A petunia mutant affected in intracellular accommodation and morphogenesis of arbuscular mycorrhizal fungi

The regulation of the arbuscular mycorrhizal (AM) symbiosis is largely under the control of a genetic programme of the plant host. This programme includes a common symbiosis signalling pathway that is shared with the root nodule symbiosis. Whereas this common pathway has been investigated in detail, little is known about the mycorrhiza-specific regulatory steps upstream and downstream of the common pathway. To get further insight in the regulation of the AM symbiosis, a transposon-mutagenized population of Petunia hybrida was screened for mutants with defects in AM development. Here, we describe a petunia mutant, penetration and arbuscule morphogenesis1 (pam1), which is characterized by a strong decrease in colonization by three different AM fungi. Penetrating hyphae are frequently aborted in epidermal cells. Occasionally the fungus can progress to the cortex, but fails to develop arbuscules. The resulting hyphal colonization of the cortex in mutant plants does not support symbiotic acquisition of phosphate and copper by the plant. Expression analysis of three petunia orthologues of the common SYM genes LjPOLLUX, LjSYMRK and MtDMI3 indicates that pam1 is not mutated in these genes. We conclude that the PAM1 gene may play a specific role in intracellular accommodation and morphogenesis of the fungal endosymbiont.     

Molecular identification and phylogeny of arbuscular mycorrhizal fungi

The fossil record and molecular data show that the evolutionary history of arbuscular mycorrhizal fungi (Glomales) goes back at least to the Ordovician (460 million years ago), coinciding with the colonization of the terrestrial environment by the first land plants. At that time, the land flora only consisted of plants on the bryophytic level. Ribosomal DNA sequences indicate that the diversity within the Glomales on the family and genus level is much higher than previously expected from morphology-based taxonomy. Two deeply divergent lineages were found and described in two new genera, Archaeospora and Paraglomus, each in its own family. Based on a fast-growing number of available DNA sequences, several systems for molecular identification of the Glomales within roots have been designed and tested in the past few years. These detection methods have opened up entirely new perspectives for studying the ecology of arbuscular mycorrhiza.     

Communities, populations and individuals of arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi in the phylum Glomeromycota are found globally in most vegetation types, where they form a mutualistic symbiosis with plant roots. Despite their wide distribution, only relatively few species are described. The taxonomy is based on morphological characters of the asexual resting spores, but molecular approaches to community ecology have revealed a considerable unknown diversity from colonized roots. Although the lack of genetic recombination is not unique in the fungal kingdom, arbuscular mycorrhizal fungi are probably ancient asexuals. The long asexual evolution of the fungi has resulted in considerable genetic diversity within morphologically recognizable species, and challenges our concepts of individuals and populations. This review critically examines the concepts of species, communities, populations and individuals of arbuscular mycorrhizal fungi.     

一种改进的丛枝菌根染色方法

研究改进了Vierheilig等描述的AM菌根染色法:将根样于20%KOH溶液中60℃水浴透明40-120 min,5%醋酸酸化5min后,用5%醋酸墨水染色液(派克纯黑书写墨水Quink),于60℃水浴染色30 min,清水浸泡脱色(14h)后即可镜检。根皮层细胞内AM真菌的丛枝结构清晰可见,并且能够明确地分辨AM真菌与其它未知真菌。此外,Quink初染后,再经过SudanⅣ复染(60℃、60 min),70%乙醇脱色5min,暗隔真菌的透明菌丝内所积聚的脂类颗粒被SudanⅣ染上鲜红色,在复式显微镜下能够观察到此类透明菌丝在根皮层组织内的存在状况。采用甘油明胶为封固剂制片,根的染色效果可以保存长久。此项技术可以对同一种植物的多个根样进行同步的透明和染色处理,而且操作简便、低毒性、成本低廉、染色效果极佳,适用于野生和栽培草本植物AM菌根的染色和制片观察。     

Metabolite profiling of mycorrhizal roots of Medicago truncatula

Metabolite profiling of soluble primary and secondary metabolites, as well as cell wall-bound phenolic compounds from roots of barrel medic (Medicago truncatula) was carried out by GC-MS, HPLC and LC-MS. These analyses revealed a number of metabolic characteristics over 56 days of symbiotic interaction with the arbuscular mycorrhizal (AM) fungus Glomus intraradices, when compared to the controls, i.e. nonmycorrhizal roots supplied with low and high amounts of phosphate. During the most active stages of overall root mycorrhization, elevated levels of certain amino acids (Glu, Asp, Asn) were observed accompanied by increases in amounts of some fatty acids (palmitic and oleic acids), indicating a mycorrhiza-specific activation of plastidial metabolism. In addition, some accumulating fungus-specific fatty acids (palmitvaccenic and vaccenic acids) were assigned that may be used as markers of fungal root colonization. Stimulation of the biosynthesis of some constitutive isoflavonoids (daidzein, ononin and malonylononin) occurred, however, only at late stages of root mycorrhization. Increase of the levels of saponins correlated AM-independently with plant growth. Only in AM roots was the accumulation of apocarotenoids (cyclohexenone and mycorradicin derivatives) observed. The structures of the unknown cyclohexenone derivatives were identified by spectroscopic methods as glucosides of blumenol C and 13-hydroxyblumenol C and their corresponding malonyl conjugates. During mycorrhization, the levels of typical cell wall-bound phenolics (e.g. 4-hydroxybenzaldehyde, vanillin, ferulic acid) did not change; however, high amounts of cell wall-bound tyrosol were exclusively detected in AM roots. Principal component analyses of nonpolar primary and secondary metabolites clearly separated AM roots from those of the controls, which was confirmed by an hierarchical cluster analysis. Circular networks of primary nonpolar metabolites showed stronger and more frequent correlations between metabolites in the mycorrhizal roots. The same trend, but to a lesser extent, was observed in nonmycorrhizal roots supplied with high amounts of phosphate. These results indicate a tighter control of primary metabolism in AM roots compared to control plants. Network correlation analyses revealed distinct clusters of amino acids and sugars/aliphatic acids with strong metabolic correlations among one another in all plants analyzed; however, mycorrhizal symbiosis reduced the cluster separation and enlarged the sugar cluster size. The amino acid clusters represent groups of metabolites with strong correlations among one another (cliques) that are differently composed in mycorrhizal and nonmycorrhizal roots. In conclusion, the present work shows for the first time that there are clear differences in development- and symbiosis-dependent primary and secondary metabolism of M. truncatula roots.     

丛枝菌根真菌与植物硫素营养研究进展

丛枝菌根真菌是土壤微生物群落的重要组成部分,是最常见的地下共生菌,对植物和土壤具有多种有益作用.本文阐述了近年来丛枝菌根真菌对植物吸收土壤硫素的最新进展,在目前耕地缺硫状况下,着重分析了丛枝菌根真菌改善植物硫素营养以及丛枝菌根真菌利用硫素的分子调控机制,总结了影响菌根硫代谢的因素,并指出该研究方向仍存在的一些问题以及未...     

我国北方农田土壤中AM真菌的多样性

AM真菌是农业生态系中一类重要的土壤微生物,它在农田土壤中的发生和分布受多种环境因素的影响。为深入了解我国北方农田土壤中AM真菌的多样性规律,于2000年在河北、山东的农田土壤中采集有代表性的土样127个。通过进一步扩繁、纯化,从中分离出AM真菌5属22种,鉴定了20个种,包括一个国内新记录种沾屑球囊霉(Glomus spurcum)。分析AM真菌的多样性特点及其影响因素发现,农田土壤中以球囊霉属(Glomus)的频度最高,其次为无梗囊霉属(Acaulospora);优势种类为幼套球囊霉(Glomus etunicatum)和摩西球囊霉(Glomus mosseae).土壤速效磷含量、pH状况主要对孢子密度产生影响,对种群分布影响不大。宿主植物类型对AM真菌的侵染状况和多样性影响较大;比较玉米(Zea mays)、甘薯(Ipomoea batatas)根区AM真菌的种群组成后发现两者有所不同,但优势种一致.     

Phosphate uptake, transport and transfer by the arbuscular mycorrhizal fungus Glomus intraradices is stimulated by increased carbohydrate availability

* The influence of carbohydrate availability to mycorrhizal roots on uptake, metabolism and translocation of phosphate (P) by the fungus was examined in axenic cultures of transformed carrot (Daucus carota) roots in symbiosis with Glomus intraradices. * 14C-labelled carbohydrates, 33P-phosphate and energy dispersive X-ray microanalysis were used to follow the uptake and transfer of C and P in the arbuscular mycorrhizal (AM) symbiosis. * The uptake of P by the extraradical mycelium (ERM) and its translocation to the mycorrhizal roots was stimulated and the metabolic and spatial distribution of P within the fungus were altered in response to increased carbohydrate availability. Sucrose supply resulted in a decrease of polyphosphates and an increased incorporation into phospholipids and other growth-related P pools and also caused elevated cytoplasmic P levels in the intraradical mycelium (IRM) within the root and higher cytoplasmic P levels in the root cortex. * These findings indicate that the uptake of P by the fungus and its transfer to the host is also stimulated by the transfer of carbon from plant to fungus across the mycorrhizal interface.     

Variable effects of arbuscular mycorrhizal fungal inoculation on physiological and molecular measures of root and stomatal conductance of diverse Medicago truncatula accessions

Association with arbuscular mycorrhizal fungi (AMF) can impact on plant water relations; mycorrhizal plants can exhibit increased stomatal conductance (g_s) and root hydraulic conductance (normalized to root dry weight, L_o), and altered expression of aquaporins (AQP). Many factors regulate such responses; however, plant intraspecific diversity effects have yet to be explored. Twenty geographically diverse accessions of Medicago truncatula were inoculated with the AMF Funneliformis mosseae or mock‐inoculated, and grown under well‐watered conditions. Biomass, g_s, shoot nutrient concentrations and mycorrhizal colonization were measured in all accessions, and L_o and gene expression in five accessions. The diverse accessions varied in physiology and gene expression; some accessions were also larger or had higher g_s when colonized by F. mosseae. In the five accessions, L_o was higher in two accessions when colonized by AMF and also maintained within a much smaller range than the mock‐inoculated plants. Expression of MtPIP1 correlated with both g_s and L_o, and when plants were more than 3% colonized, mycorrhizal colonization correlated with L_o. Accession and AMF treatments had profound effects on M. truncatula, including several measures of plant water relations. Correlations between response variables, especially between molecular and physiological variables, across genotypes, highlight the findings of this study.     

高磷土壤中丛枝菌根真菌的分离鉴定

【背景】丛枝菌根(arbuscular mycorrhiza,AM)真菌具有广泛的寄主范围、环境适应性和优良的植物促生能力。然而,土壤的高磷水平严重抑制了AM真菌生长及AM形成。【目的】分离鉴定出耐较高有效磷含量的华南土著AM真菌菌株,为菌根学研究工作提供新颖材料。【方法】采用经典形态学和分子系统学方法鉴定高磷土壤中AM真菌。【结果】从有效磷含量为53-131 (平均值±标准差为88.2±17.6) mg/kg的根区土壤中鉴定出7属25种AM真菌,包括无梗囊霉属(Acaulospora) 12种、球囊霉属(Glomus) 7种、隔球囊霉属(Septoglomus) 2种、近明球囊霉属(Claroideoglomus) 1种、根孢囊霉属(Rhizophagus) 1种、硬囊霉属(Sclerocystis) 1种和类球囊霉属(Paraglomus) 1种,其中幼套近明球囊霉(Claroideoglomus etunicatum)和蜜色无梗囊霉(Acaulospora mellea)是优势种。在(87.7±8.0) mg/kg的高磷水平下,AM真菌仍能形成丛枝和泡囊。但当有效磷含量达到(99.7±1.2) mg/kg时,菌根侵染率和丛枝丰度显著下降,但仍能够形成泡囊。【结论】从广州市南沙区有效磷含量为(88.2±17.6) mg/kg的耕地植物根区土壤中,鉴定出具有耐高磷潜力的7属25种AM真菌,幼套近明球囊霉和蜜色无梗囊霉等分离株可作为后续高磷抑制机制解析及耐高磷AM真菌菌剂研发工作的试验菌株。     

Improvement of Cupressus atlantica Gaussen growth by inoculation with native arbuscular mycorrhizal fungi

AIMS: The study aimed to determine whether inoculation with native arbuscular mycorrhizal (AM) fungi could improve survival and growth of seedlings in degraded soils of Morocco. METHODS AND RESULTS: Soil samples were collected from the rhizosphere of Cupressus atlantica trees in the N'Fis valley (Haut Atlas, Morocco). AM spores were extracted from the soil, identified and this mixture of native AM fungi was propagated on maize for 12 weeks on a sterilized soil to enrich the fungal inoculum. Then C. atlantica seedlings were inoculated with and without (control) mycorrhizal maize roots, cultured in glasshouse conditions and further, transplanted into the field. The experiment was a randomized block design with one factor and three replication blocks. The results showed that a high AM fungal diversity was associated with C. atlantica; native AM fungi inoculation was very effective on the growth of C. atlantica seedlings in glasshouse conditions and this plant growth stimulation was maintained for 1 year after outplanting. CONCLUSIONS: Inoculation of C. atlantica with AM fungi increased growth and survival in greenhouse and field. SIGNIFICANCE AND IMPACT OF THE STUDY: The data indicate that use of native species of AM fungi may accelerate reforestation of degraded soils. Further studies have to be performed to determine the persistence of these mycorrhizae for a longer period of plantation and to measure the effects of this microbial inoculation on soil biofunctioning.     

Rapid and sensitive bioassay to study signals between root exudates and arbuscular mycorrhizal fungi**

A sensitive bioassay was developed to provide a way to detect chemical signals from host plants which induce changes in hyphal growth patterns of germinated spores of arbuscular mycorrhizal (AM) fungi. The assay can be used to test host root exudates, as well as particulate fractions (root cap border cells and root mucilage), for their ability to affect AM fungal growth. Hyphal branching, induced by various host root components, can be detected as early as 4 h although results of the bioassay were usually determined after 16 to 24 h. The type of branching pattern observed was dose-dependent.     

Transport of radiocaesium by arbuscular mycorrhizal fungi to Medicago truncatula under in vitro conditions

The capacity of arbuscular mycorrhizal (AM) fungi to take up and translocate radiocaesium (Cs) to their host has been shown using the root-organ culture (ROC) system. However, the absence of photosynthetic tissues, lack of a normal root hormonal balance and incomplete source-sink relationships may bias the bidirectional transfer of elements at the symbiotic interface and complicate transport studies. Accordingly, we developed a novel culture system [i.e. the Arbuscular Mycorrhizal-Plant (AM-P) in vitro culture system], where AM fungi and an autotrophic host plant develop under strict in vitro conditions. With this system, we unambiguously demonstrated the capacity of AM fungi to transport Cs. The extraradical fungal hyphae took up 21.0% of the initial supply of 134Cs. Translocation to the plant represented 83.6% of the 134Cs taken up. Distribution of 134Cs in the host plant was 89.8% in the mycorrhizal roots and 10.2% in the shoot. These results confirm that AM fungi can take up, translocate and accumulate Cs. They further demonstrate unambiguously and for the first time that Cs can be transferred from AM fungi to host tissues. These results suggest a potential involvement of AM fungi in Cs biogeochemical cycle and in plant Cs accumulation.     

低温胁迫下丛枝菌根真菌对玉米光合特性的影响

利用盆栽试验,在15 ℃和5 ℃低温胁迫下研究了丛枝菌根（AM）真菌对玉米生长、叶绿素含量、叶绿素荧光和光合作用的影响.结果表明：低温胁迫抑制了AM真菌的侵染;接种AM真菌的玉米地上部和地下部干物质量、相对叶绿素含量高于不接种植株.与非菌根玉米相比,菌根玉米具有较高的最大荧光（F_m）、可变荧光（F_v）、最大光化学效率（F_v/F_m）和潜在光化学效率（F_v/F_o）及较低的初始荧光（F_o）,并且在5 ℃处理中差异显著.接种AM真菌使玉米叶片的净光合速率（P_n）和蒸腾速率（T_r）显著增强;低温胁迫下,菌根植株的气孔导度（G_s）显著高于非菌根植株;而胞间CO₂浓度（C_i）显著低于非菌根植株.表明AM真菌可通过提高叶绿素含量及改善叶片叶绿素荧光和光合作用来减轻低温胁迫对玉米植株造成的伤害,提高玉米耐受低温的能力,进而提高玉米的生物量,促进玉米生长.