Enhanced Production of Farnesol by \u3ci\u3eCandida albicans\u3c/i\u3e Treated with Four Azoles

The dimorphic fungus Candida albicans excretes farnesol, which is produced enzymatically from the sterol biosynthetic intermediate farnesyl pyrophosphate. Inhibition of C. albicans by four azole antifungals, fluconazole, ketoconazole, miconazole, and clotrimazole, caused elevated farnesol production (10- to 45-fold). Furthermore, farnesol production occurs in both laboratory strains and clinical isolates (J. M. Hornby et al., Appl. Environ. Microbiol. 67:2982-2992, 2001) of C. albicans

Quorum Sensing in \u3ci\u3eCandida albicans\u3c/i\u3e: Probing Farnesol’s Mode of Action with 40 Natural and Synthetic Farnesol Analogs

The dimorphic fungus Candida albicans produces extracellular farnesol (3,7, 11-trimethyl-2,6,10-dodecatriene- 1-ol) which acts as a quorum-sensing molecule (QSM) to suppress filamentation. Of four possible geometric isomers of farnesol, only the E,E isomer possesses QSM activity. We tested 40 natural and synthetic analogs of farnesol for their activity in an N-acetylglucosamine-induced differentiation assay for germ tube formation (GTF). Modified structural features include the head group, chain length, presence or absence of the three double bonds, substitution of a backbone carbon by S, O, N, and Se heteroatoms, presence or absence of a 3-methyl branch, and the bulkiness of the hydrophobic tail. Of the 40 compounds, 22 showed QSM activity by their ability to reduce GTF by 50%. However, even the most active of the analogs tested had only 7.3% of the activity of E,E-farnesol. Structure-activity relationships were examined in terms of the likely presence in C. albicans of a farnesol binding receptor protein. Includes supplemental material

Farnesol Concentrations Required To Block Germ Tube Formation in \u3ci\u3eCandida albicans\u3c/i\u3e in the Presence and Absence of Serum

Concentrations of (E,E)-farnesol needed to inhibit germ tube formation were determined for Candida albicans strains A72 and SC5314 by using six different conditions known to trigger germination. For defined media, 1 to 2 μM farnesol was sufficient. However, with serum at 2 to 20%, up to 250 μM farnesol was required. Farnesol blocked germ tube formation but did not block elongation of existing germ tubes

Farnesol restores wild-type colony morphology to 96% of \u3ci\u3eCandida albicans\u3c/i\u3e colony morphology variants recovered following treatment with mutagens

Candida albicans is a diploid fungus that undergoes a morphological transition between budding yeast, hyphal, and pseudohyphal forms. The morphological transition is strongly correlated with virulence and is regulated in part by quorum sensing. Candida albicans produces and secretes farnesol that regulates the yeast to mycelia morphological transition. Mutants that fail to synthesize or respond to farnesol could be locked in the filamentous mode. To test this hypothesis, a collection of C. albicans mutants were isolated that have altered colony morphologies indicative of the presence of hyphal cells under environmental conditions where C. albicans normally grows only as yeasts. All mutants were characterized for their ability to respond to farnesol. Of these, 95.9% fully or partially reverted to wildtype morphology on yeast malt (YM) agar plates supplemented with farnesol. All mutants that respond to farnesol regained their hyphal morphology when restreaked on YM plates without farnesol. The observation that farnesol remedial mutants are so common (95.9%) relative to mutants that fail to respond to farnesol (4.1%) suggests that farnesol activates and (or) induces a pathway that can override many of the morphogenesis defects in these mutants. Additionally, 9 mutants chosen at random were screened for farnesol production. Two mutants failed to produce detectable levels of farnesol. Candida albicans est un champignon diploïde qui subit une transition morphologique entre les levures en herbe, les hyphes et les formes pseudohyphales. La transition morphologique est fortement corrélée à la virulence et est régulée en partie par la détection du quorum. Candida albicans produit et sécrète du farnésol qui régule la transition morphologique levure-mycélium. Les mutants qui ne parviennent pas à synthétiser ou à répondre au farnésol pourraient être verrouillés en mode filamenteux. Pour tester cette hypothèse, une collection de mutants de C. albicans a été isolée qui ont modifié les morphologies des colonies, indiquant la présence de cellules hyphales dans des conditions environnementales où C. albicans ne pousse normalement que sous forme de levures. Tous les mutants ont été caractérisés pour leur capacité à répondre au farnésol. Parmi ceux-ci, 95,9% sont entièrement ou partiellement revenus à la morphologie de type sauvage sur des plaques de gélose au levure de malt (YM) complétées par du farnésol. Tous les mutants qui répondent au farnésol ont retrouvé leur morphologie hyphale lorsqu\u27ils ont été recréés sur des plaques YM sans farnésol. L\u27observation selon laquelle les mutants curatifs du farnésol sont si communs (95,9%) par rapport aux mutants qui ne répondent pas au farnésol (4,1%) suggère que le farnésol s\u27active et (ou) induit une voie qui peut supplanter bon nombre des défauts de morphogenèse de ces mutants. De plus, 9 mutants choisis au hasard ont été testés pour la production de farnésol. Deux mutants n\u27ont pas réussi à produire des niveaux détectables de farnésol

Inoculum Size Effect in Dimorphic Fungi: Extracellular Control of Yeast-Mycelium Dimorphism in \u3ci\u3eCeratocystis ulmi\u3c/i\u3e

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 \u3e106 spores per ml and as mycelia when inoculated at 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 dosedependent 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 μM. 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

Global distributed evolution of L-systems fractals

Internet based parallel genetic programming (GP) creates fractal patterns like Koch’s snow flake. Pfeiffer, http://www.cs.ucl.ac.uk/staff/W.Langdon/pfeiffer.html, by analogy with seed/embryo development, uses Lindenmayer grammars and LOGO style turtle graphics written in Javascript and Perl. 298 novel pictures were produced. Images are placed in animated snow globes (computerised snowstorms) by www web browsers anywhere on the planet. We discuss artificial life (Alife) evolving autonomous agents and virtual creatures in higher dimensions from a free format representation in the context of neutral networks, gene duplication and the evolution of higher order genetic operators

High phosphate (up to 600 mM) induces pseudohyphal development in five wild type \u3ci\u3eCandida albicans\u3c/i\u3e

A method is described for the formation of nearly 100% pseudohyphae populations of wild-type Candida albicans A72. The method employs fungal growth at 37 °C (ca. 5×106 cells/ml) in a glucose–proline–N-acetyl-glucosamine medium supplemented with up to 600 mM phosphate (KH2PO4/K2HPO4 1:1) at pH 6.5. Four other strains of C. albicans (MEN, 10261, SG5314 and CAI-4) also formed pseudohyphae under these conditions, although the phosphate response profiles differed in the concentration required for each strain to form pseudohyphae

Defined Anaerobic Growth Medium for Studying \u3ci\u3eCandida albicans\u3c/i\u3e Basic Biology and Resistance to Eight Antifungal Drugs

The polymorphic fungus Candida albicans is one of the most versatile opportunistic pathogens in humans. Many organs of the human body are potential targets for infection by this pathogen, but infection is commonly localized in the gastrointestinal tract, an environment providing anaerobic growth conditions. We describe a chemically defined anaerobic growth medium for four strains of Candida albicans (A72, SC5314, MEN, and 10261). It is a defined liquid glucose-phosphate-proline growth medium supplemented with oleic acid, nicotinic acid, and ammonium chloride. The cells did not require or respond to added ergosterol. Oleic acid and nicotinic acid are growth factors which are required only for the anaerobic growth of C. albicans. An important technical feature of this study was the use of anaerobically grown inocula to study anaerobic growth. Anaerobically, the cells grew exclusively as mycelia at 25, 30, and 37°C. The doubling time at 30°C was ca. 20 h. The cells did not produce farnesol and did not respond to exogenous farnesol, and they were resistant to the highest tested levels of amphotericin B and four of the azole antifungals. We suggest that the anaerobic growth of C. albicans may contribute to the trailing end point phenomenon and the resistance of C. albicans biofilms to antifungal drugs