Rejuvenation of a 100-year-old Sequoiadendron giganteum through in vitro meristem culture. I. Organogenic and morphological arguments

True-to-type cloning of mature trees, especially when they do not sprout from their base, remains problematic. Special attention is focused on the shoot apical meristem, since it is an obvious choice for vegetative propagation. In Sequoiadendron giganteum a meristem removed during budbreak from a 100-year-old tree regenerated a truly rejuvenated line that exhibited the same juvenile characters as the juvenile clone used as control, especially in regard to morphological traits and organogenic capacity, and as manifested by the ability to produce adventitious roots in vitro. This rejuvenation has been maintained for 3 years in both in vitro and ex vitro conditions. This result is discussed in terms of inhibitory correlative systems acting within the donor tree in situ, especially as concerns miniaturization of the explant.     

Notes on new Agaricales of Japan 3

Three new species of Agaricales are described and illustrated from eastern Honshu, Japan:Agrocybe pseudoerebia sp. nov. (sectionVelatae of subgenusAporus), forming fugaceous veil remnants around the pileal margin and relatively shorter basidiospores (less than 10 μm long) without a germ pore, was found on the ground in a broad-leaved forest;Lactarius glutininitens sp. nov. (sectionTriviales of subgenusRussularia), forming a pale grayish, strongly glutinous pileus and watery, latex without discoloration, was found on the ground in a lowland forest dominated byQuercus myrsinaefolia andQuercus serrata; Tricholoma foliicola sp. nov. (close to sectionAlbobrunnea), forming a reddish brown, hygrophanous, dry, glabrous pileus, almost adnate, densely crowded lamellae, small, ellipsoid basidiospores, and clampless hyphae, was found on leaf litter of a broad-leaved forest.     

Relationship between cumulative heat units and fruit-body emergence of ectomycorrhizal fungusTricholoma bakamatsutake in the fields

The relationship between cumulative heat units and fruit-body emergence ofTricholoma bakamatsutake was examined by using the method used for estimating dates of adult emergence of insects. Fruit-body numbers at a study site at Tateyama, Chiba Prefecture and records at a weather station near the site from 1987 to 1993 were used. For the seven years, daily average temperatures above the developmental threshold were cumulated for the days following induction of fruit-body development. The developmental threshold was taken to be 0°C, and the day when the daily minimum temperature fell to near 20°C in each year was taken as the day of induction of fruit-body development. A linear relationship between fruit-body numbers and daily average temperatures was observed as follows: Y=0.00376X + 1.873 (r=0.810), where Y is the probit of percentage of cumulative fruit-bodies, and X is the heat units above 0°C cumulated for the days following induction of fruit-body development. X for Y=5, which represents the cumulative emergence of 50% of fruit-bodies, was estimated to be 832 day-degrees.     

Detection of the ectomycorrhizal fungusTricholoma matsutake and some related species with specific ITS primers

Oligonucleotide primers (Tm1 and Tm4) were designed to amplify a 447–448 base pair fragment, comprising sections of the rDNA internal transcribed spacers (ITS) and the entire 5.8S rDNA, ofTricholoma matsutake. PCR products of predicted size were produced for six of eight isolates ofT. matsutake from across its natural range in Asia, and for isolates of some closely related fungi includingT. bakamatsutake, T. magnivelare, andT. caligatum. The closely relatedT. robustum could be discriminated fromT. matsutake by PCR fragment size. No PCR products were produced where the primers were tested against 16 species of ectomycorrhizal fungi associated withPinus spp. in the Southern Hemisphere. The specific primers were also used successfully to produce PCR products from matsutake infected roots collected in natural forests in China and Japan, and from pure culture synthesisedPinus radiata-T. matsutake material. These primers will be useful in research directed at establishing matsutake in the Southern Hemisphere, and also have the potential to be applied to the study of matsutake within its natural range.     

Cytological Studies of the Genus Nomocharis and Its Related Genera

The somatic cells of 12 taxa of the genera Nomocharis, Lilium, Fritillaria and Cardiocrinum from north-western Yunnan were karyomorphologically studied. The 12 taxa are similar to each other in chromosomal morphology at the resting nucleus stage, which belongs to the complex chromocenter type. They are also similar to each other in morphology of mitotic prophase chromosomes, which belongs to the interstitial type. Their chromosome numbers are all 2n = 24, and centromeric terminalization values range from 79.9- 82.2%. The above results show that the four genera have close relationships. These 12 taxa, however, exhibit some similarities and differences in the position of secondary constrictions seemingly showing the karyological characters of each genus in the area. Firstly, on m(sm)-chromosomes, secondary constrictions close to centromeres appear stable in one of pairs in both Nomocharis( 6 taxa) and Lilium (4 taxa); In Cardiocrinum giganteum, they were found distant from centromeres on short arms in one of pairs; None was observed in Fritillaria cirrhosa. Secondly, on st (t)- chromosomes, remarkable secondary constrictions were generally found on long arms in Nomocharis, C. giganteum, F. cirrhosa and the high mountain habitants of Lilium, such as L. souliei, while they were always observed on short arms in L. nepalense, L. taliense and L. brownii, which grow in the lower altitude region. So the Nomocharis and high mountain members of Lilium have similary distribution of secondary constrictions, and theyare considered to be most closely related.     

大白口蘑菌丝的培养优化

从大白口蘑子实体中分离得到菌种,并进行不同营养条件对大白口蘑菌丝生长的影响实验。结果表明,最佳氮源为酵母粉;最佳碳源为果糖;能明显促进菌丝生长的矿质元素为磷酸二氢钾;最佳生长调节因子为吲哚乙酸;最适温度为28－32℃;最适pH4.0－5.0。     

采收期松茸菌塘土壤理化特性及对细菌多样性的影响

【目的】探讨四川松茸菌塘土壤理化因素对细菌多样性的影响。【方法】应用DGGE(Denaturing gradient gel electrophoresis,DGGE)技术分析和比较四川省小金、雅江、木里、盐源及盐边县松茸主产区的松茸菌塘土壤与对照土壤在松茸采收期的细菌多样性,并结合不同样品土壤理化因子的差异,采用主成分分析(PCA)和多元回归树分析(MRT)方法研究土壤理化因子对细菌多样性的影响。【结果】不同样品土壤理化性质之间差异明显,菌塘土壤的总体肥力水平明显高于非菌塘土壤;不同县域之间土壤微生物多样性指数有着明显差异,且同一地区内的对照土壤微生物多样性指数与菌塘土壤之间并没有相关性;MRT结果表明,当有效铜含量为0.296 7 mg/kg≤Cu0.651 7 mg/kg时,有机质在此限量内(136.5 g/kg≤OM≤257.1 g/kg)含量越大、速效氮在该范围内(151.2 mg/kg≤AN≤277.0 mg/kg)含量越高,松茸菌塘土壤细菌多样性越大;而当Cu≥0.651 7 mg/kg时,p H在该范围内(4.800p H6.028)越小、有效铜在此限量内(0.651 7 mg/kg≤Cu2.040 0 mg/kg)含量越小、交换性钙在该限量内(9.697 cmol/kg≤Ca23.300 cmol/kg)含量越大、速效钾在该限量内(243.0 mg/kg≤AK296.7 mg/kg)含量越大,松茸菌塘土壤细菌多样性越大。【结论】有效铜、有机质、p H、速效氮、交换性钙及速效钾6个因子对菌塘土壤细菌多样性影响最大。     

Purification,Characterization, and Molecular Cloning of a Pyranose Oxidase from the Fruit Body of the Basidiomycete,Tricholoma matsutake

A new H₂O₂-generating pyranose oxidase was purified as a strong antifungal protein from an arbuscular mycorrhizal fungus, Tricholoma matsutake. The protein showed a molecular mass of 250 kDa in gel filtration, and probably consisted of four identical 62 kDa subunits. The protein contained flavin moiety and it oxidized D-glucose at position C-2. H₂O₂ and D-glucosone produced by the pyranose oxidase reaction showed antifungal activity, suggesting these compounds were the molecular basis of the antifungal property. The V _max, K _m, and k _cat for D-glucose were calculated to be 26.6 U/mg protein, 1.28 mM, and 111/s, respectively. The enzyme was optimally active at pH 7.5 to 8.0 and at 50°C. The preferred substrate was D-glucose, but 1,5-anhydro-D-glucitol, L-sorbose, and D-xylose were also oxidized at a moderate level. The cDNA encodes a protein consisting of 564 amino acids, showing 35.1% identity to Coriolus versicolor pyranose oxidase. The recombinant protein was used for raising the antibody.     

蒙古口蘑分类地位研究

利用LSU序列分析技术研究蒙古口蘑分类地位,用最大简约法和最大似然法构建系统发育树。结果表明：蒙古口蘑形成一独立分支,与白蘑属Trciholoma亲缘关系较远,蒙古口蘑应从Trciholoma属中划分出来,放入新属———白丽蘑属（新拟）Leucocalocybe,并命名为蒙古白丽蘑（新拟）Leucocalocybe mongolicum（Imai）X.D.Yu＆Y.J.Yao。     

Characterization of the carbohydrase productions of an ectomycorrhizal fungus, Tricholoma matsutake

To evaluate the potential of the production of the ectomycorrhizal fungus Tricholoma matsutake to produce carbohydrases, (1) the distribution of carbohydrase activities among the different strains (18 strains) was investigated and (2) the abilities of T. matsutake and saprophytic fungi to produce β-glucosidase were compared. The results showed that the carbohydrase productions patterns of T. matsutake still resemble one another. Moreover, this fungus exhibited markedly higher β-glucosidase than did the saprophytic mushrooms. Tricholoma matsutake showed weak production of α-amylase and α-glucosidase in a static cultur filtrate. On the other hand, glucoamylase activity was not observed. Surprisingly, we discovered that β-glucosidase demonstrated strong activity. This finding suggests that this fungus has saprotrophic abilities. The carbohydrase production systems in T. matsutake were characterized from our experimental results. Also, we point out some weak points in the carbohydrase production systems of T. matsutake.