Polyamines in dimorphic fungi

The polyamine contents of the yeast and mycelial forms of seven dimorphic fungi were examined to determine if a relationship existed between polyamine composition and fungal morphology. Although the polyamine contents differed, no consistent patterns emerged. Evidently, the observed differences in polyamine composition were not related to fungal dimorphism but rather to the growth conditions used to achieve those morphological forms.     

Protective antibodies and endemic dimorphic fungi

The host response to infection is the outcome of a complex interaction between a microbe and a host's innate and adaptive immune system. In this context, the role of antibody in the endemic mycoses is relatively poorly understood. Recently, a monoclonal antibody to a cell surface protein has been shown to be protective in a murine histoplasmosis model. The findings with Histoplasma capsulatum may provide a paradigm for antibody protection against endemic fungi. This paper reviews the recent data on protective antibody in histoplasmosis and previous data supporting a role for antibody in protective responses in other dimorphic fungi.     

Differences in DNA methylation patterns are detectable during the dimorphic transition of fungi by amplification of restriction polymorphisms

A modification of the amplified fragment length polymorphism technique was developed for the determination of DNA methylation in dimorphic fungi representative of three of the major fungal taxa: Mucor rouxii, a zygomycete; Yarrowia lipolytica, an ascomycete; and Ustilago maydis, a basidiomycete. DNA obtained from the yeast or mycelial stages of the fungi was digested with a mixture of EcoRI, and one of the isoschizomers MspI and HpaII, whose ability to cleave at the sequence CpCpGpG is affected by the methylation state. The resulting fragments were ligated to primers and subjected to a double round of amplification by the polymerase chain reaction, radiolabeled in the second round, and separated by polyacrylamide gel electrophoresis. Comparison of patterns revealed differences indicative of fragments whose methylation state did or did not change during the dimorphic transition. These results indicate the usefulness of the method for the study of DNA methylation, demonstrate the universality of DNA methylation in fungi, and confirm that differential DNA methylation occurs during fungal morphogenesis. Received: 3 May 1996 / Accepted: 19 September 1996     

二型态真菌形态调控研究现状与展望

真菌二型态(Dimorphism)是指某些真菌在外界环境因子的诱导下,其营养体可在酵母型(Yeast form)和菌丝型(Mycelium form)两种不同细胞形态间转化的能力。诱导二型态真菌形态转化的环境因子众多,不同环境因子诱导二型态转化的分子机制也不尽相同。二型态真菌形态转换常与致病性有关,在微生物发酵调控中也至关重要,研究真菌形态调控不仅有助于阐明二型态真菌的致病机理,还可丰富二型态真菌形态调控在发酵工业中的应用。本文从影响真菌二型态的外界环境因子、转化机制和真菌发酵形态控制等方面对二型态真菌形态调控研究现状及应用前景进行综述。     

Studies on the physiological bases of morphogenesis in fungi; the respiratory metabolism of dimorphic pathogenic fungi

The interconversions of mycelial and yeast-like forms (M right harpoon over left harpoon Y) in Blastomyces dermatitidis and in Blastomyces brasiliensis are characterized as examples of thermal dimorphism since the phenomena are apparently dependent only on the temperature of incubation of these two species. The change in morphology consequent upon Y --> M conversion is considered to result from the selective inhibition of cell division, without the simultaneous inhibition of growth. Such selective inhibition is viewed in a wider context as an example of the differential operation of the physicochemical apparatus of the cell in the control of form development by an organism. To analyze this differential operation, which is here dependent only on temperature, we have studied the effect of temperature on oxygen consumption by each of the dimorphic forms. In the absence of external substrate the yeast forms consume 5 to 6 times more oxygen per unit dry weight than do the M forms.The Y forms exhibit an exogenous oxidation of acetate and of glucose, as well as an oxidative assimilation of these substrates, whereas the M forms exhibit no exogenous metabolism in either a resting or starved condition. A study of the effect of a wide range of temperatures on oxygen consumption by the M forms indicates the operation of two rate-limiting processes: (a) one with an activation energy of 13,250 calories/gm. molecule over the range 5-30 degrees , and (b) reversible enzyme inactivation; the latter process assuming importance in the higher temperature range. On abrupt, large changes in temperature the balance between these two rate-limiting reactions (which it is suggested characterizes the steady state) is apparently disrupted as a result of a lag in the assumption of a rate of reversible enzyme inactivation characteristic of the new temperature. This disruption of balance is evidenced in overshoot phenomena. The effect of an analogous disruption of balance, and of increasing enzymic inactivation; on a competition between enzyme systems, competing for substrate for cell elongation and for cell division, is considered in explanation of the observed dependence of the cell division mechanism on the maintenance of an elevated temperature.     

cAMP signalling in pathogenic fungi: control of dimorphic switching and pathogenicity

Morphological changes in pathogenic fungi often underlie the development of virulence and infection by these organisms. Our knowledge of the components of the cell signalling pathways controlling morphological switching has, to a large extent, come from studies of pseudohyphal growth of the model organism Saccharomyces cerevisiae, in which control is exerted via changes in the intracellular cAMP and mitogen-activated protein kinase cascades. There is evidence that pathogenic fungi also utilize these pathways to control dimorphic switching between saprobic and pathogenic forms and, as such, the elements of these pathways have potential as drug targets.     

Genomic transition to pathogenicity in chytrid fungi

Understanding the molecular mechanisms of pathogen emergence is central to mitigating the impacts of novel infectious disease agents. The chytrid fungus Batrachochytrium dendrobatidis (Bd) is an emerging pathogen of amphibians that has been implicated in amphibian declines worldwide. Bd is the only member of its clade known to attack vertebrates. However, little is known about the molecular determinants of - or evolutionary transition to - pathogenicity in Bd. Here we sequence the genome of Bd's closest known relative - a non-pathogenic chytrid Homolaphlyctis polyrhiza (Hp). We first describe the genome of Hp, which is comparable to other chytrid genomes in size and number of predicted proteins. We then compare the genomes of Hp, Bd, and 19 additional fungal genomes to identify unique or recent evolutionary elements in the Bd genome. We identified 1,974 Bd-specific genes, a gene set that is enriched for protease, lipase, and microbial effector Gene Ontology terms. We describe significant lineage-specific expansions in three Bd protease families (metallo-, serine-type, and aspartyl proteases). We show that these protease gene family expansions occurred after the divergence of Bd and Hp from their common ancestor and thus are localized to the Bd branch. Finally, we demonstrate that the timing of the protease gene family expansions predates the emergence of Bd as a globally important amphibian pathogen.     

Ultrastructure of chitin in hyphae ofCandida albicans and other dimorphic and mycelial fungi

Summary Chitin microfibrils exposed by chemical extraction of hyphal walls ofCandida albicans, Histoplasma capsulatum, Blastomyces dermatitidis, Paracoccidiodes brasiliensis, Coprinus cinereus andMucor mucedo were of variable morphology but gave identical infrared spectra and behaved as pure chitin in chromatographic analyses. The microfibrils of the four dimorphic fungi studied were shorter than those in the mouldsC. cinereus andM. mucedo but were similar to those reported for the yeastSaccharomyces cerevisiae. InC. albicans the microfibrils in the septal plates of hyphae were predominantly tangentially orientated and were longer than those in the lateral walls. Microfibrils produced by chitin synthasein vitro were very much longer than any observed from hyphal preparations.     

Changes in cyclic nucleotide levels and dimorphic transition in Candida albicans.

The relationship between the levels of cyclic nucleotides and dimorphic transition in Candida albicans was examined. The results showed that cells of this pathogenic fungus contained both cyclic adenosine 3',5'-monophosphate (cAMP) and cyclic guanosine 3',5'-monophosphate (cGMP), the concentration of the latter being about one-tenth that of the former in stationary-phase cells of the yeast form. Our results further indicated that germ tube formation induced by incubation at 40 degrees C followed a rise in cAMP concentration in the cell with no accompanying change in cGMP content. Cysteine, which suppressed germination, also reversed the increase in intracellular cAMP concentration. Dibutyryl cAMP (1 MM) significantly promoted germination in proline medium at temperatures of 32 to 34 degrees C. These results suggested that cAMP was one of the controlling factors in the morphological transition in Candida albicans.     

Phase transition in tensionless surfaces

We study the critical behavior of the Laplacian roughening model, which describes the growth of tensionless surfaces. This type of growth phenomena is very important, for instance, in biological membranes and in molecular beam epitaxy. We summarize previous analytical and numerical results and point out their contradictions and differences, thus making clear the context of our work. Our contribution, achieved through large-scale numerical simulations, is the confirmation that the model exhibits a single continuous phase transition: the transition takes place between a continuum massless (i.e., with infinite correlation length) bilaplacian behavior and a massive propagator (finite correlation length).     

Inoculum size effect in dimorphic fungi: extracellular control of yeast-mycelium dimorphism in Ceratocystis ulmi

We studied the inoculum size effect in Ceratocystis ulmi, the dimorphic fungus that causes Dutch elm disease. In a defined glucose-proline-salts medium, cells develop as budding yeasts when inoculated at > or = 10(6) spores per ml and as mycelia when inoculated at <10(6) spores per ml. The inoculum size effect was not influenced by inoculum spore type, age of the spores, temperature, pH, oxygen availability, trace metals, sulfur source, phosphorous source, or the concentration of glucose or proline. Similarly, it was not influenced by added adenosine, reducing agents, methyl donors, amino sugars, fatty acids, or carbon dioxide. Instead, growing cells excreted an unknown quorum-sensing factor that caused a morphological shift from mycelia to budding yeasts. This yeast-promoting effect is abolished if it is extracted with an organic solvent such as ethyl acetate. The quorum-sensing activity acquired by the organic solvent could be added back to fresh medium in a dose-dependent fashion. The quorum-sensing activity in C. ulmi spent medium was specific for C. ulmi and had no effect on the dimorphic fungus Candida albicans or the photomorphogenic fungus Penicillium isariaeforme. In addition, farnesol, the quorum-sensing molecule produced by C. albicans, did not inhibit mycelial development of C. ulmi when present at concentrations of up to 100 microM. We conclude that the inoculum size effect is a manifestation of a quorum-sensing system that is mediated by an excreted extracellular molecule, and we suggest that quorum sensing is a general phenomenon in dimorphic fungi.     

A putative phospholipase C is involved in Pichia fermentans dimorphic transition

Background

Pichia fermentans DiSAABA 726 is a dimorphic yeast that reversibly shifts from yeast-like to pseudohyphal morphology. This yeast behaves as a promising antagonist of Monilia spp. in the yeast-like form, but becomes a destructive plant pathogen in the pseudohyphal form thus raising the problem of the biological risk associated with the use of dimorphic yeasts as microbial antagonists in the biocontrol of phytopathogenic fungi.

Methods

Pichia fermentans DiSAABA 726 was grown in urea- and methionine-containing media in order to induce and separate yeast-like and pseudohyphal morphologies. Total RNA was extracted from yeast-like cells and pseudohyphae and retro-transcribed into cDNA. A rapid subtraction hybridization approach was utilized to obtain the cDNA sequences putatively over-expressed during growth on methionine-containing medium and involved in pseudohyphal transition.

Results

Five genes that are over-expressed during yeast-like/pseudohyphal dimorphic transition were isolated. One of these, encoding a putative phospholipase C, is involved in P. fermentans filamentation. In fact, while the inhibition of phospholipase C, by means of 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphorylcholine (Et-18), is accompanied by a significant reduction of pseudohyphae formation in P. fermentans, the addition of exogenous cAMP fully restores pseudohyphal growth also in the presence of Et-18.

Conclusion

Phospholipase C is part of a putative “methionine sensing machinery” that activates cAMP-PKA signal transduction pathway and controls P. fermentans yeast-like/pseudohyphal dimorphic transition.

General significance

Phospholipase C is a promising molecular target for further investigations into the link between pseudohyphae formation and pathogenicity in P. fermentans.     

Phase transition in natural collagen]

The state of water protons and peptide chains in natural collagen was investigated by the method of nuclear magnetic resonance. The study of orientation and temperature dependence of collagen PMR spectra allows us to mage a conclusion about the considerable disorder of mutual orientation in the polypeptide chains at the temperature above -4 degrees C. Under these conditions it was observed that natural collagen was near to the liquid-crystal. The phasic transition at -4 degrees C was discovered, the latter was followed by an abrupt change of the character of water proton mobility and by the change of orientation of collagen fibres order. The thermal effect of transition was measured, the latter presents 18k/gr of the whole specimen.