Mycorrhizal dependency of some endemic and endangered Hawaiian plant species.

Four endemic species of Hawaiian plants were tested for their response to inoculation with a Hawaiian isolate of Glomus aggregatum (an arbuscular mycorrhizal fungus [AMF]) when grown in a native soil with or without P added to achieve different soil-solution P levels. The endangered species (Sesbania tomentosa [Fabaceae] and Colubrina oppositifolia [Rhamnaceae]) and two nonendangered species (Bidens sandvicensis and B. asymmetrica × sandvicensis [Asteraceae]) were tested. When soil-solution P levels in greenhouse trials were similar to unfertilized field soils (e.g., 0.005-0.020 mg P/L), shoots of inoculated plants were 2.1 to 7.0 times larger than noninoculated plants. Leaf tissue P levels and root biomass in these species showed similar responses to inoculation. Mycorrhizal dependencies ranging from 44 to 88% were measured when plants were grown in low-P soils and were -4-42% in soil with P levels typical of highly productive agricultural soils. A survey of P levels in a variety of native (nonagricultural) Hawaiian soils indicated the widespread occurrence of P-limited sites (mean = 0.010 mg P/L, range = <0.001-0.030 mg P/L; N = 41). The terms "ecological mycorrhizal dependency" (EMD) and "agricultural mycorrhizal dependency" (AMD) are introduced to refine the concept of mycorrhizal dependency.

Interaction of Glomus species and Vigna unguiculata in an oxisol subjected to simulated erosion.

An experiment was conducted in the greenhouse to determine the effectiveness of three species of vesicular-arbuscular (VA) mycorrhizal fungi in an oxisol subjected to simulated erosion using cowpea [Vigtta unguiculata (L.) Walp, cv.'California Black Eye'] as an indicator host. Inoculation of the eroded soil resulted in increased VA mycorrhizal colonization of roots without enhancing shoot P concentration and dry matter yields. Inoculation of the uneroded soil, however, led to significant improvement in infection level as well as in symbiotic effectiveness. Based on the results of this study, it appears that the lack of expression of mycorrhizal effectivness in the eroded soil is a result of nutrient deficiency. The results thus suggest the importance of restoring lost nutrients before the benefits of VA mycorrhizal inoculation could be effectively exploited for a successful establishment of a mycorrhizal cowpea in eroded soils.

Interaction of vesicular-arbuscular mycorrhizal fungi with erosion in an oxisol.

The development of vesicular-arbuscular mycorrhizal (VAM) symbiosis was monitored in Leucaena leucocephala grown in an Oxisol subjected to incremental simulated erosion. The density of VAM infective propagules in the soil diminished as the level of simulated erosion (removal of surface soil) was increased from 0 to 50 cm. The level of infection on L. leucocephala roots observed at harvest was not significantly influenced by simulated erosion unless removal of surface soil exceeded 25 cm. Inoculation of this soil and the uneroded soil with Glomus aggregatum enhanced the early onset of infection but did not significantly influence the level of infection observed at the time of harvest. Simulated erosion in excess of 7.5 cm of surface soil removal significantly delayed the development of VAM effectiveness monitored in terms of the P status of L. leucocephala subleaflets and also curtailed the level of maximum effectiveness observed. Decreases in VAM effectiveness were significantly correlated with decreases in soil chemical constituents. However, VAM effectiveness in a soil subjected to 30 cm of surface soil removal was not restored to a significant extent unless the soil was amended with P, even though other nutrients were restored to sufficiency levels. Our results demonstrate that the development of VAM effectiveness is the phase of the VAM symbiosis that is most adversely influenced by simulated erosion and that this effect appears to be caused primarily by insufficient P in the soil solution.

Soil Solution Phosphorus Status and Mycorrhizal Dependency in Leucaena leucocephala.

A phosphorus sorption isotherm was used to establish concentrations of P in a soil solution ranging from 0.002 to 0.807 mug/ml. The influence of P concentration on the symbiotic interaction between the tropical tree legume Leucaena leucocephala and the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum was evaluated in pot experiments. The level of mycorrhizal infection in Leucaena roots increased as the concentration of P was raised from 0.002 to 0.153 mug/ml. Higher levels of P depressed mycorrhizal infection, but the level of infection never declined below 50%. Periodic monitoring of P contents of Leucaena subleaflets indicated that significant mycorrhizal activity was detected as early as 17 days after planting, with the activity peaking 12 to 16 days thereafter. The highest level of mycorrhizal activity was associated with a soil solution P level of 0.021 mug/ml. Even though the mycorrhizal inoculation effect diminished as the concentration of P in the soil solution was increased, mycorrhizal inoculation significantly increased P uptake and dry-matter yield of Leucaena at all levels of soil solution P examined. The concentration of P required by nonmycorrhizal L. leucocephala for maximum yield was 27 to 38 times higher than that required by mycorrhizal L. leucocephala. The results illustrate the very high dependence of L. leucocephala on VAM fungi and the significance of optimizing soil solution phosphorus for enhancing the benefits of the VAM symbiosis.

Azolla filiculoides Nitrogenase Activity Decrease Induced by Inoculation with Chlamydomonas sp.

Experiments were conducted to determine the influence of Chlamydomonas sp. on nitrogen fixation (C(2)H(2) --> C(2)H(4)) in Azolla filiculoides and on the nitrogen fixation and growth of free-living Anabaena azollae 2B organisms. Inoculation of azolla medium with Chlamydomonas sp. was associated with decreased nitrogenase activity in A. filiculoides and with increases in the density of a fungal population identified as Acremonium sp. Subsequent inoculation of azolla medium with this fungus was also accompanied by a significant decrease in nitrogenase activity of A. filiculoides. However, the extent of depression of nitrogenase activity was significantly higher when azolla medium was inoculated with Chlamydomonas sp. than when it was inoculated with Acremonium sp. Inoculation of nitrogen-free Stanier medium with either Acremonium sp. or Chlamydomonas sp. did not adversely affect the growth or nitrogenase activity of free-living A. azollae. Decreased nitrogenase activity in A. filiculoides is apparently related to the adverse influence of the green alga and the fungus on the macrosymbiont. The mechanisms that might be involved are discussed.

Selective medium for recovering specific populations of rhizobia introduced into tropical soils.

Experiments were designed to evaluate the usefulness of antifungal agents and streptomycin for recovering low densities of rhizobia inoculated into tropical soils. The results showed that yeast-mannitol agar (pH 6.0) containing 500 mug of streptomycin, 400 mug of cycloheximide and 50 mug of benomyl or chlorothalonil per ml was the best selective medium.

Response of Sesbania grandiflora to Inoculation of Soil with Vesicular-Arbuscular Mycorrhizal Fungi.

A greenhouse experiment was conducted to determine the influence of two tropical isolates of Glomus fasciculatum and Glomus mosseae on the nutrient uptake and growth of Sesbania grandiflora. Inoculation of sterile soil with the fungi significantly improved growth and nutrient uptake by S. grandiflora, but the response of the legume was markedly better when the soil was inoculated with G. fasciculatum than when it was inoculated with G. mosseae. Nutrient uptake and growth of S. grandiflora in nonsterile soil was also significantly stimulated by inoculation, but the legume did not respond differently to the two endophytes under this condition.

Interaction of Rhizobium sp. with Toxin-Producing Fungus in Culture Medium and in a Tropical Soil.

Experiments were conducted to determine the influence of a toxin-producing fungus on a rhizobial population in yeast-mannitol medium and in a tropical soil. The fungus, which was isolated from a highly weathered soil (Tropeptic Eutrustox), was identified as a Metarhizum sp. The density of rhizobial populations established in yeast-mannitol medium in the absence of the fungus was 10 times higher than that established in its presence. However, the fungus did not exert similar antagonistic influence on the rhizobial population incubated with it in the sterilized test soil. Rhizobial growth activity in yeast-mannitol medium was also insensitive to the presence of the fungus when the medium was amended with 1% (wt/vol) kaolinite or montmorillonite. The results suggest that clay minerals may be responsible for protecting rhizobia against toxin-producing fungi in soil.

Effect of rice plants on nitrogenase activity of flooded soils.

In samples of flooded soil containing blue-green algae (cyanobacteria), the presence of rice plants did not influence the nitrogenase activity of the algae. Nitrogenase activity of heterotrophic bacteria was enhanced by the presence of rice plants, but this activity was not affected by changes in plant density. The rate of nitrogen fixation in the rhizosphere, however, varied significantly among the 16 rice varieties tested. A simple method was devised to test the nitrogen-fixing activity in the root zone of rice varieties, and data were obtained showing marked differences in the activities of the 16 varieties. In tests of two varieties with dissimilar rates of nitrogen fixation in their rhizospheres, the variety which had the greater root weight and lesser shoot weight and which supported greater methane formation had the greater nitrogenase activity.

Nitrogen fixation by photosynthetic bacteria in lowland rice culture.

Propanil (3',4'-dichloropropionanilide) was a potent inhibitor of the nitrogenase activity of blue-green algae (cyanobacteria) in flooded soil, but the herbicide at comparable concentrations was not toxic to rice, protozoa, and nitrogen-fixing bacteria. Ethanol-amended flooded soils treated with propanil exhibited higher rates of nitrogenase activity than those not treated with the herbicide. The enhanced nitrogenase activity in propanil-treated soils was associated with a rise in the population of purple sulfur bacteria, especially of cells resembling Chromatium and Thiospirillum. By employing propanil and a means of excluding light from the floodwater to prevent the development of phototrophs during rice growth under lowland conditions, the relative activities of blue-green algae, photosynthetic bacteria, and the rhizosphere microflora were determined. The results suggest that the potential contribution of photosynthetic bacteria may be quite high.

Further evidence for the regulation of bacterial populations in soil by protozoa.

After the addition to soil of large numbers of a cowpea Rhizobium strain, the population declined steadily until the numbers reached about 10(7)/g, and the protozoa rose to about 10(4)/g. When indigenous protozoa were suppressed by the addition of actidione to the soil, the density of the test rhizobium did not fall initially, but its abundance declined to about 10(7)/g when actidione-resistant protozoa arose in significant numbers. The addition to actidione-treated soil of an antibiotic-resistant strain of Paramecium led to a rapid decrease in the population of the rhizobium, the density reaching essentially the same value as in soil receiving neither the drug nor the paramecia. The same changes occurred with Xanthomonas campestris as test prey except that its numbers fell to about 10(5)/g of soil. These data provide further evidence for the key role of protozoa in controlling the abundance of populations of certain bacteria introduced into soil.

Protozoa as agents responsible for the decline of Xanthomonas campestris in soil.

A streptomycin-resistant mutant of Xanthomonas campestris was used to assess the persistence of the plant pathogen in soil and the changes in populations that might be important for its survival. In soil into which large numbers of the organism were introduced, a marked decline in its abundance occurred, but after about 1 week its population density reached a level of about 105 and did not continue to fall during the test period. No such marked decline was evident in sterile soil inoculated with X. campestris. The bacterium did not lose viability if starved for carbon or inorganic nitrogen. Although abundant in soil, the numbers of propagules capable of producing antibiotics or lytic enzymes active against X. campestris did not increase coincident with the pathogen's decline, and no increase in tartrate-extractable toxins was observed. Neither bdellovibrios nor bacteriophages active against the xanthomonad were found in the soil, but a marked increase in the frequency of protozoa paralleled the phase of rapid diminution in the X. campestris population. In actidione-treated soil, in which protozoan activity was severly limited, the high cell density of the pathogen was maintained. On the basis of these data, it is concluded that predation by protozoa is responsible for the abrupt fall in frequency of the bacterium in natural soil.