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PubMed2005

External ultrastructure of fruit body initiation in Morchella.

The external morphological changes occurring during initiation and early stages of fruit body development of a Morchella sp., before the development of asci, were examined by scanning electron microscopy. Four stages of primordial development were distinguished. First, disk-shaped knots of 0.5-1.5 mm were observed on the surface of the substrate. Next, the knot inflated and a primordial stem emerged from its centre. Afterward, the stem lengthened, oriented upward, and two types of hyphal elements developed: long, straight and smooth basal hairy hyphae and short stem hyphae, some of which were inflated and projected out of a cohesive layer of tightly packed hyphal elements. Finally, when the stem was 2-3 mm long, pre-apothecia emerged in the apical end, with ridges and pits having distinguished types of paraphyses. Extracelluar mucilage covered the ridge layer and helped give the tissue its shape and rigidity.

Ascomycota↗
PubMed2005

The systematic relevance of conidiogenesis modes in the gilled Agaricales.

Dikaryotic and haploid mycelia of more than 150 gilled species of euagarics were studied morphologically and by molecular phylogenetic methods. The morphological investigations revealed anamorphs in more than 90 species that were often specific at the genus or family level. Thallic conidiogenesis dominated and varied from fragmentation of normally branched hyphae to the formation of differentiated sympodially branched conidiophores. Secession modes, coiling of the conidiogenous hyphae or the swelling of the conidia were additional distinguishing features. Phylogenetic analysis of the D1-D3 domains of the nuclear gene for the ribosomal large subunit using a Bayesian Markov chain Monte Carlo approach resulted in several well-supported groups that are consistent with anamorph morphology. These results indicate that the anamorphs provide valuable characters for a natural classification of the Agaricales.

Agaricales↗
PubMed2002

Observation of woronin bodies in Arthrinium aureum by scanning electron microscopy.

Woronin bodies are cytoplasmic organelles of filamentous fungi that can be observed on one, or both sides of each septum. The goal of this paper is to illustrate the presence of them in hyphae of Arthrinium aureum by means of scanning electron microscopy and to show that they act as a safety plug to close septa pores in hypha. Results show that Woronin bodies as an immediate response to prevent a cytoplasm loss. Results support hypothesis proposed previously in literature.

Hyphae↗
PubMed2003

In-situ visualisation of hyphal structure and arrangement in mycoprotein pastes.

A novel method to examine the morphology and structure of fungal hyphae in solid pastes used for the production of meat alternative product is presented. A sample of fermentation broth was fluorescently stained with Calcofluor White and added back to the broth, mixed and then a paste made using ultra-filtration. Fibre visualisation was by fluorescence microscopy and quantification by manual image analysis. This method enables the determination of fibre length and orientation within the paste. Imaging of the hypha fibre paste proved that its structure was 'isotropic', i.e. that fibres were randomly oriented in all directions. Processing of the paste altered the orientation of the fibres, the method was able to identify and quantify the changes in fibre position.

Benzenesulfonates↗
PubMed2003

Primary aspergillosis affecting the tongue of a leukemic patient.

We describe a case of primary aspergillosis involving the tongue of a patient with acute myeloid leukemia. Intraoral aspergillosis is very rare and we found only 23 cases reported in the English literature. Clinically it was a 2-cm, ulcerated, grayish lesion on the dorsum of the tongue. Microscopically there was invasion of the epithelium, connective tissue and muscle of the tongue by fungal hyphae branching at 45 degrees angle. The large hyphae were easily seen by H & E stain, and were strongly positive for periodic acid-Schiff and Grocott methenamine. The patient was successfully treated with intravenous amphotericin B. Based on clinical, microscopic and culture data, the diagnosis of primary aspergillosis of the tongue was established. Invasive oral aspergillosis is a potentially lethal disease and it should be considered in immunosuppressed patients.

Acute Disease↗
PubMed2002

Candida attachment to oral epithelium.

Inflamed oral mucosa biopsies from patients with thrush and high candidal density were observed in a transmission electron microscope (TEM) using ultra-histochemical staining with ruthenium red for glycocalyx visualization. Fimbriae comprising the glycocalyx and enabling yeast adhesion to epithelial cells were clearly visualized by ruthenium red. All internalized portions of the yeast walls were devoid of glycocalyx, indicating that the growing tips of the hyphae mechanically penetrated the host cells. The attachment of Candida occurred in two ways: by fimbria-mediated adhesion enabling colonization of the epithelial surface, and by invasion of the superficial epithelial cells via hyphae. As the interaction between adhesin receptors and adhesins stimulates the production of proinflammatory cytokines, Candida adhesion itself is assumed to induce mucosal inflammation.

Adult↗
PubMed2004

Candida yeasts in chronic periodontitis tissues and subgingival microbial biofilms in vivo.

The frequency of Candida infection in periodontal tissues of chronic periodontitis (CP) patients and the extent of candidal penetration into gingival tissues was studied. Tissue specimens collected from 25 CP patients during periodontal flap operations of initial periodontal therapy were examined by immunohistochemistry using Candida albicans-specific antibodies. Sections were also stained with periodic acid-Schiff (PAS) and subgingival plaque samples from 17 patients were cultured. Immunoreactivity for Candida was present in four of the 25 CP specimens (16%). Only one yeast-positive specimen was found when PAS-staining was used (4%) and two yeast-positive specimens were found with plaque culture (8%). Hyphal formation seemed essential and candidal hyphae were found to extend into the periodontal connective tissue. The degree and type of inflammation adjacent to the hyphae varied from a site and a patient to another. The sensitivity of specific antibodies was superior to PAS-stain or plaque culture in detection of Candida in tissues.

Biofilms↗
PubMed2002

Antimicrobial activity of methyl cis-7-oxo deisopropyldehydroabietate on Botrytis cinerea and Lophodermium seditiosum: ultrastructural observations by transmission electron microscopy.

AIMS: To study the antifungal activity of methyl cis-7-oxo-deisopropyldehydroabietate (MCOD) against phytopathogenic fungi, Botrytis cinerea and Lophodermium seditiousm. The effect of the compound was studied by transmission electron microscopy (TEM) and the composition of sterols on both treated and untreated cultures was determined. METHODS AND RESULTS: MCOD was tested at concentrations in the range 0.003-0.5% by the agar plate dilution method. The radial growth of the colonies treated with MCOD was measured against colonies from untreated cultures. The radial growth of colonies of both fungi and the spore germination of B. cinerea were partially or completely inhibited. Fragments of active growing colonies treated and untreated with MCOD were submitted to the conventional procedure for ultrastructural observation by TEM. Observations by TEM on colonies of B. cinerea and L. seditiosum under 0.1% MCOD revealed several autophagic-like vacuoles, morphological alterations on lomasome and lipid accumulations in the apical zone of hyphae of both fungi. Observations on spore germination of B. cinerea revealed the presence of strongly stained lipid accumulations retained by vacuoles at the cell periphery of young hyphae. The sterol composition of B. cinerea and L. seditiosum was determined on MCOD treated and untreated cultures by gas-chromatography/mass-spectrometry (GC-MS) with molecular ions and fragmentation patterns characteristics of ergosterol (M+396) and dihydroergosterol (M+398) in both fungi. CONCLUSIONS: The morphological alterations are consistent with an unspecific mode of action of MCOD causing inhibition of normal growth or damaging the fungi cells. TEM observations suggest a mechanism of resistance based on the retention of MCOD by the lipid accumulation. SIGNIFICANCE AND IMPACT OF THE STUDY: The results obtained in the present work afforded a better understanding of the mode of action of a resin acid derivative on phytopathogenic fungi. The inhibition growth of both fungi by MCOD demonstrates the antifungal activity of this compound and the interest on further in vivo studies, in order to evaluate its potential as a benign alternative to conventional fungicides.

Abietanes↗
PubMed2003

Retention of fluorescent probes during aldehyde-free anhydrous freeze-substitution.

Fluorescent probes are widely used for microscopy of live-cell processes, but few such probes can also be used with classically fixed or otherwise immobilized material, and none has been used without aldehyde fixation, which can introduce artefacts of structure and probe localization. Here we show that the fluorescence patterns in fungal hyphae loaded with chloromethyl aminocoumarin (CMAC), and then anhydrously freeze-substituted, without any aldehyde fixation, are similar to those seen in living hyphae. Probe loss into the mounting medium (Spurr's resin) with CMAC and five other probes tested indicated that some unwanted solubilization of probe occurred during embedding, but nevertheless vacuoles could be imaged by their retention of probe.

Aldehydes↗
PubMed2003

Antifungal activity of human polymorphonuclear and mononuclear phagocytes against non-fumigatus Aspergillus species.

Human phagocytic defenses against non-fumigatus aspergilli were compared with those against Aspergillus fumigatus. Monocyte-derived macrophages exhibited lower phagocytic capacities against non-fumigatus aspergilli, particularly A. nidulans and A. niger, compared with A fumigatus (P < 0.05). Non-opsonized hyphae suppressed oxidative burst (as measured by superoxide anion production) of polymorphonuclear leukocytes (PMNs). Further, these cells responded with less vigorous oxidative burst to serum-opsonized hyphae of non-fumigatus Aspergillus species, particularly A. flavus and A. terreus, compared with A. fumigatus (P < or = 0.05). Similarly, PMNs induced less hyphal damage assessed by XTT colorimetric assay to non-fumigatus species, particularly A. flavus and A. nidulans, compared with A. fumigatus (P < 0.05). Thus, non-fumigatus aspergilli are generally more resistant to mononuclear and polymorphonuclear phagocytes than A. fumigatus, a finding which should be considered during management of invasive aspergillosis caused by these species.

Adult↗
PubMed2005

Calibration of quantitative real-time TaqMan PCR by correlation with hyphal biomass and ITS copies in mycelia of Piloderma croceum.

DNA-based quantification methods such as real-time TaqMan PCR allow a rapid and highly sensitive species-specific quantification of isolated fungal DNA material, but most quantification systems are only able to measure relative amounts of biomass or biomass changes during different treatments. In this experiment, an already established DNA quantification system for the ectomycorrhizal fungus Piloderma croceum, based on the ITS region of ribosomal DNA, was calibrated to absolute biomass to obtain a direct correlation between mycelial biomass and isolated ITS copies. Thin layers of sterile mycelia were cultured on slides. The mycelial biomass was calculated from measurements of the total hyphal length using image analysis, followed by determination of the mycelial volume, and multiplied by the specific weight of hyphae obtained from literature data. Using the very same mycelium, the number of ITS copies was quantified by TaqMan PCR. The mean value of 1047 (+/- 185) copies per mm hypha results in possible data for a direct conversion: one billion (10 (9)) ITS copies corresponded to 0.79 mg hyphal dry weight. For the ribosomal ITS multi-copy genes, the number of ITS copies could be calculated to approx. 152 (+/- 26) copies per dikaryotic cell. These conversion data now allow determination of the mycelial biomass of Piloderma croceum using real-time TaqMan PCR, a prerequisite for competition experiments with Piloderma croceum.

Basidiomycota↗
PubMed2002

Detection of fungal organisms in eosinophilic mucin using a fluorescein-labeled chitin-specific binding protein.

BACKGROUND: The ability to identify fungal hyphae in patients with chronic rhinosinusitis (CRS) has been inconsistent. A new fluorescein-labeled staining method targets chitin found in fungal cell walls. OBJECTIVE: We hypothesize that this method would be able to more consistently detect fungi within the mucin of CRS patients. METHODS: Fifty-four consecutive CRS surgical patients were evaluated. After ensuring sensitivity and specificity of this new method, all specimens were stained with either fluorescein-labeled chitinase or Grocott methanamine silver stain for comparison. RESULTS: All 54 specimens contained eosinophilic mucin on hematoxylin and eosin staining. One or more fungal hyphae could be visualized within the mucin of 54 (100%) of 54 specimens stained using the fluorescein-labeled chitinase. Only 41 (76%) of 54 of the specimens stained with the Grocott methanamine silver stain technique demonstrated fungi. CONCLUSION: The fluorescein-labeled chitinase-staining technique has greater sensitivity in detecting fungal organisms within eosinophilic mucin. Fungal organisms are present in the mucin of CRS patients.

Adolescent↗
PubMed2006

Identification of the mstE gene encoding a glucose-inducible, low affinity glucose transporter in Aspergillus nidulans.

The mstE gene encoding a low affinity glucose transporter active during the germination of Aspergillus nidulans conidia on glucose medium has been identified. mstE expression also occurs in hyphae, is induced in the presence of other repressing carbon sources besides glucose, and is dependent on the function of the transcriptional repressor CreA. The expression of MstE and its subcellular distribution have been studied using a MstE-sGFP fusion protein. Concordant with data on mstE expression, MstE-sGFP is synthesized in the presence of repressing carbon sources, and fluorescence at the periphery of conidia and hyphae is consistent with MstE location in the plasma membrane. Deletion of mstE has no morphological phenotype but results in the absence of low affinity glucose uptake kinetics, the latter being substituted by a high affinity system.

Amino Acid Sequence↗
PubMed2002

Co-regulation of pathogenesis with dimorphism and phenotypic switching in Candida albicans, a commensal and a pathogen.

Candida albicans, a common fungal pathogen of humans, can colonize in many diverse environments of the host and convert between a harmless commensal and a pathogen. Recent advances indicate that C. albicans uses a common set of conserved pathways to regulate dimorphism, mating and phenotypic switching. Major pathways known to regulate dimorphism include a mitogen-activated protein (MAP) kinase pathway through Cph1, the cAMP-dependent protein kinase pathway via Efg1, and Tup1-mediated repression through Rfg1 and Nrg1. The Cph1-mediated MAP kinase pathway is critical for the mating process, while all three pathways are implicated in the regulation of white-opaque switching. All these developmental pathways regulate the expression of hypha-specific and/or phase-specific genes. A high proportion of hypha-specific genes and phase-specific genes encode proteins that contribute directly or indirectly to pathogenesis and virulence of C. albicans. Therefore, virulence genes are co-regulated with cell morphogenesis. This supports a previous notion that the unique aspects of C. albicans commensalism and pathogenesis may lie in the developmental programs of dimorphism and phenotypic switching.

Candida albicans↗
PubMed2003

Signaling through protein kinases and transcriptional regulators in Candida albicans.

The human fungal pathogen Candida albicans switches from a budding yeast form to a polarized hyphal form in response to various external signals. This morphogenetic switching has been implicated in the development of pathogenicity. Several signaling pathways that regulate morphogenesis have been identified, including various transcription factors that either activate or repress hypha-specific genes. Two well-characterized pathways include the MAP kinase cascade and cAMP-dependent protein kinase pathway that regulate the transcription factors Cph1p and Efg1p, respectively. cAMP also appears to interplay with other second messengers: Ca2+, inositol tri-phosphates in regulating yeast-hyphal transition. Other, less-characterized pathways include two component histidine kinases, cyclin-dependent kinase pathway, and condition specific pathways such as pH and embedded growth conditions. Nrg1 and Rfg1 function as transcriptional repressors of hyphal genes via recruitment of Tup1 co-repressor complex. Different upstream signals converge into a common downstream output during hyphal switch. The levels of expression of several genes have been shown to be associated with hyphal morphogenesis rather than with a specific hypha-inducing condition. Hyphal development is also linked to the expression of a range of other virulence factors. This review explains the relative contribution of multiple pathways that could be used by Candida albican cells to sense subtle differences in the growth conditions of its native host environment.

Candida albicans↗
PubMed1961

The fine structure of Streptomyces violaceoruber (S. coelicolor). III. The walls of the mycelium and spores.

A study of thin sections of hyphae of Streptomyces violaceoruber in the electron microscope showed that the structure of the walls and the mode of formation of cross-walls are similar to those of Gram-positive bacteria. A beaded structure was seen in some regions of the wall, and the significance of this observation is discussed in relation to previous studies of the fine structure of bacterial cell walls. Elements of the intracytoplasmic membrane system appear to be involved in the process of cross-wall formation. The walls of the hyphae of the aerial mycelium divide into two layers before the spores are formed, and only the inner component of the wall grows inwards to form the cross-walls and so delimit the spores. The outer component remains intact for a time and acts as a sheath around the developing spores. Finally the sheath breaks and the spores are liberated. This process is contrasted with the formation of endospores in eubacteria. When the spores germinate, the walls of the germ tubes are continuous with those of the spores.

Hyphae↗
PubMed2004

In Candida albicans, the Nim1 kinases Gin4 and Hsl1 negatively regulate pseudohypha formation and Gin4 also controls septin organization.

In the development of hyphal germ tubes of Candida albicans, a band of septin forms at the base of the germ tube (basal septin band). Later, a septin ring forms, which organizes the first septum within the germ tube (septin ring). We have investigated the role of the Nim1 kinases, Gin4 and Hsl1, in the formation of these septin structures. We show that during germ tube formation, Gin4 is required for the organization of the septin ring but not the basal septin band. Hsl1 is not required for the formation of either septin rings or basal bands. Unexpectedly, we found that both gin4Delta and hsl1Delta mutants form pseudohyphae constitutively, in a fashion that in the case of gin4Delta, is partly independent of Swe1. Gin4-depleted pseudohyphae are unable to form hyphae when challenged with serum, but this can be overcome by ectopic expression of Gin4 from the MET3 promoter. Thus, Gin4 may regulate the developmental switch from pseudohyphae to hyphae.

Candida albicans↗
PubMed1960

Observations on the chromatinic bodies of Streptomyces coelicolor.

Colonies of Streptomyces coelicolor growing on cellophane and impression preparations from sporing colonies were stained for chromatin by the methods of Feulgen, DeLamater (1951), and Piéchaud (1954). The chromatinic bodies of the substrate hyphae have a great variety of configurations. During the development of the spores, elongated chromatinic structures in the young aerial hyphae separate into a number of subunits and a single round chromatinic body is included in each spore.

Chromatin↗