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PubMed2003

Pythium regulare sp. nov., isolated from the Canary Islands, its taxonomy, its region of rDNA, and comparison with related species.

Pythium regulare (CI-34) was isolated from some soil samples taken in the Canary Islands (Spain). This new species is very closely related to P. irregulare isolated from pea roots in The Netherlands by Buisman in 1927. The species of Pythium are members of the kingdom Chromista. Pythium regulare is characterized by its ornamented oogonia bearing blunt or digitate spines, and its non-sporulating type of sporangia or hyphal bodies, its aplerotic oospores, its monoclinous and diclinous antheridia that at times crowd around the oogonia. The taxonomic description of this oomycete, the PCR of the internal transcribed region (spacers ITS1, ITS2, and the gene 5.8 S) of its ribosomal nuclear DNA as well as the nucleotide sequences, and its comparison with related species are given here.

Base Sequence↗

PubMed2004

A new species of Pythium isolated from burgundian vineyards and its antagonism towards Botrytis cinerea, the causative agent of the grey mould disease.

During the course of an investigation on the pythiaceous fungi occurring in burgundian vineyards, three isolates of Pythium were found, all of which were characterised by spherical to lemoniform, proliferating sporangia. Only one out of these three isolates produced sexual characters (S-12) after prolonged incubation, but their asexual structures, temperature growth relationship and molecular characteristics were identical. Saprolegnia-type internal proliferation of the sporangia, and Phytophthora-type lemoniform, papillate sporangia were characteristic of all the three isolates. The type specimen is S-12 which is a medium slow growing saprophyte. The sequence of its ITS region of the rDNA shows a close relationship with the genus Phytophthora. Morphological and molecular characteristics of this isolate justifies its description as a new species: Pythium citrinum. When grown together with Botrytis cinerea, the causal agent of the grey mould disease of the grapevine, this new species shows a pronounced antagonism and suppresses its growth. This new species is very close to P. montanum, a recently described fungus and constitutes a "border species" linking the genera Pythium and Phytophthora. Morphological features of this new species, its antagonism with B. cinerea, the sequences of the ITS region of its nuclear ribosomal DNA, and its comparison with related species are discussed.

Amino Acid Sequence↗

PubMed2005

Characterisation of the early events in atypical tomato root colonisation by a biocontrol agent, Pythium oligandrum.

The specific oomycete-plant relationship established between a biological agent, Pythium oligandrum, and tomato (Lycopersicon esculentum Mill.) plants was examined over the first 48 h after inoculation of tomato roots with the antagonist. One of the most significant effects was the quick colonisation of cortical and vascular root areas by P. oligandrum (until 9 h post-inoculation); it was similar to invasions by the major pathogens of Pythium genus, and much faster than those by Pythium-minor pathogens. Despite the multiplication of hyphae in the root areas, fungal colonisation was associated with neither host wall disruption nor host cell alterations. The colonising hyphae looked healthy till the ninth hour after inoculation, then, they progressively became highly vacuolated. Cytological observations showed that, over the first 14 h of experiment, oomycete invasion was accompanied with rare host-induced defence reactions. Biochemical analysis evidenced an accumulation of phenolic compounds starting 3 h after inoculation. The 14th hour corresponded to the beginning of rishitin (phytoalexin) synthesis. Accumulation of biochemical host defence compounds was concomitant with early signs of hyphae alterations. During the next 34 h several host reactions were regularly amplified as evidenced by the plugging of invaded host cells with heterogeneous osmiophilic or high electron-dense (ED) materials. Fungal cell decay was accompanied with the formation of oogonia in the cortex, vascular parenchyma and xylem vessels. All these early events suggest a peculiar relationship established between P. oligandrum and the plant.

Cell Wall↗

PubMed1978

Parasitic relationships between Pythium oligandrum Drechsler and some other species of the Oomycetes class.

Parasitic relationships between Pythium oligandrum Drechsler and some phytopathogenic species of the Oomycetes class were investigated on agar plates. A high parasitic ability of Pythium oligandrum on Pythium ultimum Trow, P. debaryanum Hesse, and Aphanomyces laevis De By species was proved in this study of relationships among these fungi, commonly populating the rhizosphere of emerging sugar beet. P. oligandrum produces numerous thin haustorial threads, searching the hyphae of host species and enwrapping them during the parasitation. In a later parasitation stage the host organs were enwrapped by thicker hyphae of P. oligandrum as well. P. oligandrum grew through the colony of parasitized species mostly within 24 hours after inoculation and was able to destroy it entirely within 5--6 days. P. oligandrum produced numerous fructification organs in contradistinction to parasitized species. P. oligandrum oospores germinated abundantly when host species were present. Hyperparasitism of P. oligandrum was found both in cultivation with a single host and in common cultivation with all three host species simultaneously.

Agar↗

PubMed1979

Activity of pectinases, amylases, and saccharase in Pythium spp.

The activity of pectine-lyase, polygalacturonase, pectine-methyl-esterase, amylase, and saccharase in Pythium ultimum, Pythium oligandrum, and Pythium debaryanum was determined. Cultures of fungi were cultivated under different temperatures and pH-values within 24 hours and 15 days. The optimum temperature for production of the mentioned enzymes was found to be 24 degrees C. Furthermore, the influence of pH and age of culture on activity of enzyme was investigated. The same trend was found in all the fungus species examined.

Amylases↗

PubMed2003

Characterisation of a new species of Pythium isolated from a wheat field in northern France and its antagonism towards Botrytis cinerea causing the grey mould disease of the grapevine.

A new species, Pythium bifurcatum, isolated from soil samples taken from a wheat field in Lille in northern France is described here. The oomycete occurred thrice out of 50 samples. The type specimen is F-91, which is a slow-growing saprophyte living on vegetable debris and which can be recognised by its antheridial as well as oogonial characteristics, which are different from other known species of Pythium. When grown together with Botrytis cinerea, the causal agent of the grey mould disease of the grapevine, Pythium bifurcatum shows a pronounced antagonism and suppresses its growth. Morphological features of this new species, its antagonism to B. cinerea, the sequences of the ITS region of its nuclear ribosomal DNA, and its comparison with related species are discussed in this article.

Base Sequence↗

PubMed2003

A molecular phylogeny of Pythium insidiosum.

Sequence analysis of the ribosomal DNA internal transcribed spacers (ITS) was used to establish phylogenetic relationships among 23 isolates of Pythium insidiosum, the etiological agent of pythiosis in mammals. The isolates were divided into three distinct clades that exhibited significant geographic isolation. Clade I consisted of isolates from North, Central, and South America, while clade II contained isolates from Asia and Australia. Also present in clade II was an isolate from a patient in the USA, but the origin of the infection may have been in the Middle East. Clade III was comprised of isolates from Thailand and the USA. All 23 P. insidiosum isolates were more closely related to each other than to any other Pythium species in this study. Additionally, all Pythium isolates formed a clade separate from both outgroup species, Phytophthora megasperma and Lagenidium giganteum. The ITS sequence results tend to support the existence of geographic variants or cryptic speciation within P. insidiosum. The sequence information obtained also provides an abundance of data for applications in the diagnosis of pythiosis and identification of P. insidiosum from clinical samples.

Animals↗

PubMed2003

Identification of Pythium oligandrum using species-specific ITS rDNA PCR oligonucleotides.

Pythium oligandrum is a parasite of cultivated Agaricus bisporus. Infection results in significant yield reductions and a disease referred to as 'black compost'. In this study, P. oligandrum isolates were isolated from New Zealand mushroom composts, and their ribosomal DNA internal transcribed spacer (ITS) regions were amplified using the polymerase chain reaction (PCR). ITS nucleotide sequences obtained from New Zealand P. oligandrum isolates were compared with those previously identified P. oligandrum isolates and 23 described Pythium species. Although New Zealand P. oligandrum isolates had high ITS nucleotide identity with internationally identified P. oligandrum, the order of nucleotides in some regions varied when compared with other Pythium species. These varied nucleotides within the ITS region were used to design PCR primers (P.OLIG.F1 and P.OLIG.R04) for the specific amplification of a 384-bp fragment from P. oligandrum DNA. P.OLIG.F1 and P.OLIG.R04 were used to identify a major source of P. oligandrum inoculation on a New Zealand mushroom farm. Application of this diagnostic test will assist farming strategies implemented to prevent future P. oligandrum outbreaks. Furthermore, results presented identify a need for species resolution between P. oligandrum and P. hydnosporum.

Agaricus↗

PubMedDate not supplied

The relationships between the toxicity and the primary and secondary structures of elicitinlike protein elicitors secreted by the phytopathogenic fungus Pythium vexans.

Elicitins are toxic and signaling proteins secreted by Phytophthora spp. responsible for the incompatible reaction and systemic hypersensitive-like necroses of diverse plant species leading to resistance against fungal or bacterial plant pathogens. Such proteins were observed in the culture filtrate of another species of the Oomycete genus, Pythium vexans. Two alpha elicitinlike proteins were purified and sequenced. One of these novel elicitins (Vex2) exhibited a 100-residue sequence instead of 98 while the other (Vex1) had an N-glycosylation site, effectively glycosylated (equivalent of 16 hexose residues). In addition to the point mutations already observed in Phytophthora species, we found several novel amino acid changes. Furthermore, circular dichroism revealed some differences in their structure in solution compared with the Phytophthora elicitins that were correlated with specific point mutations. These sequences permitted the establishment of a phylogenic tree, suggesting that Pythium vexans is a species close to the Phytophthora genus. The toxicity of the Pythium vexans elictins to tobacco leaves was investigated and correlated with the occurrence of the carbohydrate moiety of one of the two isoforms, observed for the first time in an elicitin.

Amino Acid Sequence↗

PubMed1988

Comparative studies on the DNA of Pythium species and some possibly related taxa.

DNA from 14 Pythium species, Verrucalvus flavofaciens and Zoophagus insidians was characterized by CsCl-bisbenzimide density gradients in order to investigate its taxonomic potential. A few incomplete analyses were made for other species. All clearly assignable Pythium species produced three DNA bands in the gradient. Pythium undulatum along with Verrucalvus and Zoophagus produced only two bands. Another possible exception, which needs further investigation, is P. vexans. The DNA had a relatively constant banding pattern in CsCl gradients. The small number (eight) of DNA criteria that were available were subjected to cluster analysis to assess the relationships between replicates and species. This restricted database, similar in size to the number of criteria used in morphological taxonomy, provided an independent assessment of the values that have been attached to generic and subgeneric classifications. This approach enabled assessments to be made of relationships between species that have incomplete life-histories and which therefore lack features essential for traditional taxonomic decisions.

Base Composition↗

PubMed2005

Activity of some aminoglycoside antibiotics against true fungi, Phytophthora and Pythium species.

AIMS: To investigate the in vitro antifungal and antioomycete activities of some aminoglycosides against true fungi and Phytophthora and Pythium species and to evaluate the potential of the antibiotics against Phytophthora late blight on plants. METHODS AND RESULTS: Antifungal and antioomycete activities of aminoglycoside antibiotics (neomycin, paromomycin, ribostamycin and streptomycin) and a paromomycin-producing strain (Streptomyces sp. AMG-P1) against Phytophthora and Pythium species and 10 common fungi were measured in potato dextrose broth (PDB) and on seedlings in pots. Paromomycin was the most active against Phytophthora and Pythium species with a minimal inhibitory concentration of 1-10 microg ml(-1) in PDB, but displayed low to moderate activities towards other common fungi at the same concentration. Paromomycin also showed potent in vivo activity against red pepper and tomato late blight diseases with 80 and 99% control value, respectively, at 100 microg ml(-1). In addition, culture broth of Streptomyces sp. AMG-P1 as a paromomycin producer exhibited high in vivo activity against late blight at 500 microg freeze-dried weight per millilitre. CONCLUSIONS: Among tested aminoglycoside antibiotics, paromomycin was the most active against oomycetes both in vitro and in vivo. SIGNIFICANCE AND IMPACT OF THE STUDY: Data from this study show that aminoglycoside antibiotics have in vitro and in vivo activities against oomycetes, suggesting that Streptomyces sp. AMG-P1 may be used as a biocontrol agent against oomycete diseases.

Antifungal Agents↗

PubMed2000

ITS1 region of the rDNA of Pythium megacarpum sp. nov., its taxonomy, and its comparison with related species.

Pythium megacarpum sp. nov., was isolated from a soil sample taken from a wheat field in Lille in northern France. It was mistakenly described as Pythium ostracodes Drechsler [Paul, B (1994) Cryptogam. -Mycol. 15,263-271]. Despite morphological resemblance, the comparison between the internal transcribed spacer (ITS)1 regions of the rDNA of the two fungi, leaves no doubt of their different identities. This species is unique because of its large oogonia and plerotic, thick walled oospores, its monoclinous antheridia with large antheridial cells and its lack of zoospores. The character combination of P. megacarpum and the ITS1 sequence of its rDNA, justifies the creation of a new species within the genus Pythium. The fungus is closely related to P. ostracodes. The taxonomic description of this fungus and its comparison with related species, together with the PCR of the ITS1 of its ribosomal nuclear DNA as well as the nucleotide sequences of ITS1 encoding 5.8S rRNA are discussed here.

Base Sequence↗

PubMed2001

ITS region of the rDNA of Pythium longandrum, a new species; its taxonomy and its comparison with related species.

Pythium longandrum (F-73.0) was isolated, from soil samples taken in Lille in northern France. Morphologically the fungus resembles closely Pythium rostratum, however its antheridial characters are unique. The oogonia of this species are provided with hypogynous and monoclinous antheridia. The antheridial cells are inflated and are probably the largest and longest for the genus. The internal transcribed spacer region of its nuclear ribosomal DNA indicates that it is entirely different from all other species of Pythium. This new species is characterized by its spherical to elongated sporangia, smooth-walled oogonia and hypogynous to monoclinous antheridia bearing long antheridial cells closely applied to the oogonia. Morphological features of this new species, together with the sequences of the ITS region of its nuclear ribosomal DNA and comparison with related species are discussed here.

Base Sequence↗

PubMed2002

Pythium terrestris, a new species isolated from France, its ITS region, taxonomy and its comparison with related species.

Pythium terrestris (F-78) was isolated from soil samples taken in Lille in northern France. Its morphology resembles that of Pythium rostratum, and Pythium longandrum, a recently described species. However the antheridial and sporangial characteristics of this new species are unique. The fungus does not sporulate, the sporangium germinates directly into mycelium through germ tubes. The oogonia of P. terrestris are normally provided with hypogynous and monoclinous antheridia, at times the monoclinous antheridial branches wrap around the oogonia forming a complicated knot. Morphological features of this new species, together with the sequences of the internal transcribed spacer (ITS) region of its nuclear ribosomal DNA and its comparison with related species are discussed here.

Base Sequence↗

PubMed1997

Trehalose induces antagonism towards Pythium debaryanum in Pseudomonas fluorescens ATCC 17400.

Pseudomonas fluorescens ATCC 17400 shows in vitro activity against Pythium debaryanum under conditions of iron limitation. A lacZ reporter gene introduced by transposon mutagenesis into the P. fluorescens ATCC 17400 trehalase gene (treA) was induced by a factor released by the phytopathogen Pythium debaryanum. The induction of the lacZ gene was lost upon treatment of the Pythium supernatant with commercial trehalase. A trehalose concentration as low as 1 microM could induce the expression of treA. The mutation did not affect the wild-type potential for fungus antagonism but drastically decreased the osmotolerance of the mutant in liquid culture and suppressed the ability of P. fluorescens ATCC 17400 to utilize trehalose as a carbon source. A subsequent transposon insertion in treP, one of the trehalose phosphotransferase genes upstream of treA, silenced the lacZ gene. This double mutant restricted fungal growth only under conditions of high osmolarity, which probably results in internal trehalose accumulation. These data confirm the role of the disaccharide trehalose in osmotolerance, and they indicate its additional role as an initiator of or a signal for fungal antagonism.

Amino Acid Sequence↗

PubMed2000

Fatty acid competition as a mechanism by which Enterobacter cloacae suppresses Pythium ultimum sporangium germination and damping-off.

Interactions between plant-associated microorganisms play important roles in suppressing plant diseases and enhancing plant growth and development. While competition between plant-associated bacteria and plant pathogens has long been thought to be an important means of suppressing plant diseases microbiologically, unequivocal evidence supporting such a mechanism has been lacking. We present evidence here that competition for plant-derived unsaturated long-chain fatty acids between the biological control bacterium Enterobacter cloacae and the seed-rotting oomycete, Pythium ultimum, results in disease suppression. Since fatty acids from seeds and roots are required to elicit germination responses of P. ultimum, we generated mutants of E. cloacae to evaluate the role of E. cloacae fatty acid metabolism on the suppression of Pythium sporangium germination and subsequent plant infection. Two mutants of E. cloacae EcCT-501R3, Ec31 (fadB) and EcL1 (fadL), were reduced in beta-oxidation and fatty acid uptake, respectively. Both strains failed to metabolize linoleic acid, to inactivate the germination-stimulating activity of cottonseed exudate and linoleic acid, and to suppress Pythium seed rot in cotton seedling bioassays. Subclones containing fadBA or fadL complemented each of these phenotypes in Ec31 and EcL1, respectively. These data provide strong evidence for a competitive exclusion mechanism for the biological control of P. ultimum-incited seed infections by E. cloacae where E. cloacae prevents the germination of P. ultimum sporangia by the efficient metabolism of fatty acid components of seed exudate and thus prevents seed infections.

Base Sequence↗

PubMed2002

An antifungal protein from the marine bacterium streptomyces sp. Strain AP77 is specific for Pythium porphyrae, a causative agent of red rot disease in Porphyra spp.

A novel antifungal protein (SAP) was found in the culture supernatant of a marine bacterium, Streptomyces sp. strain AP77, and was purified. This protein was characterized by chemical, biochemical, and biological analyses. By using gel filtration, the molecular mass of SAP was estimated to be 160 kDa. Structural analysis of SAP by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry suggested that SAP is composed of three heterologous protein subunits of 41.7 kDa (SAP1), 21.7 kDa (SAP2), and 18.7 kDa (SAP3) at a molar ratio of 1:1:5 (or 1:1:6). N-terminal amino acid sequence analysis and a homology search revealed that SAP1, SAP2, and SAP3 exhibit 64.3, 68.4, and 86.7% similarity to three Streptomyces coelicolor polypeptides, puromycin resistance protein (Pur8), a conserved hypothetical protein, and bacterioferritin, respectively. The MIC of purified SAP against Pythium porphyrae was determined to be 1.6 microg/disk, whereas no inhibitory effect was observed at concentrations up to 100 microg/disk against most of the fungal and bacterial strains tested; the only exception was relatively strong antifungal activity against Pythium ultimum (MIC, 6.3 microg/disk). In vitro and in vivo toxicity tests demonstrated that SAP showed no toxicity against Porphyra yezoensis cells, human normal dermal fibroblasts, and mice at doses up to 700 microg/ml (for 24 h), 250 microg/ml (for 12 h), and 75 mg/kg (for 35 days), respectively. SAP was labile when it was subjected to a heated-air drying treatment, which is a great advantage in food production procedures. These results indicated that Streptomyces sp. strain AP77 might be useful as a gene source for safe transgenic Porphyra breeding for tolerance to Pythium infection.

Amino Acid Sequence↗

PubMed2002

Ability of nonpathogenic Fusarium oxysporum strain Fo47 to induce resistance against Pythium ultimum infection in cucumber.

The influence exerted by nonpathogenic Fusarium oxysporum strain Fo47 in triggering cucumber protection against infection by Pythium ultimum was investigated ultrastructurally. Macroscopic and microscopic observations of the pathogen colony in dual cultures revealed that reduction of Pythium growth was associated with marked disorders, including generalized disorganization of the host cytoplasm, retraction of the plasmalemma, and complete loss of the protoplasm. Cytochemical labeling of cellulose with an exoglucanase-gold complex showed that the cellulose component of the host cell walls was structurally preserved at a time when the host cytoplasm had undergone complete disorganization. A similar antagonistic process was observed at the root cell surface. Most striking and interesting was the finding that mycoparasitism, as evidenced by the frequent occurrence of Fo47 hyphae within nearly empty cells of the pathogen, occurred not only at the root surface but also within the invaded root tissues. The specific labeling pattern obtained with the exoglucanase-gold complex confirmed that Fo47 successfully penetrated cells of the pathogen, both in the rhizosphere and inside the root tissues. Pythium cells that could evade the first defensive line in the rhizosphere could penetrate the root epidermis, but their growth was restricted to the outermost tissues. Positive correlations between Fo47 treatment and induced resistance to infection by P. ultimum in cucumber were confirmed by (i) the reduction of pathogen viability; (ii) the elaboration of newly formed barriers, a phenomenon which was not seen in Fo47-free plants, where the pathogen proliferated in all root tissues within a few days; and (iii) the occlusion of intercellular spaces with a dense material likely enriched in phenolics. Taken together, our observations provide the first convincing evidence that Fo47 exerts a direct inhibitory effect on P. ultimum through a combination of antibiosis and mycoparasitism, in addition to being a strong inducer of plant defense reactions.

Antibiosis↗