The Beltsville method for soilless production of vesicular-arbuscular mycorrhizal fungi

A low-cost, low-maintenance system for soilless production of vesicular-arbuscular mycorrhizal (VAM) fungus spores and inoculum was developed and adapted for production of acidophilic and basophilic isolates. Corn (Zea mays) plants were grown with Glomus etunicatum, G. mosseae or Gigaspora margarita in sand automatically irrigated with modified Hoagland's solution. Sand particle size, irrigation frequency, P concentration, and buffer constituents were adjusted to maximize spore production. Modified half-strength Hoagland's solution buffered with 4-morpholine ethane-sulfonic acid (MES) automatically applied 5 times/day resulted in production of 235 G. etunicatum spores/g dry wt. of medium (341000 spores/pot) and 44 G. margarita spores/g dry wt. of medium (64800 spores/pot). For six basophilic isolates of G. mosseae, CaCO₃ was incorporated into the sand and pots were supplied with the same nutrient solution as for acidophilic isolates. The increased pH from 6.1±0.2 to 7.2±0.2 resulted in spore production ranging from 70 to 145 spores/g dry wt. (102000–210000 spores/pot). Spore production by all isolates grown in the soilless sand system at Beltsville has exceeded that of traditional soil mixtures by 32–362% in 8–12 weeks.     

Increased Sporulation of Vesicular-Arbuscular Mycorrhizal Fungi by Manipulation of Nutrient Regimens

Adjustment of pot culture nutrient solutions increased root colonization and sporulation of vesicular-arbuscular mycorrhizal (VAM) fungi. Paspalum notatum Flugge and VAM fungi were grown in a sandy soil low in N and available P. Hoagland nutrient solution without P enhanced sporulation in soil and root colonization of Acaulospora longula, Scutellospora heterogama, Gigaspora margarita, and a wide range of other VAM fungi over levels produced by a tap water control or nutrient solutions containing P. However, Glomus intraradices produced significantly more spores in plant roots in the tap water control treatment. The effect of the nutrient solutions was not due solely to N nutrition, because the addition of NH₄NO₃ decreased both colonization and sporulation by G. margarita relative to levels produced by Hoagland solution without P.     

Topsoil storage effects on primary production and rates of vesicular-arbuscular mycorrhizal development inAgropyron trachycaulum

Summary A greenhouse study was conducted to determine the effects of stockpiling prairie grassland topsoil for 3 years on mycorrhizal development and root and shoot production of slender wheatgrass. The vesicular-arbuscular mycorrhizal (VAM) fungi involved in the symbiosis were also assessed as was the decomposition potential of the soil. During the first week of growth, VAM development in grasses grown in the stockpiled soil lagged behind that observed for grasses in the undisturbed soil. However, by 3 weeks, the mycorrhizal infection in plants in the stockpiled soil had reached levels similar to that in plants in the undisturbed soil. The dominant species of VAM fungi involved in the symbiosis at 8 weeks after planting shifted fromGlomus fasciculatum in the undisturbed soil toG. mosseae in the stockpiled soil. The delay in initial VAM infection and shift in VAM fungal species did not significantly affect plant productivity which was greatest in the stockpiled soil. The greater shoot production exhibited by grasses in the stockpiled soil was attributed to higher levels of NO₃-N in the stockpiled than undisturbed soil. The potential of the soil to decay dead slender wheatgrass roots was not altered by stockpiling.     

Stimulation of Growth and Nutrient Uptake by VAM Fungi in Brassica Oleracea Var. Capitata

Cabbage (Brassica oleracea, var. capitata, cv. Hercules) seedlings were inoculated with vesicular-arbuscular mycorrhizal (VAM) fungi Glomus fasciculatum, G. aggregatum, and G. mosseae. Differential efficiency in mycorrhizal colonization and the specificity of fungal symbiont to stimulate the growth and nutrient uptake of the host were observed. In addition, there was an increase in phenol, protein, reducing sugar contents, and peroxidase activity in the VAM inoculated seedlings. Since these compounds are known to confer resistance against fungal pathogens, the use of VAM as a biological control agent to protect cabbage against several root diseases is suggested.     

Cd-tolerant arbuscular mycorrhizal (AM) fungi from heavy-metal polluted soils

Spores of arbuscular mycorrhizal (AM) fungi were isolated from two heavy-metal polluted soils in France via trap culture with leek (Allium porrum L.). Preliminary identification showed that the predominant spore type of both cultures (P2 and Cd40) belongs to the Glomus mosseae group. Their sensitivity to cadmium was compared to a laboratory reference strain (G. mosseae) by in vitro germination tests with cadmium nitrate solutions at a range of concentrations (0 to 100 mg L^–1) as well as extracts from a metal-polluted and unpolluted soils. Both cultures of AM fungi from heavy-metal polluted soils were more tolerant to cadmium than the G. mosseae reference strain. The graphically estimated EC₅₀ was 0.8 mg L^–1 Cd (concentration added to the test device) for G. mosseae and 7 mg L^–1 for P2 culture, corresponding to effective Cd concentrations of approximately 50–70 g L^–1 and 200–500 g L^–1, respectively. The extract of the metal-polluted soil P2 decreased germination of spores from the reference G. mosseae but not from P2 culture. However, the extracts of two unpolluted soils with different physico-chemical characteristics did not affect G. mosseae, whereas germination of P2 spores was markedly decreased in the presence of one of the extracts. These results indicate a potential adaptation of AM fungi to elevated metal concentrations in soil. The tested spores may be considered as metal-tolerant ecotypes. Spore germination results in presence of soil extracts show the difficulty of assessing the ecotoxic effect of metals on AM fungi without considering other soil factors that may interfere in spore germination and hyphal extension.     

Response of Stylosanthes guianensis to endomycorrhizal fungi inoculation as affected by lime and phosphorus applications

The effect of 3 different species of vesicular-arbuscular mycorrhizal fungi on the growth of Stylosanthes guianensis (Aubl.) Sw. cultivated in a sterilized acid and dystrophic soil (Quartzipsament), with 4 levels of lime (0; 0.27; 0.63 and 1.10 meq Ca²⁺/100 g air-dried soil, as Ca(OH)₂) and 2 P levels (0 and 20 mg P/kg soil, as KH₂PO₄) was evaluated under greenhouse conditions. Plants were harvested 40, 60, and 80 days after planting. Stylosanthes guianensis was highly mycotrophic, especially in soil without P fertilization. Mycotrophism was highest in plants inoculated with Acaulospora scrobiculata in soil receiving no P fertilizer and with 0.63 meq Ca²⁺/100 g air-dried soil. Shoot growth increment was as high as 5129% at the third harvest. Inoculation with Glomus macrocarpum presented intermediate results, whereas inoculation with Gigaspora margarita had no significant effect on plant growth. Root per cent colonization and shoot dry weight, as well as root percent colonization and shoot to root ratio were significantly correlated. The occurrence of S. guianensis in very acid and dystropic soils, containing toxic levels of Al³⁺, requires the association with VAM fungi for the plant tolerate such conditions.     

Effect of soil liming and vesicular-arbuscular-mycorrhizal inoculation on the growth and micronutrient content of the teff plant

In a greenhouse experiment involving an acid soil teff [Eragrostis tef (Zucc.) Trotter] plants failed to grow unless the soil was limed or inoculated with either of two vesicular-arbuscular-mycorrhizal (VAM) fungi,Glomus mosseae orGlomus macrocarpum. Plant growth increased by liming and to a lesser extent by VAM fungal inoculation. Liming also enhanced root colonization by VAM fungi. Shoot micronutrient content generally increased as a result of inoculation, and decreased by increased lime applications.     

Interaction of Vesicular-arbuscular Mycorrhizal Fungi and Phosphorus with Meloidogyne incognita on Tomato

The influence of two vesicular-arbuscular mycorrhizal fungi and phosphorus (P) nutrition on penetration, development, and reproduction by Meloidogyne incognita on Walter tomato was studied in the greenhouse. Inoculation with either Gigaspora margarita or Glomus mosseae 2 wk prior to nematode inoculation did not alter infection by M. incognita compared with nonmycorrhizal plants, regardless of soil P level (either 3 μg [low P] or 30 μg [high P] available P/g soil). At a given soil P level, nematode penetration and reproduction did not differ in mycorrhizal and nonmycorrhizal plants. However, plants grown in high P soil had greater root weights, increased nematode penetration and egg production per plant, and decreased colonization by mycorrhizal fungi, compared with plants grown in low P soil. The number of eggs per female nematode on mycorrhizal and nonmycorrhizal plants was not influenced by P treatment. Tomato plants with split root systems grown in double-compartment containers which had either low P soil in both sides or high P in one side and low P in the other, were inoculated at transplanting with G. margarita and 2 wk later one-half of the split root system of each plant was inoculated with M. incognita larvae. Although the mycoorhizal fungus increased the inorganic P content of the root to a level comparable to that in plants grown in high P soil, nematode penetration and reproduction were not altered. In a third series of experiments, the rate of nematode development was not influenced by either the presence of G. margarita or high soil P, compared with control plants grown in low P soil. These data indicate that supplemental P (30 μ/g soil) alters root-knot nematode infection of tomato more than G. mosseae and G. margarita.     

Soil Fungicides for Controlling Chytridiaceous Mycoparasites of Gigaspora margarita and Glomus fasciculatum

The percentage of azygospores of Gigaspora margarita with zoosporangia of chytridiaceous fungi (CF) was reduced significantly after agitating them in fenaminosulf before incubation in soil. Fenaminosulf did not affect zoosporangia development on chlamydospores of Glomus fasciculatum. Metalaxyl and ethazol were not effective against CF on spores of either mycorrhizal fungus. Azygospores of G. margarita were treated with fenaminosulf and used as the inoculum for pot cultures. After 19 weeks, the percentage of azygospores containing CF was reduced significantly by this treatment, whereas root colonization and sporulation by the mycorrhizal fungus were unaffected. Pot cultures of G. margarita, either drenched with fenaminosulf or not, did not differ in the percentage of azygospores containing CF. However, root colonization and sporulation by the mycorrhizal fungus was temporarily delayed when pots were drenched with fenaminosulf.     

Interaction of vascular plants and vesicular-arbuscular mycorrhizal fungi across a soil moisture-nutrient gradient

Summary Abundance and distribution of vascular plants and vesicular-arbuscular mycorrhizal (VAM) fungi across a soil moisture-nutrient gradient were studied at a single site. Vegetation on the site varied from a dry mesic paririe dominated by little bluestem (Schizachyrium scoparium) to emergent aquatic vegetation dominated by cattail (Typha latifolia) and water smartweed (Polygonum hydropiperoides). Plant cover, VAM spore abundance, plant species richness, and number of VAM fungi represented as spores, had significant positive correlations with each other and with percent organic matter. The plant and VAM spore variables had significant negative correlations with soil pH and available Ca, Mg, P and gravimetric soil moisture. Using stepwise multiple regression, Ca was found to be the best predictor of spore abundance. Test for association between plant species and VAM fungal spores indicated that the spores of Glomus caledonium are associated with plants from dry, nutrient poor sites and spores of gigaspora gigantea are positively associated with plants occurring on the wet, relatively nutrient rich sites. Glomus fasciculatum was the most abundant and widely distributed VAM fungus and it had more positive associations with endophyte hosts than the other VAM fungi. We found no relationship between beta niche breadth of plant species and the presence or absence of mycorrhizal infection. However, our data suggest that some plant species may vary with respect to their infection status depending upon soil moisture conditions that may fluctuate seasonally or annually to favor or hinder VAM associations.     

Growth stimulation ofTamarindus indica by selected VA mycorrhizal fungi

Efficient vesicular arbuscular mycorrhizal (VAM) fungi were screened and selected for a slow-growing forest tree species,Tamarindus indica L., important in tropical forestry. Seedlings were inoculated with 13 different VAM fungi, obtained from various sources around the world. Inoculated plants had greater plant height, leaf number, stem girth, biomass, phosphate and Zn²⁺ content. They also had higher amounts of mycorrhizal spores, per cent root colonization and external hyphae, as measured by per cent soil aggregation.Tamarindus indica seedlings responded best to inoculation withGigaspora margarita (ICRISAT) followed byGlomus fasciculatum.

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Résumé On a testé et sélectionné des moisissures mycorrhizales (VAM) vésiculaires, arbusculaires efficaces pour une espèce d'arbre forestier à croissance lente,Tamarindus indica L., importante en foresterie tropicale. On a inoculé des semences avec 13 moisissures VAM différentes, obtenues de diverses sources de par le monde. Les plants inoculés sont plus hauts, ont un nombre de feuilles et une circonférence de tronc plus élevés et un contenu plus élevé en biomasse, phosphate et Zn²⁺. Ils ont aussi une quantité plus élevée de spores mycorrhizales, une colonisation d'un pourcentage plus élevé de racines et d'hyphes externes, mesurés par le pourcentage d'aggrégation du sol. Les semences deTamarindus indica répondent le mieux à l'inoculation deGigaspora margarita ICRISAT suivi de celle deGlomus fasciculatum.

     

Effect of salinity on mycorrhizal onion and tomato in soil with and without additional phosphate

Summary Vesicular-arbuscular mycorrhizal fungi (VAM) are known to increase plant growth in saline soils. Previous studies, however, have not distinguished whether this growth response is due to enhanced P uptake or a direct mechanism of increased plant salt tolerance by VAM. In a glasshouse experiment onions (Allium cepa L.) were grown in sterilized, low-P sandy loam soil amended with 0, 0.8, 1.6 mmol P kg^–1 soil with and without mycorrhizal inoculum. Pots were irrigated with saline waters having conductivities of 1.0, 2.8, 4.3, and 5.9 dS m^–1. Onion colonized withGlomus deserticola (Trappe, Bloss, and Menge) increased growth from 394% to 100% over non-inoculated control plants when soil P was low ( 0.2 mmol kg^–1 NaHCO₃-extractable P) at soil saturation extract salinities from 1.1 dS m^–1 to 8.8 dS m^–1. When 0.8 and 1.6 mM P was added no dry weight differences due to VAM were observed, however, K and P concentrations were higher in VAM plants in saline treatments.Glomus fasciculatum (Gerdeman and Trappe) andGlomus mosseae (Nicol. and Gerd.) isolates increased growth of VAM tomato 44% to 193% in non-sterilized, saline soil (10 dS m^–1 saturation extract) despite having little effect on growth in less saline conditions when soil P was low. Higher tomato water potentials, along with improved K nutrition by VAM in onion, indicate mechanisms other than increased P nutrition may be important for VAM plants growing under saline stress. These effects appear to be secondary to the effects of VAM on P uptake.     

Responses of barley,pea, and maize to inoculation with different vesicular-arbuscular mycorrhizal fungi in irradiated soil

Summary The efficiency of different VAM fungi was investigated by inoculating barley, pea, and maize with different VAM fungi in irradiated soil in pots buried in the field. VAM frequency, growth and nutrient uptake were measured. In barleyGlomus epigaeus (CA) andG. macrocarpus (CA) were the most efficient out of 11 tested species and increased yield of grain by 24% and 21%, though they were not significant according to oneway analysis of variance. In pea, yield of grain was significantly increased from 46% to 104% (mean=68%) by 7 out of 10 tested species and by 105% by application of P fertilizer. The most efficient species wereG. epigaeus (CA),G. mosseae (GB), andG. etunicatus (CA). In maizeG. mosseae (GB) andG. caledonius (DK) increased total yield significantly by 59% and 47% in one experiment and in another experiment yield of cob was increased by 68% byG. mosseae (GB), 72% byG. caledonius (DK), and by 153% by application of P fertilizer. This experiment demonstrated that responsiveness to inoculation by VAM fungi differed among plant species, and that efficiency of different VAM fungi differed.     

The improvement of plant N acquisition from an ammonium-treated,drought-stressed soil by the fungal symbiont in arbuscular mycorrhizae

The ability of the external mycelium in arbuscular mycorrhiza for N uptake and transport was studied. The contribution of the fungal symbiont to N acquisition by plants was studied mainly under waterstressed conditions using ¹⁵N. Lettuce (Lactuca sativa L) was the host for two isolates of the arbuscular mycorrhizal fungi Glomus mosseae and G. fasciculatum. The experimental pots had two soil compartments separated by a fine mesh screen (60 m). The root system was restricted to one of these compartments, while the fungal mycelium was able to cross the screen and colonize the soil in the hyphal compartment. A trace amount of ¹⁵NH ₄ ⁺ was applied to the hyphal compartment 1 week before harvest. Under water-stressed conditions both endophytes increased the ¹⁵N enrichment of plant tissues; this was negligible in nonmycorrhizal control plants. This indicates a direct effect of arbuscular mycorrhizal fungi on N acquisition in relatively dry soils. G. mosseae had more effect on N uptake and G. fasciculatum on P uptake under the water-limited conditions tested, but both fungi improved plant biomass production relative to nonmycorrhizal plants to a similar extent.     

Distribution of vesicular-arbuscular mycorrhizal fungi in the natural ecosystem

Natural occurrence of vesicular-arbuscular mycorrhizal (VAM) fungi in Haryana soils showed that VAM sporulation was more intensive in the rhizosphere of nonlegumes than of legumes. Maximum number of spores (342 spores per 50 g of soil) was observed in the rhizosphere of mustard, followed by chickpea, wheat, pearl millet and pigeonpea. Four VAM generaviz. Glomus, Gigaspora, Sclerocystis andAcaulospora, were present there. Soil pH, total soil P, available P, type of soil, soil moisture and cropping season all variables influenced the VA mycorrhizal population in the natural ecosystem. Numbers of VAM spores highly correlated with the presence of total soil P and soil pH indirectly affected the VAM population through the total soil P. The spore population was abundant in sandy soils as compared to loamy sands. Drier soils had higher number of VAM spores. In summer, the VAM population in soil was less as compared to winter season.     

Alteration of growth ofSolanum opacum andPlantago drummondii and inhibition of regrowth of hyphae of vesicular-arbuscular mycorrhizal fungi from dried root pieces by manganese

Addition of MnSO₄ or MnCl₂ to a fine sandy soil from South Australia had a negative effect on shoot growth and root elongation ofSolanum opacum in the absence of significant presence of vesicular-arbuscular mycorrhiza (VAM). VAM ameliorated the reduction of plant growth by Mn, even though mycorrhizal development was decreased. Mn inhibited infection of roots by a fine endophyte less than that by some coarse endophytes. High concentrations of available Mn inhibited growth of hyphae of VAM fungi from dried root pieces, a significant source of infection by mycorrhizal fungi in the soil used.     

Arbuscular mycorrhizal contribution to heavy metal uptake by maize (Zea mays L.) in pot culture with contaminated soil

In two pot-culture experiments with maize in a silty loam (P2 soil) contaminated by atmospheric deposition from a metal smelter, root colonization with indigenous or introduced arbuscular mycorrhizal (AM) fungi and their influence on plant metal uptake (Cd, Zn, Cu, Pb, Mn) were investigated. Soil was -irradiated for the nonmycorrhizal control. In experiment 1, nonirradiated soil provided the mycorrhizal treatment, whereas in experiment 2 the irradiated soil was inoculated with spores of a fungal culture from P2 soil or a laboratory reference culture, Glomus mosseae. Light intensity was considerably higher in experiment 2 and resulted in a fourfold higher shoot and tenfold higher root biomass. Under the conditions of experiment 1, biomass was significantly higher and Cd, Cu, Zn and Mn concentrations significantly lower in the mycorrhizal plants than in the nonmycorrhizal plants, suggesting a protection against metal toxicity. In contrast, in experiment 2, biomass did not differ between treatments and only Cu root concentration was decreased with G. mosseae-inoculated plants, whereas Cu shoot concentration was significantly increased with the indigenous P2 fungal culture. The latter achieved a significantly higher root colonization than G. mosseae (31.7 and 19.1%, respectively) suggesting its higher metal tolerance. Zn shoot concentration was higher in both mycorrhizal treatments and Pb concentrations, particularly in the roots, also tended to increase with mycorrhizal colonization. Cd concentrations were not altered between treatments. Cu and Zn, but not Pb and Cd root-shoot translocation increased with mycorrhizal colonization. The results show that the influence of AM on plant metal uptake depends on plant growth conditions, on the fungal partner and on the metal, and cannot be generalized. It is suggested that metal-tolerant mycorrhizal inoculants might be considered for soil reclamation, since under adverse conditions AM may be more important for plant metal resistance. Under the optimized conditions of normal agricultural practice, however, AM colonization even may increase plant metal absorption from polluted soils.     

Effect of aromatic plant species on vesicular-arbuscular mycorrhizal establishment in Pistacia terebinthus

The inoculation of Pistacia terebinthus with vesicular-arbuscular mycorrhizal (VAM) fungi and the spread of the infection were studied using a mixed cropping system, under glasshouse conditions, with Salvia officinalis, Lavandula officinalis and Thymus vulgaris colonized by Glomus mosseae as an inoculation method. This method was compared with soil inoculum placed under the seed or distributed evenly in the soil. Indirect inoculation with all the aromatic plants tested significantly increased VAM root colonization of P. terebinthus compared with the use of soil inoculum, although the effect on plant growth was different for each one of the aromatic species used as inoculum source. Inoculation with L. officinalis and T. vulgaris were the best treatments resulting in high VAM colonization and growth enhancement of P. terebinthus.     

Influence of temperature on colonization of spring barleys by vesicular arbuscular mycorrhizal fungi

The effects of three soil temperatures on growth of spring barleys (Hordeum vulgare L.) and on their root colonization by vesicular arbuscular mycorrhizal (VAM) fungi from agricultural soils in Montana (USA) or Syria at different inoculum concentrations were tested in soil incubators in the greenhouse. The number of mycorrhizal plants as well as the proportion and intensity of roots colonized increased with higher soil temperatures. VAM fungi from Montana, primarily Glomus macrocarpum, were cold tolerant at 11°C while those from Syria, primarily G. hoi, were heat tolerant at 26°C. Inoculum potential of Montana VAM fungi was higher than Syrian VAM fungi in cool soils. Harmal, selected from Syrian barley land races, had the highest colonization by mycorrhizal fungi of the cultivars tested.Journal Series Paper: J-2532 Montana Agricultural Experiment Station.