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PCR primers that amplify fungal rRNA genes from environmental samples.

Two PCR primer pairs were designed to amplify rRNA genes (rDNA) from all four major phyla of fungi: Ascomycota, Basidiomycota, Chytridomycota, and Zygomycota. PCRs performed with these primers showed that both pairs amplify DNA from organisms representing the major taxonomic groups of fungi but not from nonfungal sources. To test the ability of the primers to amplify fungal rDNA from environment samples, clone libraries from two avocado grove soils were constructed and analyzed. These soils possess different abilities to inhibit avocado root rot caused by Phythophthora cinnamomi. Analysis of the two rDNA clone libraries revealed differences in the two fungal communities. It also revealed a markedly different depiction of the soil fungal community than that generated by a culture-based analysis, confirming the value of rDNA-based approaches for identifying organisms that may not readily grow on agar media. Additional evidence of the usefulness of the primers was obtained by identifying fungi associated with avocado leaves. In both the soil and leaf analyses, no nonfungal rDNA sequences were identified, illustrating the selectivity of these PCR primers. This work demonstrates the ability of two newly developed PCR primer sets to amplify fungal rDNA from soil and plant tissue, thereby providing unique tools to examine this vast and mostly undescribed community of organisms.

Ascomycota↗

Positive autoregulation and signaling properties of pyoluteorin, an antibiotic produced by the biological control organism Pseudomonas fluorescens Pf-5.

Pseudomonas fluorescens Pf-5, a rhizosphere bacterium, produces a suite of secondary metabolites that are toxic to seed- and root-rotting plant pathogens. Among these are the polyketide compounds pyoluteorin and 2,4-diacetylphloroglucinol. We provide evidence that pyoluteorin production is influenced by positive autoregulation. Addition of pyoluteorin to liquid cultures of Pf-5 enhanced pyoluteorin production. In addition, pyoluteorin and 2,4-diacetylphloroglucinol mutually inhibit one another's production in Pf-5. For pyoluteorin, both positive autoregulation and negative influences on production by 2,4-diacetylphloroglucinol were demonstrated at the transcriptional level by measuring activity from transcriptional fusions of an ice nucleation reporter gene (inaZ) to three separate pyoluteorin biosynthetic genes. The occurrence of pyoluteorin autoregulation in the rhizosphere was assessed on cucumber seedlings in pasteurized soil with cross-feeding experiments. In the rhizosphere, expression of a pyoluteorin biosynthesis gene by a pyoluteorin-deficient mutant of Pf-5 was enhanced by pyoluteorin produced by coinoculated cells of Pf-5. These data establish that the polyketide pyoluteorin is an autoregulatory compound and functions as a signal molecule influencing the spectrum of secondary metabolites produced by the bacterial cell.

Anti-Bacterial Agents↗

EcbI and EcbR: homologs of LuxI and LuxR affecting antibiotic and exoenzyme production by Erwinia carotovora subsp. betavasculorum.

Erwinia carotovora subsp. betavasculorum Ecb168 causes vascular necrosis and root rot of sugar beet and produces an antibiotic(s) that is antagonistic against other Erwinia spp. EcbI- mutants of Ecb168, each containing a single transposon insertion in the ecbI gene (for Erwinia carotovora subsp. betavasculorum inducer), do not produce detectable levels of extracellular protease or antibiotic(s), and express less pectate lyase activity and virulence than the wild-type strain. A plasmid containing the cloned ecbI gene complemented the EcbI- mutants for these phenotypes. Protease production by EcbI- mutants grown on agar surfaces was restored by neighboring cells of Escherichia coli containing ecbI. Production of a diffusible N-acylhomoserine lactone autoinducer by wild-type Ecb168 was detected with indicator strains of E. coli and Agrobacterium tumefaciens. EcbI- mutant strains did not produce an autoinducer detected by the indicator strains. Antibiotic production by EcbI- mutants was restored by cell-free culture supernatants of Ecb168 or E. coli containing a cloned ecbI gene. The predicted amino acid sequence of EcbI is similar to those of CarI, ExpI, and HsII, three LuxI homologs required for production of a diffusible N-acylhomoserine lactone autoinducer in Erwinia carotovora subsp. carotovora. A luxR homolog, termed ecbR (for Erwinia carotovora subsp. betavasculorum regulator), is convergently transcribed and overlaps with ecbI by 17 bp at their 3' ends. These results are consistent with the hypothesis that a quorum-sensing system related to the prototypic luxI-luxR gene pair controls antibiotic and exoenzyme production in Erwinia carotovora subsp. betavasculorum.

Agrobacterium tumefaciens↗

Effect of acidic electrolyzed water on the viability of bacterial and fungal plant pathogens and on bacterial spot disease of tomato.

Acidic electrolyzed water (AEW), known to have germicidal activity, was obtained after electrolysis of 0.045% aqueous solution of sodium chloride. Freshly prepared AEW (pH 2.3-2.6, oxidation-reduction potential 1007-1025 mV, and free active chlorine concentration 27-35 ppm) was tested in vitro and (or) on tomato foliage and seed surfaces for its effects on the viability of plant pathogen propagules that could be potential seed contaminants. Foliar sprays of AEW were tested against bacterial spot disease of tomato under greenhouse and field conditions. The viability of propagules of Xanthomonas campestris pv. vesicatoria (bacterial spot pathogen), Streptomyces scabies (potato scab pathogen), and Fusarium oxysporum f.sp. lycopersici (root rot pathogen) was significantly reduced 4-8 log units within 2 min of exposure to AEW. Immersion of tomato seed from infected fruit in AEW for 1 and 3 min significantly reduced the populations of X. campestris pv. vesicatoria from the surface of the seed without affecting seed germination. Foliar sprays of AEW reduced X. campestris pv. vesicatoria populations and leaf spot severity on tomato foliage in the greenhouse. In the field, multiple sprays of AEW consistently reduced bacterial spot severity on tomato foliage. Disease incidence and severity was also reduced on fruit, but only in 2003. Fruit yield was either enhanced or not affected by the AEW sprays. These results indicate a potential use of AEW as a seed surface disinfectant or contact bactericide.

Disinfectants↗

Studies on the cell surface of zoospores and cysts of the fungus Phytophthora cinnamomi: nature of the surface saccharides as determined by quantitative lectin binding studies.

The nature of the surface saccharides of zoospores, "partially encysted zoospores" and cysts of the root-rotting fungus Phytophthora cinnamomi, has been examined by quantitative lectin binding studies. Zoospores bound concanavalin A (Con A), but did not bind any of a variety of other lectins tested. In contrast, both cysts and "partially encysted zoospores" bound soybean agglutinin (SBA) as well as Con A. This indicates that accessible alpha-D-glucosyl/alpha-D-mannosyl-containing glycoconjugates predominate at the zoospore surface, whereas both alpha-D-glucosyl/alpha-D-mannosyl and galactosyl and/or N-acetyl-D-galactosaminosyl residues are accessible at the surface of cysts and "partially encysted zoospores." Neither Ulex europeus lectin nor wheat germ agglutinin (WGA) bound to any of the three cell preparations, indicating the absence of accessible alpha-L-fucosyl and N-acetyl-D-glucosaminosyl residues.

Binding Sites↗

[Causal factors of decline disease in exotic pine (Pinus taeda and P. elliottii) plantations].

Investigation on the cause of the decline of exotic pine plantations in Anhui Province was carried out from 1995 to 2000. The results showed that the cause might develop from the interaction of three or more sets of factors. The first group of factors called as predisposing factors were high and low temperature and rainfall, poor soil fertility, and excessive soil bulk density, soil moisture, and of stand density; the second group of factors called as inciting factors were insect defoliators, damaging diseases of needles, late spring frost, severe drought, and excessive resin-tapping, and the third group of factors called as contributing factors were several biotic agents such as shoot blight infected by Sphaeropsis sapinea, decay wood and root rot by fungi, and bark beetles and wood borers.

Causality↗

Cloning, expression, and biological activity of recombinant alpha-cinnamomin: toxicity to cranberry and other plant species.

Elicitins produced by the pathogenic fungi Phytophthora are known to exhibit The elicitin cinnamomin is of nonspecific toxicity to different solanaceous plant species. particular interest for its potential role in the hypersensitive-like cell death and in the biological response of cranberry plants to the fungal pathogen Phytophthora cinnamomi. In order to understand the biochemical steps of the Phytophthora root rot disease in cranberry, we investigated the alpha-cinnamomin-induced plant responses. Toxicity of alpha-cinnamomin, which shows a high degree of sequence homology to the alpha-elicitin group, was tested on Vaccinum macrocarpon, Nicotiana tabacum, Capsicum annuum, Lycopersicon esculentum, Lactuca sativa, and Phaseolus vulgaris plants. Gene corresponding to alpha-cinnamomin gene fused with maltose binding protein gene, was cloned into a pMALTEV expression vector, which was transformed into E. coli cells. Cells containing alpha-cinnamomin clones were cultured and extracted protein was purified on a maltose binding protein affinity column. Biological activity of alpha-cinnamomin fusion protein was examined on propagated plants and cuttings. In cranberry plants treated with cinnamomin necrotic hypersensitive-like response in the proximal areas of the leaf lamina of lower plant leaves was observed after 48-72 hr of incubation. Limited leaf necrosis observed days after application of low amounts of recombinant cinnamomin directly on the leaves of other plants indicates that the recombinant protein might be functioning as a toxin, capable of inducing aging accompanied by plant cell death.

Algal Proteins↗

[Applied studies of biological control techniques on healthful Chinese crude drugs].

Medicinal plants always suffer from diseases and pests in their cultivation and growing, and the yield and quality are reduced. At present, chemical pesticide is mainly used to control the diseases and pests. The long-term use and over-use of pesticide result in pesticide residue, pests resistance to pesticide, water and soil pollution and other environmental contamination. It is necrssary to provide the integrated pests management, including new technology of biological control for developing green drug. This project makes research on the following aspects: 1. Applied studies on bethyloid wasp (Scleroderma guani) control for stem borer; 2. To develop Trichoderma spp. and use it to control the diseases of medicinal plants; 3. Applying Agro-antibiotics 120 to control phytophthora root rot of ginseng; 4. Researches on application of entomopathogenic nematodes against ten-spotted lema (Lema decempuntata) et al.

Animals↗

The use of RAPD to characterize Bipolaris sorokiniana isolates.

Bipolaris sorokiniana is a phytopathogenic fungus causing diseases of cereal crops such as common root rot, the leaf spot disease, seedling blight, and black point of the grain. Random-amplified polymorphic DNA (RAPD) assay was used to investigate the genetic diversity of 20 isolates collected from different cultivars in wheat-producing regions in Brazil. Seventy primers, with random nucleotide sequences, were tested. Reproducibility to amplify the genomic DNA of isolates was found for 30 of the 70 primers tested, generating between 1 and 17 fragments ranging from 0.35 to 2.0 kb (average size). The degree of similarity between samples was calculated through simple association and the dendrogram was assessed using the unweighted pair group method with arithmetical average. After the RAPD analyses 19 isolates were closely grouped, having a similarity coefficient of >or= 78%. Isolate I017 showed very low similarity coefficients, ranging between 38 and 46%. The RAPD analyses provided important information as to the degree of genetic variability and the relationship between the isolates investigated, revealing polymorphism and establishing electrophoretic profiles useful to characterize the phytopathogen.

Ascomycota↗

[Biological preparations with different mechanism of action for protecting potato against fungal diseases].

Mycological analysis throughout the vegetation period of potato (Solanum tuberosum) made it possible to study in detail the structure of micromycete community, to determine typical dominant (frequency, more than 60%), typical common (frequency, 30 to 60%), typical rare (frequency, 10 to 30%), and casual (frequency, less than 10%) species and to estimate changes in the microorganism community caused by plant protection preparations with different mechanisms of action. It was shown that, as a result of occurrence of resistant forms, synthetic preparations against fungal pathogens of potato (such as TMTD, Ridomil gold MC, and Cupricol) were only slightly more effective than biological preparations (Trichodermin and AgroChit), with the former considerably changing the natural saprophytic mycological community. An increase in the soil pool of Trichoderma harzianum as a result of application of a biological preparation based on this antagonistic fungus correlated with its effectiveness against the soil pathogen Fusarium sp., which causes root rots. A chitosan-based elicitor preparation more effectively suppressed the development of early (Alternaria sp. and Macrosporium sp.) and late (Phytophthora sp.) blights of leaves and had a weaker effect on the soil microflora.

Alanine↗

Physiological studies on Phymatotrichum omnivorum. IX. Synthesis of indole acetic acid in vitro.

The cotton root rot fungus, Phymatotrichum omnivorum, synthesized indole acetic acid (IAA) in culture medium containing tryptophan as the only source of nitrogen. Synthesis of IAA was not affected by illumination, and maximum amounts of the auxin were produced at 30 days. The optimum hydrogen ion concentration and temperature for IAA production were 6.5 and 28 degrees C, respectively. Indole ethanol (IEA) was also detected in tryptophan media, primarily during the active growth of the organism.

Culture Media↗

[Identification with roseofungin of an antibiotic produced by Streptomyces roseoflavus strain A-23/791 and its undifferentiated variant].

Isolation, purification and chemical identification of an antibiotic produced by strain IMV A-23/791 of Streptomyces roseoflavus and its adifferentiated variant are described. The antibiotic inhibits the growth of many phytopathogenic fungi, dermatophytes and yeast-like fungi and is capable of stimulating the growth and development of plants. The antibiotic is effective in control of cucumber root rot under hydroponic cultivation conditions. It was shown that the adifferentiated proactinomycetous variant 4-76 was more productive than the initial strain A-23/791. The antibiotic isolated from the variant mycelium is more active. It was shown that the antibiotic can be isolated from the biomass of the adifferentiated variant with a more simple method in the authors' modification providing higher antibiotic yields and the use of lower amounts of the solvents. The UV, IR, mass and luminescence spectra and mobility in paper and thin-layer chromatography have shown the carbonyl-conjugated pentaene A-23/791 to be identical to roseofungin described earlier. Based on the data obtained strain A-23/791 should be classified with Streptomyces roseoflavus var, roseofungini Nikitina, 1968, a variant of Streptomyces roseoflavus Arai, 1951.

Antifungal Agents↗

[Fungicidal activity of imbricin and its salts].

The fungicidal activity of imbricin and its salts against fungal phytopathogens was studied. The biological tests revealed that the compounds had a fungicidal activity and inhibited the development of a number of plant diseases. Imbricin showed a marked fungicidal activity when used as a disinfectant of wheat seeds and in the control of root rot of cucumbers. It was demonstrated that imbricin and its salts had the growth regulating activity in plant cell culture.

Ascomycota↗

Antagonistic effects of several bacteria on Fusarium oxysporum, the causal agent of root and crown rot of onion under field conditions.

Onion (Allium cepa) is one of the most important vegetable crop which is commonly used as a food supplement. This plant is found to be vulnerable to various pathogenic infections during its growth development. Among different onion diseases, root and crown rot,caused by Fusarium oxysporum f.sp. cepa, s considered an importantfungal disease. In this study, the inhibitory effect of Bacillus cereus (isolates 22 and 52), B. subtilis (isolate 126), Pseudomonas fluorescens (isolates 48 and CHAO), benomyl fungicide and a combination of isolates CHAO and 22 and isolate 52 and benomyl were investigated on disease development under the field condition. This experiment was carried out in a randomize complete blocks with 10 treatments and three repetitions. Grouping of treatments was done at 5% level using Duncan multiple comparison test. It was also demonstrated that isolate 126 was the most effective antagonist with regard to crop yield but other treatments despite showing significant on plant growth factors were less effective in increasing crop yield.

Bacillus cereus↗

Phytophthora pseudosyringae sp. nov., a new species causing root and collar rot of deciduous tree species in Europe.

In several studies of oak decline in Europe, a semi-papillate homothallic Phytophthora taxon was consistently isolated, together with other Phytophthora species, from rhizosphere soil samples. It was also found associated with necrotic fine roots and stem necroses of Fagus sylvatica and Alnus glutinosa. Due to morphological and physiological similarities, the semi-papillate isolates were previously identified as P. syringae by various authors. The morphology, physiology and pathogenicity against fine roots of Quercus robur, Q. petraea and F. sylvatica, bark of A. glutinosa, leaves of Ilex aquifolium and apple fruits of this Phytophthora species are described and compared with those of related and similar Phytophthora species, namely P. ilicis, P. psychrophila, P. quercina, P. citricola and P. syringae. The phylogenetic placement on the basis of ITS and mtDNA sequence data was also examined. Isolates of this taxon produce colonies with stellate to rosaceous growth patterns and limited aerial mycelium on various agar media. Antheridia are predominantly paragynous. In water culture catenulate hyphal swellings and semi-papillate caducous sporangia, that are usually limoniform, ellipsoid or ovoid, are formed abundandly, mostly in lax or dense sympodia. This taxon is a moderately slow growing, low temperature species with optimum and maximum temperatures around 20 and 25 degrees C, respectively. Tested isolates are moderately aggressive to fine roots of oaks and beech, highly aggressive to holly leaves and apple fruits, and slightly pathogenic to alder bark. Thirteen tested isolates had an identical and distinct ITS sequence which was more similar to that of P. ilicis and P. psychrophila than any other known taxa. On the basis of their unique combination of morphological characters, colony growth patterns, cardinal temperatures for growth, growth rates, pathogenicity to oaks, beech, alder, apple and holly, their host range, and ITS and mtDNA sequences the semi-papillate caducous Phytophthora isolates from oaks, beech and alder are clearly separated from related and similar Phytophthora spp., and described as a new species, P. pseudosyringae sp. nov.

Alnus↗

[An immunoregulatory effect of endoglucanase preparations on plant resistance to fungal infections].

Effects of enzymatic preparations--pectomacerin, hemicellulase, and Trichoderma viride 13/10 cellulase--on the plant immunological status were studied by examples of the pathosystems carrot root-white rot agent (Sclerotinia libertiana) and carrot root-black rot agent (Rhizopus nigricans). It was demonstrated that these preparations reduced the damage of infections, namely, decreased the permeability of cell membranes in infected tissue and stimulated its defense responses, which manifested itself in a stable elevation in the content of phenolic compounds and formation of tissue protective barriers.

Adjuvants, Immunologic↗

The causal agents of damping-off disease of buglosse from Iran.

Iran is considered a major genetic for medicinal plant in the world. Because of this significant diversity and historical background in identification and utilization to remedy human and animal diseases, export of medicinal plant can help to strengthen local as well as natural economy. Buglosse (Fig. 1) is one of the most important and common medicinal plants in Iran and exist as Echium amoneum and Borago officinalis. This work was conducted in order to identify the causal agent(s) of damping off disease in buglosse. Plant disease samples were taken from Esfahan and Tehran provinces. Symptoms on original plant including root, crown rot, dark tissue, pith and hallow root were collected in order to isolate disease agent(s). Symptomatic root and crown tissues after surface sterilization with 96% ethanol were transferred on to PDA and WA media and also on moist filter paper in petri dishes. Two fungal colonies grew from tissue segments and spore culture was subsequently purified. The fungal isolate identified as Rhizoctonia solani based on the following test. Hyphal tip was removed from colony margin placed on PDA and PSA media and incubated in dark. Colony diameter of one hundred hyphae measured and nucleus was stained according to Bandoni (1979), Kronland and Stanghellini (1988). It was observed that in each cell of hyphae there are more than two nuclei. Single spore culture were obtained from macroconidia of Fusarium isolate. After 24 hr of incubation, growing single spore were transferred to KCL medium to detect spore chains. Fungal isolates transferred to PSA and PDA media for sporulation. After 7 days colonies appeared as white cream to pinkish on top and cream to dark pink at the bottom of petri dish with abundant micro and macro conidia. Colonies were snow white, felting shape, with ample causal hyphae on PSA medium. On KCL medium, fungal growth was superficial and colonies were colorless with long macroconidia and individual sausage-shape macroconidia being thinner one side and having maximum four septa. Microconidia were long double compartment round on both side, straight to slightly curved. Base on morphology and dimension of conidia and production of chlamidospore the funguses identify as Fusarium solani.

Borago↗

C.L. Shear: Gifted Mycologist, Plant Pathologist, and APS Founder.

Cornelius Lott Shear was one of the most influential plant pathologists of the early twentieth century. He was first and foremost an excellent mycologist who did pioneering research on pathogenic fungi and, as a senior pathologist with the USDA's Bureau of Plant Industry, studied important crop diseases and offered useful control measures. Shear's successful research enhanced his reputation among his fellow pathologists and allowed him to embark on what was perhaps his most significant contribution to plant pathology, his pivotal role in the creation of the American Phytopathological Society in 1908. Shear felt that an independent society dedicated to the unique needs of plant pathologists would facilitate communication and cooperation among practitioners. Between his scientific research and his role in the creation of APS, Shear stands out for the enormous impact he had on his science.

American Phytopathological Society↗