies of triploidplus tree clones. Seven trisomics and one hypotriploid were used to assign the chromosomes to the RFLP linkage groups constructed previously. The Southern blots containing their genomic DNA were hybridized with the labeled DNA clones corresponding to the loci in the linkage map. The additional dosage in autoradiographs showed that the cloned DNA fragment was located on the extra chromosome in the trisomics. On the other hand, the extra chromosome in two trisomics and the chromosome lacking the triplet in the hypotriploid were cytologically identified as chromosome 10 by consistent presence of a secondary constriction in the proximal region of its short arm. As a result, three linkage groups were assigned to their respective chromosomes, namely chromosome 10 and two other chromosomes.     

A histological comparison of the development of pollen and female gametophytes in fertile and sterile Cryptomeria japonica

To determine a possible mechanism causing male and female sterility in Cryptomeria japonica male and female cones were collected from a C. japonica, tree, ShinDai2, that lacks pollen release and fertile seeds and specimens were processed to examine the development of pollen and female gametophytes using light microscopy and field emission scanning electron microscopy. Pre-meiotic development proceeded normally, but the formation of aberrant meiotic products was observed in cones of both sexes. In sterile microsporangia, heterogeneous microspore populations ranging from monads to polyads gave rise to mature pollen grains of non-uniform size. These pollen grains were covered with an amorphous layer and adhered to each other. In addition, they remained in the microsporangia and were not released even after the onset of pollen dissemination from fertile trees. In the ovules of sterile female cones, megaspores with abnormal shapes, numbers, and sizes formed, and the development of female gametophytes was arrested at the free nuclear or archegonium formation stages. These gametophytes collapsed, and no fertile embryo was generated. Results indicate that meiotic defects are important in the sterility mechanism.     

Density,biomass, and guild structure of arboreal arthropods as related to their inhabited tree size in aCryptomeria japonica plantation

Arboreal arthropods in a 15-year-old plantation ofCryptomeria japonica were surveyed using the smoking method under open conditions and also in the enclosed condition in which the whole above-ground part of a tree was covered by a cloth bag. Per tree, the number of individuals collected was 8200–14000, with a biomass of 340–1700 mg d.wt. Collembola and Acarina were major components in number, while Diplopoda, Collembola, and Araneae occupied the larger part of the biomass. About 60–70% of total numbers of individuals dropped within two hours after the open fumigation. Clear power-form regressions between total numbers of individuals and biomasses of all animals and their host tree size (stem diameter at clear length,D _B ) showed they were approximately proportional toD _B ². The guilds of scavengers and tourists demonstrated the most significant correlations between their numbers and biomasses and the tree size. Numbers and biomasses of Collembola, Diptera, and Araneae revealed remarkable dependence on the tree size. From these regressions, numbers of individuals and biomasses per unit ground area were estimated for all arthropods to be 3755/m² and 165.87 mg d.wt./m², respectively. Power-form regressions were observed between numbers and biomasses of prey and predators. The number-and biomass ratios of all predators to all prey tended to decrease with increasing tree size. A similar trend was observed in the corresponding ratios of parasitic Hymenoptera to lepidopteran larvae, while those of Araneae to Collembola were almost constant, irrespective of tree size.     

Measurement of serum IgE antibodies against Japanese cedar pollen (Cryptomeria japonica) in Japanese monkeys (Macaca fuscata) with pollinosis.

IgE antibodies against allergens of Japanese cedar (Cryptomeria japonica, CJ) pollen in the serum of seven Japanese monkeys (Macaca fuscata) with pollinosis were measured by fluorometric indirect enzyme-linked immunosorbent assay (ELISA). All of the monkeys were found to have specific IgE to the crude pollen antigen. The specific IgE levels were well correlated with those determined by the Pharmacia CAP system. IgE antibodies were then assayed with two kinds of purified allergens (Cry j I and Cry j II) by the ELISA. We found that five monkeys had specific IgE to both allergens, although the other two had IgE only to Cry j I or Cry j II; there is different immune responsiveness to the two major allergens in the monkeys.     

Seasonal changes in abundance and spatial distribution of the soil arthropods in a Japanese cedar (Cryptomeria japonica D.Don) plantation,with special reference to collembola and acarina

Seasonal changes in abundance and distribution pattern of soil micro-arthropods were studied in connection with a few environmental factors in a Japanese cedar (Cryptomeria japonica D. Don) plantation. The soil arthropods were sampled from three different depths at intervals of two months for two years. Of the collected animals (total 51000–155000 m⁻²), Collembola (20000–76000 m⁻²), oribatid mites (19000–55000 m⁻²) and carnivorous mites (6200–21000 m⁻²) were the numerically dominant animal groups. Low seasonal variations in abundance indicated their seasonal stability in population levels. The trends in seasonal fluctuation were similar among these groups and between the two years, showing bimodal pattern with little peaks in early summer and winter. The pattern of seasonal fluctuation in abundance of carnivorous mites (P _d) was significantly synchronized with that in the total abundance of Collembola and oribatid mites (P _τ). Thus, the number-ratios (P _d/P _τ) were fairly constant, ranging from 0.10 to 0.25. Seasonal changes in vertical distribution of the three animal groups showed a similar pattern for both years. The downward migrations were shown to be more affected by low temperatures in winter accompanied by snow coverage rather than by the desiccation of the surface soil in summer. All the three groups demonstrated as a whole slightly aggregated patterns of horizontal distribution throughout the two years. Temporal increases in the patchiness indices were observed from summer to autumn when the moisture content of the surface soil was low.     

Immunocytochemical localization of the allergenic proteins in the pollen of Cryptomeria japonica

Applying an immunocytochemical method, a localization of the protein Cry j I in the Cryptomeria japonica pollen, which is the major allergen responsible for Japanese cedar pollinosis, is investigated with the monoclonal and polyclonal antibodies produced from the protein. The protein that reacts to the polyclonal antibody localizes on the sexine, nexine, between nexine and intine layers, orbicles, cell wall of a generative cell, Golgi body and Golgi vesicles. The allergenic protein contained in the exine and orbicles of Japanese cedar pollen can diffuse or dissolve easily from there into the mucus covering of the eye and nose, causing a response in less than 1 min after exposure. Since the orbicles have a diameter of about 430 nm, they can pass easily through the pores of most protective masks to reach the sensitive tissues of the patient. The proteins react to the monoclonal antibodies (J1BO1 and J1BO7) and localize on the Golgi body, sexine, nexine and orbicles (but not between the nexine and intine layers), and on the generative cell wall. In the young pollen grain, numerous allergenic protein particles contained in the orbicles and sexine layer, but there is only a small amount of the protein between the nexine and intine layers, since the intine layer is not yet complete at this stage. More will be accumulated there during developmental maturation. The allergenic protein is also found on the tapetal materials remaining in the young anther. Since the materials forming the exine layer and orbicles come from tapetal tissue, it is assumed that some of the allergenic protein is produced in the tapetum and localized in the orbicles and pollen wall during maturation, and that the rest of the allergenic protein is produced in the Golgi body in the mature pollen grain.     

Selection and microculture of single embryogenic cell clusters in japanese conifers: Picea jezoensis, Larix leptolepis and Cryptomeria japonica

Summary A microculture method for single embryogenic cell clusters was established for several Japanese conifer species, namely Picea jezoensis, Larix leptolepis and Cryptomeria japonica. Individual small, dense cell clusters were identified and picked up by a micromanipulator and cultured in 50 μl liquid medium in a 96-well culture plate. In all three species studied, a majority of the cell clusters actively proliferated within the wells. Maturation of somatic embryos was successful when the newly proliferated cell clusters were transferred to solid abscisic acid-containing medium. Thus, the small, dense cell clusters could be a useful morphological marker for cells capable of proliferation and differentiation.     

Regeneration of transgenic Cryptomeria japonica D. Don after Agrobacterium tumefaciens-mediated transformation of embryogenic tissue

A genetic transformation procedure for Cryptomeria japonica was developed after co-cultivation of embryogenic tissues with the disarmed Agrobacterium tumefaciens strain C58/pMP90, which harbours the visual reporter gene sgfp and two selectable marker genes, hpt and nptII. We were able to generate eight and three independent transgenic lines per gram of embryogenic tissue after selection on hygromycin and kanamycin medium, respectively. Transgenic plants were regenerated through somatic embryogenesis in 4 lines out of these 11 lines. Green fluorescent protein fluorescence was observed under fluorescent microscopy. Integration of the genes into the genome was confirmed by polymerase chain reaction analysis of embryogenic tissues and Southern blot analysis of regenerated plantlets.     

Estimation of leaf area at the level of branch,tree and stand inCryptomeria japonica

The frequency distribution of diameter (x) in foliage shoot segments, ø(x), was examined in 18 branches at different height levels of three trees in a 25-yr-old sugi (Cryptomeria japonica D. Don) plantation. The ø-x relationships were approximated by power-form equations, in which the exponent differed among the branches from ?0.6 to ?4.2. Leaf area (S _B) and leaf weight (W _B) of a branch were estimated on the basis of the ø-x relationship, and the dependency of specific leaf area (SLA ₀) and density (ρ ₀) of a foliage shoot segment on itsx. SLA _B value of a branch defined byS _B/W _B ranged from 27 to 80 cm² g. d.w.^?1 according to the exponent in the function of ø(x). Total leaf area (u) and leaf weight (wl) of a tree were estimated by summation ofS _B andW _B for seven sample trees. TheSLA _T value of a tree defined byu/wl ranged from 65 to 76 cm² g d.w.^?1 and increased with stem diameter at clear length (D _B). By use of the allometric equations betweenu andD _B,LAI of the plot was estimated to be 17.3 ha ha^?1 (half of the total surface area of needles). By a process similar to that used for calculatingLAI, the amount of woody tissues included in sugi foliage was evaluated to be about 10% of the stand foliage biomass.     

Spatial genetic structure within two populations of a self-incompatible perennial,Chionographis japonica var.japonica (Liliaceae)

Intrapopulational spatial genetic structure was examined in two populations ofChionographis japonica var.japonica, a self-incompatible perennial, by spatial autocorrelation analysis of enzyme polymorphism. Although most spatial autocorrelation indices (Moran'sI) in the shortes distance class were significantly positive, most in the other distance classes did not significantly deviate from the values expected from random distributions of genotypes in both populations. This contrasts with a spatial genetic pattern previously reported for a population of the predominantly selfing congener,C. japonica var.kurohimensis, indicating that pollen-mediated gene flow highly impedes genetic substructuring within populations of outcrossingC. japonica var.japonica. Genetic similarity in very proximate distance found in outcrossingC. japonica var.japonica is probably due to restricted dispersal of seeds.     

Age structure and regeneration of old growthCryptomeria japonica forests on Yakushima Island

The regeneration process ofCryptomeria japonica forests was studied from surveys of age and initial growth of the coniferous stumps in plots of 2.66 ha in total on Yakushima Island, south Japan. The conifers germinated during particular regeneration periods each of which was shorter than 100 years. The periods repeated themselves several times in each plot, and conifers of each period formed patches. Both tree cutting and natural gap formation of the canopies initiated the regenerations. The saplings that were thought to have germinated before the initiation of the regeneration grew slowly. After the initiation, the saplings that germinated earlier grew more quickly. For the three codominant conifers:Cryptomeria japonica, Tsuga sieboldii, andAbies firma, the shade tolerance was inversely correlated with maximum age. Even the most shade-tolerantA. firma was a gap-dependent species in regeneration periods, and there were no species differences in the colonizing times during the regeneration period. Shade-intolerant long-lived species and shade-tolerant short-lived species coexisted as climax species, not as alternate species in the sere of succession.     

Monitoring airborne concentrations of sporangia ofPhytophthora infestans in relation to tomato late blight in Emilia Romagna,Italy

Late blight caused byPhytophthora infestans is one of the most important diseases of tomato crops in Emilia Romagna. In total, 7–8 chemical sprays are applied per season to control the disease without taking into account the risk of infection. An integrated warning service for the prediction of tomato late blight was set up in order to define a more rational disease control strategy. As a part of the warning service, a number of sites monitoringPh. infestans airborne sporangia was set up for a timely disease prediction in the regional tomato growing areas. The results of 3 years monitoring ofPh. infestans sporangia are presented. They suggest that concentrations of airborne sporangia rapidly increase 1–2 weeks prior the blight onset in the field.     

Live crown tops of Japanese cedar (Cryptomeria japonica) approach one another in height at urban forest sites

Since the 1980s, we have found Japanese cedar, Cryptomeria japonica (L. f.) D. Don, trees with crown tops affected by dieback in isolated urban forests on the low altitude plain. To clarify future growth of C. japonica in these forests, we investigated their growth, decline levels and water status. The live crown-top heights from the ground (LCTHs) per diameter at breast height (dbh) were lower in forests with C. japonica top dieback than in forests with no top dieback. Furthermore, in a forest with C. japonica top dieback, the LCTHs were similar between trees although dbh and decline levels were different. Moreover, water status near the top of the live foliage was very similar although decline levels were different, suggesting that in urban forests, where C. japonica top dieback is observed, the LCTH is subject of restriction. A sudden increase in temperatures since the 1980s may be restricting the LCTHs of C. japonica in urban forests. We conclude that LCTHs of C. japonica in urban forests are becoming lower and more equal in each forest. If temperature continues to increase, height restriction will become more severe and LCTHs of C. japonica in urban forests will become even lower.