Processed Manure as Carrier to Introduce Trichoderma harzianum: Population Dynamics and Biocontrol Effect on Rhizoctonia solani

Manure pellets produced from processed swine faeces can be used as carrier material for the biocontrol fungus Trichoderma harzianum. The antagonist can grow and sporulate on the processed manure powder as the sole source of carbon and nutrients. The incorporation of conidia in pellets of the processed manure was shown to be feasible on a laboratory scale. Survival of the fungus in the pellets during storage was satisfactory. The population dynamics of T. harzianum were studied using a benomyl-resistance marker after introduction of conidia into soil. The antagonist could colonize and spread through a number of non-sterile soils and was able to establish a stable population over a period exceeding 125 days. Under sterile conditions, the propagation of T. harzianum in soil was much greater than under non-sterile conditions. The incorporation of antagonist conidia in pellets was found to be essential for the successful colonization of non-sterile soil. In growth chamber experiments, application of T. harzianum via processed manure pellets reduced damping-off of sugar beet seedlings caused by Rhizoctonia solani in artificially and naturally infested soil. In artificially infested soil, T. harzianum reduced the population of R. solani and protected beet seedlings from damping-off 3 weeks after introduction. The application of T. harzianum to naturally infested soil increased the number of healthy beet seedlings more than two-fold.     

Growth of Arthrobotrys superba from a birch wood resource base into soil determined by radioactive tracing

The ability of a nematode-trapping fungus to establish in field soil is an important characteristic when considering its use as a biological control agent. The outgrowth of the nematode-trapping fungus Arthrobotrys superba from wood was recorded by labelling the fungus with [(14)C]3-O-methylglucose and [(32)P]orthophosphoric acid and by using the soil sprinkling method. The fungus reached a distance of 7-8 cm during 25 days in heat-treated (60 degrees C) soil, detected by either radioactive tracing or the soil sprinkling technique. The two labelled compounds were co-distributed at all sampling times (r(2)=0.946) which indicates that the glucose pool (as methylglucose) and phosphorus content were correlated throughout the mycelium. In natural, non-heat-treated soil the fungus reached a distance of 1.5 cm from one disc of birch wood after 30 days, while it reached 3.2 cm during the same period when the food base was a pile of five inoculated discs. The experiments showed, for the first time, that a nematophagous fungus, A. superba, can grow out into soil from a piece of wood and supported by nutrients translocated from the resource base to the edge of the mycelium.     

Environmental factors and growth of Histoplasma capsulatum in soil

Summary Environmental factors influencing the growth ofHistoplasma capsulatum in soil have been studied. The role of temperature and moisture in the growth of the fungus was found to be critical. The fungus can tolerate very low temperatures, if the soil moisture content is high, but cannot withstand temperatures of 40° C or above for an extended period.Dry, sterile chicken manure and an extract of unsterile chicken manure showed an inhibitory effect on the growth of the fungus. However, the relationship between bird manure andH. capsulatum has not been satisfactorily clarified.Morphological studies ofH. capsulatum in soil showed that the fungus grows within the upper two inches of the soil and a majority of sporulation occurs within the upper one-half inch of soil. Morphological studies of the fungus, in the presence of chicken manure and chicken manure extract, showed an increased number of macroconidia and microconidia and decreased mycelial production.The growth ofH. capsulatum in soil is markedly affected by soil pH above 10 and below 5. No growth was observed on soil cultures outside these values. There was no definite pH range within these values in which the fungus grew more abundantly.     

Phialophora graminicola, a dark septate fungus, is a beneficial associate of the grass Vulpia ciliata ssp. ambigua

The influence of three organic compounds and bakers' dry yeast on growth of external mycelium and phosphorus uptake of the arbuscular mycorrhizal fungus Glomus intraradices Schenck & Smith (BEG 87) was examined. Two experiments were carried out in compartmentalized growth systems with root-free sand or soil compartments. The sand and soil in the root-free compartments were left untreated or uniformly mixed with one of the following substrates (0.5 mg g⁻¹ soil): bakers' dry yeast, bovine serum albumin, starch or cellulose. Effects of the organic substrates on biomass and hyphal length density of the arbuscular mycorrhizal fungus were examined by using specific fatty acid signatures in combination with direct microscopy. Micro-organisms other than the arbuscular mycorrhizal fungus were measured by fatty acid signatures, and radioactive ³³P labelling of the root-free soil was used to determine arbuscular mycorrhizal hyphal phosphorus uptake. In general, hyphal growth of G. intraradices was enhanced by yeast and bovine serum albumin, whereas the carbon sources, starch and cellulose, depressed fungal growth. By analysing the fatty acid 16:1ω5 from phospholipids (indicating mycelium) and neutral lipids (indicating storage structures) it was shown that increased fungal growth due to yeast was mainly in vegetative hyphae and less in storage structures. Arbuscular mycorrhizal hyphal phosphorus uptake was decreased by cellulose, but unaffected by the other substrates compared with the control. This means that both growth and phosphorus transport by the arbuscular mycorrhizal fungus were decreased under cellulose treatment. However, the composition of the microbial community varied under different substrate conditions indicating a possible interactive component with arbuscular mycorrhizal hyphal growth and phosphorus uptake.     

STUDIES ON SYSTEMIC FUNGICIDES. I. FUNGICIDAL PROPERTIES OF THE ARYLOXYALKYLCARBOXYLIC ACIDS

A method is described for assessing the systemic activity of compounds in checking the infection of broad bean ( Vicia faba ) by the fungi Botrytis cinerea or B. fabae. Treatment consisted in allowing the roots of young seedlings to stand in a solution containing 10 p.p.m. of the chemical for 2–3 weeks. The plants, together with controls, were then inoculated and when symptoms had had time to develop, the degree of chocolate spot infection was assessed. Several methods of disease assessment were examined and are critically discussed.
Certain phenoxyalkylcarboxylic acids tested by this method consistently gave a reduction in the mean size of fungus lesions on the bean leaves, clearly indicating systemic fungicidal action. The most promising substances were 2:4:6-trichlorophenoxyacetic, pentachlorophenoxyacetic and pentachlorophenoxy iso butyric acids. Further experiments with these compounds involving soil treatment, stem injection and spray application are described, and in most cases systemic fungicidal activity was clearly demonstrated. Certain compounds caused visible damage to the plants or resulted in a reduction in growth.
The results presented indicate that phenoxy acids can be fairly readily translocated in bean plants and that they tend to accumulate in actively growing tissues. It is considered unlikely, however, that they persist for long periods in plant tissue.
In the soil, the compounds appeared to remain effective for a considerable time, particularly the less soluble pentachloro acids, suggesting that soil application might provide a safe and useful method of treatment.     

The Exotic Legume Tree Species Acacia holosericea Alters Microbial Soil Functionalities and the Structure of the Arbuscular Mycorrhizal Community

The response of microbial functional diversity as well as its resistance to stress or disturbances caused by the introduction of an exotic tree species, Acacia holosericea, ectomycorrhized or not with Pisolithus albus, was examined. The results show that this ectomycorrhizal fungus promotes drastically the growth of this fast-growing tree species in field conditions after 7 years of plantation. Compared to the crop soil surrounding the A. holosericea plantation, this exotic tree species, associated or not with the ectomycorrhizal symbiont, induced strong modifications in soil microbial functionalities (assessed by measuring the patterns of in situ catabolic potential of microbial communities) and reduced soil resistance in response to increasing stress or disturbance (salinity, temperature, and freeze-thaw and wet-dry cycles). In addition, A. holosericea strongly modified the structure of arbuscular mycorrhizal fungus communities. These results show clearly that exotic plants may be responsible for important changes in soil microbiota affecting the structure and functions of microbial communities.     

Influence of cadmium and mercury on activities of ligninolytic enzymes and degradation of polycyclic aromatic hydrocarbons by Pleurotus ostreatus in soil

The white-rot fungus Pleurotus ostreatus was able to degrade the polycyclic aromatic hydrocarbons (PAHs) benzo[a]anthracene, chrysene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenzo[a,h]anthracene, and benzo[ghi]perylene in nonsterile soil both in the presence and in the absence of cadmium and mercury. During 15 weeks of incubation, recovery of individual compounds was 16 to 69% in soil without additional metal. While soil microflora contributed mostly to degradation of pyrene (82%) and benzo[a]anthracene (41%), the fungus enhanced the disappearance of less-soluble polycyclic aromatic compounds containing five or six aromatic rings. Although the heavy metals in the soil affected the activity of ligninolytic enzymes produced by the fungus (laccase and Mn-dependent peroxidase), no decrease in PAH degradation was found in soil containing Cd or Hg at 10 to 100 ppm. In the presence of cadmium at 500 ppm in soil, degradation of PAHs by soil microflora was not affected whereas the contribution of fungus was negligible, probably due to the absence of Mn-dependent peroxidase activity. In the presence of Hg at 50 to 100 ppm or Cd at 100 to 500 ppm, the extent of soil colonization by the fungus was limited.     

Effect of inoculation method and plant growth medium on endophytic colonization of sorghum by the entomopathogenic fungus <Emphasis Type="Italic">Beauveria bassiana</Emphasis>

A study was conducted to determine the effect of inoculation method and plant growth medium on colonization of sorghum by an endophytic Beauveria bassiana. Colonization of leaves, stems, and roots by B. bassiana was assessed 20-days after application of the fungus. Although B. bassiana established as an endophyte in sorghum leaves, stems, and roots regardless of inoculation method (leaf, seed, or soil inoculation), plant growth medium (sterile soil, non-sterile soil, or vermiculite) apparently influenced colonization rates. Seed inoculation with conidia caused no stem or leaf colonization by the fungus in non-sterile soil but did result in substantial endophytic colonization in vermiculite and sterile soil. Leaf inoculation did not result in root colonization, regardless of plant growth medium. Endophytic colonization was greater in leaves and stems than roots. Endophytic colonization by B. bassiana had no adverse effects on the growth of sorghum plants. Leaf inoculation with a conidial suspension proved to be the best method to introduce B. bassiana into sorghum leaves for plants growing in either sterile or non-sterile soil. Further research should focus on the virulence of endophytic B. bassiana against sorghum stem borers.     

Biologically available versus exchangeable magnesium in soil

Summary Equal weight of dry mycelium of the fungusAspergillus niger was found equivalent with equal magnesium uptake from synthetic nutrient (as standard series) and from solutions with soil as magnesium source. The quantities of magnesium remaining in the nutrient solutions after growth were also the same in cultures with or without addition of soil. It may be concluded that the method employed will give a correct picture of the ability of the fungus to release magnesium from the soil.During growth the fungus releases magnesium in excess of the maximum content of magnesium exchangeable with ammonium ions. This excess was found to be derived mainly from the clay fraction of the soil, although the silt fraction gives a considerable supplement. This is emphasized by the results from up to five repeated cultivations of the fungus in the same soil or its clay-/silt-fractions.Microbiologically available magnesium was determined after removal of the exchangeable fraction in 13 soils after three years of exhaustion by crop plants. The results showed a marked correlation (r=0.90) between the magnesium uptake by the fungus and the higher plant, and in addition the actual figures for magnesium uptake were on a comparable magnitude. Analysis of the soils after the third year of experiment showed that the fraction of magnesium available toAspergillus niger was not exhausted but only reduced to approximately one-half of the original content.     

Biosynthesis and beneficial effects of microbial gibberellins on crops for sustainable agriculture

Soil microbes promote plant growth through several mechanisms such as secretion of chemical compounds including plant growth hormones. Among the phytohormones, auxins, ethylene, cytokinins, abscisic acid and gibberellins are the best understood compounds. Gibberellins were first isolated in 1935 from the fungus Gibberella fujikuroi and are synthesized by several soil microbes. The effect of gibberellins on plant growth and development has been studied, as has the biosynthesis pathways, enzymes, genes and their regulation. This review revisits the history of gibberellin research highlighting microbial gibberellins and their effects on plant health with an emphasis on the early discoveries and current advances that can find vital applications in agricultural practices.     

Secondary metabolites produced by a root‐inhabiting sterile fungus antagonistic towards pathogenic fungi

Aims: A sterile red fungus (SRF) isolated from cortices of roots of both wheat (Triticum aestivum cv. Gamenya) and ryegrass (Lolium rigidum cv. Wimmera) was found to protect the hosts from phytopathogens and promote plant growth. In this work, the major secondary metabolites produced by this SRF were analysed, and their antibiotic and plant‐growth‐promoting activities investigated. Methods and Results: Two main compounds, veratryl alcohol (VA) and 4‐(hydroxymethyl)‐quinoline, were isolated from the culture filtrate of the fungus. In antifungal assays, VA inhibited the growth of Sclerotinia sclerotiorum and Pythium irregulare even at low amounts, while high concentrations (>100 μg per plug) of 4‐(hydroxymethyl)‐quinoline were needed. Both metabolites revealed weak inhibition of Rhizoctonia solani. Furthermore, both compounds showed a growth promotion activity on canola (Brassica napus) seedlings used as bioassays. Conclusions: Isolation and characterization of the main secondary metabolites from culture filtrates of a root‐inhabiting sterile fungus are reported. The biological assays indicate that these secondary metabolites may have a role in both plant growth regulation and antifungal activity. Significance and Impact of the Study: This study provides a better understanding of the metabolism of a cortical fungus that may have a useful role in the biological suppression of root‐infecting soil‐borne plant pathogens.     

Application of Aspergillus niger-treated agrowaste residue and Glomus mosseae for improving growth and nutrition of Trifolium repens in a Cd-contaminated soil

The microbial transformation of sugar beet (SB) agrowaste with or without rock-phosphate (RP) has utility for the improvement of plant growth in a Cd (5 microg g-1) artificially contaminated soil, particularly when the soil is co-inoculated with arbuscular mycorrhizal (AM) fungus Glomus mosseae isolated from a Cd-polluted area. Under such Cd-polluted conditions, the limited growth, mineral nutrition, symbiotic developments (nodulation and AM-colonization) and soil enzymatic activities were stimulated using SB or SB+RP as soil amendments and G. mosseae as inoculant. G. mosseae enhanced plant establishment in a higher extent in amended soil; it is probably due to the interactive effect increasing the potential fertility of such compounds and its ability for decreasing Cd transfer from soil to plant. The amount of Cd transferred from soil solution to biomass of AM-colonized plants ranged from 0.09 microg Cd g-1 (in SB+RP-amended soil) to 0.6 microg Cd g-1 (in non-amended soil). Nodule formation was more sensitive to Cd than AM-colonization, and both symbioses were stimulated in amended soils. Not only AM-colonization but also amendments were critical for plant growth and nutrition in Cd-polluted soil. The high effectiveness of AM inoculum increasing nutrients and decreasing Cd in amended soil indicated the positive interaction of these treatments in increasing plant tolerance to Cd contamination.     

Production of phleichrome by Cladosporium phlei as stimulated by diketopiperadines of Epichloe typhina

Epichloe typhina is an endophytic fungus, while Cladosporium phlei is a pathogenic fungus of the timothy plant (Phleum pretense L.). We found two activities in the culture filtrate of E. typhina: one stimulated the pathogenic fungus, C. phlei, to produce phleichrome and the other inhibited its growth. The active ingredients that stimulated the production of phleichrome and inhibited the growth of C. phlei were isolated and characterized. The isolated compounds were identified as cyclo-(L-Pro-L-Leu) and cyclo-(L-Pro-L-Phe), which were stimulatory compounds, and p-hydroxybenzaldehyde, which was the growth inhibitory compound, based on an analysis of their spectral data. Of the two stimulatory compounds, cyclo-(L-Pro-L-Phe) showed higher activity. However, when 500 microg of cyclo-(L-Pro-L-Phe) was spotted on the TLC plate for bio-autography, a growth inhibitory zone was identified in the central red region, which contained phleichrome. On the other hand, phleichrome showed antifungal activity against E. typhina in the light, so it is assumed that there might be antagonism between the endophytic fungus, E. typhina, and the pathogenic fungus, C. phlei.     

Acibenzolar-S-methyl-Induced Systemic Resistance against Anthracnose and Powdery Mildew Diseases on Cucumber Plants without Accumulation of Phytoalexins

In a study to elucidate the possible involvement of phytoalexins in acibenzolar-S-methyl (ASM)-induced systemic resistance in cucumber plants ( Cucumis sativus L.), the phenolic compounds were extracted from ASM-treated and inoculated plants and compared with those from Milsana^®-treated plants previously reported to accumulate phytoalexins in cucumber. The glycoside-linked phenolic compounds from cucumber leaves were tested for their antifungal activity to the growth of pathogens which were most effective against the cucumber anthracnose fungus Colletotrichum orbiculare , followed by the scab fungus Cladosporium cucumerinum but ineffective against the Corynespora leaf spot fungus Corynespora cassiicola . Nevertheless, the accumulations of active compounds appeared to increase with the growth stages of cucumber plants irrespective of ASM treatment. In ASM-pretreated cucumber plants either inoculated with anthracnose or the powdery mildew fungus, there was no increase in phytoalexin-like phenolics.     

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 10⁵ 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.     

Quantification in soil and the rhizosphere of the nematophagous fungus Verticillium chlamydosporium by competitive PCR and comparison with selective plating

A competitive PCR (cPCR) assay was developed to quantify the nematophagous fungus Verticillium chlamydosporium in soil. A gamma-irradiated soil was seeded with different numbers of chlamydospores from V. chlamydosporium isolate 10, and samples were obtained at time intervals of up to 8 weeks. Samples were analyzed by cPCR and by plating onto a semiselective medium. The results suggested that saprophytic V. chlamydosporium growth did occur in soil and that the two methods detected different phases of growth. The first stage of growth, DNA replication, was demonstrated by the rapid increase in cPCR estimates, and the presumed carrying capacity (PCC) of the soil was reached after only 1 week of incubation. The second stage, an increase in fungal propagules presumably due to cell division, sporulation, and hyphal fragmentation, was indicated by a less rapid increase in CFU, and 3 weeks was required to reach the PCC. Experiments with field soil revealed that saprophytic fungal growth was limited, presumably due to competition from the indigenous soil microflora, and that the PCR results were less variable than the equivalent plate count results. In addition, the limit of detection of V. chlamydosporium in field soil was lower than that in gamma-irradiated soil, suggesting that there was a background population of the fungus in the field, although the level was below the limit of detection. Tomatoes were infected with the root knot nematode (RKN) or the potato cyst nematode (PCN) along with a PCN-derived isolate of the fungus (V. chlamydosporium isolate Jersey). Increases in fungal growth were observed in the rhizosphere of PCN-infested plants but not in the rhizosphere of RKN-infested plants after 14 weeks using cPCR. In this paper we describe for the first time PCR-based quantification of a fungal biological control agent for nematodes in soil and the rhizosphere, and we provide evidence for nematode host specificity that is highly relevant to the biological control efficacy of this fungus.     

Growth, respiration and nutrient acquisition by the arbuscular mycorrhizal fungus Glomus mosseae and its host plant Plantago lanceolata in cooled soil

Although plant phosphate uptake is reduced by low soil temperature, arbuscular mycorrhizal (AM) fungi are responsible for P uptake in many plants. We investigated growth and carbon allocation of the AM fungus Glomus mosseae and a host plant (Plantago lanceolata) under reduced soil temperature. Plants were grown in compartmented microcosm units to determine the impact on both fungus and roots of a constant 2.7 °C reduction in soil temperature for 16 d. C allocation was measured using two (13)CO(2) pulse labels. Although root growth was reduced by cooling, AM colonization, growth and respiration of the extraradical mycelium (ERM) and allocation of assimilated (13)C to the ERM were all unaffected; the frequency of arbuscules increased. In contrast, root respiration and (13)C content and plant P and Zn content were all reduced by cooling. Cooling had less effect on N and K, and none on Ca and Mg content. The AM fungus G. mosseae was more able to sustain activity in cooled soil than were the roots of P. lanceolata, and so enhanced plant P content under a realistic degree of soil cooling that reduced plant growth. AM fungi may therefore be an effective means to promote plant nutrition under low soil temperatures.     

Influence of soil organic matter decomposition on arbuscular mycorrhizal fungi in terms of asymbiotic hyphal growth and root colonization

Soil organic matter is known to influence arbuscular mycorrhizal (AM) fungi, but limited information is available on the chemical components in the organic matter causing these effects. We studied the influence of decomposing organic matter (pure cellulose and alfalfa shoot and root material) on AM fungi after 30, 100, and 300 days of decomposition in nonsterile soil with and without addition of mineral N and P. Decomposing organic matter affected maize root length colonized by the AM fungus Glomus claroideum in a similar manner as other plant growth parameters. Colonized root length was slightly increased by both nitrogen and phosphorus application and plant materials, but not by application of cellulose. In vitro hyphal growth of Glomus intraradices was increased by soil extracts from the treatments with all types of organic materials independently of mineral N and P application. Pyrolysis of soil samples from the different decomposition treatments revealed in total 266 recognizable organic compounds and in vitro hyphal growth of G. intraradices in soil extract positively correlated with 33 of these compounds. The strongest correlation was found with 3,4,5-trimethoxybenzoic acid methyl ester. This compound is a typical product of pyrolysis of phenolic compounds produced by angiosperm woody plants, but in our experiment, it was produced mainly from cellulose by some components of the soil microflora. In conclusion, our results indicate that mycelia of AM fungi are influenced by organic matter decomposition both via compounds released during the decomposition process and also by secondary metabolites produced by microorganisms involved in organic matter decomposition.     

Behaviour of the fungus Rhizoctonia solani Kiihn in the soil

Rhizoctonia solani was found able to grow as a saprophyte through natural unsterilized soil. Its rate of growth under different soil conditions in glass tumblers was studied by the Rossi-Cholodny soil-plate method. Growth was most rapid at the lowest soil-moisture content tested, viz. 30 % saturation, and was accelerated by forced aeration of the soil. The maximum distance to which mycelial growth could be supported on the food reserves of the agar inoculum alone was some 5 cm., as shown by extent of growth through tubes of moist sand, but in 23 days the fungus grew 21–24 cm through tubes of soil. Removal of the agar disk 2 days after inoculation of the tubes reduced growth through sand by more than half, but through soil by only a small proportion. In soil, Rhizoctonia was able to cause 100% damping-off of radish seedlings planted at a radial distance of 4 cm. from the agar inoculum, and some 40 % damping-off at a distance of 9 cm. The depressing effect of additions of 1 % ground-wheat straw or dried grass to the soil upon growth of the fungus was attributed to (1) the negligible cellulose-decomposing ability of Rhizoctonia, (2) nitrogen starvation of the mycelium, through rapid utilization of the available soil nitrogen by the cellulose-decomposing micro-organisms multiplying upon the fresh organic material, (3) fungistatic action on Rhizoctonia of the respiratory carbon dioxide produced by the cellulose-decomposers.     

The Saprophytic Status of Fusarium oxysporum Schl: Causing Vascular Wilt of Oil Palm

The droplet plating method described, especially when used inconjunction with a baiting technique, facilitates the demonstrationof Fusarium oxysporum in soil. This fungus was present in soilfrom wilt-free areas of oil palm plantations as well as in soilabout wilt-diseased palms. Hyphal fusions between soil isolatesand isolates from wilted palms could be demonstrated. The funguspersisted for periods of at least ¹ year in naturally infestedsoils under a variety of moisture conditions; it also survivedin inoculated alien soil forms of Ieast ¹year. Soil forms ofthe fungus had a high competitive saprophytic ability, competitivelycolonizing sterilized soils and sand, and organic materialsadded to soils and mixed cultures. The pathogenic isolate, aswell as the soil isolates, exhibited characters belonging tosoil-inhabiting fungi, namely continued persistance in soil:tolerance, in respect of growth and reproduction, to antagonism:and the ability competitively to colonize dead organic materialin soils. The pathogenic form is considered to be a soil inhabitant.