Effect of onion (Allium cepa) root exudates on the hyphal growth of Gigaspora margarita

 The effect of root exudates from onions differing in P status on spore germination and hyphal growth of arbuscular mycorrhizal fungi was investigated. Onion (Allium cepa) was grown in solution culture at different phosphorus concentrations (0, 0.1, 1.0, 8.0 and 24.0 mg P l^–1) and root exudates were collected. When spores of the arbuscular mycorrhizal fungus, Gigaspora margarita were incubated with these root exudates, spore germination was only slightly affected but hyphal growth was greatly affected, particularly with exudates from P-deficient plants. This suggests that the P nutrition of host plants influences the composition of root exudates and thereby the hyphal growth of arbuscular mycorrhizal fungi. Accepted: 25 June 1995     

Studies of iron transport by arbuscular mycorrhizal hyphae from soil to peanut and sorghum plants

 The influence of an arbuscular mycorrhizal (AM) fungus on phosphorus (P) and iron (Fe) uptake of peanut (Arachis hypogea L.) and sorghum (Sorghum bicolor L.) plants was studied in a pot experiment under controlled environmental conditions. The plants were grown for 10 weeks in pots containing sterilised calcareous soil with two levels of Fe supply. The soil was inoculated with rhizosphere microorganisms only or with rhizosphere microorganisms together with an AM fungus (Glomus mosseae [Nicol. & Gerd.] Gerdemann & Trappe). An additional small soil compartment accessible to hyphae but not roots was added to each pot after 6 weeks of plant growth. Radiolabelled P and Fe were supplied to the hyphae compartment 2 weeks after addition of this compartment. After a further 2 weeks, plants were harvested and shoots were analysed for radiolabelled elements. In both plant species, P uptake from the labelled soil increased significantly more in shoots of mycorrhizal plants than non-mycorrhizal plants, thus confirming the well-known activity of the fungus in P uptake. Mycorrhizal inoculation had no significant influence on the concentration of labelled Fe in shoots of peanut plants. In contrast, ⁵⁹Fe increased in shoots of mycorrhizal sorghum plants. The uptake of Fe from labelled soil by sorghum was particularly high under conditions producing a low Fe nutritional status of the plants. These results are preliminary evidence that hyphae of an arbuscular mycorrhizal fungus can mobilise and/or take up Fe from soil and translocate it to the plant. Accepted: 6 March 1998     

Formation of appressoria by the arbuscular mycorrhizal fungus <Emphasis Type="Italic">Gigaspora margarita</Emphasis> on roots of <Emphasis Type="Italic">Allium cepa</Emphasis> is linked with root age

Spores of the arbuscular mycorrhizal fungus, Gigaspora margarita, were placed near the root tip, the middle of the root (equal distance from root base and root tip), or the root base (close to the shoot) of the first primary root of 9-day-old onion. Two weeks later, the number and position of appressoria and the appressoria with penetrating hyphae were determined in the first and the newly formed second primary roots. The total number of appressoria was not significantly different among the treatments. Inoculation near the root tip of the first primary root resulted in the formation of a large number of appressoria on the first primary root and the formation of about three times fewer appressoria on the second primary root. Inoculation near the base of the first primary root resulted in the formation of no appressoria on the first primary root, whereas many appressoria were formed on the second primary root. Our results suggest that the root age is a determinant of the appressorium formation.     

EST-library construction using spore RNA of the arbuscular mycorrhizal fungus Gigaspora rosea

RNA was extracted from activated spores of the arbuscular mycorrhizal fungus Gigaspora rosea. Double-stranded cDNA was synthesised, digested and cloned into the vector lambda-ZAP express. Of the 1,500 clones obtained, 1.5% carried inserts of the rRNA gene cluster. After excision, inserts from 50 randomly selected clones were sequenced. Database searches revealed that 62% of the clones had similarities to already known sequences. These mainly code for proteins involved in translation and protein processing, replication and the cell cycle and cell signal transduction. One fragment probably belonged to a metallothionein-encoding gene which may be involved in heavy-metal binding. The method presented is an easy and rapid way to obtain short fragments of coding regions for expressed sequence tag libraries. Accepted: 28 November 2000     

Response of 11 eucalyptus species to inoculation with three arbuscular mycorrhizal fungi

 Numerous publications have reported growth stimulation of Eucalyptus following ectomycorrhizal inoculation in nursery or field conditions. Although Eucalyptus species can also form arbuscular mycorrhiza, their dependency on this type of mycorrhiza is still debatable. This paper presents information on the effect of inoculation of arbuscular mycorrhizal fungi on eucalypt growth. Twenty weeks after mycorrhizal inoculation, Eucalyptus seedlings' stem dry weight could be increased up to 49% compared to non-inoculated control plants. Intensity of root colonization by a given fungus depended on the host species, but it was not related to a plant growth response. Leaf phosphorus concentration of non-inoculated Eucalyptus seedlings varied greatly between species. Increases in leaf phosphorus concentration following mycorrhizal infection were not necessarily associated with plant growth stimulation. The most mycorrhiza-dependent Eucalyptus species tended to be those having the highest leaf phosphorus concentration in the absence of a fungal symbiont. These mycorrhiza-dependent Eucalyptus species seem to have greater phosphorus requirements and consequently to rely more on the symbiotic association. Accepted: 1 September 1995     

The presence of the arbuscular mycorrhizal fungus Glomus intraradices influences enzymatic activities of the root pathogen Aphanomyces euteiches in pea roots

 Fungal enzyme activities were quantified in an interaction study between the fungus Glomus intraradices and the pea pathogen Aphanomyces euteiches. Fungal and host enzymes were separated by polyacrylamide gel electrophoresis and the activity of A. euteiches–specific glucose-6-phosphate dehydrogenase (Gd), phosphoglucomutase and peptidase (PEP) enzymes were quantified by densitometry. The activity of A. euteiches–specific enzymes increased until 14 days after inoculation with A. euteiches, and then decreased. The plants preinoculated with G. intraradices showed no symptoms of severe root rot even though the pathogen was present and active in these plants. Thus, plants preinoculated with G. intraradices were more tolerant of infection with A. euteiches than non-mycorrhizal plants. This effect was evident even though the A. euteiches infection levels of mycorrhizal and non-mycorrhizal plants were the same. A. euteiches enzyme activities in the mycorrhizal plants were different to those in non-mycorrhizal plants. The peaks of PEP and Gd enzyme activity of A. euteiches were lower and the development of A. euteiches PEP activity was later in the mycorrhizal plants than in the non-mycorrhizal plants. Accepted: 14 November 1996     

Phosphorus amendment inhibits hyphal branching of the VAM fungus Gigaspora margarita directly and indirectly through its effect on root exudation

 The effect of solution phosphorus (P) concentration upon growth of pregerminated spores of the vesicular-arbuscular mycorrhizal fungus Gigaspora margarita was examined in vitro. P at 1 mM significantly inhibited branching of the primary germ tube. The number of branches and the total hyphal length were both significantly inhibited at 10 mM P. In addition, germinated spores exposed to exudates produced by Ri T-DNA-transformed roots of Daucus carota L. grown in the presence of P showed significantly less hyphal branching than those exposed to exudates produced by P-stressed roots. These phenomena could contribute to the observed inhibition of mycorrhiza formation by high P. Accepted: 31 July 1996     

Influence of two legume species on hyphal production and activity of two arbuscular mycorrhizal fungi

 Two arbuscular mycorrhizal (AM) fungi (Glomus mosseae and G. intraradices) were compared for abundance of intraradical and soil-borne hyphae in association with Astragalus sinicum, a small-seeded, and Glycine max, a large-seeded legume. A. sinicum was more responsive than G. max to mycorrhizal formation, especially at early growth stages. Biomass allocation was greater in roots than shoots for mycorrhizal A. sinicum, while the opposite was true for G. max. Hyphal development in root and soil compartments was estimated by trypan blue staining and after staining for succinate dehydrogenase (SDH) or alkaline phosphatase (ALP) activity. Total fungal abundance increased steadily in roots and soil with time to a maximum 8 weeks after planting. SDH- and ALP-active AM hyphae increased in roots during plant growth but decreased in soil at later harvests. Mycorrhizal root mass in A. sinicum and G. max increased about 14-fold and 2.5-fold, respectively, but total length of soil hyphae produced per plant differed little, so that the pattern of AM soil to root abundance of the two fungi varied considerably with the host plant. Accepted: 23 July 1997     

Interactions between a mycophagous Collembola,dry yeast and the external mycelium of an arbuscular mycorrhizal fungus

 A plant growth system with root-free hyphal compartments was used to examine the interactions between a mycophagous Collembola (Folsomia candida Willem), dry yeast and an arbuscular mycorrhizal (AM) fungus [Glomus caledonium (Nicol. & Gerd.) Trappe and Gerdemann] in terms of Collembola reproduction, AM-hyphal length and AM-hyphal P transport. Collembola reproduction was unaffected by AM mycelium, but a supplement of dry yeast increased the Collembola population size. The addition of dry yeast increased AM-hyphal P transport by increasing hyphal length. Collembola without yeast affected neither AM-hyphal growth nor AM-hyphal P transport, whereas Collembola with yeast decreased AM-hyphal P transport by 75% after 8 weeks. The hyphal density of G. caledonium remained unaffected by Collembola except after 4 weeks in combination with yeast, when a 33% reduction was observed. The results of this experiment show that the interaction between F. candida and the external mycelium of G. caledonium is limited under the conditions imposed. Accepted: 27 February 1996     

Further root colonization by arbuscular mycorrhizal fungi in already mycorrhizal plants is suppressed after a critical level of root colonization

An established arbuscular mycorrhizal symbiosis suppresses further mycorrhization. It is not clear whether the observed suppressional effect is linked with the level of root colonization or not. In the present work we studied the effect of the degree of root colonization by the arbuscular mycorrhizal fungus Glomus mosseae on further root colonization by G. mosseae. At different time points barley plants grown in split-root compartments were pre-inoculated on one half of the split-root system with G. mosseae. Sequential inoculation resulted in different colonization levels. Thereafter, the second half of the split root system was inoculated. The results indicate an enhanced suppression of root colonization on the second side of the split-root system when colonization levels increased on the first side.     

Effect of phosphate and the arbuscular mycorrhizal fungus Glomus intraradices on disease severity of root rot of peas (Pisum sativum) caused by Aphanomyces euteiches

 The effects of inorganic phosphate levels and the presence of arbuscular mycorrhiza on disease severity of Aphanomyces euteiches in pea roots were studied. Disease severity on roots and epicotyl as well as the oospore number within infected root tissue were correlated with the phosphorus (P) level in the growth medium. The arbuscular mycorrhizal fungus Glomus intraradices increased P uptake and the P concentration in the plant but reduced disease development in peas. Polyacrylamide gel electrophoresis followed by densitometry of glucose-6-phosphate dehydrogenase specific to A.euteiches was used to measure the activity of the pathogen in roots. The enzyme activity increased with disease severity and disease incidence, except in plants supplemented with P at the highest level, where a peak in activity was seen 12 days after inoculation with the pathogen, followed by a decrease in activity. The epicotyl of mycorrhizal plants showed a reduction in disease severity although this part of the plants was not mycorrhizal. Thus, an induced systemic factor may be responsible for increased resistance in mycorrhizal plants. Accepted: 21 August 1998     

Production and specificity of polyclonal antibodies against soluble proteins from the arbuscular mycorrhizal fungus Glomus intraradices

Rabbit polyclonal antibodies were produced against a soluble protein fraction from a vesicle and spore mixture of the arbuscular mycorrhizal fungus (AMF) Glomus intraradices. The protocol for isolation of vesicles and spores from plant roots was optimized to minimize debris contamination. Protein extract purification and preparation for immunization was adapted to increase protein content and immunogenicity. Active antisera were produced starting from the second boost immunization. Antibodies obtained were specific for surface antigens of AMF and revealed different patterns of soluble protein antigens in G. intraradices, G. constrictum and an unidentified Glomus species. Accepted: 6 December 2000     

Differential activation of H+-ATPase genes by an arbuscular mycorrhizal fungus in root cells of transgenic tobacco

In arbuscular mycorrhizas, H⁺-ATPase is active in the plant membrane around arbuscules but absent from plant mutants defective in arbuscule development (Gianinazzi-Pearson et al. 1995, Can J Bot 73: S526–S532). The proton-pumping H⁺-ATPase is encoded by a family of genes in plants. Immunocytochemical studies and promoter-gusA fusion assays were performed in transgenic tobacco (Nicotiana tabacum L.) to determine whether the periarbuscular enzyme activity results from de-novo activation of plant genes by an arbuscular mycorrhizal fungus. The H⁺-ATPase protein was localized in the plant membrane around arbuscule hyphae. The enzyme was absent from non-colonized cortical cells. Regulation of seven H⁺-ATPase genes (pma) was compared in non-mycorrhizal and mycorrhizal roots by histochemical detection of β-glucuronidase (GUS) activity. Two genes (pma2, pma4) were induced in arbuscule-containing cells of mycorrhizal roots but not in non-mycorrhizal cortical tissues or senescent mycorrhiza. It is concluded that de-novo H⁺-ATPase activity in the periarbuscular membrane results from selective induction of two H⁺-ATPase genes, which can have diverse roles in plant-fungal interactions at the symbiotic interface. Received: 23 October 1999 / Accepted: 7 February 2000     

Direct application of carbendazim and propiconazole at field rates to the external mycelium of three arbuscular mycorrhizal fungi species: effect on 32P transport and succinate dehydrogenase activity

 The influence of the systemic fungicides propiconazole (Tilt 250E) and carbendazim (Bavistin) at field application rates on the functioning of three arbuscular mycorrhizal fungi was studied. Short-term fungal ³²P transport and succinate dehydrogenase (SDH) activity in external hyphae of Glomus intraradices Schenck and Smith, G. claroideum Schenck and Smith and G. invermaium Hall in symbiosis with pea (Pisum sativum L.) were measured. In the experimental system used, the hyphae grew into two root-free hyphal compartments (HCs). The fungicides were applied to each HC 24 days after sowing and ³²P was added to one HC of each pot. Four days later, the fungicide effect on fungal P transport was measured as the difference in ³²P content of treated and untreated plants. SDH activity in fungal hyphae was determined in the HCs given no ³²P. Carbendazim severely inhibited ³²P transport and SDH activity in external hyphae at an application rate of 0.5 μg g^–1 soil. The ergosterol inhibitor propiconazole affected none of these parameters. The fungicides had similar effects on all three fungal species, although P transport efficiency and SDH activity differed markedly between the fungi. Accepted: 12 December 1996     

White clover living mulch increases the yield of silage corn via arbuscular mycorrhizal fungus colonization

A field experiment was conducted to investigate the effects of white clover living mulch on the arbuscular mycorrhizal (AM) fungus colonization of corn roots and the yield of silage corn. The following seven treatments were setup in a field that had been kept bare by rotary tillage from August 2003 to July 2004: two white clover living mulch treatments without phosphorus (P) application, with the white clover shoots clipped and removed or allowed to lie in place before sowing corn; one no-tillage treatment without P application; and four rotary tillage treatments with different P application rates. White clover was broadcasted in the living mulch treatments in August 2004. In June 2005, the white clover shoots in the living mulch treatments were clipped. After tilling the four rotary tillage treatments, corn was sown in all the treatments. The fallow period before sowing corn was 0 month (living mulch treatments) and 22 months (no-tillage and rotary tillage treatments). At knee high stage, the AM fungus colonization of the corn roots and the P concentrations of the corn shoots in both the living mulch treatments were increased relative to those in the other treatments. The yield of corn tended to increase in the no-tillage and rotary tillage treatments with an increase in the P application rate. On the other hand, the yields of corn in the living mulch treatments without the P application were not significantly different from the maximum yield among the no-tillage and rotary tillage treatments. These results suggested that the white clover living mulch increased the yield of corn by facilitating the AM fungus colonization and improving the P nutrition of corn.     

The use of arbuscular mycorrhizae to control onion white rot (Sclerotium cepivorum Berk.) under field conditions

 A field experiment was carried out to determine the effects of the inoculation of onion (Allium cepa L.) with Glomus sp. Zac-19 on the development of onion white rot (Sclerotium cepivorum Berk.) and on onion production. Mycorrhization delayed onion white rot epidemics by 2 weeks and provided a significant protection against the disease for 11 weeks after onion transplanting, as compared with nonmycorrhizal controls. Mycorrhizal plants showed an increase of 22% in yield, regardless of the presence of the white rot pathogen. Accepted: 8 January 1996