Especies oportunistas de importancia clínica de los géneros bipolaris shoemaker y curvularia boedijn: su caracterización bajo los nuevos criterios taxonómicos / Opportunistic species of clinical relevance in the bipolaris shoemaker & curvularia boedijn genera: its characterization under newest taxonomic criteria

Los géneros fúngicos Bipolaris y Curvularia,incluyen numerosas especies fitopatógenas,saprotrofas y algunas oportunistas emergentes enel ser humano como en otros animales. La distribuciónecológica de ambos taxas es cosmopolita enla gran mayoría de sus integrantes, encontrándosefrecuentemente en hojas, tallos y raíces de diferentespastos, en especial Poaceae, pero tambiénen una gran variedad de dicotiledoneas, aire y sueloen todos los continentes. Cochliobolus, Bipolarisy Curvularia integran un complejo de especiestaxonómicamente confuso, debido a los constantescambios en la nomenclatura de algunos de sus integrantesasexuales (Bipolaris y Curvularia), loscuales se han diferenciado principalmente en basea la morfología de sus conidios, situación a vecesmuy dificultosa debido a que en ambos génerosalgunas especies presentan similares característicasconidiales. Una cuidadosa identificación y unanominación precisa de las especies es crucial parael acceso a la información referente en la literaturamoderna, que gracias a la biología molecularha resuelto muchas de las dudas de la taxonomíabasada solo en el fenotipo.Las especies de ambos géneros (principalmenteCurvularia) pueden afectar a pacientesinmunocomprometidos e inmunocompetentes,en especial B. cynodontis, C. australiensis C.hawaiensis y C. spicifera, causando cuadros clínicosdiversos ya sea superficiales o profundos enmuchas áreas geográficas...

The fungal genera Bipolaris and Curvularia,include numerous plant pathogenic species,saprophitic and some emerging opportunistic inman and other animals. The ecological distributionof both taxa is cosmopolitan in the vast majority ofits members, often being in leaves, stems and rootsof different grasses, especially Poaceae, but also ina variety of dicotyledonous, air and soil on all continents. Cochliobolus, Bipolaris and Curvulariaintegrate a complex of species taxonomically confusingdue to the constant changes in the nomenclatureof some of its asexual members (Bipolarisand Curvularia), which are differentiated mainlybased on the morphology of its conidia, a situationsometimes very difficult because in both generaconidial of some species have similar characteristics. Careful identification and a precise nominationof species is crucial for access to informationconcerning in modern literature, actually thanks tomolecular biology has solved many of the doubtsof taxonomy based only on the phenotype.The species of both genera (mainly Curvularia)can affect immunocompetent or debilitatepatients, especially B. cynodontis, C. australiensis,C. hawaiensis and C.spicifera, causing varioussuperficial or systemic clinical cases in manygeographic areas...

The mycological and molecular study of Hortaea werneckii isolated from blood and splenic abscess.

Hortaea werneckii is an environmental dematiaceous fungus found in the halophilic environment. It causes tinea nigra. We report the isolation of H. werneckii from blood and splenic abscess of two patients with acute myelomonocytic leukaemia. H. werneckii grew at room temperature but not at 37 degrees C, it was identified by biochemical tests, growth characteristics and the presence of conspicuous collarette intercalary on dividing yeast cells. The use of specific oligonucleotide primer Hor-F (5'-TGGACACCTTCA TAACTCTTG-3') and Hor-R (5'-TCACAACGCTTAGAGACGG-3') confirmed the two isolates were H. werneckii. The sequence for 281 nucleotide of HW299 and HW403 were 99% identical but differed only in one nucleotide. In vitro anti-fungal susceptibility testing showed that the isolates were resistant to amphotericin B and flucytosine.

Production of a monoclonal antibody specific to the genus Trichoderma and closely related fungi, and its use to detect Trichoderma spp. in naturally infested composts.

Studies of the interactions between hyperparasitic fungi and their hosts are severely hampered by the absence of methods that allow the unambiguous identification of individual genera in complex environments that contain mixed populations of fungi, such as soil or compost. This study details the development of a monoclonal antibody (MF2) that allows the detection and recovery of Trichoderma spp. in naturally infested composts, and the visualization of hyperparasitic strains of Trichoderma during antagonistic interactions with their hosts. Murine monoclonal antibody MF2, of immunoglobulin class M (IgM), was raised against a protein epitope of a glycoprotein antigen(s) specific for species of the genus Trichoderma and for the closely related fungi Gliocladium viride, Hypomyces chrysospermus, Sphaerostilbella spp. and Hypocrea spp. MF2 did not react with antigens from Gliocladium catenulatum, Gliocladium roseum, Nectria ochroleuca and Clonostachys spp., nor with a range of unrelated soil- and compost-borne fungi. Extracellular production of the MF2 antigen was constitutive. Western-blotting analysis showed that MF2 bound to a ladder of proteins with apparent molecular masses in the range 35-200 kDa. Immunofluorescence studies showed that MF2 bound strongly to the cell walls of hyphae and phialides and the intercalary and terminal chlamydospores of Trichoderma spp., whereas immunogold electron microscopy revealed strong binding of MF2 to the cell walls and septa of hyphae and to the cell walls of phialoconidia. In immunofluorescence studies of dual cultures of Trichoderma and Rhizoctonia solani, only the cell walls of the hyperparasite, which coiled around the host, were stained by MF2. The specificity of MF2 enabled the development of a combined baiting-ELISA technique for the detection of Trichoderma spp. in naturally infested composts. The specificity of this technique was confirmed by phylogenetic analysis based on sequences of the ITS1-5.8S-ITS2 rRNA-encoding regions of the isolates.

Compartmentalization and transport in beta-lactam antibiotic biosynthesis by filamentous fungi.

A proper description of the biosynthesis of fungal beta-lactam antibiotics requires detailed knowledge of the cell biology of the producing organisms. This involves a delineation of the compartmentalization of the biosynthetic pathways, and of the consequential transport steps across the cell-boundary plasma membrane and across organellar membranes. Of the enzymes of the penicillin biosynthetic pathway in Penicillium chrysogenum and Aspergillus nidulans, delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase (ACVS) and isopenicillin N synthase (IPNS) probably have a cytosolic location. Acyl-coenzyme A:isopenicillin N acyltransferase (IAT) is located in microbodies. Of the two enzymes that may be involved in activation of the side chain, acetyl-coenzyme A synthetase (ACS) is located in the cytosol, and phenylacetyl-coenzyme A ligase (PCL) is probably located in the microbody. All enzymes of the cephalosporin biosynthesis pathway in Cephalosporium acremonium probably have a cytosolic location. The vacuole may play an ancillary role in the supply of precursor amino acids, and in the storage of intermediates. The distribution of precursors, intermediates, end- and side-products, the transport of nutrients. precursor, intermediates and products across the plasma membrane, and the transport of small solutes across organellar membranes, is discussed. The relevance of compartmentalization is considered against the background of recent biotechnological innovations of fungal beta-lactam biosynthesis pathways.

Fine structure and cytochemical study of the interaction between Fonsecaea pedrosoi and rat polymorphonuclear leukocyte.

The induction of a granulomatous reaction is frequently observed in subcutaneous mycoses. Our previous studies demonstrated that Fonsecaea pedrosoi was able to survive and proliferate in tissue macrophages and that activated macrophages were fungistatic but not fungicidal. By contrast, our present studies revealed that neutrophils were able to kill F. pedrosoi cells in periods shorter than 20 min. Several phases of the interaction process were analysed by light and electron microscopy. The kinetic analysis demonstrated no significant difference during the first hour of F. pedrosoi-neutrophil interaction. Electron microscopy images showed that neutrophils readily associated with and killed extracellular fungi; however, few fungi were ingested. During this process the activation of respiratory burst took place as evaluated by light and electron microscopy. Cytochemical activity of acid and alkaline phosphatase was detected in low levels during the host cell parasite interaction.

Subcutaneous mycosis in a cat due to Staphylotrichum coccosporum.

A 5.5-year-old, male, feline leucosis virus-positive cat developed a concurrent dermatophytosis due to Microsporum canis and a subcutaneous infection due to Staphylotrichum coccosporum. St. coccosporum caused mycetoma-like lesions. The fungal elements revealed features like those seen in phaeohyphomycosis. Until now St. coccosporum has been described to be non-pathogenic. The pathogenicity of St. coccosporum was corroborated by experimental infection.

Biochemical characterization and ultrastructural localization of two extracellular trypsins produced by Metarhizium anisopliae in infected insect cuticles.

Proteinase 2 (Pr2) is a fungal (Metarhizium anisopliae) serine proteinase which has a tryptic specificity for basic residues and which may be involved in entomopathogenicity. Analytical and preparative isoelectric focusing methods were used to separate two trypsin components, produced during growth on cockroach cuticle, with isoelectric points of 4.4 (molecular mass, 30 kDa) and 4.9 (27 kDa). The catalytic properties of the proteases were analyzed by their kinetic constants and by a combination of two-dimensional gelatin-sodium dodecyl sulfate-polyacrylamide gel electrophoresis and enzyme overlay membranes. Both Pr2 isoforms preferentially cleave at the carboxyl sides of positively charged amino acids, preferring arginine; the pI 4.4 Pr2 isoform also possessed significant activity against lysine. Compared with the pathogen's subtilisin-like enzyme (Pr1), the pI 4.4 Pr2 isoform shows low activity against insoluble proteins in a host (Manduca sexta) cuticle. However, it degrades most cuticle proteins when they are solubilized, with high-molecular-weight basic proteins being preferentially hydrolyzed. Polyclonal antibodies raised against each Pr2 isoform were isotype specific. This allowed us to use ultrastructural immunocytochemistry to independently visualize each isoform during penetration of the host (M. sexta) cuticle. Both isoforms were secreted by infection structures (appressoria) on the cuticle surface and by the penetrant hyphae within the cuticle. The extracellular sheath, which is commonly observed around fungal cells, often contained Pr2 molecules. Intracellular labelling was sparse.

Taxol from Pestalotiopsis microspora, an endophytic fungus of Taxus wallachiana.

Pestalotiopsis microspora was isolated from the inner bark of a small limb of Himalayan yew, Taxus wallachiana, and was shown to produce taxol in mycelial culture. Taxol was identified by spectroscopic and chromatographic comparisons with authentic taxol. Optimal taxol production occurred after 2-3 weeks in still culture at 23 degrees C. [14C]Acetate and [14C]phenylalanine served as precursors for fungal [14C]taxol. These observations on P. microspora are discussed in relation to the biological importance of taxol production by fungi in general.

Ovadendron sulphureo-ochraceum endophthalmitis after cataract surgery.

PURPOSE: We examined an 82-year-old woman with delayed-onset endophthalmitis caused by an opportunistic pathogen, Ovadendron sulphureo-ochraceum. METHODS: Tissue obtained during vitrectomy was cultured and examined by light and electron microscopy. An enucleation specimen was examined by light microscopy. RESULTS: The patient had fungal endophthalmitis, with O. sulphureo-ochraceum present in the lens capsule. The eye developed a necrotizing scleritis secondary to O. sulphureo-ochraceum. The patient failed to respond to intravitreous, subconjunctival, and systemic amphotericin B, and the eye was enucleated. CONCLUSION: In this case of O. sulphureo-ochraceum as a human pathogen, the organism caused endophthalmitis after cataract extraction.

A novel bioactive delta lactone FD-211. Taxonomy, isolation and characterization.

During our screening program for natural product drugs effective against multidrug-resistant mammalian cells, we have discovered a new delta lactone FD-211 from the fermantation broth of Myceliophthora lutea TF-0409. FD-211 had a broad spectrum activity against cultured tumor cell lines, including adriamycin-resistant HL-60 cells.

Scopulariopsis endocarditis associated with Duran ring valvuloplasty.

Fungal endocarditis is rare and is usually caused by Aspergillus and Candida species. We present a patient with endocarditis caused by Scopulariopsis brevicaulis. The patient had a history of mitral valve disease and, 1 year earlier, had undergone valvuloplasty with the placement of a prosthetic Duran ring in the mitral valve position. S. brevicaulis was cultured from samples of a large vegetation on the mitral valve apparatus. The mitral valve was replaced with a St. Jude mechanical prosthesis. The patient was treated with amphotericin B but was later switched to oral itraconazole when antibiotic tests indicated susceptibility to that agent. We believe this is the 1st reported case of endocarditis caused by Scopulariopsis.

Does the eukaryotic large ribosomal subunit have a channel passing through it?

The large ribosomal subunit of the thermophilic fungus Thermomyces lanuginosus was treated with 2.96 M NH4Cl to remove specific complements of ribosomal proteins, and the core particles thereby derived were imaged by bright field transmission electron microscopy, and recurring views computed by single particle electron image analysis. A new characteristic projection was elucidated which showed a large depression or channel passing through the subunit. Such a channel has been perceived in the prokaryotic large ribosomal subunit under certain conditions and has been postulated to be the exit pathway for the nascent polypeptide chain, but its existence has not hitherto been demonstrated in eukaryotes.

Presence of double-stranded RNA and virus-like particles in Phaffia rhodozyma.

Four double-stranded RNA (dsRNA) molecules were isolated from Phaffia rhodozyma UCD 67-385. Their molecular sizes were approximately 4.3, 3.1, 0.9 and 0.75 kilobase pairs (kbp) as determined by agarose-gel electrophoresis and they were designated as L, M, S1 and S2, respectively. By differential centrifugation in sucrose gradients, these dsRNAs copurified with isometric virus-like particles 36 nm in diameter. A cured strain, UV-S2, lacking the S2-dsRNA was obtained from P. rhodozyma UCD 67-385 by ultraviolet (UV) light treatment. UV-S2 strain contains identical virus-like particles to those from the wild-type strain, as determined by electron microscopy, suggesting that the S2-dsRNA was not essential for the expression of mycovirus structural polypeptides. On the other hand, both the UCD 67-385 and UV-S2 strains were able to kill P. rhodozyma UCD 67-383, a strain without dsRNAs. These results suggest that the dsRNA molecules also encode a killer system. Finally, the UV-S2 strain maintains killer ability, which suggests that S2-dsRNA is not involved in the killer phenotype expression.

Scedosporium inflatum osteomyelitis in a dog.

Scedosporium inflatum Malloch et Salkin was found to cause osteomyelitis in a 6-year-old spayed female beagle. The previously healthy dog suddenly developed right-forelimb lameness. Bony changes consisting of proliferation with some lysis were noted on radiographic examinations. Microscopic observations of stained sections of tissue obtained by biopsy of the distal humerus revealed the presence of septate branching hyphae. Cultures inoculated with tissue from a later biopsy yielded a mold subsequently identified as S. inflatum. Tissue sections stained with specific Scedosporium fluorescent-antibody conjugate were positive, further substantiating the diagnosis. Although the dog was treated with oral itraconazole, no improvement in the animal's condition was noted, and it was euthanized. Autopsy revealed dissemination of the etiologic agent to the lungs.

Fatal disseminated infection caused by Myceliophthora thermophila, a new agent of mycosis: case history and laboratory characteristics.

We report a case of human infection caused by the hyphomycete Myceliophthora thermophila. A 7-year-old male with neurofibromatosis (type 1) was diagnosed in 1987 with acute myeloblastic leukemia associated with the chromosomal abnormality monosomy 7. The patient experienced multiple serious infections over a three-year period before expiring in 1990 while in the end stage of leukemia. Autopsy findings included fungal vegetations of the left atrium, ascending aorta, and pulmonary arteries and fungal invasion of both lungs. Cultures yielded M. thermophila. We believe that this is the first reported fatality caused by M. thermophila.

[Mycetoma with black granules: first case in a native of Israel]. / Myzetom mit schwarzen Granula: erstmals bei einem gebürtigen Israeli.

We present a case of black grain mycetoma caused by Madurella mycetomatis. The diagnosis was made on the basis of conventional light microscopy and mycological studies. After 12 months of oral therapy with ketoconazole, the patient showed partial response. This is the first native Israeli case of black grain mycetoma.

Cell surface redox potential as a mechanism of defense against photosensitizers in fungi.

The phytotoxin cercosporin, a singlet oxygen-generating photosensitizer, is toxic to plants, mice, and many fungi, yet the fungi that produce it, Cercospora spp., are resistant. We hypothesize that resistance to cercosporin may result from a reducing environment at the cell surface. Twenty tetrazolium dyes differing in redox potential were used as indicators of cell surface redox potential of seven fungal species differing in resistance to cercosporin. Resistant fungi were able to reduce significantly more dyes than were sensitive fungi. A correlation between dye reduction and cercosporin resistance was also observed when resistance levels of Cercospora species were manipulated by growth on different media. The addition of the reducing agents ascorbate, cysteine, and reduced glutathione (GSH) to growth media decreased cercosporin toxicity for sensitive fungi. None of these agents directly reduced cercosporin at the concentrations at which they protected fungi. Spectral and thin-layer chromatographic analyses of cercosporin solutions containing the different reducing agents indicated that GSH, but not cysteine or ascorbate, reacted with cercosporin. Resistant and sensitive fungi did not differ in endogenous levels of cysteine, GSH, or total thiols. On the basis of data from this and other studies, this report presents a model which proposes that cercosporin resistance results from the production of reducing power at the surfaces of resistant cells, leading to transient reduction and detoxification of the cercosporin molecule.

Eumycetoma caused by Curvularia lunata in a dog.

Curvularia lunata was cultured from black granules found in granulomatous tumefactions excised from the subcutis of a three year old Medium Schnauzer dog. Draining sinuses were present in some of the tumefactions. Accordingly the diagnosis of eumycotic mycetoma was made. This diagnosis was confirmed by histopathological examination. During the four years following the first surgical intervention, several more similar tumefactions were excised on three different occasions. The dog died of chronic renal failure at the age of 8 years. There was no bone involvement or visceral diffusion of the fungus. The granules were examined by scanning electron microscopy. Immunoglobulins in the dog's serum, assessed by a qualitative test, proved to be equal to immunoglobulins in the serum of a control dog. Precipitating antibodies against C. lunata were not found. The dog was treated for 150 days with itraconazole. In spite of good initial results, recurrence of the fungal lesions were observed after the treatment's interruption. Further treatment with itraconazole for 45 days proved ineffective. No side effects of the drug were observed. This is, to the best of our knowledge, the first case in which C. lunata is identified as the causative agent of an animal eumycetoma.

Subcutaneous phaeohyphomycosis caused by Bipolaris hawaiiensis: a case report

Os autores descrevem um caso de feohifomicose com lesao verrucosa no halux esquerdo. O paciente nao apresentava sinais clinicos de deficiencia imunologica. Os exames direto e histopatologico mostraram celulas leveduriformes e poucas hifas septadas, demacioides, essenciais ao diagnostico desta micose. O cultivo em lamina permitiu identificar o hifomiceto como Biopolaris hawaiiensis.