J.L.F. Kock

The distribution of 3-hydroxy oxylipins in fungi

One of the best-kept secrets by fungi especially yeast is the function of the different shapes and surface structures of their vegetative and sexual cells. They definitely do not produce these shapes (e.g. round, elongated, kidney, needle, hat, saturnoid, etc.) and surfaces (e.g. smooth, rough, hairy, warty, etc.) for our curiosity or to be classified, but surely produce these for their own benefit. This mini-review will show that a large variety of 3-hydroxy oxylipins are widely distributed in the fungal domain and closely associated with these surface ornamentations. In concert with nano-scale surface structures, they probably play a role in cell aggregation as well as spore release from sexual structures such as asci.

Arachidonic acid stimulates cell growth and forms a novel oxygenated metabolite in Candida albicans

Infection of human tissues by Candida albicans has been reported to cause the release of arachidonic acid (AA), eicosanoids and other proinflammatory mediators from host cells. Therefore, we investigated the interaction of this pathogen with AA. AA stimulated cell growth at micromolar concentrations when used as a sole carbon source. Moreover, it selectively inhibited the antimycin A-resistant alternative oxidase. [1-(14)C]AA was completely metabolised by C. albicans. Only one-seventh of the radioactivity metabolised was found in CO(2), whereas two-thirds occurred in carbohydrates suggesting a predominant role of the glyoxalate shunt of citrate cycle. About 1% of radioactivity was found in polar lipids including eicosanoids. A novel AA metabolite, which revealed immunoreactivity with an antibody against 3(R)-hydroxy-oxylipins, was identified as 3, 18-dihydroxy-5,8,11,14-eicosatetraenoic acid. Using immunofluorescence microscopy, endogenous 3(R)-hydroxy-oxylipins were found in hyphae but not in yeast cells. Such compounds have recently been shown to be connected with the sexual stage of the life cycle of Dipodascopsis uninucleata. Together, we propose that infection-mediated release of AA from host cells may modulate cell growth, morphogenesis and invasiveness of C. albicans by several modes. A better understanding of its role is thus promising for novel approaches towards the treatment of human mycoses.

Production of 3R-hydroxy-polyenoic fatty acids by the yeast Dipodascopsis uninucleata

Various fatty acids were fed to the yeast Dipodascopsis uninucleata UOFS Y 128, and the extracted samples were analyzed for the accumulation of 3-hydroxy metabolites with the help of electron impact gas chromatography-mass spectrometry. Fatty acids containing a 5Z,8Z-diene system (5Z,8Z,11Z-eicosatrienoic, 5Z,8Z,11Z,14Z-eicosatetraenoic, and 5Z,8Z,11Z,14Z,17Z-eicosapentaenoic acids) yielded the corresponding 3-hydroxy-all-Z-eicosapolyenoic acids. Moreover, linoleic acid (9Z,12Z-octadecadienoic acid) and 11Z,14Z,17Z-eicosatrienoic acid were converted to the 3-hydroxylated metabolites of shorter chain length, e.g., 3-hydroxy-5Z,8Z-tetradecadienoic acid and 3-hydroxy-5Z,8Z,11Z-tetradecatrienoic acid, respectively. In contrast, no accumulation of a 3-hydroxy metabolite was observed with oleic acid (9Z-octadecenoic acid), linolelaidic acid (9E,12E-octadecadienoic acid), γ-linolenic acid (6Z,9Z,12Z-octadecatrienoic acid), and eicosanoic acid as substrate. These findings pinpoint that the 3-hydroxylation of a fatty acid in Dipodascopsis uninucleata requires a 5Z,8Z-diene system either directly or following initial incomplete β-oxidation. Following analysis of the enantiomer composition, the arachidonic acid metabolite was identified as 3R-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid, which rules out a normal β-oxidation as biosynthetic route to this new class of oxylipins.

A novel oxylipin-associated 'ghosting' phenomenon in yeast flocculation.

Research on the distribution of oxylipins (3-hydroxy fatty acids) in flocculant strains of the yeast Saccharomyces cerevisiae led to the uncovering of a novel ‘ghosting’ phenomenon observed during assumed lectin-mediated aggregation. We found that intracellular oxylipin-containing osmiophilic layers migrate through yeast cell walls in a ‘ghostlike’ fashion without visually affecting the cell wall structure or the layers. This migration resulted in the binding of these layers to cell walls of adjacent cells. Consequently, ‘ghosting’ seems a prerequisite for flocculation to occur. However, ‘ghosting’ alone may not be sufficient to ensure flocculation.

Hydroxy long-chain fatty acids in fungi

Hydroxy long-chain fatty acids occur widely in animals and plants and have important physiological activities in these eukaryotes. There are indications that these compounds are also common and important in fungi. The occurrence of hydroxy-polyunsaturated fatty acids (hydroxy-PUFAs) is of biotechnological importance, because these compounds are potentially high-value lipid products with medical applications. This review pays particular attention to the production of hydroxy-PUFAs by yeasts and other fungi. Hydroxy-PUFAs derived from lipoxygenase activity appear to be present in most fungi, while hydroxy-PUFAs from cyclooxygenase activity (i.e. prostaglandins) have mainly been implicated in the Oomycota and in yeasts from the genus Dipodascopsis. The occurrence of other hydroxy long-chain fatty acids in fungi is also discussed briefly; these include hydroxy fatty acids that are generally associated with cytochrome P-450 monooxygenase activity (i.e. terminal and sub-terminal hydroxy acids and diols derived from the corresponding epoxides) as well as 2-hydroxy-fatty acids and 3-hydroxy-fatty acids.

Production of gamma-linolenic acid by Mucor circinelloides and Mucor rouxii with acetic acid as carbon substrate

SummaryThe production of gamma-linolenic acid (GLA) by Mucor circinelloides CBS 203.28 and M. rouxii CBS 416.77 in fed-batch cultures operated in pH-stat mode with acetic acid as carbon substrate and titrant compared favourably with the performance of M. circinelloides in batch culture on glucose. On acetic acid M. circinelloides accumulated up to 39.8 mg GLA/g biomass, with a crude oil content of 28% containing 91% neutral lipids. The GLA content of the neutral lipid fraction was 15.6%.

Developing a rapid statistical identification process for different yeast species

Abstract A yeast identification procedure was developed with the aid of statistical methods. In this process cellular fatty acids of ten yeast species grown on yeast nitrogen base medium were extracted from yeast cells by saponification and analyzed as methyl esters by gas-liquid chromatography. Each species produced a distinctive fatty acid ‘fingerprint’ characterized by certain fatty acid compositions. A statistical procedure was developed in order to transform the ‘fingerprints’ into a yeast profile which resulted in a marked reduction in variation between profiles of the same yeast species. With this identification method, it was possible to identify the ten species within 4 h after they were obtained from cultures of yeasts grown for 48 h on glucose yeast nitrogen base medium as compared with 7–10 days for the more conventional methods.

An acetylsalicylic acid-sensitive aggregation phenomenon in Dipodascopsis uninucleata

Aggregation of ascospores has been discovered in the yeast Dipodascopsis uninucleata. When this yeast is cultivated to reach the sexual reproductive stage, small ascospores are individually released from the tip of a sac-like ascus which then aggregate in orderly clusters. Acetylsalicylic acid (ASA) inhibited ascospore release and subsequent ordered aggregation process. We suggest that novel ASA-sensitive oxidised fatty acids (3 R-hydroxy-oxylipins) and small hooks located on the surface of these ascospores, are involved.

Differentiation of yeast species, and strains within a species, by cellular fatty acid analysis. 1. Application of an adapted technique to differentiate between strains of Saccharomyces cerevisiae

A fatty acid analysis technique was adapted to give colourless, moisture-free, completely esterified fatty acid extracts from yeasts. Capillary columns with different stationary phases spanning a range of polarity were used ensure optimal resolution of the yeast fatty acids. 15 fatty acids were identified in 13 strains of Saccharomyces cerevisiae by comparison of relative retention times, methyl ester mass spectra and picolinyl ester mass spectra with those of reference standards. Fatty acids were identified by interpretation of picolinyl ester mass spectra in the absence of suitable reference standards. The various strains showed district variations in the mean relative percentages of the fatty acids present in the cellular material. These variations were strain-specific and the fatty acid profiles could be satisfactorily reproduced. Percentage deviation for 10 fatty acids, C14: 0, C14: 1(9), C15: 0, C15: 1(9), C16: 1(11), C18: 0, C18: 1(9) and C18: 1(11) was good and the variation in the mean relative percentages of these fatty acids between strains afforded unambiguous strain differentiation after subjecting the data to an ANOVA. Minor fatty acids (relative percentages ⩽ 1%) were important contributors to the differentiating ability of the technique.

Candida albicans or Candida dubliniensis

Candida albicans is increasing as an opportunistic pathogen causing candidemia and candidiasis worldwide. In addition, other non‐albicans Candida species are now also associated with pertinent infections. These include the closely related C. dubliniensis, which shares many phenotypic similarities with C. albicans. These similarities pose problems in the identification of isolates and have previously led to misidentification of these species. As a result, several identification techniques based on phenotypic and genotypic characteristics have been developed to differentiate between these Candida species. This review will focus on the similarities and differences between these two Candida species highlighting different identification methods and their advantages and disadvantages.