The influence of inorganic nutrient fertilization on the growth,nutrient composition and vesicular-arbuscular mycorrhizal colonization of pretransplant rice (Oryza sativa L.) plants

Summary The effects of P, N and Ca+Mg fertilization on biomass production, leaf area, root length, vesiculararbuscular mycorrhizal (VAM) colonization, and shoot and root nutrient concentrations of pretransplant rice (Oryza sativa L.) plants were investigated. Mycorrhizal plants generally had a higher biomass and P, N, K, Ca, Mn, Fe, Cu, Na, B, Zn, Al, Mg, and S shoot-tissue nutrient concentrations than non-mycorrhizal plants. Although mycorrhizal plants always had higher root-tissue nutrient concentrations than non-mycorrhizal plants, they were not significantly different, except for Mn. N fertilization stimulated colonization of the root system (colonized root length), and increased biomass production and nutrient concentrations of mycorrhizal plants. Biomass increases due to N were larger when the plants were not fertilized with additional P. P fertilization reduced the colonized root length and biomass production of mycorrhizal plants. The base treatment (Ca+Mg) did not significantly affect biomass production but increased the colonized root length. These results stress the importance of evaluating the VAM rice symbiosis under various fertilization regimes. The results of this study suggest that pretransplant mycorrhizal rice plants may have a potential for better field establishment than non-mycorrhizal plants.     

Selection of efficient vesicular-arbuscular mycorrhizal fungi for wetland rice (Oryza sativa L.)

Summary We tested the response of the wetland rice cultivar Prakash to inoculation with ten vescular-arbucular mycorrhizal (VAM) fungi (three selected from the first screening and seven isolated from local paddy fields) in a pot experiment under flooded conditions in order to select the most efficient mycorrhizal fungi to inoculate the rice nursery. A sandy clay loam soil was used as the substrate, fertilized with the recommended N and K levels (100 kg N ha^–1 as ammonium sulphate and 50 kg K ha^–1 as muriate of potash) and half the recommended level of P (25 kg ha^–1 as super phosphate). The inoculation was made into dry nursery beds and the beds were flooded when the seedlings were about 25 cm high, in 15 days. Twenty-eight-day old seedlings were transferred to pots filled with well puddled soil flooded with 5 cm of standing water. Based on the increase in grain yield and total biomass, Glomus intraradices and Acaulospora sp. were considered efficient and suitable for inoculation into rice nurseries.     

Growth response of field-grown Siratro (Macroptilium atropurpureum Urb.) and Aeschynomene american L. to inoculation with selected vesicular-arbuscular mycorrhizal fungi

Summary The effect of inoculation with a selected isolate of Glomus etunicatum Becker and Gerdemann and one of G. intraradices Schenck and Smith on the growth and nutrient content of Macroptilium atropurpureum Urb. cv. Siratro and Aeschynomene americana L., at applied P levels of 10, 30, 60, and 120 kg ha^-1, was studied under field conditions. At all P levels and for all harvests, the shoot dry mass of Siratro and A. americana were greater for the plants inoculated with the vesicular-arbuscular mycorrhizal (VAM) fungi than the control plants. Differences between the VAM fungus-inoculated and the control plants were most marked between 30 and 90 kg ha^-1 of applied P and diminished at 120 kg ha^-1. At the first harvest of Siratro, the plants inoculated with G. etunicatum had a greater shoot dry mass than those inoculated with G. intraradices, for all levels of applied P. However, for subsequent harvest of Siratro and for the one harvest of A. americana the response of shoot dry mass to the two VAM fungi was equivocal. Fungal inoculation gave at least a 30% saving in the amount of P fertilizer required (40 kg ha^-1) for the maximum yield. The plants inoculated with VAM fungi had a greater tissue concentration and total content of P and N than the control plants at low and intermediate levels of applied P. The percentage of root colonized by VAM fungi for the inoculated plants of the two legumes increased linearly with P additions up to 60 kg ha^-1. The conclusion is that under amended (limed and fertilized) soil conditions, inoculation with selected VAM fungi can improve the establishement and growth of forage legumes in fields that contain ineffective populations of native VAM fungi.     

Effect of fungicides on three vesicular-arbuscular mycorrhizal fungi associated with winter wheat (Triticum aestivum L.)

Summary We studied the effects of field application rates of three fungicides on spore germination and root infection in three species of Glomus. Bavistin (carbendazim) prevented germination of spores of G. monosporum and G. mosseae whereas Calixin (tridemorph) had no effect. Tilt Turbo (propiconazole) inhibited spore germination less than Bavistin. Spores of G. geosporum germinated in the presence of all three fungicides. Foliar applications of Bavistin alone and in a triple-spray programme significantly reduced infection of all three Glomus species in wheat roots grown in conditions of low P availability (2 mg P 1^–1). Two fungicides with triazole derivatives, Sportak and Tilt Turbo, also decreased infection and the former significantly reduced spore production of all three Glomus species. Calixin-treated plants, however, showed increased infection levels, resulting in greater yields and greater foliar-P concentrations than control plants. Milgo (ethirimol) was not detrimental to the vesicular-arbuscular mycorrhizal (VAM) symbiosis when applied as a foliar spray.     

Reduction of bacterial growth by a vesicular-arbuscular mycorrhizal fungus in the rhizosphere of cucumber (Cucumis sativus L.)

Summary Cucumber was grown in a partially sterilized sand-soil mixture with the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum or left uninoculated. Fresh soil extract was places in polyvinyl chloride tubes without propagules of mycorrhizal fungi. Root tips and root segments with adhering soil, bulk soil, and soil from unplanted tubes were sampled after 4 weeks. Samples were labelled with [³H]-thymidine and bacteria in different size classes were measured after staining by acridine orange. The presence of VAM decreased the rate of bacterial DNA synthesis, decreased the bacterial biomass, and changed the spatial pattern of bacterial growth compared to non-mycorrhizal cucumbers. The [³H]-thymidine incorporation was significantly higher on root tips in the top of tubes, and on root segments and bulk soil in the center of tubes on non-mycorrhizal plants compared to mycorrhizal plants. At the bottom of the tubes, the [³H]-thymidine incorporation was significantly higher on root tips of mycorrhizal plants. Correspondingly, the bacterial biovolumes of rods with dimension 0.28–0.40×1.1–1.6 m, from the bulk soil in the center of tubes and from root segments in the center and top of tubes, and of cocci with a diameter of 0.55–0.78 m in the bulk soil in the center of tubes, were significantly reduced by VAM fungi. The extremely high bacterial biomass (1–7 mg C g^-1 dry weight soil) was significant reduced by mycorrhizal colonization on root segments and in bulk soil. The incorporation of [³H]-thymidine was around one order of magnitude lower compared to other rhizosphere measurements, probably because pseudomonads that did not incorporate [³H]-thymidine dominated the bacterial population. The VAM probably decreased the amount of plant root-derived organic matter available for bacterial growth, and increased bacterial spatial variability by competition. Thus VAM plants seem to be better adapted to compete with the saprophytic soil microflora for common nutrients, e.g., N and P, compared to non-mycorrhizal plants.     

Physiological enhancement of early growth of rice seedlings (Oryza sativa L.) by production of phytohormone of N2-fixing methylotrophic isolates

Three plant-growth promoting, N₂-fixing methylotrophic strains isolated from rice cultivars (Oryza sativa L.), viz, Methylobacterium sp. CBMB20, Enterobacter sp. CBMB30, Burkholderia sp. CBMB40, were selected, and their activities in promoting the early growth of rice were studied. Seeds treated with the methylotrophic strains improved seed germination, seedling vigor index (SVI), and biomass of rice seedlings. The methylotrophic population in the treated seedlings increased in the vegetative stages when compared to seeding stages. Treated seedlings showed a higher accumulation of plant hormones viz trans-zeatin riboside, isopentenyladenosine, and indole-3-acetic acid than untreated seedlings. Plant hormones were detected immunologically using the phytodetek kit. Conformational evidence suggested that cytokinins were produced by the epiphytic bacteria colonizing the plants rather than by the plants themselves. In addition, the inoculated early stage rice seedlings also exhibited a wide range of acetylene reduction activity. The results suggest the potential use of these bacteria to stimulate germination, SVI, and biomass production, which is mediated by production of plant hormone accumulation and nitrogen fixation.     

Soil pH,rhizobia, and vesicular-arbuscular mycorrhizae inoculation effects on growth and heavy metal uptake of alfalfa (Medicago sativa L.)

Summary The interaction between soil pH and inoculation with rhizobia and vesicular-arbuscular mycorrhizae (VAM) was studied in an industrially polluted soil contaminated with high levels of Zn and Cd. A silt loam soil (pH 6.7) was amended with Ca(OH)₂ or elemental S to adjust the soil pH to 4.3, 5.3, 6.0, and 7.2. Alfalfa (Medicago sativa L.) was planted in each treated soil an subsequently inoculated with Rhizobium meliloti and/or a mixed VAM spore population. Alfalfa growing in soils at a pH of 4.3 and 5.3 failed to survive as a result of soil acidity and heavy metal toxicity. At the three higher pH values, growth and foliar N and P were significantly increased by inoculation with rhizobia or VAM. The greatest increase was observed when both VAM and rhizobia were inoculated together into the soil. With a soil pH of 6.0 and 6.7, the available heavy metal concentration in the soil was high and the VAM significantly decreased heavy metal uptake from these soils. The foliar concentration of Zn was reduced from 455 to 306 g g^–1 by inoculation with VAM (pH 6.0). At the highest soil pH (7.2), however, available heavy metal concentrations were generally lower and NAM significantly increased the heavy metal uptake. The influence of VAM on heavy metal uptake thus appears to be partly a function of the available heavy metal content in the soil.     

Effects of herbicides on three vesicular-arbuscular fungi associated with winter wheat (Riticum aestivum L.)

Summary We studied the effects of field application rates of four herbicides (Avenge, Ceridor, Dicurane, and Harrier) on spore germination and infection of wheat roots by three species of Glomus grown under conditions of low P availability. Low concentrations of Ceridor (bifenox, mecoprop) and Harrier (mecoprop, ioxynil, clopyralid) inhibited spore germination while higher concentrations were stimulatory. Avenge (difenzoquat methyl sulphate) prevented spore germination completely, while Dicurane (chlortoluron) had no effect. The herbicide applications had no significant effect on the infection rates of any of the three fungi except Harrier on G. geosporum. The herbicide treatments did, however, affect plant growth and ear yields. Ceridor and Harrier increased ear yields, while Dicurane showed marked phytotoxic effects.     

Ecology of soil microflora and mycorrhizal symbionts in degraded forests at two altitudes

Summary Microbial populations were estimated in four different forest stands at different regenerational stages, two each at higher and lower altitudes. The fungal and bacterial populations showed marked seasonal variations at both altitudes. Quantitatively, the bacterial population was higher than the fungal population. Although 25 fungal species were isolated at the lower altitude, only 15 were obtained at the higher altitude. Penicillium chrysogenum and Trichoderma viride were dominant at the lower and higher altitudes, respectively. In the more degraded forest stand at the lower altitude both the fungal and the bacterial population showed a significant positive correlation with organic C (r=0.658 and 0.735, respectively), whereas in the less degraded forest stand there was a significant correlation only between the fungal population and organic C (r=0.835). At the higher altitude, however, a highly significant correlation (P<0.05) was observed between the fungal population, soil moisture and organic C in both the forest stands. Disturbance to the soil and vegetation adversely affected the microbial population, and also affected endogonaceous spores. At the lower altitude, plants in the more degraded forest stand were more mycotrophic compared to those in the less degraded stand. The level of mycorrhizal infection showed a highly positive correlation with soil moisture, organic C, total N, and available P. The spore population, however, was correlated negatively with these parameters. Three different endogonaceous genera, Glomus, Gigaspora, and Acaulospora, were identified during the course of investigation. Glomus, however, was dominant.     

Phosphatase activity and cytokinin content in cowpeas (Vigna unguiculata) inoculated with a vesicular-arbuscular mycorrhizal fungus

We examined the effect of a vesicular-arbuscular mycorrhizal (VAM) fungus Glomus pallidum Hall on the phosphatase activity and cytokinin concentration in cowpea [Vigna unguiculata (L.) Walp] roots at successive stages of plant growth. Both acid and alkaline phosphatase activity were significantly (P=0.05) higher in mycorrhizal than in non-mycorrhizal roots 30 days after inoculation. Similarly, the cytokinin content was significantly increased in mycorrhizal roots compared to non-mycorrhizal roots. Our study suggests that these biochemical changes may improve the growth of mycorrhizal cowpea plants.     

Variable effectivity of three vesicular-arbuscular mycorrhizal endophytes in Hedysarum coronarium and Medicago sativa

Summary Plants of Hedysarum coronarium L. and Medicago sativa L., inoculated with Glomus caledonium, were grown for 6 weeks in soils with increasing amounts of available phosphorus (P). H. coronarium showed no or very low levels of infection even at very low soil P content (4 ppm), while in M. sativa the highest soil P level (20 ppm) was associated with the lowest percentage of infection. Medicago sativa and H. coronarium grown for 16 weeks in a sandy soil showed clear effects of Glomus inoculation on shoot growth and P uptake. In M. sativa inoculation increased shoot growth and P uptake in all treatments considerably, while the P concentration in the shoot was depressed by G. caledonium. In H. coronarium inoculation with G. mosseae and G. occultum had only a modest impact on shoot growth; G. caledonium even depressed shoot growth significantly. Glomus inoculation had a positive impact on the P concentrations in the shoots of H. coronarium. The growth-depressing effect of G. caledonium on H. coronarium is discussed.     

Inoculation response in kidney beans (Phaseolus vulgaris,L.) to vesicular-arbuscular mycorrhizal fungi and rhizobia in non-sterilized soil

The effect of dual inoculation on three local cultivars (Miss Kelly, Portland Red, Round Red) of red kidney beans (Phaseolus vulgaris, L.) with four strains of Rhizobium leguminosarum bv. phaseoli and three species of vesicular-arbuscular mycorrhizal (VAM) fungi was examined in a clay loam soil. Rhizobial strains B 17 and B 36, each paired with Glomus pallidum or G. aggregatum, were the most effective pairings for cv. Miss Kelly. Inoculation of Miss Kelly with any of these pairings significantly (P=0.05) increased growth, number of nodules, nodule dry weight, mycorrhizal colonization, and shoot N and P content than other pairings. The growth response by cv. Portland Red was significantly improved by pairings of B 36 or B 17 with any of the three VAM fungi. For both cultivars (Miss Kelly and Portland Red), CIAT 652 or T 2 paired with VAM fungi did not give a positive growth response. In contrast, for cv Round Red the T 2 rhizobial strain in combination with any of the three VAM fungi showed a significant (P=0.05) growth improvement in all parameters. Our results suggest that while dual inoculation of VAM fungi and rhizobia significantly improves the growth response by red kidney beans, the best pairings of VAM fungus and rhizobia for each cultivar need to be carefully selected.     

Impact of simulated erosion on the abundance and activity of indigenous vesicular-arbuscular mycorrhizal endophytes in an Oxisol

Summary The influence of simulated erosion on the abundance and activity of indigenous vesiculararbuscular mycorrhizal (VAM) populations was evaluated in an Oxisol. Surface-soil losses in excess of 7.5 cm were generally associated with significant decreases in the numbers of total and active VAM propagules and in the symbiotic effectiveness of the active propagules. Surface-soil removal not exceeding 7.5 cm was associated with decreased propagule abundance without adverse effects on VAM colonization of roots and symbiotic effectiveness of the fungi. The extent of VAM colonization of roots and the degree of symbiotic effectiveness observed at this level of simulated erosion were significantly higher than those observed in the soil not subjected to simulated erosion. This stimulation is attributed to the removal of antagonistic biotic factors as the top 7.5 cm of soil was removed. It is concluded that propagules lost during erosional soil losses must be replaced before legumes grown on relatively highly weathered and severely eroded tropical soils could appreciably benefit from the VAM symbiosis.Hawaii Institute of Tropical Agriculture and Human Resources Journal Series No. 3234     

Increased emergence of spring wheat after inoculation with Pseudomonas chlororaphis isolate 2E3 under field and laboratory conditions

Inoculation at the time of planting with Pseudomonas chlororaphis strain 2E3 increased the emergence of spring wheat by 8% and 6% at two different sites in northern Utah. Isolate 2E3 strongly inhibited the growth of the wheat pathogen Fusarium culmorum on artificial media. A second isolate of P. chlororaphis (strain O6) also inhibited fungal growth on artifical media but did not increase emergence of the spring wheat at the same field sites. Inoculation of winter wheat by 2E3 did not promote emergence when planted into field soil sterilized by fumigation with methyl bromide. Under laboratory conditions, emergence of spring wheat in sterilized soils from both sites was at least 90%. In the soils that were not sterilized, emergence was below 25% in soil from one site and below 50% in soil from the other. Treating seeds with 2E3 significantly improved emergence in a sterile soil-containing matrix that had been inoculated with the wheat pathogen Fusarium culmorum. Consequently, we propose that increases in wheat emergence can be attributed to the suppression by 2E3 of pathogenic organisms present in the native field soils. A strain of Rhizoctonia solani, shown to the pathogenic on winter wheat, was isolated from one of these soils.     

Co-selection of compatible rhizobia and vesicular-arbuscular mycorrhizal fungi for cowpea in sterilized and non-sterilized soils

Summary We selected two isolates of Rhizobium for cowpea (Vigna unguiculata) with sterilized soil tests and two different isolates by non-sterilized soil testing. The four rhizobia were then paired individually with either Glomus pallidum, Glomus aggregatum, or Sclerocystis microcarpa in separate, sterilized, or non-sterilized soil experiments. The purpose of the experiments was to determine the effect of soil sterilization on the selection of effective cowpea rhizobia, and to see whether these rhizobia differed in their effects on cowpea growth when paired with various vesicular-arbuscular mycorrhizal (VAM) fungi. Our experiments showed that the rhizobia selected in sterilized soil tests produced few growth responses in the cowpea compared to the other introduced rhizobia, irrespective of pairing with VAM fungi in sterilized or non-sterilized soil. In contrast, the two rhizobia initially selected by non-sterilized soil testing significantly improved cowpea growth in non-sterilized soil, especially when paired with G. pallidum. Our results suggest that it is important to select for effective rhizobia in non-sterilized soil, and that pairing these rhizobia with specific, coselected VAM fungi can significantly improve the legume growth response.     

Uptake and metabolism of isouron in rice (Oryza sativa L.) seedlings

Summary Isouron, a pre-emergence herbicide, inhibited the growth of rice seedlings in a nutrient solution; the critical concentration was about 0.10 mg l^-1. Isotope studies showed that isouron, added to the nutrient solution, was observed by the rice roots and translocated rapidly to the shoot. In rice seedlings, isouron was converted to six known metabolites, 3-(5-(1-dimethyl-2-hydroxy-ethy)-3-isoxazolyl]urea, 3-(5-tert-butyl-3-isoxazolyl)-1-methylurea, 3-(5-tert-butyl-3-isoxazolyl)ure, 3-amino-5-tert-butylisoxazole, 3-[5-(1,1-dimethyl-2-hydroxyethyl)-3-isoxazolyl]-1-dimethylurea, and 3-[5-(1,1-dimethyl-2-hydroxyethyl)-3-isoxazolyl]-1-methylurea, and two unknown metabolites.     

Effect of indigenous and introduced vesicular-arbuscular mycorrhizal fungi on growth and phosphorus uptake of Trifolium pratense and on inorganic phosphorus fractions in a cambisol

Summary Five selected vesicular-arbuscular mycorrhizal (VAM) fungi and the native population of a cambisol were tested in sterilized soil conditions, with Trifolium pratense as host plant. Indigenous fungi were the most effective in enhancing plant growth and P uptake, which were correlated with a higher root colonization. Selected fungi did not spread further in the root after 4 months from sowing, occupying less than 10% at the end of the experiment; inoculation with Glomus fasciculatum E3 yielded a higher dry-matter production than any other VAM species, but did not significantly increase shoot P concentration above that of the non-mycorrhizal control. Interactions between indigenous and introduced VAM fungi were studied in unsterilized soil. Results from fresh and dry weights of shoots and the percentage of fungal infection showed that the native endophytes competed more efficiently in colonizing the root. Inoculation with selected VAM species did not improve plant growth. Sterilization altered the inorganic P fractions of the soil, particularly those extracted with NH₄F and NaOH. Sterilized soil contained less inorganic P than unsterilized soil, but more soluble P. By the end of the experiment in sterilized soil, P extracted with NH₄Cl, NH₄F and NaOH and total inorganic P were significantly different among inoculation treatments, suggesting that VAM fungi may differ in their ability to take up P.     

Effects of inoculation with N2-fixing cyanobacteria on the nitrogenase activity in soil and rhizosphere of wetland rice (Oryza sativa L.)

Summary A greenhouse experiment was conducted with wetland rice (Oryza sativa cv. IR-50) in a clay-loam soil (Fluventic Eutrochrept) to study the effect of cyanobacterial inoculation a mixed culture of Aulosira fertilissima, Nostoc muscorum, Nostoc spp., and Anabaena spp., applied at the rate of 0.15 g (dry weight pot^-1 or 43 kg ha^-1) on acetylene reduction activity in soil and the root system (excised root), and the grain and straw yield. The effects of applying P (40 kg ha^-1), N (60 kg ha^-1), and P+N to the soil were also evaluated. Cyanbacterial inoculation significantly increased (more than 200% on average) photo-dependent acetylene reduction activity in soils, particularly where the indigenous activity was considerably low, i.e. under unfertilized and N-fertilized conditions. The effect of inoculation was prominent at the maximum tillering and grain formation stages of the crop. This inoculation benefit was, however, marginal in P-applied soils (P and P+N), where the indigenous activity was stimulated more than threefold. The inoculation led to a remarkable increase in root-associative acetylene reduction activity after the maximum tillering stage of the crop, particularly with applied N but for other treatments this inoculation effect was not significant. Cyanobacterial inoculation also increased the grain and straw yield of the crop when N was not applied. The grain and straw yield was significantly correlated with the acetylene reduction activity in flooded soils and in the root system during the tillering and maximum tillering stages of rice growth, respectively.     

Influence of humic acids on laurel growth,associated rhizospheric microorganisms,and mycorrhizal fungi

Summary Increasing concentrations of humic acids were tested in order to determine their effects on the microbial rhizosphere and the growth of laurel (Laurus nobilis L.). Plants that were treated with 300 mg kg^-1 of humic acids had the heaviest weights of both fresh and dry shoots; however, doses of 3000 mg kg^-1 were highly phytotoxic and inhibited the growth of laurel. Total aerobic bacteria and actinomycetes were stimulated by doses of 1500 and 3000 mg kg^-1 of humic acids at the first harvest. Nevertheless, at the end of the experiment no significant differences were found among the different doses. The number of fungi living in the laurel rhizosphere was not affected by any concentration of humic acids. Vesicular-arbuscular mycorrhizal (VAM) colonization was only slightly affected by the addition of increasing concentrations of humic substances to the soil, while the hyphal growth of Glomus mosseae was reduced.     

Differential colonization of Azospirillum lipoferum on roots of two varieties of rice (Oryza sativa L.)

Summary Seedlings of rice (IR42 and IR50) were aseptically dipped into Azospirillum lipoferum strain 34H suspension under dark, and the presence of bacteria on the differentiating regions of rice roots was observed by scanning electron microscopy. The bacterium did not colonize the root tips of IR42, while it colonized this region in the case of IR50, within 24 h after inoculation. In the early stages, most of the bacteria were embedded in the ruptured mucigel below the root cap cells of IR42. Mucigel was hardly detectable in IR50. While the root hair primordia of IR50 were colonized heavily with the bacterium within 24 h, the root hairs of IR42 were colonized 48 and 72 h after inoculation. This phenomenon in relation to plant varietal differences was discussed.