Glomus fasciculatum alleviates transplantation shock of micropropagated Sesbania sesban

Investigations were carried out using the vesicular arbuscular mycorrhizal fungus, Glomus fasciculatum, to improve the success in transplanting micropropagated plantlets of Sesbania sesban. Plantlets were developed from somatic embryos and/or adventitious buds (induced from various explants on Gamborg's medium supplemented with 6-benzylaminopurine), in the presence of 10^–7 m α-naphthaleneacetic acid and 5×10^–6 m gibberellic acid. Subsequent to nodulating the roots with Rhizobium, plantlets were transplanted into sterile garden soil and inoculated with or without G. fasciculatum. Only 30% of plantlets transferred to soil without G. fasciculatum survived. In contrast, all the plantlets inoculated with G. fasciculatum survived. Histochemical study revealed the presence of intracellular hyphae with well-developed arbuscules and intercellular hyphae with vesicles, suggesting that G. fasciculatum formed a good mycorrhizal association with S. sesban roots. These observations showed that mycorrhizal association helped to increase the potential of micropropagated plantlets to successfully withstand transplantation shock. Received: 6 January 1997 / Revision received: 28 August 1997 / Accepted: 5 September 1997     

Survival of the external mycelium of a VAM fungus in frozen soil over winter

The present investigation examines (1) whether the external VAM mycelium survives winter freezing to act as a source of inoculum in the spring, and (2) whether soil disturbance reduces the infectivity of the external VAM mycelium following freezing of the soil. Sealed pouches of fine nylon mesh were placed in pots containing soil inoculated with a Glomus species. The mesh was impervious to roots but not to hyphae. Following two 3-week growth cycles of maize in the pots, the pouches were transplanted to the field. Pouches were removed from the field once during the 4 months when the soil was frozen, and once after spring thaw. Measurements were made of VAM spore density, hyphal length and viability in the pouches. Bioassays for infectivity were conducted on all pouches. Some VAM hyphae survived freezing and remained infective following winter freezing, in the absence of plant roots. Soil disturbance did not reduce the infectivity of hyphae following exposure to freezing temperatures. We observed a change in the distribution of viable cytoplasm within hyphae over winter, which we hypothesize represents an adaptation allowing hyphae to survive freezing temperatures. We suggest that the effect of disturbance on hyphal infectivity may be related to this seasonal change in the distribution of hyphal viability.     

Phosphorus effect on phosphatase activity in endomycorrhizal maize

Success of a mycorrhizal symbiosis is influenced by the availability of phosphorus (P) in the soil. Maize ( Zea mays L. cv. Great Lakes 586) plants were grown under five different levels of soil P, either in the presence or absence of formononetin or the vesicular‐arbuscular mycorrhizal (VAM) fungus Glomus intraradices Schenck and Smith. We detected physiological differences in mycorrhizal roots very early in the development of symbiosis, before the onset of nutrient‐dependent responses. Under low P levels, VAM roots accumulated a greater shoot dry weight (13%), root P concentration (15%) and protein concentration (30%) than non-VAM roots, although root growth was not statistically significantly different. At higher P levels, mycorrhizal roots weighed less than non-VAM roots (10%) without a concomitant host alteration of growth or root P concentration. Mycorrhizal colonization decreased as soil P increased. Formononetin-treatment enhanced colonization of the root by G. intraradices and partially overcame inhibition of VAM colonization by high soil P concentrations. This is the first report that formononetin improves root colonization under high levels of soil P. Acid phosphatase (ACP) and alkaline phosphatase (ALP) activities were closely related to the level of fungal colonization in corn roots. ACP activity in corn roots responded more to soil P availability than did ALP activity (38% more). These results suggest that ACP was involved in the increased uptake of P from the soil, while ALP may be linked to active phosphate assimilation or transport in mycorrhizal roots. Thus, soil P directly affected a number of enzymes essential in host-endophyte interplay, while formononetin enhanced fungal colonization.     

Vesicular mycorrhizal colonization of seedlings of Pinaceae and Betulaceae after spore inoculation with Glomus intraradices

 Although Pinaceae and Betulaceae have been reported to contain Glomus–type root endophytes, its ecological importance and the conditions influencing this symbiosis are poorly understood. Seedlings of Abies lasiocarpa, Alnus rubra, Pinus contorta, Pinus ponderosa, Pseudotsuga menziesii, and Tsuga heterophylla were inoculated with Glomus intraradices to determine the vesicular-arbuscular mycorrhizae (VAM) development and responsiveness of these hosts. The role of companion VAM host plants on mycorrhizal colonization and nutrient uptake by Pseudotsuga menziesii was also examined by growing seedlings of Pseudotsuga menziesii in dual culture with VAM hosts Thuja plicata or Calamagrostis rubescens. After 8 weeks, no seedlings were colonized. At 16 weeks, 8 of 17 Thuja plicata seedlings grown with Pseudotsuga menziesii and all 18 inoculated Thuja plicata seedlings grown alone were colonized with vesicles and hyphae. Two of 17 inoculated Pseudotsuga menziesii seedlings grown in dual culture with Thuja plicata were colonized with abundant vesicles and hyphae. No ectomycorrhizal seedlings grown in monoculture were colonized. At 9 months, all 10 Calamagrostis rubescens and all 10 inoculated Pseudotsuga menziesii seedlings grown in dual culture were colonized by vesicles and hyphae. Two of 10 inoculated Pseudotsuga menziesiiand 1 of 10 inoculated Pinus ponderosa seedlings grown in monoculture were similarly colonized. The mean phosphorus content in the needles of colonized Pseudotsuga menziesii seedlings grown with Calamagrostis rubescens was about twice as high as in noncolonized Pseudotsuga menziesiiseedlings grown with Calamagrostis rubescens. Tissue nitrogen did not differ between these treatments. The results show that Glomus intraradices colonization of Pinaceae is most successful when a VAM host is present, although some vesicular colonization of Pinaceae occurred in the absence of a VAM host. Accepted: 24 September 1997     

Multiple mycorrhizal associations of individual calcicole host plants in the alpine grass-heath zone

Summary Mycorrhizal colonization of the fibrous roots of alpine grasses and perennial herbs in microhabitats on rendzina soil were examined. Various host plants were associated simultaneously with more than one species of vesicular-arbuscular mycorrhizal fungi. All dominant graminoids had a high degree of endomycorrhizal infection. Septate endophytes (Phialophora sp., Rhizoctonia sp.) often occurred together with Acaulospora sp., Glomus tenue, G. tortuosum and Scutelispora calospora on individual hosts.     

丛枝菌根真菌侵染番茄离体毛状根双重培养体系的建立

利用发根农杆菌Agrobacterium rhizogenes A4诱导樱桃番茄Micro-Tom产生毛状根,并在此基础上成功建立了丛枝菌根(AM)真菌根内球囊霉Glomus intraradices与转移Ri T-DNA番茄离体毛状根的双重培养体系。该真菌侵染14d左右,菌丝开始形成多级分支,17-21d时,一些菌丝顶端膨大,长出新生孢子。接种后3个月时,每皿孢子数量达到600-800个。新形成的孢子无需休眠,可直接侵染番茄离体根。成功的番茄双重培养为番茄菌根生理分子机制的研究提供了理想的实验体系。     

Nitrogen limitation of production in a Californianannual grassland: The contribution of arbuscularmycorrhizae

Nutrient availability limits plant production acrossa wide range of terrestrial ecosystems. In this studyof a Californian annual grassland community, theinfluence of arbuscular mycorrhizal (AM) associationson plant nutrient acquisition was investigated usingfactorial combinations of nitrogen (N), phosphorus (P)and benomyl fungicide. N additions resulted in asignificant increase in shoot biomass demonstratingthat plant productivity in these soils was N-limited. The effect of P additions consistently depended onfungicide treatment. In the absence of benomyl, shootP accumulation was unaltered by P addition. In thepresence of benomyl, P addition significantlyincreased shoot P accumulation and was associated witha consistent trend towards enhanced shoot biomass.The induction of P-deficiency with benomylapplication suggests that effective P acquisition byAM may contribute to the strong N-limitation ofproduction observed in many terrestrial ecosystems.     

Glomus intraradices causes differential changes in amino acid and starch concentrations of in vitro strawberry subjected to water stress

The effect of colonization of tissue-cultured strawberry ( Fragaria × ananassa Duch. cv. Kent) plantlets in vitro by the arbuscular mycorrhizal fungus (AMF) Glomus intraradices on plantlet response to poly(ethylene glycol) (PEG)-8000-induced water stress was investigated. The plantlets were inoculated axenically and co-cultured with the AMF for 4 wk, then transferred to 15% PEG-8000 solutions for 4, 8 and 12 h. Relative water content, water potential, osmotic potential, leaf conductance for water vapour diffusion and photosynthetic efficiency as estimated by chlorophyll a fluorescence were all affected by the PEG treatment and its duration but not by the presence of the intraradical phase of the AMF. However, distinct differences in PEG-induced changes in amino acid content were observed between nonmycorrhizal and mycorrhizal plantlets. In the latter, the treatment with PEG caused a substantial decrease in asparagine levels in leaves that was accompanied by a marked increase in asparagine concentration in roots. The opposite was observed in nonmycorrhizal plantlets. Furthermore, concentrations of aspartic acid, serine, threonine, amino- N -butyric acid, alanine and starch increased in roots of mycorrhizal and decreased in nonmycorrhizal plantlets. Our results suggest the presence of a mobile pool of asparagine that can be translocated from leaves to roots or vice versa in response to PEG-induced water stress, depending on the mycorrhizal status of the plantlets. These opposite patterns suggest different strategies of mycorrhizal and nonmycorrhizal plantlets to water stress, which seem to involve different adjustments in nitrogen and carbon metabolism.     

Development of two endomycorrhizal symbioses on soybean and comparison with phosphorus fertilization

Summary Soybean (Glycine max L. Merr. cv. Amsoy 71) plants were grown in a greenhouse in a soil very low in plant-available P, and plants were harvested 5 times over a 21-week growth period. Soybeans were inoculated with one of two species of VAM fungi or received daily one of three nutrient solutions of different P concentrations (0.0, 0.2, or 1.0mMP). Until week 9, the dry weights, leaf areas and developmental stage of soybeans inoculated withG. fasciculatum orG. mosseae were similar to the 1.0 or 0.2mMP-treated plants, respectively. Phosphorus concentrations were significantly lower in VAM plants at weeks 6 and 9 as compared to non-VAM soybeans given 1.0mMP, suggesting P input in VAM plants was immediately used for new growth. Total P input for VAM plants was linear over 21 weeks, and the average rate of P uptake for these plants was 0.19mg P d⁻¹. Estimated specific P uptake rates (SPUR) for the mycorrhizae (VAM roots) were twice that of the control (0.0mMP) roots. The calculated SPURs forG. fasciculatum andG. mosseae hyphae were 95 and 120μg P g⁻¹ VAM d⁻¹ respectively, a 4 to 5 fold increase over non-inoculated roots, indicating more attention must be paid to P assimilation by VAM fungi in P-fixing substrates. Contribution from the Western Regional Research Center, USDA-ARS (CRIS No. 5325-20580-003).     

Characterization and localization of plant phenolics likely involved in the pathogen resistance expressed by endomycorrhizal roots

Phytochemical characterization of the major phenolic compounds and their ultrastructural localization were carried out on onion roots (Allium cepa L.) colonized by two vesicular-arbuscular mycorrhizal (VAM) fungi: Glomus intraradix Schenck & Smith and G. versiforme (Karst.) Berch. Free and wall-bound forms of phenolic components were quantified in relation to the duration of symbiosis. Both ferulic and p-coumaric acids, as well as N-feruloyltyramine were identified as the major phenolic metabolites bound to the cell walls of VAM onion roots. Results from mycorrhized and control plants suggest the presence of a mechanism leading to the oxidative condensation of phenols, the latter process depending on the presence or absence of symbiosis. Bioassays reveal that N-feruloyltyramine induces the branching of hyphae and reduces total fungal development. The overall results lead us to suggest that the progressive binding of phenolic compounds in VAM roots is directly involved in the control of VAM endophytic establishment and development, as it gradually reduces the plasticity and elasticity of the symbiotic matrix. Phenolic compounds bound to cell walls could also be indirectly responsible for the resistance of VAM roots to pathogenic fungi, since they result in increased resistance by the cell wall to the action of digestive enzymes.Presented in part at the 31st Annual Meeting and Symposium of the Phytochemical Society of North America, Fort Collins, Colo., June 1991     

Accumulation of cyclohexenone derivatives in barley, wheat and maize roots in response to inoculation with different arbuscular mycorrhizal fungi

Glomus intraradices , Glomus mosseae, and Gigaspora rosea leads to the accumulation of cyclohexenone derivatives. Mycorrhizal roots of all plants accumulate in response to all three fungi blumenin [9-O-(2′-O-glucuronosyl)-β-glucopyranoside of 6-(3-hydroxybutyl)-1,1,5-trimethyl-4-cyclohexen-3-one], 13-carboxyblumenol C 9-O-gentiobioside, nicoblumin [9-O-(6′-O-β-glucopyranosyl)-β-glucopyranoside of 13-hydroxy-6-(3-hydroxybutyl)-1,1,5-trimethyl-4-cyclohexen-3-one] and another, as yet unidentified, cyclohexenone derivative. The accumulation of all four compounds in three tested mycorrhizal plants colonized by the three arbuscular mycorrhizal fungi indicates no fungus-specific induction of these compounds. Accepted: 6 October 1999     

Mycorrhizal relationships in bottomland hardwood forests of the southern United States

Mycorrhizae are important in the functioning of forest ecosystems worldwide, and play a critical role in water uptake, nutrient acquisition, and prevention of feeder root disease. The majority of mycorrhizal research has been conducted on upland sites, especially in coniferous ecosystems and in commercial agricultural production. However, the maintenance and restoration of bottomland hardwood (BLH) forest ecosystems in the southern United States is of increasing concern. Both ectomycorrhizae and endomycorrhizae are present in BLH forests, although the dominance of one or the other type depends primarily on both the tree species and the hydrologic regime. Ectomycorrhizae tend to be more sensitive to flooding, while endomycorrhizal infection can be present even in permanently flooded soils. The mycorrhizae of sweetgum (Liquidambar styraciflua), green ash (Fraxinus pennsylvanica), and the oaks (Quercus spp.) have been studied most due to their economic importance.Considerable work is still needed to better understand mycorrhizal relationships in BLH ecosystems and associated trees, both with respect to infectivity and nutrient cycling. Such information may be necessary for restoration of BLH forests on old agricultural fields, or to maintain the productivity of BLH forests after harvest. This paper summarizes studies on mycorrhizae relationships in BLH forests and suggests future work necessary for understanding the role mycorrhizae can have in managing these ecosystems.     

Purification and identification of an IgE suppressor from strawberry in an in vitro immunization system

We purified and identified an IgE suppressor from the strawberry 'Toyonoka', based on the decrease of IgE production in in vitro immunization (IVI). Gel filtration experiment indicated that fractions in a 15-48 kDa range and <10 kDa have an IgE suppressive activity. Furthermore, the fraction in 15-48 kDa was subjected to chromatofocusing and found to have activities at isoelectric points, pI 6.0, 7.0, and 8.0-9.2. We focused on the active fractions of pI 8.0-9.2 and the purified a large amount of strawberry extracts by cation exchange resins in batch. A purified 39 kDa protein showed homology to plant glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in N-terminal amino acid sequence and had GAPDH enzymatic activity. Nucleotide sequence and deduced amino acid sequence of the obtained cDNA clone of the protein matched with the sequence of Fragaria x ananassa GAPDH in the GenBank with >98% identical nucleotides and >99% identical amino acids, respectively. The purified strawberry GAPDH suppressed total IgE production in IVI in a dose-dependent manner. From these results, we identified GAPDH as IgE suppressor in the strawberry. Our study may be applicable to the development of new methods to relieve allergic conditions using GAPDH and the screening of other functional factors for human health.     

Growth and photosynthetic activity of micropropagated strawberry plants inoculated with endomycorrhizal fungi (AMF) and growing under drought stress

The plants produced by in vitro methods are free of any microflora contrary to natural systems where plants are colonized by symbiotic fungi. The present paper reports the experiments carried out to evaluate the role of arbuscular endomycorrhizal fungi in development of micropropagated strawberries and their photosynthetic activity (measured by chlorophyll fluorescence) under drought conditions. Mycorrhization strongly affected growth and tolerance to water deficiency of the plants cultivated in greenhouse. Wilting of not-mycorrhized plants was accompanied by drastic increase of Fo and Tfm and decrease of Fm. At the same time, the value of these parameters for mycorrhized plants did not change. Drastic decrease in the value of parameters Fv/Fm, Fv/Fo and Fo/Fm for plants without AMF appeared at the end of dry period. Rise of Fs and decrease Rfd was noted only for not-mycorrhized plants. The plants colonized by fungi, fully recovered their photosynthetic activity when watering was restored.     

Greenhouse irrigation method influences growth and quality of in vitro propagated Cryptocoryne wendtii plantlets

Effects of free-draining, non-draining and hydroponic tank culture on ex vitro establishment and quality of micropropagated microcuttings of the ornamental aquarium plant Cryptocoryne wendtii De Wit. were examined. After 16 weeks microcutting survival and shoot production was similar between treatments. Plants cultured hydroponically in either non-draining flats or tanks exhibited significantly greater leaf production and elongation than those grown in free-draining plugs. Root production and elongation were significantly reduced in free-draining plugs, presumably due to increased water stress. Plantlet growth and quality was increased when microcuttings were maintained in truly hydroponic systems. This presumably resulted from reduced water stress.     

The impact of wildfire on vesicular-arbuscular mycorrhizal fungi and their potential to influence the re-establishment of post-fire plant communities

Wildfires are a typical event in many Australian plant communities. Vesicular-arbuscular mycorrhizal (VAM) fungi are important for plant growth in many communities, especially on infertile soils, yet few studies have examined the impact of wildfire on the infectivity of VAM fungi. This study took the opportunity offered by a wildfire to compare the infectivity and abundance of spores of VAM fungi from: (i) pre-fire and post-fire sites, and (ii) post-fire burned and unburned sites. Pre-fire samples had been taken in May 1990 and mid-December 1990 as part of another study. A wildfire of moderate intensity burned the site in late December 1990. Post-fire samples were taken from burned and unburned areas immediately after the fire and 6 months after the fire. A bioassay was used to examine the infectivity of VAM fungi. The post-fire soil produced significantly less VAM infection than the pre-fire soil. However, no difference was observed between colonization of plant roots by VAM fungi in soil taken from post-fire burned and adjacent unburned plots. Soil samples taken 6 months after the fire produced significantly more VAM than corresponding soil samples taken one year earlier. Spore numbers were quantified be wet-sieving and decanting of 100-g, air-dried soil subsamples and microscopic examination. For the most abundant spore type, spore numbers were significantly lower immediately post-fire. However, no significant difference in spore numbers was observed between post-fire burned and unburned plots. Six months after the fire, spore numbers were the same as the corresponding samples taken 1 year earlier. All plants appearing in the burned site resprouted from underground organs. All post-fire plant species recorded to have mycorrhizal associations before the fire had the same associations after the fire, except for species of Conospermum (Proteaceae), which lacked internal vesicles in cortical cells in the post-fire samples.     

Changes in the population of infective endomycorrhizal fungi in a rice-based cropping system

The numbers of infectious propagules of indigenous vesicular-arbuscular mycorrhizal (VAM) fungi were determined at different stages of the rice-based cropping systems in two irrigated rice fields of varying strata and in a rainfed field. The most-probable-number method was used to estimate the infective VAM fungal population. On the irrigated farms the mycorrhizal inoculum was consistently less in the poorly drained low-lying field with a rice-rice cropping pattern than in the field in the better-drained upper stratum with a rice-corn-mung-bean pattern. The population of infective VAM fungi was generally low after the wet season rice crop when the field was inundated for a long period, increased during fallow in the presence of weeds, and was highest upon the maturity of the dry-season corn or rice crop. In the rainfed area the highest endophytic population was found at maturity of the mungbean crop and the lowest after land preparation prior to rice seeding.