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PubMed2004

Role of Ca2+/calmodulin signaling pathway on morphological development of Candida albicans.

A human fungal pathogen, Candida albicans, varies from the yeast form to the hyphal form due to various external signals. This morphogenetic transformation has been implicated in the development of pathogenicity. In this report, we show that calmodulin inhibitors (TFP and W-7) and an adenylatecyclase inhibitor (MDL-12-330A) suppressed the hyphae formation of C. albicans. Furthermore, the expression of hyphae-specific mRNAs located downstream from the RAS1-cAMP pathway was inhibited by these inhibitors. Suppression of hyphae formation by TFP or W-7 was not inhibited by the addition of cAMP, and these inhibitors did not affect the amount of cAMP in C. albicans. These results suggest that the Ca2+/calmodulin pathway contributes to hyphae formation and is related to the RAS1-cAMP pathway.

Adenylyl Cyclase Inhibitors↗
PubMed1995

Use of DAPI for anastomosis group typing of strains of the fungus Rhizoctonia solani.

Strains of Rhizoctonia solani, a common soil-borne, pathogenic fungus of plants, are assigned to one of 11 anastomosis groups (AGs) based on the occurrence of imperfect fusions (anastomoses) between hyphae of a non-typed strain and a tester strain of one of the 11 AG's. Imperfect fusion is characterized by the death of one or more cells in each of the hyphae involved in the fusion. Although hyphae from branches of the same strain of R. solani may fuse with each other (self-fusion), cell death does not occur. Cell death is accompanied by nuclear degradation and granulation, or plasmolysis of the cytoplasm, which often is not visible using brightfield microscopy. When the DNA-binding fluorochrome DAPI (4',6-diamidino-2-phenylindole) is used and the hyphal fusions viewed under fluorescence microscopy, no nuclei are observed in fused hyphal cells from two strains of the same AG of R. solani. Because DAPI reacts only with living nuclei, lack of staining is presumptive evidence that the fused cells are dead as a result of imperfect fusion. The use of DAPI reduces the time required for making AG determinations compared to standard methods because it eliminates the need to assess cell wall dissolution and cytoplasmic fusion. Also, it is not necessary to trace the hyphae involved in the fusion to their respective origins to ensure that self-fusion has not occurred.

Cell Death↗
PubMed2000

The response of arbuscular mycorrhizae to fertilization, mowing, and removal of dominant species in a diverse oligotrophic wet meadow.

In a wet oligotrophic meadow located in the Czech Republic, a factorial experiment with treatments consisting of fertilization, mowing, and removal of the dominant species (Molinia caerulea) was established in 1994. In 1997 Holcus lanatus, Molinia caerulea, Potentilla erecta, Prunella vulgaris, and Ranunculus auricomus were examined for arbuscular mycorrhizal (AM) hyphae, arbuscles, and vesicles three times over the season. Time had a significant effect on AM in all five species. Except for Molinia arbuscles, a modal effect occurred, with the second sampling having a greater level of AM structures than the first and the third. Fertilization had the greatest effect on AM levels by decreasing the level of Holcus hyphae and vesicles, Potentilla vesicles, Prunella hyphae, and Ranunculus hyphae and vesicles. Mowing significantly increased the number of Potentilla vesicles, and the removal of dominant species had no significant effects. Interactions between time and treatments were common. Significant effects to the arbuscle:vesicle ratio were infrequent, and those that occurred were related to changes over the season. Seasonal effects appear to have a more powerful effect on AM structures and the arbuscle:vesicle ratio than do treatment effects. In a second experiment, Ranunculus auricomus, R. acris, and R. nemorosus, sampled four times over the season, showed significant changes in AM colonization. Overall, AM structures either declined over the season or increased from April to May and then declined. There was no AM colonization response to a spring fertilization in the three species. It is postulated that the patterns observed are due to phosphorus availability and seasonal changes in soil moisture and rates of root growth and turnover.

Journal Article↗
PubMed2001

A better potassium hydroxide preparation? In vivo diagnosis of tinea with confocal microscopy.

BACKGROUND: Traditional diagnostic testing for dermatophyte infection currently requires skin scraping for light microscopy and/or fungal culture or skin biopsy. Immunofluorescent microscopy can also be used with calcofluor stain. All of these tests can be time-consuming to perform, require a waiting period for results, and are invasive. We investigated the use of a real-time, noninvasive, confocal microscope in visualizing dermatophyte hyphae in vivo. OBSERVATIONS: Confocal microscopic imaging of active tinea can clearly identify dermatophyte hyphae within the upper epidermis after potassium hydroxide application. The hyphae appear as bright linear branching objects not found in uninvolved skin. CONCLUSIONS: It is possible to immediately and painlessly image dermatophyte hyphae in active lesions of tinea by means of a confocal microscope. With further improvement, imaging devices may be available to physicians to instantly and noninvasively evaluate a variety of skin disorders in microscopic detail.

Adult↗
PubMed2000

Cloning and Identification of Genes Related with Morphogenesis of Candida albicans.

A Candida albicans cDNA library was constructed and screened by differential hybridization. In hybridization using probes derived from population of yeast cells or hyphae, 67 recombinant phages exhibited more intense signal with the probe derived from hyphae than with the probe from yeast cells. One phage behaved vice versa. Physical map analysis and nucleotide sequence analysis suggested that cDNA of the CX1, a clone specific for yeast form, coded for cytochrome P450 L1A1 (Lanosterol 14alpha-demethylase). Its specific expression pattern was confirmed by Northern analysis. Inhibition of serum on the expression of CX1 cDNA was observed. CX2 cDNA was one of those giving intensive signal with hyphae probes. The cDNA sequence contained a tandem repeat sequence, which was also found in ALS1, another Candida albicans gene identified, whose expression was related with morphogenesis. Northern analysis proved that it was expressed intensively with hyphae probes, however the expression could not be detected in those strongly hybridized to yeast cell probes. The locations of both cDNA on chromosome were analyzed.

Journal Article↗
PubMed2006

[Study on endophyte and the growth stages of Ophiopogon japonicus].

OBJECTIVE: To study on the relationship between the endophyte and the life cycle of Ophiopogon japonicus in Zhejiang. METHODS: Sample roots of Ophiopogon japonicus at different growth stages were thoroughly washed and cut into small fragments, then cleared (removing cytoplasmic contents from cells) using hot 10% KOH and stained with acid fuchsin (alternative stain). The hyphae, the arbuscular and the vesicular of endophyte were examined. RESULTS: The hyphae appeared and grew in the seedling stage, the hyphae grew into arbuscular in the root tuber generating stage and vesicular in the stage of root tuber expanding period. CONCLUSION: The endophytes in Ophiopogon japonicus appeared in forms of hyphae, arbuscular and vesicular at different growth stages to meet the needs of Ophiopogon japonicus developing.

Fungi↗
PubMed1991

[Studies on the effects of fungi on the course of seed germination of Dendrobium lohohens and Dendrobium candidum].

This paper reports the results of studies on the effects of fungi isolated from the protocorms of Orchidaceae on the course of seed germination of D. lohohens and D. candidum. The experiments revealed that these fungi could promote seed germination: the seed germination rate of D. lohohens was 20%, that of D. candidum was 64%, and that of control was zero. The changes of cell ultrastructure in the course of fungus-infected seed germination of D. candidum was also studied. After hyphae of M. osmundicola penetrate the embryo from the suspensor, organelles of embryonic cells infected with fungi will disappear. When the hyphae wall is thickened and enclosed by the embryo's, the hyphae will be digested and used as the source of nutrition for seed germination. Also, the metabolism of cells without infection, but adjacent to those cells penetrated by hyphae is enhanced.

Fungi↗
PubMed1991

Ultrastructure of the pseudo-cleistothecia produced by the Trichophyton mentagrophytes complex.

The internal structure and the formation processes of pseudo-cleistothecia in Arthroderma vanbreuseghemii, a member of the Trichophyton mentagrophytes complex, were observed by light and electron microscopy. The structure of pseudo-cleistothecia, obtained by single spore culture of A. vanbreuseghemii, developed as a clump of hyphae producing microconidia which increased gradually in size. Echinulations appeared on the surrounding hyphae, which developed into peridial hyphae with characteristic constrictions. These pseudo-cleistothecia contained the microconidia. During the mating of A. vanbreuseghemii and Arthroderma simii, pseudo-cleistothecia structures developed from organs similar to the gametangia. These pseudo-cleistothecia contained interwoven hyphae but no microconidia.

Microscopy, Electron↗
PubMed1980

A histological and ultrastructural comparison of the sulfur granule of the actinomycosis and actinobacillosis.

The sulfur granules of actinomycosis and actinobacillosis were studied histologically and ultrastructurally. The sulfur granule of actinomycosis was large, and oval or horsehoe-shaped. In it, there were a number of gram-positive, filamentous or short rod-like hyphae beneath clubs. Ultrastructurally, the center of the sulfur granule was composed of a cluster of hyphae, and the periphery was surrounded by clubs arranged radially. The clubs were made of electron-dense fine granules and had a degenerated hypha at the center. These granules varied in amount from small to large. As for the host reaction, neutrophil infiltration appeared first around the sulfur granule. Then epithelioid cells proliferated and phagocytized hyphae. Finally, proliferating fibroblasts enclosed them and formed tiny granulomas. The sulfur granule of actinobacillosis was small and lobulated. In it, a number of gram-negative, short rod-like bacteria were present beneath clubs. Ultrastructurally, the center of the sulfur granule contained intact or degenerated bacteria, and the periphery was composed of radially projecting clubs. These clubs were made of electron-dense amorphous material, in which several layers of tubular structures surrounded the bacterium conentrically. The host reaction was almost the same as that of actinomycosis. The proliferation of fibroblasts was prominent. There was a strong tendency for these cells to form many tiny granulomas.

Actinobacillosis↗
PubMed1993

Ion channel activity and tip growth: tip-localized stretch-activated channels generate an essential Ca2+ gradient in the oomycete Saprolegnia ferax.

The plasma membrane of tip-growing hyphae of the oomycete Saprolegnia ferax contains stretch-activated (SA) Ca(2+)-permeable and Ca(2+)-activated K+ ion channels. Patch clamp measurements on protoplasts derived from specific regions of hyphae demonstrated that SA channels were most abundant in the tip. Gadolinium (Gd3+) inhibited SA channel activity and stopped tip growth. The Ca(2+)-sensitive fluorochrome INDO 1 revealed a tip-high gradient of free cytoplasmic Ca2+ in growing hyphae. This gradient could be dissipated with the addition of Gd3+. The calcium gradient returned and growth resumed when Gd3+ was washed out. This implies a fundamental requirement for growth for Ca2+ influx through the SA channels. Ca(2+)-activated K+ channels were distributed evenly along the hyphae. These channels were inhibited by tetraethylammonium concentrations which caused a rapid but transient decrease in growth. We suggest that the SA channels at the apex act as feedback sensors, responding to membrane stretching at the tip. They are an obligate requirement for tip growth. The Ca(2+)-activated K+ channels may act to maintain turgor pressure, but are not obligatory for growth.

Calcium↗
PubMedDate not supplied

Proventriculitis and ventriculitis associated with zygomycosis in ostrich chicks.

This report describes eight cases of proventriculitis and ventriculitis in ostrich chicks less than 2 months old. Clinical signs included acute onset of lethargy and anorexia in three cases, and chronic weight loss with lethargy and anorexia in four cases; no history was available in one case. There was limited antibiotic therapy in two cases; a third case was treated for giardiasis. Concurrent bacterial, yeast, and viral infections were common. Lymphoid depletion and/or necrosis of bursa, thymus, and spleen suggested severe immune challenge or immunosuppression in many cases. Histologically, there was severe ulcerative proventriculitis and ventriculitis with intralesional fungal hyphae. In two chicks with granulomatous pneumonia, similar fungal hyphae were also observed in the lung. Fungal hyphae were rarely septate, with irregular, non-parallel walls, and ranged in diameter from 7 to 20 microns. Occasional globoid distentions of the hyphae were present. Fungi were identified morphologically as species in the Zygomycetes class; in one case a Mucor sp. was cultured. Zygomycetes appear to be potentially serious opportunistic pathogens of ostrich chicks.

Animals↗
PubMed1996

Identification of the reactive subunits of Aspergillus umbrosus involved in the antigenic response in farmer's lung.

BACKGROUND: Farmer's lung (FL) is the most common form of extrinsic allergic alveolitis and the fungus Aspergillus umbrosus has frequently been associated with FL among Finnish farmers. IgG and IgA class antibodies against Aspergillus umbrosus are found in the serum of FL patients and healthy exposed farmers. So far the immunologically reactive subunits of Aspergillus umbrosus are unknown. OBJECTIVES: The purpose of the study was to identify and localize the reactive subunits of Aspergillus umbrosus that are involved in the antigenic response in FL. METHODS: The distribution of the primary antibody on the outer wall layers of 4- and 11 day-old fungi Aspergillus umbrosus was studied by pre-embedding immunolabelling with fluorescein isothiocyanate (FITC)- and gold-conjugated antibodies. The surface carbohydrates of the outer wall layers were studied using FITC-conjugated lectins. RESULTS: FITC-conjugated antibodies from FL patient sera reacted with spores (conidia), phialides and hyphae, whereas the fruiting body, cleistothecia, remained unstainable. Immunofluorescence labelling was confirmed by immunolabelling electron microscopy, and gold-conjugated IgG antibodies were found on the outer cell walls of spinulose conidia, and in a less degree on the outer cell walls of hyphae and phialides. Only single gold particles were found scattered on the surfaces of cleistothecia. The surface carbohydrates of Aspergillus umbrosus conidia, vesicles, phialides and hyphae had a high affinity to GlcNAc beta 1- and Man alpha 1-/Glc alpha 1-specific lectins and only GlcNAc beta 1-specific lectins also reacted with the outer cell walls of cleistothecia. Blocking of the antigenic sites of Aspergillus umbrosus by preincubating antigen-coated enzyme linked immunosorbent assay (ELISA) plates with lectins caused some inhibition, indicating that sites with the highest affinity for lectins were not always antigenic sites. CONCLUSION: Gold-and FITC -conjugated antibodies were distributed mostly on the outer wall layers of conidia, hyphae and phialides of Aspergillus umbrosus and GlcNAc beta 1-linked lectins had the strongest affinity to the outer wall layers.

Antibody Formation↗
PubMed1996

[Studies on the septal ultrastructure of Armilaria mellea].

The structure and development of septum in Armillaria mellea were studied under the electronmicroscope. Hyphal septum was developed by inside wall of hyphae in cross direction, leaving a small pore in the center. Then the septal plate arround the pore became swollen, and formed a septal pore apparatus called dolipore septum. With subsequent development of endoplasmic reticulum septal pore caps appeared on each side of the pore. Organelles, such as mitochondrium and Golgi body in hyphal cells were all related with the development of septa. Thin-walled narrow hyphae of the medualla in mature rhizomorph were in a state of dormancy, and their septa were similar to those of the hyphae growing from liquid fermentation. Both of them were short of septal pore caps, but the latter had larger septal pores, the protoplasm flow from cell to cell through the pore was visible, and fusion rate between the hyphae was much higher. The relationship between the structural changes and functions of septa in A. mellea was also systematically discussed in this paper.

Agaricales↗
PubMed1997

Germ tube growth of Candida albicans.

The clinical pathogen Candida albicans is a budding yeast that is capable of forming a range of polarized and expanded cell shapes from pseudohyphae to true nonconstricted hyphae. Filamentous forms consist of contiguous uninucleated compartments that are partitioned by septa. It has long been held that the so-called "dimorphic transition" from a budding to a filamentous form may aid the fungus to penetrate epithelia and may therefore be a virulence factor. This review summarized new information regarding the physiology and ecology of hyphal growth in C. albicans. New evidence has demonstrated that hyphae of C. albicans have a sense of touch so that they grow along grooves and through pores (thigmotropism). This may aid infiltration of epithelial surfaces during tissue invasion. Hyphae are also aerotropic and can form helices when contacting solid surfaces. Growing evidence supports the view that hyphal growth is a response to nutrient deprivation, especially low nitrogen and that filamentous growth enables the fungus to forage for nutrients more effectively. Further insights into the growth of C. albicans have come from the analysis of genes and mutations of Saccharomyces which have begun to reveal the molecular mechanisms underlying the mechanisms of bud site selection, cell polarity and signal transduction pathways that lead to pseudohyphal development in this and other organisms. For example, it is now clear that a MAP-kinase cascade, homologous to the mating pathway in Saccharomyces, regulates filamentous growth in both fungi. However, this must be only one of several overlapping or separate signal transduction pathways for hyphal development because filamentous growth still occurs in mutants of Candida and Saccharomyces which are blocked in this pathway. Cell cycle analyses have shown that hyphal phase cell cycle of Candida is distinct from that in budding and pseudohyphal formation and so pseudohyphal growth of Saccharomyces is not a true model of germ tube growth in Candida. Pseudohyphal growth in both Candida and Saccharomyces involves synchronous division of mother cells and their daughters. In contrast, during germ tube growth of Candida, cytoplasm is unequally partitioned at cytokinesis so that apical cells inherit more cytoplasm and sub-apical cells have a single nucleus but are extensively vacuolated. As a result, apical cells grow and divide while sub-apical cells are apparently arrested in the cell cycle until they can regenerate sufficient cytoplasm to re-enter the cell cycle. Although current studies still fall short of verifying the status of yeast-hypha dimorphism as a virulence factor, they suggest that the cell biology of germ tube growth of C. albicans is well suited for the invasive growth of the fungus in vivo.

Candida albicans↗
PubMed1998

The role of bronchial biopsy and washing in the diagnosis of allergic bronchopulmonary aspergillosis.

The diagnosis of allergic bronchopulmonary aspergillosis (ABPA) is usually established on the basis of clinical, serologic, and radiologic criteria, of which eight are commonly cited. Although histologic abnormalities are not usually considered, a study of lobectomy/lung biopsy specimens provided evidence that the combination of mucoid impaction of bronchi with "allergic" mucin and fungal hyphae is diagnostic of ABPA. We attempted to explore the use of surgical specimens in the diagnosis of ABPA by examining endobronchial biopsy specimens and washings. Bronchial biopsy specimens obtained from 1990 to 1995 that contained fungal hyphae were reviewed and assessed for the presence of "allergic" mucin and bronchial wall abnormalities. We also reviewed cytologic specimens and clinical, serologic, and radiologic findings of the corresponding patients. Five biopsy specimens had fungal hyphae and associated "allergic" mucin. In some patients, the mucin was small in amount and contained degenerated eosinophils that resembled macrophages. None of the patients was thought to have ABPA clinically. All of the five had bronchial mucoid impaction at bronchoscopy. After additional investigation, only one patient fulfilled all of the eight traditional criteria for ABPA; one other patient fulfilled six criteria. Although some tests were not completed in the remaining three patients, follow-up and/or response to steroids was consistent with ABPA in all of the three. Four patients had bronchial washing performed at the time of biopsy. Cytologic examination showed one to be normal except for inflammatory cells that did not include eosinophils; three others showed mucus containing eosinophils intermixed with Charcot-Leyden crystals and scattered hyphae. We conclude that patients with ABPA might have atypical manifestations of disease and not meet standard diagnostic criteria. Bronchoscopy with bronchial biopsy and washing plays a role in detecting these patients. The presence of "allergic" mucin might be overlooked in the small biopsy fragments obtained by bronchoscopy.

Adult↗
PubMed2006

Proteomic analysis of Candida albicans yeast and hyphal cell wall and associated proteins.

Candida albicans is an important human pathogen that causes systemic infections, predominantly among populations with weakened immune systems. The morphological transition from the yeast to the hyphal state is one of the key factors in C. albicans pathogenesis. Owing to their location at the host-pathogen interface, the cell wall and associated proteins are of interest, especially with respect to the yeast to hyphal transition. This study entailed the proteomic analysis of differentially regulated proteins involved in this transition. The protein profiles of C. albicans DTT/SDS-extractible proteins and the cyanogen bromide (CNBr)/trypsin-extractable proteins of a cell wall-enriched fraction from yeast and hyphae were compared. In total, 107 spots were identified from the DTT/SDS-extractible cell wall-enriched fraction, corresponding to 82 unique proteins. Of these DTT/SDS-extractible proteins, 14 proteins were upregulated and 10 were downregulated in response to hyphal induction. Approximately 6-9% of total cell wall-protein-enriched fraction was found to be resistant to DTT/SDS extraction. Analysis of the DTT/SDS-resistant fraction using a CNBr/trypsin extraction resulted in the identification of 29 proteins. Of these, 17 were identified only in the hyphae, four were identified only in the yeast, and eight were identified in both the yeast and hyphae.

Amino Acid Sequence↗
PubMed2006

The alpha-tubulin gene AmTuba1: a marker for rapid mycelial growth in the ectomycorrhizal basidiomycete Amanita muscaria.

The apical extension of hyphae is of central importance for extensive spread of fungal mycelium in forest soils and for effective ectomycorrhiza development. Since the tubulin cytoskeleton is known to be important for fungal tip growth, we have investigated the expression of an alpha-tubulin gene from the ectomycorrhizal basidiomycete Amanita muscaria (AmTuba1). The phylogenetic analysis of protein sequences revealed the existence of two subgroups of alpha-tubulins in homobasidiomycetes, clearly distinguishable by defined amino acids. AmTuba1 belongs to subgroup1. The AmTuba1 transcript level is related to mycelial growth rate. Growth induction of carbohydrate starved (non-growing) hyphae resulted in an enhanced AmTuba1 expression as soon as hyphal growth started, reaching a maximum at highest mycelial growth rate. Bacterium-induced hyphal elongation also leads to increased AmTuba1 transcript levels. In mature A. muscaria/P. abies ectomycorrhizas, where fungal hyphae are highly branched, and slowly growing, AmTuba1 expression were even lower than in carbohydrate-starved mycelium, indicating a further down-regulation of gene expression in symbiosis. In conclusion, our analyses show that the AmTuba1 gene can be used as a marker for active apical extension in fly agaric, and that alpha-tubulin proteins are promising tools for the classification of fungi.

Amanita↗
PubMed2003

Morphological characterization of the mycorrhiza formed by Helianthemum almeriense Pau with Terfezia claveryi Chatin and Picoa lefebvrei (Pat.) Maire.

This work presents the first anatomical description of the mycorrhizal systems of Helianthemum almeriense, and of the structure and ultrastructure of the mycorrhizae formed by this plant species with the ascomycetes Terfezia claveryi and Picoa lefebvrei. Four different mycorrhizal systems are described, the club-shaped mycorrhiza being the most abundant. The type of mycorrhiza formed depended on the mycorrhiza culture conditions, but not on the fungal species. For both fungal species, H. almeriense formed an endomycorrhiza in natural field conditions, an ecto- and ectendomycorrhiza without a sheath in pot cultures, and an ectomycorrhiza with a characteristic sheath and Hartig net in in vitro cultures. This is the first report of a typical sheath in Helianthemum-desert truffle mycorrhizal associations. The results support the idea that culture conditions can induce changes in mycorrhiza morphology and that there is no clear barrier between the two main types of mycorrhiza organization in Helianthemum species. The ultrastructural study confirmed the regular presence of T. claveryi intracellular hyphae in direct contact with the host wall, a localization which seems to be a characteristic of the T. claveryi mycorrhiza organization. The P. lefebvrei mycorrhiza organization was characterized by intracellular hyphae with large amounts of electron-dense globules, probably with a lipidic content, and a warty ornamentation on the wall of the root external hyphae.

Ascomycota↗