A tripartite culture system for endomycorrhizal inoculation of micropropagated strawberry plantlets in vitro

The objective of the current investigation was to develop a reliable method to obtain vesicular arbuscular mycorrhizae (VAM) in micropropagated plantlets and to determine their influence on growth. An in vitro system for culturing the VA mycorrhizal fungus Glomus intraradices with Ri T-DNA-transformed carrot roots or nontransformed tomato roots was used in this study as a potential active source of inoculum for the colonization of micropropagated plantlets. After root induction, micropropagated plantlets grown on cellulose plugs (sorbarod) were placed in contact with the primary mycorrhizae in growth chambers enriched with 5000 ppm CO₂ and fed with a minimal medium. After 20 days of tripartite culture, all plantlets placed in contact with the primary symbiosis were colonized by the VAM fungus. As inoculum source, 30-day-old VA mycorrhizal transformed carrot roots had a substantially higher infection potential than 5-, 10-or 20-day-old VAM. Colonized plantlets had more extensive root systems and better shoot growth than control plants. The VAM symbiosis reduced the plantlet osmotic potential. This response may be a useful pre-adaptation for plantlets during transfer to the acclimatization stage.     

Mycorrhizas improve nitrogen nutrition of Trifolium repens after 8 yr of selection under elevated atmospheric CO2 partial pressure

Altered environmental conditions may change populations of arbuscular mycorrhizal fungi and thereby affect mycorrhizal functioning. We investigated whether 8 yr of free-air CO2 enrichment has selected fungi that differently influence the nutrition and growth of host plants. In a controlled pot experiment, two sets of seven randomly picked single spore isolates, originating from field plots of elevated (60 Pa) or ambient CO2 partial pressure (pCO2), were inoculated on nodulated Trifolium repens (white clover) plants. Fungal isolates belonged to the Glomus claroideum or Glomus intraradices species complex, and host plants were clonal micropropagates derived from nine genets. Total nitrogen (N) concentration was increased in leaves of plants inoculated with fungal isolates from elevated-pCO2 plots. These isolates took up nearly twice as much N from the soil as isolates from ambient-pCO2 plots and showed much greater stimulation of biological N2 fixation. The morpho-species identity of isolates had a more pronounced effect on N2 fixation and on root length colonized than isolate identity. We conclude that rising atmospheric pCO2 may select for fungal strains that will help their host plants to meet increased N demands.     

Sucrose synthase levels do not limit or regulate carbon transfer in the arbuscular mycorrhizal symbiosis

Abstract

Sucrose synthase (SuSy) is the main sucrose breakdown enzyme in plant sink tissues, including nodules, and is a possible candidate for the diversion of plant carbon to arbuscular mycorrhizal (AM) fungi in roots. We tested the involvement of SuSy in AM symbiosis of Glomus intraradices and Pisum sativum (pea). We observed that peas deficient in the predominant root isoform of SuSy were colonized successfully by AM fungi similar to wild-type roots. SuSy protein levels did not increase in roots as AM symbiosis developed, although SuSy protein levels did increase in nodules as the rhizobium symbiosis developed. Our results lead us to conclude that, unlike nodule symbiosis, SuSy protein does not limit or regulate carbon transfer in the AM symbiosis.     

The liverwort Marchantia foliacea forms a specialized symbiosis with arbuscular mycorrhizal fungi in the genus Glomus

Microscopic evidence suggests that fungi forming endosymbioses with liverworts in the Marchantiales are arbuscular mycorrhizal (AM) fungi from the Glomeromycota. Polymerase chain reaction amplification of ribosomal sequences confirmed that endophytes of the New Zealand liverwort, Marchantia foliacea, were members of the genus Glomus. Endophytes from two Glomus rDNA phylotypes were repeatedly isolated from geographically separated liverwort samples. Multiple phylotypes were present in the same liverwort patch. The colonizing Glomus species exhibited substantial internal transcribed spacer sequence variation within phylotypes. This work suggests that certain liverwort species may serve as a model for studying DNA sequence variation in colonizing AM phylotypes and specificity in AM-host relationships.     

丛枝菌根真菌四个中国新记录种

从梅和滨海植物根区土壤中分离得到丛枝菌根真菌4个中国新记录,即沙生球囊霉Glomus arenarium、莫顿球囊霉G. mortonii、三壁球囊霉G. trimurales和苍耳管柄囊霉Funneliformis xanthium。对其形态特征进行重新描述和图解。这4个种的标本保存在中国科学院菌物标本馆（HMAS）和厦门市园林植物园标本馆中。     

Mycorrhiza and root hairs in barley enhance acquisition of phosphorus and uranium from phosphate rock but mycorrhiza decreases root to shoot uranium transfer

Some phosphate rocks (PR) contain high concentrations of uranium (U), which are potentially toxic via accumulation in soils and food chains, and plant uptake of U is likely to be influenced by characteristics of roots and associated microorganisms. The relative importance of root hairs and mycorrhiza in U uptake from PR was studied using a root hairless barley (Hordeum vulgare) mutant (Brb) and its wild type (WT). Both plant genotypes were grown in pots with Glomus intraradices BEG 87, or in the absence of mycorrhiza, and three P treatments were included: nil P, 2% (w/w) PR and 50 mg KH(2)PO(4)-P kg(-1) soil. Mycorrhiza markedly increased d. wts and P contents of Brb amended with nil P or PR, but generally depressed d. wts of WT plants, irrespective of P amendments. Mycorrhiza had contrasting effects on U contents in roots and shoots, in particular in Brb where mycorrhiza increased root U concentrations but decreased U translocation from roots to shoots. The experiment supports our understanding of arbuscular mycorrhiza as being multifunctional by not only improving the utilization of PR by the host plant but also by contributing to the phytostabilization of uranium.     

Arbuscular mycorrhizal associations in Lycopodiaceae

This study characterizes the molecular and phylogenetic identity of fungi involved in arbuscular mycorrhizal (AM) associations in extant Huperzia and Lycopodium (Lycopodiaceae). Huperzia and Lycopodium are characterized by a life cycle with long-lived autotrophic sporophytes and long-lived mycoheterotrophic (obtain all organic carbon from fungal symbionts) gametophytes. 18S ribosomal DNA was isolated and sequenced from Glomus symbionts in autotrophic sporophytes of seven species of Huperzia and Lycopodium and mycoheterotrophic Huperzia gametophytes collected from the Páramos of Ecuador. Phylogenetic analyses recovered four Glomus A phylotypes in a single clade (MH3) that form AM associations with Huperzia and Lycopodium. In addition, phylogenetic analyses of Glomus symbionts from other nonphotosynthetic plants demonstrate that most AM fungi that form mycoheterotrophic associations belong to at least four specific clades of Glomus A. These results suggest that most mycoheterotrophic plants that form AM associations do so with restricted clades of Glomus A. Moreover, the correspondence of identity of AM symbionts in Huperzia sporophytes and gametophytes raises the possibility that photosynthetic sporophytes are a source of carbon to conspecific mycoheterotrophic gametophytes via shared fungal networks.     

Arbuscular mycorrhizal fungi can transfer substantial amounts of nitrogen to their host plant from organic material

Nitrogen (N) capture by arbuscular mycorrhizal (AM) fungi from organic material is a recently discovered phenomenon. This study investigated the ability of two Glomus species to transfer N from organic material to host plants and examined whether the ability to capture N is related to fungal hyphal growth. Experimental microcosms had two compartments; these contained either a single plant of Plantago lanceolata inoculated with Glomus hoi or Glomus intraradices, or a patch of dried shoot material labelled with (15)N and (13)carbon (C). In one treatment, hyphae, but not roots, were allowed access to the patch; in the other treatment, access by both hyphae and roots was prevented. When allowed, fungi proliferated in the patch and captured N but not C, although G. intraradices transferred more N than G. hoi to the plant. Plants colonized with G. intraradices had a higher concentration of N than controls. Up to one-third of the patch N was captured by the AM fungi and transferred to the plant, while c. 20% of plant N may have been patch derived. These findings indicate that uptake from organic N could be important in AM symbiosis for both plant and fungal partners and that some AM fungi may acquire inorganic N from organic sources.     

Arbuscular mycorrhizal colonization on carbon economy in perennial ryegrass: quantification by 13CO2/12CO2 steady-state labelling and gas exchange

Effects of the arbuscular mycorrhizal fungus (AMF) Glomus hoi on the carbon economy of perennial ryegrass (Lolium perenne) were investigated by comparing nonmycorrhizal and mycorrhizal plants of the same size, morphology and phosphorus status. Plants were grown in the presence of CO2 sources with different C isotope composition (delta13C -1 or -44). Relative respiration and gross photosynthesis rates, and belowground allocation of C assimilated during one light period ('new C'), as well as its contribution to respiration, were quantified by the concerted use of 13CO2/12CO2 steady-state labelling and 13CO2/12CO2 gas-exchange techniques. AMF (G. hoi) enhanced the relative respiration rate of the root + soil system by 16%, inducing an extra C flow amounting to 3% of daily gross photosynthesis. Total C flow into AMF growth and respiration was estimated at < 8% of daily gross photosynthesis. This was associated with a greater amount of new C allocated belowground and respired in mycorrhizal plants. AMF colonization affected the sources supplying belowground respiration, indicating a greater importance of plant C stores in supplying respiration and/or the participation of storage pools within fungal tissues. When ontogenetic and nutritional effects were accounted for, AMF increased belowground C costs, which were not compensated by increased photosynthesis rates. Therefore the instantaneous relative growth rate was lower in mycorrhizal plants.     

Carbonic anhydrase and neuronal enzymes in cultured glomus cells of the carotid body of the rat

Summary The cellular localization of carbonic anhydrase (CAH) in the carotid body of the rat was investigated by means of Hansson's cobalt-precipitation technique in cultures of dissociated cells. In both young (2-day-old) and old (77-day-old) cultures, the parenchymal glomus (type-I) cells were selectively stained by this technique, and in addition expressed tyrosine hydroxylase and neuron-specific enolase as revealed by immunofluorescence. Enzymic reaction product of CAH appeared to be predominantly intracellular since staining was more intense and occurred more rapidly following permeabilization of the cell membranes with Triton X-100; its formation was inhibited by the CAH-inhibitor acetazolamide (1–10 M) or by increasing the pH from 5.8 to 7.5. Cryostat sections of the carotid bifurcation revealed intense CAH-reaction product in cell clusters of the carotid body, in a few cells of the nodose ganglion, and in red blood cells. Neuronal cell bodies of the petrosal ganglion and superior cervical ganglion (SCG) were largely non-reactive. The SCG is known to contain clusters of small intensely fluorescent (SIF) cells, which were also non-reactive when grown in dissociated cell culture. Thus, although glomus and SIF cells are often considered to be similar cell types, functional CAH-activity appears unique to glomus cells, and this may be important for the physiological response of the carotid body to certain chemosensory stimuli.     

European and African maize cultivars differ in their physiological and molecular responses to mycorrhizal infection

Physiological and molecular responses to phosphorus (P) supply and mycorrhizal infection by Glomus intraradices were compared in European (River) and African (H511) maize (Zea mays) cultivars to examine the extent to which these responses differed between plants developed for use in high- and low-nutrient-input agricultural systems. Biomass, photosynthetic rates, nutrient and carbohydrate contents, mycorrhizal colonization and nutrient-responsive phosphate transporter gene expression were measured in nonmycorrhizal and mycorrhizal plants grown at different inorganic phosphorus (P(i)) supply rates. Nonmycorrhizal River plants grew poorly at low P(i) but were highly responsive to mycorrhizal infection; there were large increases in biomass, tissue P content and the rate of photosynthesis and a decline in the expression of phosphate transporter genes. Nonmycorrhizal H511 plants grew better than River plants at low P(i), and had a higher root : shoot ratio. However, the responses of H511 plants to higher P(i) supplies and mycorrhizal infection were much more limited than those of River plants. The adaptations that allowed nonmycorrhizal H511 plants to perform well in low-P soils limited their ability to respond to higher nutrient supply rates and mycorrhizal infection. The European variety had not lost the ability to respond to mycorrhizas and may have traits useful for low-nutrient agriculture where mycorrhizal symbioses are established.