Comparative activities of glycolytic enzymes in yeast and mycelial forms of Candida albicans

Through use of a synthetic defined medium which allows for the exclusive growth of yeast or mycelial forms of Candida albicans the activity of several major glycolytic enzymes in these forms were examined and compared. The results indicate vast metabolic differences between the forms. These data are discussed in relationship to the phenomenon of morphogenesis in C. albicans which in turn relates to problems in immunology and pathogenics of this important opportunistic organism.     

Effect of tetracycline on the virulence of Y and M forms of Candida albicans in experimentally induced infections

The tetracyclines increase the virulence of Y and M forms ofC. albicans in endoperitoneal infection in mice but not in rabbits inoculated intradermally; there is no significant difference in the mortality grade of mice provoked by Y or M forms. The effects of the tetracyclines are dose dependent; the pathogenicity ofC. albicans forms is influenced to a different degree by different tetracyclines.

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Riassunto La tetraciclina aumenta la virulenza delle forme Y e M dellaC. albicans nelle infezioni endoperitoneali nei topini, ma non nei conigli inoculati per via intradermica; nei topini, tuttavia, non è stata rilevata una significativa differenza nella virulenza delle due fasi morfologiche.E' stato infine rilevato che la patogenicità dellaC. albicans è in rapporto alla dose di antibiotico complessivamente inoculata ed è influenzata in vario grado da differenti tetracicline.

     

Importance of some factors on the dimorphism of Candida albicans

The passage between the yeast and mycelial forms of Candida albicans B 311-10 was studied by using the minimal syntehtic medium of Shepherd et al. [19] modified without biotin and with low glucose concentrations. It was observed that biotin, aminoacids and particularly pH are not important factors in the dimorphism of C. albicans. The only factor of notable importance in the passage of yeast form to mycelial form in C. albicans was glucose concentration.     

Candida albicans enhances initial biofilm growth of Cutibacterium acnes under aerobic conditions

Candida albicans and Cutibacterium acnes are opportunistic pathogens that co-colonize the human body. They are involved in biofilm-related infections of implanted medical devices. The objective of this study was to evaluate the ability of these species to interact and form polymicrobial biofilms. SEM imaging and adhesion assays showed that C. acnes adhesion to C. albicans did not have a preference for a specific morphological state of C. albicans; bacteria adhered to both hyphal and yeast forms of C. albicans. C. albicans did not influence growth of C. acnes under anaerobic growth conditions, however under aerobic growth condition, C. albicans enhanced early C. acnes biofilm formation. This favorable impact of C. albicans was not mediated by secreted compounds accumulating in the medium, but required the presence of metabolically active C. albicans. The ability of these microorganisms to interact together could modulate the physiopathology of infections.     

Rapid enzyme-linked immunosorbent assay (ELISA) for detection of IgG antibodies to Candida albicans

A rapid enzyme-linked immunosorbent assay (ELISA) where the performance time was shortened to 4h was compared with counter-immunoelectrophoresis (CIE) and a standard ELISA procedure for the detection of IgG antibodies to Candida albicans in 61 patients with suspected invasive candidosis. Using a C. albicans cytoplasmic antigen the rapid ELISA compared well with CIE and the standard ELISA. Seventeen sera that reacted with two concentrations of C. albicans antigen in CIE were also positive in both forms of ELISA. Four sera that were CIE-negative were positive in the standard ELISA and three were also positive in the rapid ELISA. The rapid ELISA provides a sensitive and reproducible test for routine serological investigation of different forms of candidosis.     

New milk medium for germ tube and chlamydoconidia production byCandida albicans

A new medium consisting of UHT milk, tween 80 and agar is described for the development of both germ tube and chlamydoconidia byCandida albicans. In total 172 isolates from clinical specimens, includingC. albicans (112),C. guilliermondii (4),C. krusei (3),C. parasilopsis (16).C. tropicalis (28),Torulopsis glabrata (6) andTrichosporon beigellii (3), were examined in this medium by using the standard method. A higher percentage (98.2%) of germ tube production byC. albicans was found in this medium than in undiluted serum (90.2%). In addition, onlyC. albicans was found to be able to produce a high percentage of chlamydoconidia (95.5%) after 48 hours' incubation. In comparison with the conventional medium, corn meal tween 80 agar (21.4%), this new medium gives a significantly higher percentage and abundance of chlamydoconidia production. Being simple, cheap and easy to prepare, the new milk medium is proposed as very practical in the clinical mycology laboratory.     

Pathogenicity of the Y form as compared to M form in experimentally inducedCandida albicans infections

Experimental pathogenicity of the Y form and the M form ofC. albicans separated with a high degree of purity has been evaluated. Experiments are described in which the two morphological forms ofC. albicans were separately inoculated intradermally, intraperitoneally and intravenously in mice and rabbits. In suitable conditions, higher pathogenicity was significantly provoked by the Y form ofC. albicans.

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Riassunto E' stato effettuato uno studio sperimentale sulla patogenicità esercitata nei topini e nei conigli dalle forme morfologiche Y e M dellaC. albicans, separate tra di loro con un alto grado di purezza. Le due forme Y e M sono state inoculate, separatamente, nei conigli per via endovenosa oppure intradermica, nei topini per via intraperitoneale.Nelle condizioni sperimentali seguite, laC. albicans presenta una piú alta patogenicità della forma Y rispetto la forma M.

     

On the Immunochemical and Biochemical Studies of Fungi

The nature of the cross reaction of the mycelial mannan of Trichophyton rubrum and galactomannan isolated from the culture medium of Aspergillus fumigatus with antisera of Saccharomyces cerevisiae and Candida albicans is described. Cross-reactivity of polysaccharides of both T. rubrum and A. fumigatus was weak with antisera of C. albicans and S. cerevisiae, but the galactomannan of A. fumigatus showed slightly stronger activity than the mannan of T. rubrum which possesses more closely related chemical structure of the mannans of S. cerevisiae and C. albicans.     

A simple medium for isolation and identification of Candida albicans directly from clinical specimens

A simple and specific medium consisting of chitosan, trypticase, Tween-80 and agar is devised to isolate the organisms directly from the clinical specimens and to produce germ tubes and chlamydospores for rapid differentiation and identification of Candida albicans from other closely related Candida species. By manipulating the incubating conditions, the specific phase of the organism can be produced in liquid or on solid medium at different time intervals to study the physiology of the organism.Many methods and media have been proposed in the past for identification of Candida albicans and to differentiate this from the closely related species of Candida (5–8, 15). Taschdjian, Burchall&Kozinn (15) showed that C. albicans produces germ tube within an hour or two when it is grown in human or animal serum or serum substitutes. The specificity of this germ tube test was later confirmed by various workers by using different media (3–5). The distinctive feature that differentiates C. albicans from other species is the production of chlamydospores (14). However, in all these studies three types of media were required to isolate the organisms from clinical specimens and to produce germ tubes and chlamydospores for identification. Recently studies have shown that a single medium can be employed to produce both structural components of the organism from the primary isolation medium but the preparation of the medium is more exhaustive (1) and time consuming (13) than the medium to be described here. The present investigation was therefore undertaken to develop a simple and specific medium to isolate the organism directly from the clinical specimens and to produce various morphological phases of Candida albicans to differentiate from other closely related Candida species for clinical diagnosis and to provide a medium to study the physiology and metabolism of the organism under in vitro conditions.Supported in part by Grant CA 20917, National Cancer Institute, National Institutes of Health and ALSAC.     

Study of two forms of the adult Stenocranus minutus

• 1 In studies on Stenocranus minutus, two winged forms of adults have been observed, macropterous and sub-macropterous. Macropterous individuals, with extra-long wings, have not been previously observed or described. Probably only the macropterous form disperses by flight.
• 2 The two forms differ in their wing lengths, intensity and extent of pigmentation.
• 3 The rate of maturation differs in the two forms. In macropters, maturation is delayed; the period of oviposition is shorter and they are less fecund.
• 4 Macropters do not live as long as sub-macropters.
• 5 The macropters weigh less than the sub-macropters.
• 6 Under laboratory conditions, both forms can hop but the macropterous forms fly for a longer time than the sub-macropters.
• 7 At 25°C, more macropterous forms were produced on wilting Dactylis than on green, succulent Dactylis.
• 8 Dispersal of field population was inferred from changes in proportions of the two forms and by simultaneous occurrence of macropters in aerial suction traps.
• 9 The macropterous forms of S.minutus migrate a few weeks after emergence while still immature and migration coincides with drying up of host plant, Dactylis glomerata L.
• 10 Relatively high temperature, low wind speeds and a small number of hours of sunlight seem to favour dispersal by flight in S.minutus.

     

Homologs of the yeast neck filament associated genes: isolation and sequence analysis of Candida albicans CDC3 and CDC10

Morphogenesis in the yeast Saccharomyes cerevisiae consists primarily of bud formation. Certain cell division cycle (CDC) genes, CDC3, CDC10, CDC11, CDC12, are known to be involved in events critical to the pattern of bud growth and the completion of cytokinesis. Their products are associated with the formation of a ring of neck filaments that forms at the region of the mother cell-bud junction during mitosis. Morphogenesis in Candida albicans, a major fungal pathogen of humans, consists of both budding and the formation of hyphae. The latter is thought to be related to the pathogenesis and invasiveness of C. albicans. We have isolated and characterized C. albicans homologs of the S. cerevisiae CDC3 and CDC10 genes. Both C. albicans genes are capable of complementing defects in the respective S. cerevisiae genes. RNA analysis of one of the genes suggests that it is a regulated gene, with higher overall expression levels during the hyphal phase than in the yeast phase. Not surprisingly, DNA sequence analysis reveals that the proteins share extensive homology at the amino acid level with their respective S. cerevisiae counterparts. Related genes are also found in other species of Candida and, more importantly, in filamentous fungi such as Aspergillus nidulans and Neurospora crassa. A database search revealed significant sequence similarity with two peptides, one from Drosophila and one from mouse, suggesting strong evolutionary conservation of function.     

Pseudo-isoenzyme forms of liver prenyl transferase

Prenyl transferase of pig liver exists in two forms, A and B, the latter being more acidic than form A. The ratio of these two forms in initial extracts of the organ varies according to the presence or absence of a thiol reducing agent in the extraction medium. In the presence of 10 mM β-mercaptoethanol the two forms exist as an equilibrium mixture of ca. 70% form A and 30% form B. The latter, when separated and rechromatographed with β-mercaptoethanol reverts to approximate the same 7:3 ratio of forms A and B. In the absence of a thiol reducing agent form B does not change, but form A may be converted almost completely to form B after treatment with oxidized glutathione.     

Fungicidal and Cytotoxic Activity of a Capsicum chinense Defensin Expressed by Endothelial Cells

Plant defensins are antimicrobial peptides that exhibit mainly antifungal activity against a broad range of plant fungal pathogens. However, their actions against Candida albicans have not been extensively studied. The mRNA for γ-thionin, a defensin from Capsicum chinense, has been expressed in bovine endothelial cells. The conditioned medium of these cells showed antifungal activity on germ tube formation (60–70% of inhibition) and on the viability of C. albicans (70–80% of inhibition). Additionally, C. albicans was not able to penetrate transfected cells. Conditioned medium from these cells also inhibited the viability (80%) of the human tumor cell line, HeLa.     

A simple liquid medium for chlamydospore formation in Candida albicans

A new, relatively simple and inexpensive liquid medium was devised to produce all structural forms ofC. albicans. Optimum conditions to induce the yeast cells, germ tubes, pseudohyphae and chlamydospores along with the methods to obtain them are described.Supported in part by Grant CA 20917, National Cancer Institute, National Institutes of Health and ALSAC.     

Immunodiagnosis of Candida-infections. I. Sensitivity of the antigens

Attemps were made to prepare a sensitive antigen from C. albicans suitable for detecting humoral antibodies and hypersensitivity in deep-seated candidiasis, in patients at risk of invasive candidiasis and in allergic states caused-by Candida.5343 persons suffering from systemic, bronchial, vaginal candidiasis, bronchial asthma, chronic bronchitis, polyarthritis nodosa, ulcus cruris, malignancy, rhinitis pollinosa, vasomotorica, and non infected miners, farmers and blood donors were investigated on the presence of antibodies and hypersensitivity against 8 different antigen preparations.The extracellular protein and mannan- protein isolated from the cultivation medium of C. albicans proved the most sensitive for specific anticandida antibodies. The mannan, especially the mannan isolated from the cell surface of C. albicans determined best for the allergy.Comparison made of commercial Candidine showed similar activity. The whole cell C. albicans antigen as well as the mixed Candida antigen reacted much weakly.Comparison made of autoantigen, C. albicans and mixed C. albicans antigen proved the highest sensitivity of the autoantigen.     

Utility of albicans ID plate for rapid identification of Candida albicans in clinical samples

Albicans ID (bioMérieux, Marcy l'Etoile, France) is a ready-to-use medium that contains a chromogenic substrate that allows rapid detection and specific identification of Candida albicans. We have evaluated its clinical performance by culturing 846 clinical specimens from pregnant women and neonates. A 99.2% sensitivity and a 100% specificity were observed in the identification of C. albicans isolates from primary culture.     

The preparation and kinetic properties of multiple forms of chicken brain acetylcholinesterase

A method for preparing various forms of acetylcholinesterase (A ChE) from chicken brain has been developed and they have been characterized in terms of kinetic parameters such as K_m, rate constant (k), turnover number (k_p), specificity constant (k_sp), V_max and half-life (t_1/2). The solubility experiments show that, there are two major forms of A ChE i.e. water-soluble and membrane-bound A ChE (MBA ChE). The MBA ChE shows several subforms, and on the basis of percentage activity only three MBA ChE forms have been selected for complete characterization by various kinetic parameters. It was found that these three forms of MBA ChE demonstrate significant differences in their kinetic properties.     

Studies on the Antigenic Activities of Yeasts

In order to identify the antigenic determinant groups of the mannan of C. albicans serotype A, six kinds of manno-oligosaccharides of up to 7 units in chain-length connected by α1→2 linkages were prepared from the partial acetolysate of the parent mannan. In the precipitation-inhibition test of anti-C. albicans serotype A serum with its homologous mannan, inhibitory power of the oligosaccharides was of the following order: heptaose→: hexaose>pentaose>tetraose>triose> biose, and the amounts for 50%-inhibition of the former four oligosaccharides were 0.08, 0.10, 0.50 and 3.0 μmole respectively, and the inhibitory power of the latter two oligomers at 0.5 μmole were 8 and 5% respectively. On the other hand, the cross-inhibition test of anti-C. albicans serotype A serum with the heterologous mannan of C. albicans serotype B afforded the result that the order of inhibitory activities was hexaose>heptaose>pentaose>tetraose>triose> biose, and that the amounts for 50%-inhibition were 0.05, 0.08, 0.1, 0.45, 0.50 and 3.0μmole respectively. Furthermore, the results of inhibition test on the anti-C. albicans serotype A serum absorbed with the mannan of C. albicans serotype B revealed that the biose, triose and tetraose did not show significant inhibitory power in the range employed, whereas the pentaose, hexaose and heptaose did not significantly affect the inhibitory activities. Thus, it was concluded that the antigenic determinants of the mannan of C. albicans serotype A are α1→2 linked hexaose or heptaose moieties. Based on the above facts, the serological differences between two antigenic mannans of C. albicans serotype A and B may reside at least in the length of the antigenic determinants in which the former is longer than the latter considering the length of the α-D-manno-pyranosyl residue.     

Capsicum annuum L. trypsin inhibitor as a template scaffold for new drug development against pathogenic yeast

A 6,000 Da peptide, named CaTI, was isolated from Capsicum annuum L. seeds and showed potent inhibitory activity against trypsin and chymotrypsin. The aim of this study was to determine the effect of CaTI on Saccharomyces cerevisiae, Candida albicans, Candida tropicalis and Kluyveromyces marxiannus cells. We observed that CaTI inhibited the growth of S. cerevisiae, K. marxiannus as well as C. albicans and induced cellular agglomeration and the release of cytoplasmic content. No effect on growth was observed in C. tropicalis but morphological changes were noted. In the spot assay, different degrees of sensitivity were shown among the strains and concentrations tested. Scanning electron microscopy showed that S. cerevisiae, K. marxiannus and C. albicans, in the presence of CaTI, exhibited morphological alterations, such as the formation of pseudohyphae, cellular aggregates and elongated forms. We also show that CaTI induces the generation of nitric oxide and interferes in a dose-dependent manner with glucose-stimulated acidification of the medium mediated by H⁺-ATPase of S. cerevisiae cells.