Fungi isolated from Sclerotia ofSclerotium cepivorum and from soil and their effects upon the pathogen

Summary Data are presented on the antagonistic effects of the fungi isolated from sclerotia ofSclerotium cepivorum and from nonrhizosphere soil taken from around the roots of infected onions upon mycelial growth and sclerotial germination ofS. cepivorum. Most of the isolated fungi especiallyPenicillium species were antagonistic to mycelial growth. Sclerotial germination was slightly inhibited by diffusates of these fungal isolates. Testing the antifungal effect of someAllium extracts against the fungal isolates by the inhibition zone method showed that garlic extract has the greatest antifungal effects and onion extract is the least potent. However, spore germination tests indicated that onion extract completely inhibits the spore germination of all test fungi. The role of host-plant extracts in stimulating sclerotial germination is discussed.     

Fungal microflora of seeds of Pisum sativum L. and its control

Five varieties ofPisum sativum L. were tested for seedborne fungi. Tests were conducted by standard International Seed Testing methods. It was observed that fungi likeAlternaria, Aspergillus, Rhizopus, Mucor andFusarium were associated with all the five varieties tested.Fusarium andRhizopus were dominant in all the varieties. They were also responsible for reduction in germination percentage of seeds. Early December was selected for testing the efficacy of certain fungicides (namely, Agrosan G.N., Ceresan, Copper carbonate, Tafasan, Tillex, Stardex and Sulphur) against seedborne fungi ofPisum sativum L., Agrosan G.N. and Ceresan gave the better control of seedborne pathogens without any adverse effect on germination.     

Antifungal activity of berberine iodide, a constituent ofFumaria indica

Berberine iodide, an isoquinoline alkaloid was isolated fromFumaria indica and its efficacy was tested against spore germination of some plant parasitic as well as saprophytic fungi. The alkaloid significantly curbed spore germination ofCurvularia lunata, Erysiphe cichoracearum, E. pisi, Fusarium udum andPenicillium species. Complete inhibition (100%) of spore germination was observed inE. cichoracearum andPenicillium species at 1.5 g/L.Albugo candida andAlternaria species were not affected by the chemical.     

The effect of volatile and gaseous metabolites of swelling seeds on germination of fungal spores

Effects of volatile and gaseous metabolites of swelling seeds of pea, bean, wheat, corn, cucumber, tomato, lentil, carrot, red pepper and lettuce on germination of spores of five genera of fungi were found to depend rather on the fungal than on the plant genus. Germination of spores ofBotrytis cinerea, Mucor racemosus andTrichoderma viride was most severely inhibited. Spores ofVerticillium dahliae were less sensitive and germination of spores ofFusarium oxysporum was inhibited only in two cases. On the other hand, exudates of pea and bean stimulated germination of spores ofFusarium oxysporum. Also spores ofTrichoderma viride germinated better in an atmosphere enriched with exuded metabolites of swelling lettuce seeds. When carbon dioxide produced by the swelling seeds was absorbed in potassium hydroxide, spores ofTrichoderma viride andVerticillium dahliae did not germinate at all, the inhibitory effects of volatile and gaseous exudates on germination of spores ofMucor racemosus were accentuated, and also the percentage of germinated spores ofFusarium oxysporum decreased. Germination of spores ofBotrytis cinerea was not influenced. Absorption of volatile and gaseous metabolites in a solution of potassium permanganate decreased in most cases their inhibitory effects, particularly inBotrytis cinerea.     

Co-occurrence of Arum- and Paris-type morphologies of arbuscular mycorrhizae in cucumber and tomato

Colonization by arbuscular mycorrhizal (AM) fungi was investigated in cucumber (Cucumis sativus), tomato (Lycopersicon esculentum) and Clethra barbinervis (Ericales) grown in field-collected soil known from previous studies to generate Paris-type arbuscular mycorrhizae in C. barbinervis. Spores of Paraglomus, Acaulospora, Glomus, and Gigaspora were found in the soil. Formation of hyphal coils and arbusculate coils of Paris-type mycorrhizae and of arbuscules of Arum-type mycorrhizae in roots raised in this soil in the growth chamber were compared with the detection of DNA of AM fungi from the same root systems using Glomales-specific primers. Only Paris-type mycorrhizae with extensive arbusculate coils developed in C. barbinervis, but cucumber and tomato developed both Paris- and Arum-types in the same root systems. Glomaceae and Archaeosporaceae and/or Paraglomaceae were detected strongly in the DNA from both cucumber and tomato roots, in which Arum-type mycorrhizae were observed. In contrast, DNA of Glomaceae was detected more sparingly in C. barbinervis, in which Paris-type mycorrhizae dominated. Acaulosporaceae and Gigasporaceae were strongly detected in the DNA from both C. barbinervis and tomato, whereas they were more weakly detected in cucumber. These results indicate that the morphology of colonization is strongly influenced by the selection of fungi to colonize the host plant from among those in the soil environment.     

Effect ofAgrobacterium radiobacter and its polysaccharide on emergence and damping-off of tomato plants

Agrobacterium radiobacter B6 and agrobacteran (an exopolysaccharide of the succinoglycan group) stimulated seed germination and tomato plant emergence. The germination was most stimulated by dipping the seeds in 0.1 % agrobacteran for 30 min whereas plant emergence in garden soil was best with 0.4 % agrobacteran at 10–20°C. Treatment of the plants withA.radiobacter cells and agrobacteran solution at 30–35°C. had a lower effect.A.radiobacter cells applied on seed surface protected the plants against damping off in garden soil artificially inoculated with the fungiRhizoctonia solani andPythium ultimum; in soils contaminated withFusarium solani 0.1 to 0.2% agrobacteran had a higher protective effect than the bacterization. The difference can be attributed to the varying density ofA.radiobacter population in plant rhizosphere in the presence of different plantpathogenic fungi, different interactions of microorganisms in the rhizosphere and different mode of penetration of the pathogens into plant roots.     

Inoculation of pines in a nursery withPisolithus tinctorius andThelephora terrestris in Korea

Summary Mycelial inocula of the two ectomycorrhizal fungi,Pisolithus tinctorius (Pt) andThelephora terrestris (Tt) were artificially introduced into fumigated nursery soil before seed sowing. At the end of the first growing season, Pt stimulated the height growth ofPinus densiflora, P. thunbergii, P. rigida, andP. rigida×taeda by 55, 36, 69, and 37%, respectively, compared with control seedlings (with no fumigation and no inoculation). When growth stimulation was expressed by dry weight, Pt increased dry weight ofP. densiflora andP. rigida×taeda by 143% and 128%, respectively, over control seedlings. Total absorption (per plant top) of nitrogen and phosphorus of inoculatedP. densiflora seedlings was also increased by 188 and 144%, respectively, even though the concentration of these two elements in the tissue was not significantly different from that of control seedlings.Tt failed to stimulate growth of the inoculated plants possibly due to its lower level of infection (40–60% mycorrhizal tips) compared with the relatively high infection (over 80%) of Pt.Pinus koraiensis did not respond to inoculation during the first growing season, probably due to its inherent characteristics of producing a small number of short roots and its slow shoot growth during the first few years after germination.This study was supported by the International Foundation for Science in Stockholm, Sweden.     

In vitro germination ofErythrorchis ochobiensis (Orchidaceae) in the presence ofLyophyllum shimeji, an ectomycorrhizal fungus

In vitro germination of a myco-heterotrophic orchid,Erythrorchis ochobiensis, was tested in the presence of ectomycorrhizal fungi,Lyophyllum shimeji andTricholoma fulvocastaneum. Lyophyllum shimeji stimulated the germination after incubation for 1.5 mo. Although most germinated seeds did not grow further after 3 mo, several seeds developed into small protocorms but showed amorphous profiles. Fungal mycelia were observed in the germinated seeds and protocorms, but pelotons were not detected. Since the seeds did not germinate axenically, it may be suggested that the fungus has the ability to stimulate germination.     

Studies on some fungi isolated from the rhizosphere of tomato plants and the consequent prospect for the control of Verticillium wilt

Summary Biological control of Verticillium wilt disease with antagonistic micro-organisms was studied. Antagonism of some fungi, isolated from tomato rhizosphere, toVerticillium albo-atrum R & B. was observedin vitro. A clearly defined zone, in which the growth of the pathogen was inhibited, was observed withPenicillium spp. (includingPenicillium chrysogenum Thom) andFusarium culmorum (S.G. Sm) Sacc., whileTrichoderma viride pers. ex Fries,Gliocladium spp. andPenicillium vermiculatum Dangeard, suppressed the growth ofV. albo-atrum by penetrating, and overgrowing it. OnlyT. viride andP. vermiculatum culture filtrate added to the Dox's agar, reduced the radial growth ofV. alboatrum. Root-dip application of culture filtrates ofT. viride andP. chrysogenum was found to be most effective in controlling the disease, followed by other species ofPenicillium andGliocladium spp. WhileFusarium culmorum provided no control. Improvement of plant height and vigour with a better yield due to culture filtrate treatment occurred. Root-dip application of antagonistic fungal propagules (T. viride, P. chrysogenum) to tomato seedlings was also very effective in controlling wilt in tomato plants grown inV. albo-atrum infested soil. Dedicated to the memory of the late Prof. Ivor Isaac with whom I had the pleasure of working     

Streptomyces plicatus as a model biocontrol agent

Three hundred and seventy two isolates belonging to the genusStreptomyces were isolated and screened for chitinase production.Streptomyces plicatus was found to be the best producer. The highest chitinase production were incubated for 3 d at 30 °C on buffered culture medium (pH 8.0) containing chitin plus sucrose and calcium nitrate as carbon and nitrogen sources.S. plicatus chitinase had a highly significant inhibitory effect on spore germination, germ tube elongation and radial growth ofFusarium oxysporum f.sp.lycopersict., Altrernaria alternata andVerticillium albo-atrum, the causal organisms ofFusarium wilt, stem canker andVerticillium wilt diseases of tomato. Application ofS. plicatus to the root system of tomato plants before transplantation markedly protected tomato plants against the tested phytopathogenic fungiin vivo.     

Effect of different west african species and strains ofArthrobotrys nematophagous fungi onMeloidogyne species

Different species ofArthrobotrys nematophagous fungi and several strains ofA. oligospora have been studied for their antagonistic effects against nematodes of the genusMeloidogyne, important pests of vegetables. All fungi trappedM. mayaguensis andM. incognita juvenilesin vitro but had no effect on the juveniles ofM. javanica. In pot experiments withM. mayaguensis, all fungi reduced the nematode populations and stimulated the growth of tomato seedlings. In a field trial, a strain of A.oligospora, isolated in Senegal and incorporated into compost blocks, was efficient in increasing the tomato seedling growth. The introduction of nematophagous fungi in compost blocks as a biological biocontrol technique against phytophagous nematodes adapted for developing countries is discussed.     

Interactions between<Emphasis Type="Italic"> Trichoderma pseudokoningii</Emphasis> strains and the arbuscular mycorrhizal fungi<Emphasis Type="Italic"> Glomus mosseae</Emphasis> and<Emphasis Type="Italic"> Gigaspora rosea</Emphasis>

The interaction between Trichoderma pseudokoningii (Rifai) 511, 2212, 741A, 741B and 453 and the arbuscular mycorrhizal fungi Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe BEG12 and Gigaspora rosea Nicolson & Schenck BEG9 were studied in vitro and in greenhouse experiments. All T. pseudokoningii strains inhibited the germination of G. mosseae and Gi. rosea except the strain 453, which did not affect the germination of Gi. rosea. Soluble exudates and volatile substances produced by all T. pseudokoningii strains inhibited the spore germination of G. mosseae. The germination of Gi. rosea spores was inhibited by the soluble exudates produced by T. pseudokoningii 2212 and 511, whereas T. pseudokoningii 714A and 714B inhibited the germination of Gi. rosea spores by the production of volatile substances. The strains of T. pseudokoningii did not affect dry matter and percentage of root length colonization of soybean inoculated with G. mosseae, except T. pseudokoningii 2212, which inhibited both parameters. However, all T. pseudokoningii strains decreased the shoot dry matter and the percentage of AM root length colonization of soybean inoculated with Gi. rosea. The saprotrophic fungi tested seem to affect AM colonization of root by effects on the presymbiotic phase of the AM fungi. No influence of AM fungi on the number of CFUs of T. pseudokoningii was found. The effect of saprotrophic fungi on AM fungal development and function varied with the strain of the saprotrophic species tested.     

Inhibition of fungi,actinomycetes and bacteria by Stachybotrys atra

The effect ofStachybotrys atra on fungi, actinomycetes and bacteria was studied in agar culture. Of the 27 genera and 52 species of fungi, 6 species of actinomycetes and 5 genera and 10 species of bacteria tested,S. atra was found to inhibit 95.7 % fungi, all the actinomycetes and 83.3 % of bacteria.Aspergillus ustus, 2 unidentified species ofFusarium, Rhizobium trifolii andPseudomonas spp. were however not inhibited. In experiments in soil,S. atra inhibited the growth ofTrichoderma viride. S atra was ineffective whereT. viride andS. atra were inoculated simultaneously or whereT. viride was added 7 days earlier. Besides,S. atra was not found to controlMacrophomina phaseoli infection on cotton. Soil culture ofS. atra showed phytotoxic effect on cotton seedlings.S. atra produced a dark brown to black thermostable toxic metabolite in the medium. The culture filtrate retarded the growth ofM. phaseoli in agar culture. Despite the importance ofS. atra as a fungistatic agent with wide antimicrobial spectrum, the use ofS. atra as a protectant against plant pathogens seems to be limited because of its phytotoxicity on cotton.     

Isolation of antifungal compounds from someZygophyllum species and their bioassay against two soil-borne plant pathogens

Roots and aerial shoots ofZygophyllum coccineum L.,Z. album L. andZ. dumosum Boiss were extracted with solvents of increasing polarity and tested against spore germination ofVerticellium albo-atrum andFusarium oxysporum f.sp.lycopersici. The effectiveness of each extract depends on the solvent, plant species, organ and test fungus. Petroleum ether extracts were ineffective as fungistatics while the methanol extracts possessed a high inhibitory effect toward spore germination suppression, depending on the fungus and plant species. Water or ethyl acetate root extracts ofZ. album caused a marked suppression of spore germination of the test fungi. Six saponins were isolated from the effective extracts on silica gel and Sephadex columns. They were identified by acid hydrolysis, alkaline saponification, IR and FAB MS. Quinovic acid was the dominant aglycone. Kaempferol was isolated from the flavonoid fraction ofZ. dumosum for the first time. Some of the isolated saponins and kaempferol provided successful control against the test plant pathogens under greenhouse experiment when used as seed treatment.     

Influence of colonisation by an arbuscular mycorrhizal fungus on the growth of seedlings of<Emphasis Type="Italic"> Banksia ericifolia</Emphasis> (Proteaceae)

Tap and primary lateral roots of seedlings of the putatively non-mycorrhizal Banksia ericifolia became marginally colonised when grown in an established mycelium of an arbuscular mycorrhizal (AM) fungus in the laboratory. A similar degree of colonisation was found in seedlings from an open woodland. All colonies lacked arbuscules. Two factors influencing colonisation and associated growth of host plants were examined experimentally: concentration of P in the soil and organic energy associated with the fungus. While some inoculated seedlings were slightly smaller when colonised by AM fungi, the results were inconsistent and never statistically significant. Seedlings take up insignificant quantities of soil P during early growth, even in the presence of abundant added P. Though colonisation was minor in all cases, an existing mycelium, whether or not connected to a companion plant, slightly increased the amount of root of B. ericifolia colonised by an AM fungus. All seedlings grew slowly. Shoots were significantly larger than roots, until the initiation of proteoid roots which commenced at about 40 days after germination, with both relatively high and low P supply.     

Biological species ofArmillaria and their mycoparasitic associations withRhodophyllus abortivus in Hokkaido

Specimens of basidiomes and/or rhizomorphs ofArmillaria mellea complex and basidiomes ofRhodophyllus abortivus, developing on the same decaying stumps or stems of forest trees, were collected in three forests in Hokkaido. Normal basidiomes ofR. abortivus were found near to, but free from, the rhizomorphs and/or basidiomes ofArmillaria, while abnormal basidiomes, as carpophoroid forms, were developed on the rhizomorphs ofArmillaria. Of three mycoparasiticArmillaria isolates found withR. abortivus, one was identified asA. gallica and two asA. jezoensis. The isolates ofR. abortivus showed excellent mycelial growth and rhizomorph formation on PDA. However, on MDA, RMDA and BMDA, they showed poor aerial mycelia growth and no rhizomorphs. In the contrapositional cultures, the growth ofA. gallica was completely inhibited byR. abortivus on PDA but only slightly inhibited on MDA and RMDA. On the other hand, mutual inhibition at a distance was observed on BMDA. The mycelial growth and rhizomorph formation inA. jezoensis were severely inhibited by the colony ofR. abortivus on PDA, but only slightly inhibited on MDA. On RMDA and BMDA, the colonies of twoArmillaria species andR. abortivus showed mutual inhibition at a distance and apparent rhizomorph formation by bothArmillaria species.     

Seasonal variation in allelopathic potential of<Emphasis Type="Italic">Artemisia princeps</Emphasis> var.<Emphasis Type="Italic">orientalis</Emphasis> on plants and microbes

We investigated seasonal variations in allelopathic potential ofArtemisia princeps var.orientalis. Aqueous and meth-anol extracts and volatile substances were prepared in the laboratory from samples collected monthly (April through October). Their impacts were then assessed on the germination and seedling growth ofLactuca sativa andAchyranthes japonica. The allelopathic potential varied with the time of sample collection and the concentration tested. For example, germination ofL. sativa was not inhibited by the aqueous extract but seedling growth (shoots and roots) was, with its seasonal effect being significant. ForA. japonica, seed germination was not inhibited at lower concentrations (except for August samples). However, at higher concentrations and in certain months (especially July), germination was more negatively affected. The degree of seedling growth inhibition also differed by month and by extract concentration, with roots being impacted more than shoots. Volatile substances also had a time-dependent influence on the germination and seedling elongation ofA. japonica. In a separate experiment, the ethyl-acetate and water fractions of a crude methanol extract were prepared monthly fromA. princeps var.orientalis. Here, we examined their antimicrobial activities against three gram-positive bacteria (Bacillus cereus, Bacillus subtilis, andStaphylococcus aureus), two gramnegative bacteria (Escherichia coli andPseudomonas fluorescens), and one lactic acid bacterium,Lactobacillus plantar urn. The ethyl-acetate fraction that was sampled in September was remarkably potent againstB. cereus andB. subtilis, whereas the water fraction collected in August and September showed great antimicrobial activity against the grampositive and -negative bacteria. In contrast,L. plantarum was not inhibited by the water fraction, regardless of the sampling month. Likewise, the ethyl-acetate and water fractions collected in April and October had the lowest levels of antimicrobial activity.     

Comparative structural study ofQuercus serrata andQ. acutissima formed byPisolithus tinctorius andHebeloma cylindrosporum

Ectomycorrhizas were synthesized in pots and growth pouches betweenQuercus serrata, Q. acutissima, and two ectomycorrhizal fungi,Pisolithus tinctorius andHebeloma cylindrosporum. Root morphology and the structure of the mantle and Hartig net were compared using light, fluorescence, scanning and transmission electron microscopy.P. tinctorius initially colonized root cap cells, and eventually produced a highly branched lateral root system with a complete mantle, whereasH. cylindrosporum promoted root elongation with few hyphae on the root apex surface indicating that interaction between roots differs with fungal species. Hartig net structure and hyphal inclusions varied between all the combinations tested. There were structural differences between mycorrhizas ofH. cylindrosporum/Q. acutissima grown in soil and growth pouches, which indicate that the growth pouch environment can induce artefacts in roots. Fruit bodies ofH. cylindrosporum developed in pots withQ. acutissima. AlthoughP. tinctorius has been used to inoculate oak seedlings in the nursery, results of this study indicate thatH. cylindrosporum may also be an effective ectomycorrhizal fungus forQ. serrata andQ. acutissima.     

Individual and combined effects of berberine and santonin on spore germination of some fungi

Berberine and santonin were isolated from rhizomes ofBerberis aristata and unexpanded flower buds ofArtemisia maritima, respectively. Efficacy of these two chemicals individually as well as of their mixtures, was tested against spore germination of some saprophytic and obligate fungi. While berberine individually was effective against most of the fungi,Helminthosporium spp. were least affected even at the highest dose (1500 ppm). Santonin was equally effective against several fungi. Mixture of both alkaloids found to be more effective than individual ones. Keeping the dose of berberine constant and santonin at two different concentrations (viz. 250 and 500 ppm) the spore germination ofHelminthosporium, oryzae was stimulated. Increasing concentration of santonin inhibited the spore germination of all other fungi tested,Colletotrichum capsici being affected only by 20 and 5% (at berberine concentration of 250 and 500 ppm, respectively). On the other hand, santonin being constant and berberine at different concentrations, the mixture was effective against all the fungi.     

Studies on the fungus flora of three grains in Egypt

In wheat, corn and sorghum grains twenty-six genera and seventy-seven species including sixteen ofAspergillus and twenty-one ofPenicillium were identified.In grain samples adjusted to various moisture levels (up to 18.8 % on a dry-weight basis) and stored for 4 months at 8, 20 and 30 C seed-borne fungi were monthly identified and counted, and the germinability of the grains was tested. In the control samples (moisture content 7.1–8.2 %) temperature had no effect on the fungal counts and on the precentage germination. With the rise of temperature and moisture content the fungal counts markedly increased and the germinability declined.The list of fungi dominant in one or more of the experiments, included eight species ofAspergillus, six ofPenicillium, two ofFusarium and one each ofAlternaria andCurvularia. The order of dominance of these fungi varied according to the type of grain, the length of storage and the treatment.A. niger was the dominant organism in the control samples of the three grains. It could grow successfully at high moisture contents but above 15 %, it was usually overgrown by other fungi. In wheatP. citrinum andA. sydowii, in cornP. citrinum andA. terreus, and in sorghumA. terreus andA. niger were the dominant fungi at moisture contents above 15 % after four months storage at 30 C. When stored at 8 CPenicillium species tended to predominate over other fungi in grains with high moisture contents.In surface-sterilized grains adjusted to 15 % moisture content, inoculated with six dominant fungi separately and incubated at 30 C,A. niger, P. citrinum andP. variabile caused severe deterioration in the three grains;A. ochraceus in wheat only andF. moniliforme in wheat and corn;C. spicifer was slightly injurious to the three grains.