Fungus Rhizoctonia Solani Research Articles

Dolichin, a New Chitinase-like Antifungal Protein Isolated from Field Beans (Dolichos lablab)

An antifungal protein, possessing a molecular weight of 28 kDa and an N-terminal sequence resembling chitinases, has been purified from the seeds of the field bean Dolichos lablab. The procedure involved extraction with aqueous buffer, affinity chromatography on Affi-gel blue gel, and ion exchange chromatography on CM-Sepharose. The protein, designated dolichin, exhibited antifungal activity against the fungi Fusarium oxysporum, Rhizoctonia solani, and Coprinus comatus. Dolichin was capable of inhibiting human immunodeficiency virus (HIV) reverse transcriptase and α- and β-glucosidases which are glycohydrolases implicated in HIV infection. It had very low ribonuclease and cell-free translation-inhibitory activities.

Biotransformation of the phytoalexin camalexin by the phytopathogen Rhizoctonia solani

The unusual metabolism of the cruciferous phytoalexin camalexin by virulent and weakly virulent isolates of the root rot fungus Rhizoctonia solani Kuhn is reported. This biotransformation proceeded via 5-hydroxycamalexin, which was further biotransformed into more polar metabolites. Importantly, the metabolites resulting from transformation of camalexin were significantly less toxic to the pathogen than camalexin. Thus, it was concluded that R. solani can detoxify camalexin through oxidation of the indole ring. The chemistry involved in the structure determination of the intermediates of this pathway, their synthesis as well as antifungal activity is described.

Effects of culture parameters on the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4-dichlorophenol (2,4-DCP) by selected fungi

In order to enhance 2,4-D and 2,4-DCP degradation by four selected fungi ( Cunningliamella elegans, C. echinnlata, Rhizoctonia solani and Verticillimn lecanii), three culture parameters (initial chemical concentration, amounts of glucose and nitrogen) were varied. The levels of both xenobiotics in the culture media were monitored by HPLC analysis after five days of cultivation. The best results were obtained at low initial concentration (20 mg.L −1) vs 100) and with low amounts of glucose (5 g.L −1 vs 10) and nitrogen (2.4 mM vs 24). When these two elements were lacking from the culture media, biodegradation was not suppressed. but took place to a lesser extent. Thus, initial chemical concentration and amounts of carbon and nitrogen, in the culture medium, were shown to strongly influence the extent of 2,4-D and 2,4-DCP removal by fungi.

Continuity of air-filled pores and invasion thresholds for a soil-borne fungal plant pathogen, Rhizoctonia solani

Fundamental knowledge of the way fungi explore the pore volume within a soil is crucial if we are to understand how soil physical conditions affect population dynamics and invasion of many important fungal parasites and saprophytes within soil. In this study, spread of the fungus Rhizoctonia solani Kühn (AG4), an economically important soil-borne plant pathogen and saprophyte, was quantified in relation to tortuosity and continuity of the air-filled pore space. Samples containing fine or coarse sand were equilibrated at matric potentials ranging from −1 to −7 kPa and inoculated with R. solani. We quantified the colony extent and biomass of R. solani spreading from a localized source of inoculum using microscopy and a monoclonal antibody-based immunosorbent assay. Air permeability rapidly increased when the air-filled pore space became continuous below a threshold matric potential of −2.0 kPa for the fine sand and −1.0 kPa for the coarse sand. Fungal spread was limited at high matric potentials with a colony extent of 6.6 and 4.6 mm for the fine and coarse sand, respectively, but increased to 32.6 and 28.5 mm, with a sharp transition below a threshold matric potential of −2.2 and −1.3 kPa. The average colony biomass dropped markedly at the same threshold matric potential. We conclude that the ability of the fungus to invade soil depends on the connectivity and tortuosity of the air-filled pore volume. At near-saturated conditions, the spread was spatially constrained to a relatively small volume, forming small dense colonies. In a well connected air-filled pore volume, larger colonies with low biomass density were formed. The broader implications for invasion of soil by a fungus and for transmission of fungal diseases are discussed.

Cell wall alterations in hypocotyls of bean seedlings protected from Rhizoctonia stem canker by a binucleate Rhizoctonia isolate

The influence exerted by the non-pathogenic binucleate Rhizoctonia (np-BNR) isolate 232-CG in stimulating plant defence reactions in young bean plants inoculated with the root rot fungus Rhizoctonia solani (AG-4) was examined using light and electron microscopy and further investigated by gold cytochemistry. Severe necrotic lesions on hypocotyles of diseased beans were observed, and the pathogen invaded the cortical tissue causing extensive damage including cell disorganization and cell wall degradation. In contrast, these host reactions were not seen in bean plants inoculated with the non-pathogenic BNR or in plants that were inoculated with BNR and subsequently challenge-inoculated with R. solani. Microscopic examination of hypocotyls inoculated with the non-pathogenic BNR, showed a different host reaction typical of plant defence reactions. In these samples, epidermal and outer cortical cells were impregnated with an electron-dense material. Histochemical assays of this material confirmed the substantial presence of phenols, pectic substances and suberin. Electron microscope observations clearly showed that in non-pathogenic BNR-inoculated plants, fungal cells were confined to the epidermal layer which was darkly stained. Gold cytochemistry confirmed the presence of pectic substances in the electron dense material. The possibility that pectic oligogalacturonides released after hydrolysis by the non-pathogenic BNR enzymes may act as elicitors of defence responses is discussed. The present ultrastructural observations corroborate that non-pathogenic BNR isolates may function as potential inducers of plant disease resistance.

Multiple Mitochondrial Viruses in an Isolate of the Dutch Elm Disease FungusOphiostoma novo-ulmi

The nucleotide sequences of three mitochondrial virus double-stranded (ds) RNAs, RNA-4 (2599 nucleotides), RNA-5 (2474 nucleotides), and RNA-6 (2343 nucleotides), in a diseased isolate Log1/3–8d2(Ld) of the Dutch elm disease fungusOphiostoma novo-ulmihave been determined. All these RNAs are A-U-rich (71–73% A + U residues). Using the fungal mitochondrial genetic code in which UGA codes for tryptophan, the positive-strand of each of RNAs 4, 5, and 6 contains a single open reading frame (ORF) with the potential to encode a protein of 783, 729, and 695 amino acids, respectively, all of which contain conserved motifs characteristic of RNA-dependent RNA polymerases (RdRps). Sequence comparisons showed that these RNAs are related to each other and to a previously characterized RNA, RNA-3a, from the sameO. novo-ulmiisolate, especially within the RdRp-like motifs. However, the overall RNA nucleotide and RdRp amino acid sequence identities were relatively low (43–55% and 20–32%, respectively). The 5′- and 3′-terminal sequences of these RNAs are different, but they can all be folded into potentially stable stem-loop structures. Those of RNA-4 and RNA-6 have inverted complementarity, potentially forming panhandle structures. Their molecular and biological properties indicate that RNAs 3a, 4, 5, and 6 are the genomes of four different viruses, which replicate independently in the same cell. These four viruses are also related to a mitochondrial RNA virus from another fungus,Cryphonectria parasitica, recently designated the type species of theMitovirusgenus of the Narnaviridae family, and to a virus from the fungusRhizoctonia solani. It is proposed that the fourO. novo-ulmimitochondrial viruses are assigned to theMitovirusgenus and designatedO. novo-ulmimitovirus (OnuMV) 3a-Ld, 4-Ld, 5-Ld, and 6-Ld, respectively. Northern blot analysis indicated thatO. novo-ulmiLd nucleic acid extracts contain more single-stranded (ss, positive-stranded) RNA than dsRNA for all three newly described mitoviruses.O. novo-ulmiRNA-7, previously believed to be a satellite-like RNA, is shown to be a defective RNA, derived from OnuMV4-Ld RNA by multiple internal deletions. OnuMV4-Ld is therefore the helper virus for the replication of both RNA-7 and another defective RNA, RNA-10. Sequence comparisons indicate that RNA-10 could be derived from RNA-7, as previously suggested, or derived directly from RNA-4.

Does water and phosphorus uptake limit leaf growth of Rhizoctonia-infected wheat seedlings?

Wheat seedlings infected with a pure inoculum of the root-rotting fungus Rhizoctonia solani were grown in pots designed to fit in pressure chambers, to allow the effects of the Rhizoctonia infection on leaf growth to be studied while maintaining the leaves at elevated water status. Wheat was grown to the third leaf stage in soil inoculated with three different levels of Rhizoctonia, and the pots were then pressurised for seven days to maintain the leaf xylem at the point of bleeding (ie. the leaves were at full turgor). The reduction in leaf expansion caused by Rhizoctonia was not overcome by pressurisation, indicating that a reduced supply of water to the leaves was not responsible for reduced leaf growth. The addition of phosphorus to pots marginally deficient in P did not increase the leaf growth of Rhizoctonia-infected plants, despite increased P uptake to the leaves. These results indicate that a reduced supply of water to the leaves and a supply of phosphorus that was bordering on deficient was not the cause of the growth reduction in seedlings with Rhizoctonia infection. The nature of this reduced growth remains uncertain but may involve growth regulators produced by the fungus, or by the plant as a result of the infection process. The mechanism of these growth reductions is of interest as it may provide a key to the development of plant resistance mechanisms.

The use of fatty acid signatures to study mycelial interactions between the arbuscular mycorrhizal fungus Glomus intraradices and the saprotrophic fungus Fusarium culmorum in root-free soil

The saprotrophic fungus Fusarium culmorum, Penicillium hordei, Rhizoctonia solani and Trichoderma harzianum and the arbuscular mycorrhizal fungus Glomus intraradices were examined for content of phospholipid fatty acids (PLFA) and neutral lipid fatty acids (NLFA). The AM fungus differed from the saprotrophic fungi especially by its content of the fatty acid 16:1ω5 which was absent in the saprotrophs. The fatty acid 18:2ω6,9 was the dominant fatty acid of the saprotrophic fungi while it was negligible in mycelium of G. intraradices. Specificity in content of fatty acids made it possible to quantify G. intraradices. and F. culmorum simultaneously in soil. Furthermore, a compartmented growth system made it possible to study mycelial interactions in the absence of roots. We measured hyphal spread of both fungi, hyphal 33P transport of G. intraradices and sporulation of F. culmorum. The two fungi did not interact according to the parameters used in this study. We conclude that fatty acid signatures may be a valuable tool when studying interactions between AM fungi and other fungi in root-free soil.

Interactions between root‐infecting fungi and plant density in an annual legume, Kummerowia stipulacea

Summary Population dynamics of an annual legume, Kummerowia stipulacea, were followed in permanent plots in a mown field for 4 years. There was a large reduction in the number of plants within seasons. Laboratory studies showed that biotic factors were responsible for seed death in the soil. Using soil from the field plot location, we isolated fungi pathogenic to K. stipulacea seeds and seedlings. Two pathogenic fungi (Rhizoctonia solani and Pythium irregulare) were used in a glasshouse study of the effects of pathogens and intraspecific competition on plant mortality and growth. Our null hypothesis was that death of individual plants due to disease would be balanced by increased growth of survivors. A factorial experiment was established with five soil treatments (three controls, P. irregulare and R. solani) and seven seed densities. Inoculation by both R. Solani and P. irregulare led to a reduction in final numbers of plants, with the reduction greatest for populations inoculated with R. solani. Average biomass per plant was reduced with increased sowing density in all treatments except the R. solani inoculation, where surviving plants in high‐density pots may be larger because of the reduced density resulting from pathogen‐induced death. No treatment differences in size hierarchies were evident. Surviving plants in both inoculation treatments could not take full advantage of the reduced density, and thus total biomass and number of trifoliate leaves per population (a surrogate for seed production) were reduced compared to controls, leading us to reject the null hypothesis. The reduction was more severe at high sowing densities. These results suggest that disease has the potential to reduce seed production and thus plant population numbers, if populations are seed limited. Shapes of yield‐sowing density relationships were asymptotic for all treatments; thus, disease‐induced ‘overcompensation’ leading to fluctuating population dynamics was not suggested.

Fluorescent Pseudomonas spp. as biocontrol agents against forage legume root pathogenic fungi

Native fluorescent pseudomonads isolated from the rhizosphere of Lotus corniculatus were screened in vitro for their antagonistic activity against the phytopathogenic fungi Pythium ultimum and Rhizoctonia solani. About 12% of the bacterial isolates inhibited one or both fungi in vitro. Isolates which exhibited the greatest antagonistic activity were assayed in vivo against the pathogens. Three Pseudomonas fluorescens strains were selected from these assays: UP61, UP143 and UP148. These strains produced HCN and siderophores, but addition of iron to the medium did not affect the antagonistic activity. Lytic enzymes such as chitinase and β-1,3-glucanase were not detected. The simultaneous inoculation of Pseudomonas strains and Rhizobium loti B816 did not affect nitrogen fixation efficiency in L. corniculatus plants. Sterile peat was successful as a carrier for these P. fluorescens strains.

The incompatible interaction between Phytophthora parasitica var. nicotianae race 0 and tobacco is suppressed in transgenic plants expressing antisense lipoxygenase sequences.

Nicotiana tabacum 46-8 cultivar displays an incompatible interaction with race 0 of Phytophthora parasitica var. nicotianae (Ppn), a fungal pathogen of most tobacco cultivars. At the plant level, incompatibility is characterized by the induction of lipoxygenase (LOX, EC = 1.13.11.12) activity and localized hypersensitive cell death before defense gene activation. To evaluate the involvement of LOX in the onset of plant defense, tobacco 46-8 plants were genetically engineered using full-length or partial-length antisense (AS) tobacco LOX cDNA constructs. AS expression strongly reduced elicitor- and pathogen-induced LOX activity. Eight independent AS-LOX lines were selected and assayed for their response to Ppn. After root or stem inoculation with race 0, all AS-LOX lines but one displayed a compatible phenotype whereas control transformed plants, not containing the AS-LOX cassette, showed the typical incompatible reaction. The presence of the fungus in transgenic lines was demonstrated by PCR amplification of a Ppn-specific genomic sequence. A linear relationship was found between the extent of LOX suppression and the size of the lesion caused by the fungus. The AS-LOX plants also showed enhanced susceptibility toward the compatible fungus Rhizoctonia solani. The results demonstrate the strong involvement of LOX in the establishment of incompatibility in plant-microorganism interactions, consistent with its role in the defense of host plants.

Effect of physical conditions on the spatial and temporal dynamics of the soil‐borne fungal pathogen Rhizoctonia solani

The transmission of infection by many soil‐borne fungal parasites of plants depends on the ability of the fungus to grow on or through soil. Progress in analysing the effects of soil physical factors on the temporal and spatial dynamics of fungal growth has been hindered by technical difficulties of quantifying fungal biomass in soil and heterogeneity in soil properties. In this paper we use a combination of a monoclonal antibody‐based immunosorbent assay and microscopy to analyse the effects of soil physical properties on the spatial and temporal dynamics of colonies of the economically important fungus Rhizoctonia solani Kühn growing in two dimensions and three dimensions in a sand. Combinations of different particle‐size distributions and matric potential are used to manipulate the air‐filled pore volume and pore‐size distribution independently of each other. Temporal dynamics are measured by the change in fungal biomass over time whereas spatial dynamics relate to fungal spread and are measured by the colony size, the rate of colony expansion and the biomass distribution within colonies. We show that the fungus spreads more than three times further over surfaces than through sand, even though the same amount of biomass is produced in each case. Pore‐size distribution and air‐filled pore space both affected the extent and rate of fungal spread in three dimensions within sand, with more rapid and extensive spread in a coarse sand compared with a fine sand at identical air‐filled pore volume. The spread of fungal hyphae along surfaces was affected neither by differences in surface texture nor by air‐filled volume, and was substantially more homogeneous than for three‐dimensional spread. We argue that the relative impermeability of sand surfaces to penetration by hyphae might be influenced by the ability of the fungus to branch within a confined space rather than simply to penetrate the pores. The broader epidemiological and ecological consequences of preferential spread by parasitic and saprophytic fungi along surfaces rather than through the dense soil volume are discussed.

Thiourea derivatives of α-aminoacids. Synthesis and characterization of Ni(II), Cu(II) and Pt(II) complexes with l-valinate derivatives. Antifungal activity

The preparation of metal complexes with 2-[(3,3′-diethylthioureido)phenylmethylamino]-3-methylbutyric acid methyl ester, Hetv, or 3-methyl-2′-{[morpholine-4-carbothioylimino)phenylmethyl]amino}-butyric acid methyl ester, Hmtv, has been achieved. The complexes obtained with formulae of Ni(etv) 2, Cu(mtv) 2·2H 2O and Pt(Hmtv)Cl 2, were studied and characterized by elemental analysis, electrical conductivity in solution, IR and UV–VIS spectra, FAB+ and electron impact (EI) mass spectrometry, 1H- and 13C-NMR, ESR, magnetic susceptibility, atomic absorption spectrometry and thermal analysis. Their antifungal activity was studied against the plant pathogenic fungi Colletotrichum fragariae, Rhizoctonia solani and Phoma betae.

Evaluation of methyl iodide as a soil fumigant in container and small field plot studies

Methyl iodide was evaluated as a soil fumigant as a potential replacement for the widely used soil fumigant methyl bromide. In container trials, methyl iodide was significantly more effective than methyl bromide against the plant parasitic nematodes Meloidogyne incognita, Heterodera schachtii and Tylenchulus semipenetrans and the plant pathogenic fungus Rhizoctonia solani. In small field plots, soil populations of root-knot nematodes were no longer detected after methyl iodide fumigation at an application rate of 112 kg ha-1. However, after growing a susceptible lima bean host for two months, substantial root-knot galling occurred, while Rhizobium nodulation was absent. At 168 kg ha-1 of methyl iodide, root-knot galling was reduced to less than 1%, and no Pythium propagules were recovered on selective detection media. These efficacy data support the conclusion that methyl iodide is a likely candidate for replacing methyl bromide as a soil fumigant. © 1998 SCI.

The complete nucleotide sequence and characterization of the linear DNA plasmid pRS64-2 from the plant pathogenic fungus Rhizoctonia solani.

The complete nucleotide sequence of one of three linear DNA plasmids (pRS64-2) from the plant pathogenic fungus Rhizoctonia solani was determined. The pRS64-2 DNA consisted of 2877 nucleotides. The nucleotide sequences of the middle 2.2-kb regions of the other two plasmids (pRS64-1 and pRS64-3) were also determined. Comparison of the nucleotide sequences among the three plasmid DNAs indicated the presence of four regions with more than 86% sequence homology, suggesting the development of three plasmid DNAs from a common ancestor. A computer-based study of the pRS64-2 DNA-folding at both termini predicted hairpin loop structures. The hairpin loops consisted of the left- and right-hand terminal 113 and 105 nucleotides, respectively, and had no sequence homology. They had not undergone flip-flop inversion. The hairpins form cruciform base-paired structures.

Unprecedented detoxification of the cruciferous phytoalexin camalexin by a root phytopathogen

The unusual metabolism of the phytoalexin camalexin by the root rot fungus Rhizoctonia solani Kuhn is reported. This transformation proceeded via 5-hydroxycamalexin, further transformed into 5-hydroxyindole-3-carbonitrile and 5-hydroxy-2-formamidophenyl-2′-thiazolylketone. Most importantly, the metabolites resulting from transformation of camalexin were significantly less toxic to the fungus than camalexin. The metabolism of camalexin ( 5 ) by the plant pathogen Rhizoctonia solani and the antifungal activities of 5–8 are reported.

Microbial linear plasmids.

While plasmids were originally considered to be generally circular until almost two decades ago, linear elements were reported to exist as well. They are now known to be common genetic elements in both, pro- and eukaryotes. Two types of linear plasmids exist, the so-called hairpin plasmids with covalently closed ends and those with proteins bound to their 5' termini. Hairpin plasmids are common in human-pathogenic Borrelia spirochetes, in which they are instrumental in escape from the immunological response; cryptic hairpin elements are present in mitochondria of the plant pathogenic fungus Rhizoctonia solani. Plasmids with 5' attached proteins constitute the largest group. In actinomycetous bacteria they are conjugative and usually confer advantageous phenotypes, e.g. formation of antibiotics, degradation of xenobiotics, heavy-metal resistance and growth on hydrogen as the sole energy source. In contrast, the majority of linear plasmids from eukaryotes are cryptic, with only a few exceptions. In some yeasts a killer phenotype may be associated, the most thoroughly investigated elements being those from Kluyveromyces lactis killer strains. In Neurospora spp. and in Podospora anserina, senescence and longevity respectively are correlated with linear plasmids. This review focuses on the biology of linear plasmids, their environmental significance and their use as tools in molecular and applied microbiology.

Rhizoctonia Foliar Blight Reactions and Seed Yields in Soybean

Rhizoctonia foliar blight (RFB) is caused by the fungus Rhizoctonia solani Kühn anastomosis group 1, intraspecific groups IA and IB. The disease affects more than half of the soybean [Glycine max (L.) Merr.] acreage in Louisiana. Although less‐susceptible cultivars are recommended, reliable estimates of cultivar susceptibility have been difficult to obtain because of several factors, including uneven distribution of RFB in fields, differences in plant developmental stage at time of infection, and the requirement for free moisture in RFD development. The objective of this study was to screen commercial soybean cultivars grown throughout the southern USA, as well as advanced breeding genotypes, for field reaction to RFB in south Louisiana. Ninety‐one soybean genotypes, representing Maturity Groups IV, V, VI, and VII, were evaluated at two locations in 1993 and 1994. Disease ratings were made with a severity scale that included pods as well as foliage. A range of reactions to RFB was observed among cultivars, but most were severely damaged. Significant negative correlations between RFB severity and yield occurred among cultivars in all Maturity Groups. Results identified several cultivars, notably Buckshot 66 and Pioneer variety 9593, with promising levels of resistance.

The identification and characterization of four laccases from the plant pathogenic fungus Rhizoctonia solani

The identification and characterization of four laccases from the plant pathogenic fungus Rhizoctonia solani

The identification and characterization of four laccases from the plant pathogenic fungusRhizoctonia solani

Four distinct laccase genes, lcc1, lcc2, lcc3 and lcc4, have been identified in the fungus Rhizoctonia solani. Both cDNA and genomic copies of these genes were isolated and characterized. Hybridization analyses indicate that each of the four laccase genes is present in a single copy in the genome. The R. solani laccases can be divided into two groups based on their protein size, intron/exon organization, and transcriptional regulation. Three of these enzymes have been expressed in the fungus Aspergillus oryzae. Two of the recombinant laccases, r-lcc1 and r-lcc4, as well as the native lcc4 enzyme were purified and characterized. The purified proteins are homodimeric, comprised of two subunits of approximately 66kDa for lcc4 and 50-100kDa for the recombinant lcc1 protein. These laccases have spectral properties that are consistent with other blue copper proteins. With syringaldazine as a substrate, lcc4 has optimal activity at pH7, whereas lcc1 has optimal activity at pH6.