丛枝菌根真菌和根瘤菌对白三叶氮同化的影响

为揭示丛枝菌根真菌(AMF)和根瘤菌在白三叶氮(N)同化中的作用,该研究对白三叶进行单一或联合接种隐类球囊霉(Paraglomus occultum)和三叶草根瘤菌(Rhizobium trifolii),分析其对白三叶的生长、光合作用、叶片N和氨基酸含量以及N同化相关酶活性的影响。结果表明:(1)单一接种AMF或根瘤菌以及联合接种AMF和根瘤菌均显著增加了白三叶的株高、匍匐茎长度、叶片数、地上部生物量、总生物量、叶绿素b和总叶绿素含量、稳态光量子效率和叶片N含量,这种增强效应是联合接种>单一AMF>单一根瘤菌>未接种处理。(2)联合接种AMF和根瘤菌显著增加了白三叶叶片中丙氨酸、精氨酸、天冬酰胺、天冬氨酸、谷氨酰胺、谷氨酸和组氨酸的含量,显著提升了叶片N同化相关酶如硝酸还原酶、亚硝酸还原酶、谷氨酰胺合成酶、谷氨酸合成酶、谷氨酸脱氢酶、天冬酰胺合成酶和天冬氨酸转氨酶的活性,显著促进AMF对白三叶根系的侵染。综上认为,联合接种AMF和根瘤菌通过激活N同化相关酶活性有效促进N同化,产生更多氨基酸,进一步促进白三叶植株生长; 联合接种AMF和根瘤菌具有协同作用,有效促进了白三叶的N同化。     

丛枝菌根真菌群落对白三叶草生长的影响

不同施肥处理影响AMF(Arbuscular mycorrhizal fungi)群体结构,然而不同AMF群体结构对植物的生长以及养分吸收的影响尚未见报道,试验利用盆栽实验研究了7种不同来源的丛枝菌根真菌(AMF)群落对白三叶草生长和N、P、K以及微量元素Cu、Zn、Mn的吸收的影响。7种AMF群落分离自长期定位施肥试验地,分别为NPK、OM、CK、1/2OM、NP、NK和PK。每年施肥量是300kg N/hm2,135kg P2O5/hm2,300kg K2O/hm2。有机肥处理的N、P、K养分量与试验地NPK处理含量相同,原料以粉碎的麦秆为主,加上适量的大豆饼和棉仁饼,有机肥经堆制发酵后施用。试验土壤采用封丘试验地土壤,经灭菌处理。试验结果表明,接种不同AMF群落均能促进三叶草的生长,对养分吸收则表现不同。分离自CK试验地的AMF群落对三叶草侵染率显著低于其它6种AMF群落。分离自1/2OM和OM试验地的AMF群落较分离自NPK、CK、NP和NK的AMF群落显著促进了三叶草对P的吸收;各种AMF群落都促进了对N和K的吸收;分离自OM、CK、1/2OM、NP、NK试验地的降低了三叶草植株N含量;分离自NPK试验地的AMF群落提高了三叶草植物K含量;对于Cu、Zn、Mn元素的吸收,不同处理存在较大的差异。AMF群落对三叶草生长以及养分吸收贡献不同,这与不同施肥管理下不同AMF群落的优势种属的侵染率、养分转化以及菌丝发育及分布有关。     

Taxon-specific PCR primers to detect two inconspicuous arbuscular mycorrhizal fungi from temperate agricultural grassland

Taxon-specific polymerase chain reaction (PCR) primers enable detection of arbuscular mycorrhizal fungi (AMF, Glomeromycota) in plant roots where the fungi lack discriminative morphological and biochemical characters. We designed and validated pairs of new PCR primers targeted to the flanking regions of the variable domain 1 of the nuclear ribosomal large subunit RNA gene to specifically detect Acaulospora paulinae and an undescribed member of the Diversisporaceae. These two fungal taxa, sporulating late in soil-trap cultures and showing small, faintly coloured spores and weakly staining intraradical structures, were frequently found in roots of Trifolium repens from a high-input agricultural grassland. The newly developed PCR primers may thus enable studies on two inconspicuous AMF taxa that appear to have been overlooked in previous molecular AMF community analyses and for which no specific PCR primers have been published.     

Xyloglucanases in the interaction between saprobe fungi and the arbuscular mycorrhizal fungus Glomus mosseae

We studied the production of xyloglucanase enzymes of pea and lettuce roots in the presence of saprobe and arbuscular mycorrhizal (AM) fungi. The AM fungus Glomus mosseae and the saprobe fungi Fusarium graminearum, Fusarium oxysporum-126, Trichoderma harzianum, Penicillium chrysogenum, Pleurotus ostreatus and Aspergillus niger were used. G. mosseae increased the shoot and root dry weight of pea but not of lettuce. Most of the saprobe fungi increased the level of mycorrhization of pea and lettuce, but only P. chrysogenum and T. harzianum inoculated together with G. mosseae increased the dry weight of pea and lettuce respectively. The AM and saprobe fungi increased the production of xyloglucanases by plant roots. The level of xyloglucanase activities and the number of xyloglucanolytic isozymes in plants inoculated with G. mosseae and most of the saprobe fungi tested were higher than when both microorganisms were inoculated separately. The possible relationship between xylogucanase activities and the ability of AM and saprobe fungi to improve the dry weight and AM root colonization of plants was discussed.     

Increased photosynthetic acclimation in alfalfa associated with arbuscular mycorrhizal fungi (AMF) and cultivated in greenhouse under elevated CO2

Medicago sativa L. (alfalfa) can exhibit photosynthetic down-regulation when grown in greenhouse conditions under elevated atmospheric CO₂. This forage legume can establish a double symbiosis with nitrogen fixing bacteria and arbuscular mycorrhizal fungi (AMF), which may increase the carbon sink effect of roots. Our aim was to assess whether the association of alfalfa with AMF can avoid, diminish or delay the photosynthetic acclimation observed in previous studies performed with nodulated plants. The results, however, showed that mycorrhizal (M) alfalfa at the end of their vegetative period had lower carbon (C) discrimination than non-mycorrhizal (NM) controls, indicating photosynthetic acclimation under ECO₂ in plants associated with AMF. Decreased C discrimination was due to the acclimation of conductance, since the amount of Rubisco and the expression of genes codifying both large and small subunits of Rubisco were similar or slightly higher in M than in NM plants. Moreover, M alfalfa accumulated a greater amount of soluble sugars in leaves than NM plants, thus favoring a down-regulation effect on photosynthetic rates. The enhanced contents of sugars in leaves coincided with a reduced percentage of arbuscules in roots, suggesting decreased sink of carbohydrates from shoots to roots in M plants. The shorter life cycle of alfalfa associated with AMF in comparison with the NM controls may also be related to the accelerated photosynthetic acclimation in M plants. Further research is needed to clarify to what extent this behavior could be extrapolated to alfalfa cultivated in the field and subjected to periodic cutting of shoots under climatic change scenarios.     

Flavonoids from shoots and roots of Trifolium repens (white clover) grown in presence or absence of the arbuscular mycorrhizal fungus Glomus intraradices

White clover (Trifolium repens) plants were grown in the presence or absence of the arbuscular mycorrhizal fungus Glomus intraradices. Flavones, 4',5,6,7,8-pentahydroxy-3-methoxyflavone and 5,6,7,8-tetrahydroxy-3-methoxyflavone, as well as two flavones 3,7-dihydroxy-4'-methoxyflavone and 5,6,7,8-tetrahydroxy-4'-methoxyflavone never previously reported in plants, were isolated. The known 3,5,6,7,8-pentahydroxy-4'-methoxyflavone, 2',3',4',5',6'-pentahydroxy-chalcone, 6-hydroxykaempferol, 4',5,6,7,8-pentahydroxyflavone and 3,4'-dimethoxykaempferol were also obtained. Analysis of extracts obtained from roots and shoots revealed that the compositions of the flavonoid mixtures varied with growing conditions. Quercetin, acacetin and rhamnetin accumulated in roots of inoculated plants, whereas they were not detected in non-inoculated plants.     

Polyphosphate dynamics in mycorrhizal roots during colonization of an arbuscular mycorrhizal fungus

Inorganic polyphosphate (poly P) has been considered to be a translocatable form of phosphate (Pi) in arbuscular mycorrhizal fungi (AMF). Here we examined time-course changes in poly P content during the AMF colonization process. Onion (Allium cepa) plants were cultured with or without inoculation with Gigaspora margarita for 2-8 wk with periodic sampling. Poly P in the extracts, purified through gel filtration, was quantified by the reverse reaction of polyphosphate kinase. The length of poly P in mycorrhizal roots appeared to be shorter than in extraradical hyphae or in spores of the AMF, indicating that AMF depolymerize poly P before providing Pi to the host. The poly P content increased as colonization proceeded, and was highly correlated with the weight of the colonized roots. These results support the model that AMF supply Pi to the host through the poly P pool, and that the poly P content of a mycorrhizal root can be a good indicator of the Pi-supplying activity of AMF.     

丛枝菌根真菌数个种的基因组和转录组研究概况

丛枝菌根真菌（AMF）在自然界分布广泛，能与大部分维管植物的根系形成菌根共生体。它们在调节植物群落结构和全球的碳、氮、磷循环等方面发挥着重要的生态功能，也是农林、环境领域最具应用前景的微生物类群。受限于培养方法、研究手段等，长期以来对AMF基因组、转录组特征的认识非常有限。最近10年，AMF基因组和转录组的相关研究在高通量测序技术的推动下取得了较快发展；研究结果也显著提高了对AMF遗传发育、代谢生理、共生机制等的认识。本文综述了目前已完成测序的AMF种类的基因组、转录组信息。结果发现，已测序的AMF种类普遍具有基因组大、转座子丰富、AT碱基含量高、含大量未知功能基因与特异性基因、缺少部分共生相关基因等特点。在转录层面，总结了不同AMF种类、AMF不同共生结构、共生阶段以及与不同寄主植物共生时的转录本特征。结果发现，不同种类AMF的转录本大小差异明显。不同共生阶段或不同共生结构中的AMF转录本也具有较大的差异，且差异表达的基因大部分与养分交易、信号转导等密切相关。相比之下，同种AMF与不同寄主植物共生时的转录本表现出较高的保守性。最后，本文提出了本领域需要重点关注的研究方向，包括AMF纯培养技术的革新、AMF基因功能的解析、非模式AMF类群的研究以及对AMF蛋白组的研究。     

Associations between arbuscular mycorrhizal fungi and Rhynchrelyrum repens in abandoned quarries in southern China

The association between arbuscular mycorrhizal fungi (AMF) and Rhynchrelyrum repens was investigated. In six abandoned quarries in the Pearl River Delta area, R. repens was found to be associated with nine AMF species, including Glomus versiforme, G. brohultii, G. microaggregatum, G. clarum and G. claroideum, Acaulospora delicata, A. mellea, A. mollowae and Entrophospora infrequens. The genus Glomus and the species G. brohultii were recorded at the highest frequencies. Three typical arbuscular mycorrhizal (AM) structures, i.e. hyphae, vesicles and arbuscules, were found in the roots of the R. repens specimens collected from all the quarries investigated. Vesicles were the most frequently recorded structure. Results of a container-based experiment showed that R. repens had very high mortality (83.3%) in the absence of AMF in soil containing sufficient P (phosphorus); this indicates that R. repens is an obligate mycotroph. The presence of AMF significantly increased the biomass accumulation of R. repens seedlings (p < 0.01). It was also observed that AMF colonization was related to soil P and K (potassium) utilization by R. repens seedlings. It is, therefore, important to inoculate with AMF when using R. repens for the restoration of damaged ecosystems.     

Increasing phosphorus removal in willow and poplar vegetation filters using arbuscular mycorrhizal fungi

Fast growing woody species are increasingly used in vegetation filters for wastewater treatment. Their efficiency in phosphorus (P) removal notably depends on plant uptake and storage in aboveground tissues. In this study, Populus NM5 (P. nigra × P. maximowiczii), Salix miyabeana (SX64) and Salix viminalis (5027) were planted in pots to evaluate the influence of colonization by arbuscular mycorrhizal fungi (AMF) Glomus intraradices on P uptake using two different P concentrations in irrigation water. Based on analysis of the foliar and woody components, our results show that the two treatments (inoculation with G. intaradices and P-irrigation) interact differently with total P content. Foliar P content is principally enhanced by the P-irrigation concentration, whereas the mycorrhizal colonization increases stem P content. In the presence of G. intraradices, both S. miyabeana and S. viminalis showed a 33% increase in stem P content. The latter finding is mainly due to an increase in biomass production, without modification of the P concentration, indicating that AMF associations affect P use efficiency. Thus, using arbuscular mycorrhizal fungi for phytoremediation strategies may increase biomass productivity and hence improve pollutant uptake.     

Variation for aluminium tolerance in white clover

Aluminium (Al) tolerance of fourteen white clover (Trifolium repens L.) cultivars from eleven countries was compared in the greenhouse in the Wainui silt loam (Typic Dystrochrept) to which Al had been added at nine levels (0, 2.5, 5, 20, 50, 150, 250, 500 and 750 mg kg⁻¹ of soil) as Al₂ (SO₄)₃ and incubated for 30 days. None of the white clover cultivars, including those either referred to as Al-tolerant, Dusi and Pathfinder, or from countries that have large areas of acid soils, El Lucero M.A.G., Bayucua, Bage and Zapican, showed greater Al-tolerance than ‘Grasslands Huia’ white clover. Subsequent screening for Al-tolerance can therefore be restricted to germplasm with wide agronomic adaptation.     

Glomus intraradices dominates arbuscular mycorrhizal communities in a heavy textured agricultural soil

Arbuscular mycorrhizal fungal (AMF) spore communities were surveyed in a long-term field fertilization experiment in Switzerland, where different amounts of phosphorus (P) were applied to soil. Plots receiving no P as well as plots systematically fertilized in excess to plant needs for 31 years were used to test the hypothesis that application of P fertilizer changes the composition and diversity of AMF communities. AMF spores were isolated from the field soil, identified, and counted so as to quantify the effect of P fertilization on AMF spore density, composition, and diversity. Trap cultures were established from field soil with four host plants (sunflower, leek, maize, and Crotalaria grahamiana), and the spore communities were then analyzed in substrate samples from the pots. Altogether, nine AMF species were detected in the soil. No evidence has been acquired for effect of P fertilization on spore density, composition, and diversity of AMF in both the field soil and in trap cultures. On the other hand, we observed strong effect of crop plant species on spore densities in the soil, the values being lowest under rapeseed and highest under Phacelia tanacetifolia covercrop. The identity of plant species in trap pots also significantly affected composition and diversity of associated AMF communities, probably due to preferential establishment of symbiosis between certain plant and AMF species. AMF spore communities under mycorrhizal host plants (wheat and Phacelia in the fields and four host plant species in trap pots) were dominated by a single AMF species, Glomus intraradices. This resulted in exceptionally low AMF spore diversity that seems to be linked to high clay content of the soil.Electronic supplementary material Supplementary material is available for this article at and accessible for authorised users.     

The influence of Rhizobium and arbuscular mycorrhizal fungi on nitrogen and phosphorus accumulation by Vicia faba

BACKGROUND AND AIMS: The aim of this study was to investigate the effects of the interactions between the microbial symbionts, Rhizobium and arbuscular mycorrhizal fungi (AMF) on N and P accumulation by broad bean (Vicia faba) and how increased N and P content influence biomass production, leaf area and net photosynthetic rate. METHODS: A multi-factorial experiment consisting of four different legume-microbial symbiotic associations and two nitrogen treatments was used to investigate the influence of the different microbial symbiotic associations on P accumulation, total N accumulation, biomass, leaf area and net photosynthesis in broad bean grown under low P conditions. KEY RESULTS: AMF promoted biomass production and photosynthetic rates by increasing the ratio of P to N accumulation. An increase in P was consistently associated with an increase in N accumulation and N productivity, expressed in terms of biomass and leaf area. Photosynthetic N use efficiency, irrespective of the inorganic source of N (e.g. NO3- or N2), was enhanced by increased P supply due to AMF. The presence of Rhizobium resulted in a significant decline in AMF colonization levels irrespective of N supply. Without Rhizobium, AMF colonization levels were higher in low N treatments. Presence or absence of AMF did not have a significant effect on nodule mass but high N with or without AMF led to a significant decline in nodule biomass. Plants with the Rhizobium and AMF symbiotic associations had higher photosynthetic rates per unit leaf area. CONCLUSIONS: The results indicated that the synergistic or additive interactions among the components of the tripartite symbiotic association (Rhizobium-AMF-broad bean) increased plant productivity.     

丛枝菌根真菌与深色有隔内生真菌对姜瘟病的防效及抗病机理初探

【背景】青枯劳尔氏菌(Ralstonia solanacearum,R.S)引发的姜瘟病是生姜产业发展的瓶颈问题。丛枝菌根真菌(arbuscular mycorrhiza fungi, AMF)与深色有隔内生真菌(dark septate endophytes,DSE)是两类重要的共生微生物。【目的】前期研究发现,AMF与DSE可提高生姜对姜瘟病的抗性,但其抗病机制尚不清楚,极大地限制了利用这两类共生真菌对该病的防治。【方法】在温室条件下做盆栽试验,以生姜组培苗为材料,设立接种AMF、DSE和不接种AMF、DSE的对照(CK)处理,并在上述处理下的植物生长4周后淋入病原菌液,病原菌接种1周后,通过测定菌根侵染率、发病率、叶绿素含量、光合指标、磷(P)含量、防御性酶活性及丙二醛(malondialdehyde, MDA)含量,研究AMF和DSE互作对病原菌侵染后生姜生长和生理生化指标的影响。【结果】AMF和DSE分别使姜瘟病发病率降低了45.27%和52.04%(P<0.05)。AMF+DSE组合处理抑病效果更好,发病率较对照降低60.87%(P<0.05)。AMF、DSE及...     

植物根际促生菌及丛枝菌根真菌协助植物修复重金属污染土壤的机制

植物修复是一种前景广阔的重金属污染土壤的主要修复技术,在微生物的协助下效果更为显著。植物根际促生菌可通过分泌吲哚-3-乙酸(IAA)、产铁载体、固氮溶磷等方式促进植物生长、改善植物重金属耐受性,从而有效提高重金属污染土壤的植物修复效率。菌根真菌是土壤-植物系统中重要的功能菌群之一,可侵染植物根系改变根系形态和矿质营养状况,通过菌丝体吸附重金属,也可产生球囊霉素、有机酸、植物生长素等次生代谢产物改变重金属生物有效性。植物根际促生菌与丛枝菌根真菌可对植物产生协同促生作用,在重金属污染土壤修复中具有一定应用潜力。目前,国内外关于植物根际促生菌和丛枝菌根真菌互作已有大量研究,而二者的相互作用机理仍处于探索阶段。本文综述了近年来国内外植物根际促生菌和丛枝菌根真菌在重金属污染土壤植物修复中的作用机制,并对其研究前景进行展望。     

Flavonoids exclusively present in mycorrhizal roots of white clover exhibit a different effect on arbuscular mycorrhizal fungi than flavonoids exclusively present in non-mycorrhizal roots of white clover

Abstract

The flavonoids 5,6,7,8,9-hydroxy chalcone, 3,7-hydroxy-4′methoxy flavone, 5,6,7,8-hydroxy-4′-methoxy flavone and 3,5,6,7,4′-hydroxy flavone can be detected only in non-mycorrhizal roots of white clover, but not in mycorrhizal roots, whereas the flavonoids acacetin, quercetin and rhamnetin are only present in mycorrhizal roots. We tested the effect of several concentrations of these compounds on spore germination, hyphal growth, hyphal branching, formation of clusters of auxiliary cells and of secondary spores of the arbuscular mycorrhizal fungi Gigaspora rosea, Gigaspora margarita, Glomus mosseae and Glomus intraradices. Our results indicate that depending on the flavonoid, the tested compounds are involved at different stages in the regulation of mycorrhization. This hypothesis is strengthened by their differing effect on several AM fungal growth parameters. Furthermore, our study provides more data on the AM fungus genus/species specificity of flavonoids.     

矿区分离丛枝菌根真菌对万寿菊吸Cd潜力影响

盆栽试验研究土壤不同施Cd水平(0、5、20、50μg/g)下,接种矿区污染土壤中丛枝菌根真菌对万寿菊根系侵染率、植株生物量及Cd吸收与分配的影响。结果表明:接种丛枝菌根真菌显著提高Cd胁迫下万寿菊的根系侵染率和植株生物量;随着施Cd水平提高,各处理植株Cd浓度显著增加。各施Cd水平下万寿菊地上部Cd吸收量远远高于根系Cd吸收量,在土壤施Cd量达到50μg/g时,接种处理地上部Cd吸收量是根系的3.48倍,对照处理地上部Cd吸收量是根系的1.67倍;同一施Cd水平下接种处理植株Cd吸收量要显著高于对照。总体上,试验条件下污染土壤中分离的丛枝菌根真菌促进了万寿菊对土壤中Cd的吸收,并在一定程度上增加Cd向地上部分的运转,表现出植物提取的应用潜力。     

Auxin-stimulated somatic embryogenesis from immature cotyledons of white clover

Cotyledons from immature embryos of white clover (Trifolium repens L.) cv. Osceola were exposed to 2,4-D or NAA to induce somatic embryogenesis. NAA at 10 or 20 mg 1^–1 was very inefficient at stimulating embryogenesis, while concentrations of 30 or 40 mg 1^–1 resulted in death of the explant tissue. Continuous exposure of cotyledons to 40 mg 1^–1 2,4-D resulted in somatic embryos which were arrested at the globular stage, or which underwent cycles of secondary embryogenesis, never proceeding beyond the globular stage. A 10 day exposure time to 2,4-D at the same concentration led to formation of somatic embryos, most of which had poorly developed cotyledons. Almost 10% of the somatic embryos converted into plants following transfer to medium devoid of growth regulators. Attempts to improve morphology of somatic embryos by using shorter exposure times to 2,4-D at 40 mg 1^–1, or by maintaining the 10 day exposure time while varying the concentration of 2,4-D, were not successful. Plants were obtained from all parents evaluated, although at different frequencies.     

Adventitious shoot production from immature embryos of white clover

Cotyledonary-stage embryos of Haifa white clover, collected 13 days after cross-pollination, were induced to form adventitious shoots primarily from the hypocotyl region. The culture medium used for the production of adventitious shoots contained 5 M thidiazuron and 0.5 M -naphthaleneacetic acid. Numerous shoot meristems were produced within the first week, discrete shoots developed by week three, small plantlets by week eight, and whole plants in soil by week ten. 95–100% of all embryos, regardless of genotype, produced adventitious shoots within four weeks with an average production of 17.5 shoots per embryo. The majority of shoots (on average 77%) were easily converted to whole plants in soil. The white clover regeneration system described is prolific, rapid and effective on a large number of genotypes.Abbreviations BA N⁶-benzylaminopurine - MS medium Murashige & Skoog medium (1962) - NAA -naphthaleneacetic acid - thidiazuron N-phenyl-N-1,2,3-thiadiazol-5-ylurea