Ectomycorrhiza formation ofTricholoma matsutake isolates on seedlings ofPinus densiflora in vitro

Tricholoma matsutake forms ectomycorrhizas withPinus densiflora under field conditions. The present study aimed to test the ability ofT. matsutake isolates to form mycorrhizas with aseptic seedlings ofP. densiflora in vitro. Pine seeds were germinated aseptically on a nutrient agar medium, and pairs of 1-wk-old seedlings were transplanted into polymethylpentene bottles containing autoclaved sphagnum moss/vermiculite substrate. The substrate was saturated with nutrient medium containing glucose. At the same time, the bottles were inoculated with aT. matsutake isolate. Three mo after inoculation, the fungus formed a sheath and Hartig net on the pine lateral roots. Ectomycorrhizas were also confirmed on 4-6-mo-old seedlings which showed the same or slinghtly better growth than the control plants. These results indicate that culturedT. matsutake mycelium can form true ectomycorrhizas withP. densiflora seedlings in vitro.     

赤松不定根和愈伤组织与松口蘑外生菌根菌的相互作用

本实验报道了以简单的离体培养方式来诱导赤松不定根和愈伤组织与松口蘑的菌根反应。不定根和愈伤组织均起源于无菌苗的下胚轴,接种2周后,菌丝体开始包围不定根。接种3周后,菌丝体出现在不定根皮层细胞间,哈蒂氏网型的形成也同时被确认。在愈伤组织培养物中,细胞间也能观察到菌丝体及拟-哈蒂氏网结构。这是第一个离体条件下成功地诱导赤松培养组织与松口蘑形成外生菌根的报道。     

土生空球菌及其在火炬松上形成的菌根

土生空球菌(cenococcum geophilum)是一种寄主范围最广、研究最多的外生菌根真菌。本文首次报道了该菌在中国的发现及该菌与火炬松所形成的菌根,全面描述了该菌根的形态学和组织学特征,并对该菌进行了分离培养,描述该菌在纯培养下的特征,对该菌在火炬松上形成的菌根的特征进行了比较,同时讨论了该菌在造林中的重要性。     

Comparative studies of ectomycorrhiza formation in Alnus glutinosa and Pinus resinosa with Paxillus involutus

 Mycorrhiza ontogeny and details of Hartig net and mantle structure were compared in ectomycorrhizas synthesized in growth pouches between the broad host range fungus Paxillus involutus and the tree species European black alder (Alnus glutinosa) and red pine (Pinus resinosa). In Alnus glutinosa, a paraepidermal Hartig net was restricted to the proximal (basal) portion of first-order laterals; the hypodermal layer appeared to be a barrier to fungal penetration. Phi-thickenings were present in some cortical cells but these were not related to lack of fungal ingress into the cortex. The mantle was often present close to the root apex but in many roots it was loosely organized and patchy. In several instances, the mantle formed around the root apex was only temporary; renewed root growth occurred without the formation of a mantle. In Pinus resinosa, the Hartig net developed between cortical cell layers of monopodial and dichotomously branched first–order laterals. Fungal hyphae in the Hartig net exhibited a complex labyrinthine mode of growth. The mantle had a pseudoparenchymatous structure and covered the root, including apices of dichotomously branched roots. The Paxillus–Pinus resinosa interaction had all the characteristics of a compatible ectomycorrhizal association. The Paxillus–Alnus glutinosa interaction, however, showed only aspects of superficial ectomycorrhizas, including the presence of a minimal (sometimes absent) and mostly proximal Hartig net and variable mantle development. Sclerotia were produced in the extraradical mycelium of Paxillus involutus when associated with either Alnus glutinosa or Pinus resinosa. Accepted: 22 October 1998     

In situ and in vitro colonization of Cathaya argyrophylla (Pinaceae) by ectomycorrhizal fungi

Cathaya argyrophylla, a critically endangered conifer, is found to grow at four isolated areas located in subtropical mountains of China. To examine the involvement and usefulness of mycorrhizas for sustaining the population of this tree, we compared the root system, morphology, and structure of mycorrhizal roots of C. argyrophylla, which were collected from a natural stand and an artificial stand, each grown at a different location. More mycorrhizal roots were found for trees from an artificial stand. The presence of extramatrical mycelium, mantle, and Hartig net revealed that C. argyrophylla formed an ectomycorrhizal association in both sampling sites. Starch granules were found in mycorrhizal roots collected only from a natural stand. The aseptic synthesis of C. argyrophylla and Cenococcum geophilum was established for the first time in vitro. Typical ectomycorrhizas formed on seedlings on RM medium containing 0.1 g/l glucose, 5 weeks after inoculation. By light microscopy, the synthesized mycorrhizas showed a thin mantle from which emanated extramatrical hyphae and highly branched Hartig net. A simple, rapid, and convenient mycorrhiza synthesis system was developed, which facilitates further studies on ectomycorrhizal development of C. argyrophylla.     

Fluctuation in free and conjugated polyamines in Scots pine seedlings after changes in temperature and daylength

Free, soluble and insoluble conjugated polyamines from the needles, roots and stem of five month old Scots pine (Pinus sylvestris L.) seedlings inoculated with Suillus variegatus (Fr.) O. Kuntze and seedlings without inoculation were analysed during decrease in daylength and temperature. Temporary changes in free, soluble and insoluble conjugated polyamine pools caused by a decrease in daylength or temperature were observed. Inoculation of pine seedlings affected significantly the polyamine levels of five month old pine seedlings. The roots of inoculated seedlings contained significantly higher levels of free and soluble conjugated purtrescine and free, soluble conjugated and insoluble conjugated spermidine than the roots of noninoculated seedlings. The needles of inoculated seedlings contained significatly higher concentrations of free putrescine and soluble conjugated spermidine but lower amount of free spermine than the needles of noninoculated seedlings. The stems of inoculated seedlings contained higher concentrations of free putrescine but lower amounts of insoluble conjugated spermine. Changes in polyamine levels in noninoculated seedlings were observed after shortening of the daylength, whereas in inoculated ones changes were induced mainly by the decrease in temperature. The possible role of polyamines in the initial stage of cold hardening process is discussed.     

濒危植物紫斑牡丹胚离体培养

以MS为基本培养基 ,添加不同浓度的BA、NAA及其组合 ,对濒危植物紫斑牡丹的胚进行体外培养。实验结果显示在没有添加任何激素的培养基上 ,离体胚生长类似种子萌发时胚的生长模式 ,子叶扩大伸长 ,达到约种子大小时停顿 ,根伸长生长极其显著 ;BA促进子叶膨大生长 ,当BA浓度增大时根生长受到抑制 ;NAA促进愈伤组织形成 ,抑制根生长。 0 .1mg·L- 1BA促进紫斑牡丹胚的胚轴、根生长 ,有利于幼苗形成。     

Conservation of rare and endangered plants using in vitro methods

Summary Many botanic gardens now have tissue culture laboratories for the micropropagation of plants that are difficult to propagate by conventional horticultural techniques. In many cases the work centers on rare and endangered species. Examples of the use of different techniques including micropropagation, in vitro seed germination, dual culture with symbiotic fungi, and regeneration from callus are discussed with reference to their application to plant germplasm conservation. Presented in the Session-in-Depth Cell Culture of Endangered Species at the 1991 World Congress on Cell and Tissue Culture, Anaheim, California, June 16–20, 1991.     

Biology of the ectomycorrhizal fungal genus, Rhizopogon

The morphology and anatomy of ectomycorrhizas of Rhizopogon arctostaphyli , R. ellenae , R. flavofibrillosus , R. occidentalis , R. rubescens , R. smithii , R. subcaerulescens and R. truncatus synthesized on Ponderosa pine ( Pinus ponderosa ) in glasshouse conditions using spore slurries, are described and compared. All species produced a well-developed Hartig net, and a well-developed fungal mantle. The mantles of R. arctostaphyli , R. smithii and R. subcaerulescens ectomycorrhizas were two-layered with outer mantle hyphae of wider diameter than inner mantle hyphae. The mantle of R. subcaerulescens ectomycorrhizas also had distinctive peg-like structures (cystidia) along peripheral hyphae. Rhizopogon truncatus ectomycorrhizas were tuberculate in morphology and had a rind-like mantle enclosing adjacent roots. In addition, several species exhibited crystal inclusions in the outer mantle, presumably at the interface between mantle and soil.     

Activity of different ectomycorrhizal types studied by vital fluorescence

Ectomycorrhizal root tips were sampled in a Norway spruce (Picea abies L. Karst) stand (Höglwald, Bavaria) during one growing season and analysed by fluorescence microscopy. After staining with fluorescein-diacetate, tissue fluorescence was used as an estimate of activity of six identified ectomycorrhizal types on Picea abies (Tylospora sp.-Picea abies, Piceirhiza nigra, Piceirhiza gelatinosa, Xerocomus badius-P. abies, Russula ochroleuca-P. abies, Cenococcum geophilum) and two unidentified mycorrhizas. There were differences among ectomycorrhizal types in the FDA-hydrolysing activity of the various tissue layers of the mycorrhizas: outer and inner hyphal sheath, Hartig net and stele. The differential activity in tissues was judged to be an estimate of physiological activity of the fungal symbiont and of the life span of the mycorrhiza. The differences in species activity may influence the overall activity of mycorrhizas in a forest stand.     

O-coumaric acid ester,a potential early signaling molecule in Pinus pinea and Pisolithus arhizus symbiosis established in vitro

During ectomycorrhizal (ECM) establishment, biochemical signals lead to the development of complex structures in both the plant and the fungus that ultimately result in the formation of an ectomycorrhiza. The cross-talk between partners begins before physical contact. Our objective was to investigate the chemical nature of the signals during the first stages of in vitro mycorrhization of Pinus pinea with Pisolithus arhizus. For this purpose a double-phase solid–liquid medium was expressly developed for the co-culture in order to simplify the extraction and further molecules analysis. O-coumaric acid ester was identified using HPLC–UV and LC–DAD–MS on the second day of co-culture and its presence was detected for up to 10 days. These results contribute to the characterization of biochemical signals during pre-colonization involving conifer species and an ECM fungus, and demonstrate the suitability of the double-phase medium developed for the growth of both organisms and for the analysis of released chemical mediators.