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[Correlation among different Candida tropicalis, Candida rhagii and Salmonella cholerae suis 0 6,7 antigen].

The immune sera for Candida tropicalis (C. benhamii) CBS-5701, Candida tropicalis (C. vulgaris) CBS-2310 and C. rhagii (C. tropicalis v. rhagii) CBS-618 agglutinated Salmonella cholarae suis 211 (0 6,7). The immune serum for S. cholerae suis agglutinated C. tropicalis (C. benhamii), C. tropicalis (C. vulgaris) and C. rhagii (C. tropicalis v. rhagii). Absorption and agglutination cross tests demonstrated common antigen factors in the tested species and in S. cholerae suis (0 6,7).

Absorption↗

In vitro susceptibilities of sucrose-negative Candida tropicalis, Candida lusitaniae, and Candida norvegensis to amphotericin B, 5-fluorocytosine, miconazole, and ketoconazole.

The MICs and minimal lethal concentrations of four antimycotics, amphotericin B, 5-fluorocytosine, miconazole nitrate, and ketoconazole, were determined for 25 yeast isolates representing species uncommonly implicated in candidiasis. A microdilution procedure was employed with complex and synthetic media. The isolates, in general, were susceptible to the same antimicrobial agents shown to be effective against Candida albicans, but differences between some of the species in relative susceptibilities to the antifungal agents were noted. Isolates of atypical sucrose-negative Candida tropicalis were similar in their susceptibility patterns to typical isolates of the species. Relative resistance to amphotericin B, miconazole nitrate, and ketoconazole was noted for two Candida lusitaniae isolates, but all strains were susceptible to 5-fluorocytosine. Candida norvegensis isolates were more resistant to miconazole and ketoconazole than C. albicans clinical isolates. The microtiter system was satisfactory for determining minimal inhibitory concentrations, but the system is not recommended for detecting finite differences in drug susceptibilities or for detecting drug synergism.

Amphotericin B↗

Application of CHROMagar Candida for rapid screening of clinical specimens for Candida albicans, Candida tropicalis, Candida krusei, and Candida (Torulopsis) glabrata.

CHROMagar Candida is a new differential culture medium that allows selective isolation of yeasts and simultaneously identifies colonies of Candida albicans, C. tropicalis, and C. krusei. We evaluated the use of this medium with 316 yeast isolates including 247 isolated directly on CHROMagar from clinical material. Over 95% of stock and clinical isolates of C. albicans, C. tropicalis, and C. krusei were correctly identified on the basis of colony morphology and pigmentation on CHROMagar. Additionally, CHROMagar also allowed the identification of C. (Torulopsis) glabrata at a similar level of accuracy. The overall agreement between two observers in reading the CHROMagar plates was 95%. Growth of Candida sp. isolates on CHROMagar had no adverse effect on antifungal MICs or Vitek identification results. In parallel, cultures of 548 stool and rectal swab specimens set up on CHROMagar and Sabouraud glucose agar (SGA) were positive in 234 instances. CHROMagar was positive and SGA was negative for 11 specimens, and CHROMagar was negative and SGA was positive for 18 specimens. A single yeast species was isolated on both media from 162 specimens, and in 146 (90%) of these specimens the same species was detected on both CHROMagar and SGA. A total of 43 of the 234 positive cultures contained mixtures of yeast species. Twenty (47%) of these mixed cultures were detected only on CHROMagar. CHROMagar is extremely useful in making a rapid presumptive identification of common yeast species. This capability plus the ability to detect mixed cultures of Candida spp. promises to improve and streamline the work flow in the mycology and clinical microbiology laboratory.

Adult↗

Comparison of in vivo activity of fluconazole with that of amphotericin B against Candida tropicalis, Candida glabrata, and Candida krusei.

Fluconazole (UK-49,858) is a new oral bis-triazole antifungal agent with demonstrated activity against Candida albicans. Because of the increasing importance of infections due to other species of Candida, we studied the efficacy of fluconazole in a rat model of established systemic candidiasis, using clinical isolates of C. tropicalis, C. glabrata, and C. Krusei. In normal rats, oral fluconazole at both 20 and 80 mg/kg per day for 7 days reduced both kidney and liver titers of C. tropicalis and C. glabrata compared with those in control animals and was only slightly inferior to amphotericin B. Both fluconazole and amphotericin B were ineffective in reducing kidney titers of C. krusei, but amphotericin B was more effective than fluconazole in reducing liver titers. Fluconazole showed no increased efficacy at the higher dose of 80 mg/kg per day compared with 20 mg/kg per day in any experiment. These results suggest that oral fluconazole may be useful in the treatment of established disseminated candidiasis caused by species other than C. albicans. Further in vivo studies are needed, however, to define minimum effective doses and length of therapy and to test additional Candida isolates.

Amphotericin B↗

DNA relatedness, karyotyping and gene probing of Candida tropicalis, Candida albicans and its synonyms Candida stellatoidea and Candida claussenii.

Isolates of Candida albicans with varied phenotypes, including sucrose-negative variants (C. stellatoidea, serotypes A and B) and avirulent germ tube-negative forms (C. claussenii) showed significant (greater than 90%) DNA relatedness to classical C. albicans, but insignificant relatedness to C. tropicalis and sucrose-negative C. tropicalis. A transverse alternating-field gel electrophoresis procedure (TAFE) showed discrete karyotype patterns among the phenotypic variants of C. albicans including the sucrose-negative C. stellatoidea. The number of chromosome-sized DNA bands for C. tropicalis (7 bands) were within the range of bands observed for C. albicans (5 to 10 bands). The general DNA-migration pattern for C. albicans appeared distinct from that of C. tropicalis. An aspartyl proteinase (PrA) gene probe from C. albicans hybridized with chromosomal DNA from C. albicans, C. claussenii and C. stellatoidea but not with that from C. tropicalis.

Aspartic Acid Endopeptidases↗

Secreted aspartic proteases of Candida albicans, Candida tropicalis, Candida parapsilosis and Candida lusitaniae. Inhibition with peptidomimetic inhibitors.

The frequency of Candida infections has increased in recent years and it has been accompanied by a significant rise in morbidity and mortality. The secretion of aspartic proteases by Candida spp. was demonstrated to be one of the virulence determinants. Candida albicans is classified as the major human pathogen in the genus Candida. However, other species of this genus have been found to cause an increasing number of candidiases. We isolated secreted aspartic proteases (Saps) of C. albicans (Sap2p), C. tropicalis (Sapt1p), C. parapsilosis (Sapp1p), and C. lusitaniae (Saplp) from culture media. All the isolated proteases were N-terminally sequenced. Their specific proteolytic activities and sensitivity to series of peptidomimetic inhibitors modified in the type of scissile bond replacement as well as in the N- and C-termini were analyzed. The most divergent substrate specificity was observed for the Sap of C. tropicalis. The specificity of Sap of C. lusitaniae is most closely related to that of Sap of C. parapsilosis. We designed and prepared an inhibitor containing phenylstatine isoster that was equipotent towards all four proteases within the range of 10-10-10-9 M. The HIV-1 protease inhibitors ritonavir, saquinavir, indinavir, and nelfinavir were also tested for the inhibition of four Saps. Only ritonavir and saquinavir inhibited Sap2p, Sapt1p, Sapp1p, and Saplp in micromolar concentrations.

Amino Acid Sequence↗

[Effects of carnitine acetyltransferase gene knockout on long chain dicarboxylic acid production and metabolism of Candida tropicalis].

Candida tropicalis can assimilate n-alkane as a sole carbon source and produce dicarboxylic acids (DCAs). The synthesis of DCAs is thought to be reduced by beta-oxidation. Carnitine acetyltransferase (CAT) is the major enzyme to transfer DCAs into beta-oxidative pathway, then DCAs would be catalyzed to generate ATP to supply cells with energy. A homologous recombination plasmid was constructed, in which CAT gene was disrupted by inserting hygromycin B resistance gene. This plasmid was used to transform Candida tropicalis wild type strain F10-1, and one single CAT gene knockout strain was obtained. Comparing with the wild type, the recombinant increased DCA13 yield and molar conversion of alkane by 13.0% and 11.8%, respectively, and decreased unnecessary consumption of DCAs in beta-oxidation.

Candida tropicalis↗

Extracellular aspartic proteinases from Candida albicans, Candida tropicalis, and Candida parapsilosis yeasts differ substantially in their specificities.

Extracellular aspartic proteinases have been implicated for some time as virulence factors associated with Candida opportunistic fungal infections. Our present knowledge of the enzymatic properties of these proteinases is rather limited. Information on their substrate specificity is important for understanding their roles in invasive Candida infections. We have isolated aspartic proteinases from each of the three Candida yeasts, Candida albicans, Candida tropicalis, and Candida parapsilosis, and investigated the specificities of these proteinases using a library of synthetic substrates and testing inhibition by pepstatin A. The specificities of these aspartic proteinases are different from those of major human proteinases, including gastric pepsins, renal renin, and cathepsin D. For the peptide substrate, Lys-Pro-Ala-Leu-Phe*Phe(p-NO2)-Arg-Leu, the values of kcat/Km were 2.95 x 10(6) M-1 s-1 for cleavage by Candida albicans proteinase, 1.60 x 10(6) M-1 s-1 for cleavage by Candida tropicalis proteinase, and 0.59 x 10(6) M-1 s-1 for Candida parapsilosis proteinase. Substantial differences in specificity among the Candida yeast proteinases were identified. For example, Candida tropicalis shows large changes in the kcat/Km value depending on the acidobasic character of the residue occupying the P2 position (1.6 x 10(6) M-1 s-1 for Leu, 0.47 x 10(6) M-1 s-1 for Lys, and 0.05 x 10(6) M-1 s-1 for Asp at P2, respectively). Candida parapsilosis by comparison is tolerant of these substitutions at P2 and is highly restrictive at position P4.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

Random amplified polymorphic DNA for strain delineation within Candida tropicalis.

Candida tropicalis DNA was used as a template in a polymerase chain reaction (PCR) utilizing a 10-mer primer to generate random amplified polymorphic DNA (RAPD). RAPD patterns associated with 25 primers were obtained for six epidemiologically-unrelated isolates, then a subset of six primers were selected to screen a panel of 18 isolates of C. tropicalis and six isolates of Candida paratropicalis, a species that resembles C. tropicalis but has a sucrose-negative phenotype. The panel, which included nine epidemiologically-related isolates from an outbreak of sternal wound infections, was typed without knowledge of each isolate's origin. The RAPD profiles of the epidemiologically-related isolates were identical to very similar; in contrast, the profiles of most unrelated isolates showed more dissimilarity. While RAPD profiles of C. albicans and Candida parapsilosis differed substantially from those of C. tropicalis, the profiles obtained for C. paratropicalis were consistent with it being a variant of C. tropicalis.

Base Sequence↗

Variations in DNA subtype and antifungal susceptibility among clinical isolates of Candida tropicalis.

Candida tropicalis has been known to be a major cause of invasive Candida infection. Numerous reports have documented C. tropicalis as the most common species of Candida other than C. albicans. The epidemiology and antifungal susceptibility of C. tropicalis are poorly defined. A series of 89 clinical isolates of C. tropicalis from 56 patients hospitalized at seven different U.S. medical centers were analyzed by restriction endonuclease analysis of genomic DNA (REAG) using the restriction enzymes Sfil and BssHII followed by pulsed-field gel electrophoresis (PFGE). The MICs of the isolates for amphotericin B, 5-fluorocytosine (5FC), fluconazole, itraconazole, and D0870 were determined by microbroth dilution testing. A total of 49 different DNA types were identified among the 89 isolates. Generally, each DNA type represented an individual patient, and serial isolates from the same patient were the same DNA type. Small clusters of patients infected with the same DNA type of C. tropicalis suggested possible nosocomial transmission. The MICs of the various antifungal agents were amphotericin B 0.5 to 2.0 micrograms/ml (MIC90 = 2.0 micrograms/ml), 5FC 0.25 to 1.0 microgram/ml (MIC90 = 0.5 microgram/ml), fluconazole 0.25 to 8.0 micrograms/ml (MIC90 = 1.0 microgram/ml), itraconazole 0.03 to 1.0 microgram/ml (MIC90 = 0.5 microgram/ml), and D0870 0.007 to 0.12 microgram/ml (MIC90 = 0.03 microgram/ml). These data support previous observations that infections caused by C. tropicalis frequently originate from the patient's own endogenous flora. Clusters of a single strain in individual hospitals also suggests that limited nosocomial transmission may occur.

Amphotericin B↗

Transformation of fatty acids catalyzed by cytochrome P450 monooxygenase enzymes of Candida tropicalis.

Candida tropicalis ATCC 20336 can grow on fatty acids or alkanes as its sole source of carbon and energy, but strains blocked in beta-oxidation convert these substrates to long-chain alpha,omega-dicarboxylic acids (diacids), compounds of potential commercial value (Picataggio et al., Biotechnology 10:894-898, 1992). The initial step in the formation of these diacids, which is thought to be rate limiting, is omega-hydroxylation by a cytochrome P450 (CYP) monooxygenase. C. tropicalis ATCC 20336 contains a family of CYP genes, and when ATCC 20336 or its derivatives are exposed to oleic acid (C(18:1)), two cytochrome P450s, CYP52A13 and CYP52A17, are consistently strongly induced (Craft et al., this issue). To determine the relative activity of each of these enzymes and their contribution to diacid formation, both cytochrome P450s were expressed separately in insect cells in conjunction with the C. tropicalis cytochrome P450 reductase (NCP). Microsomes prepared from these cells were analyzed for their ability to oxidize fatty acids. CYP52A13 preferentially oxidized oleic acid and other unsaturated acids to omega-hydroxy acids. CYP52A17 also oxidized oleic acid efficiently but converted shorter, saturated fatty acids such as myristic acid (C(14:0)) much more effectively. Both enzymes, in particular CYP52A17, also oxidized omega-hydroxy fatty acids, ultimately generating the alpha,omega-diacid. Consideration of these different specificities and selectivities will help determine which enzymes to amplify in strains blocked for beta-oxidation to enhance the production of dicarboxylic acids. The activity spectrum also identified other potential oxidation targets for commercial development.

Amino Acid Sequence↗

Development of two species-specific fingerprinting probes for broad computer-assisted epidemiological studies of Candida tropicalis.

Candida tropicalis has emerged as the second most frequent colonizing Candida species, and it has been documented in nosocomial infections. To develop an effective fingerprinting system for this species, which is amenable to computer-assisted analyses and which provides information on the genetic relatedness of independent isolates, two DNA fragments, Ct3 (18,000 bp) and Ct14 (20,000 bp), were cloned from a genomic library of Sau3AI partial digestion products. Both probes generate complex Southern blot patterns containing 8 to 20 bands, when hybridized to EcoRI- or EcoRI-HaeIII-digested DNA of independent C. tropicalis isolates. The two probes show no cross-hybridization and are both species specific for C. tropicalis. A comparison of the capacity of the two probes to identify the same strain in different isolates, and differentiate unrelated strains, using computer-assisted computation of similarity coefficients and the genesis of dendrograms, suggests that while Ct14 is more effective in grouping independent isolates, Ct3 is more effective in discriminating small differences in the patterns of highly related isolates and is therefore a more effective probe for determining microevolution within a clonal population and substrain shuffling in recurrent infections. Because of their alternative attributes, it is suggested that both probes be used in fingerprinting studies of C. tropicalis.

Candida↗

Comparison of pulsed-field gel electrophoresis with isoenzyme profiles as a typing system for Candida tropicalis.

Candida species are important nosocomial pathogens, particularly in immunocompromised and critically ill patients. A variety of methods have been used to differentiate strains, but an optimal system has not been established. We compared methods for typing a panel of nine related isolates of Candida tropicalis from an outbreak of sternal wound infections as well as four unrelated control isolates of this species. (The genetic relationships of the nine isolates in the panel had been confirmed previously by restriction fragment analysis.) Typing was undertaken without knowledge of an isolate's origin. Karyotyping by contour-clamped homogeneous electric field (CHEF) gel electrophoresis failed to distinguish between outbreak and control isolates. However, when chromosome-sized DNA was digested with SfiI, EagI, SacII, or NaeI and the fragments were separated by CHEF electrophoresis, the outbreak isolates were readily identified. The isoenzyme profiles of the outbreak isolates were identical and were distinctly different from those of the control isolates. While both isoenzyme profiles and the modified CHEF procedure were discriminatory, the latter is recommended as a relatively convenient and reproducible technique for comparison of types of C. tropicalis.

Acid Phosphatase↗

Phosphorus-31 and carbon-13 nuclear magnetic resonance study of glucose and xylose metabolism in agarose-immobilized Candida tropicalis.

Candida tropicalis can ferment both hexose and pentose sugars. Here, we have used 31P and 13C nuclear magnetic resonance spectroscopy to study the capacity of this yeast species to metabolize glucose or xylose when immobilized in small (< 1-mm-diameter) agarose beads. Immobilized C. tropicalis metabolizing glucose showed rapid initial growth within the beads. A corresponding drop in the intracellular pH (from 7.8 to 7.25) and hydrolysis of intracellular polyphosphate stores were observed. Although the initial rate of glucose metabolism with immobilized C. tropicalis was similar to the rate observed previously in cell suspensions, a decrease by a factor of 2.5 occurred over 24 h. In addition to ethanol, a significant amount of glycerol was also produced. When immobilized C. tropicalis consumed xylose, cell growth within the beads was minimal. The intracellular pH dropped rapidly by 1.05 pH units to 6.4. Intracellular ATP levels were lower and intracellular Pi levels were higher than observed with glucose-perfused cells. Consumption of xylose by immobilized C. tropicalis was slower than was previously observed for oxygen-limited cell suspensions, and xylitol was the only fermentation product.

Candida↗

Germ-tube formation by oral strains of Candida tropicalis.

Candida species isolated from the mouths of healthy children and of patients with denture stomatitis included strains of Candida tropicalis that formed germ tubes when incubated in serum. Twenty-six germ-tube-forming strains of C. albicans and of C. tropicalis were subcultured weekly for 9 wk on blood agar and on Sabouraud's agar and the ability of each subculture to form germ tubes was measured. All the strains of C. albicans formed almost as many germ tubes after nine weekly subcultures as they did when first isolated. By contrast, although all 26 strains of C. tropicalis formed germ tubes when first isolated, all had lost the ability to do so after six serial weekly subcultures. Germ-tube formation should not be the sole criterion for the identification of oral C. albicans strains.

Adult↗

Recombination after protoplast fusion in the yeast Candida tropicalis.

Candida tropicalis protoplasts obtained by snail enzyme treatment were induced to fuse by the use of polyethylene-glycol. Heterokaryons formed by two auxotrophic strains were selected by complementation on minimal medium. These heterokaryons were unstable and readily dissociated into their nuclear components. Under appropriate conditions, the parental nuclei of an heterokaryon fused. The homokaryon so obtained was unstable and segregated into various types of auxotrophic and prototrophic recombinants.

Candida↗

[Increased flavinogenesis in 1 of the strains of Candida tropicalis].

Candida tropicalis IBFM 303 is capable of the active production and liberation of flavins during the oxidation of solid paraffins with the carbon chain consisting of 20 carbon atoms and more. Unlike other known strains, this strain does not accumulate considerable amounts of the vitamin neither on media containing liquid n-paraffins not on media with glucose, even under the conditions of iron deficiency.

Alkanes↗

[Demonstration of 2 phosphate transport systems in Candida tropicalis].

Candida tropicalis has two phosphate transport systems, one of which is constitutive and has a low affinity for its substrate (Kmapp 1.2.10(-3) M). The other one characterized by a high affinity for H2PO4- (Kmapp = 4.5.10(-6) M) appears only under phosphate starvation conditions. The regulation of the latter would act on the one hand on the synthesis of binding proteins for P1 (repression-derepression) and on the other on the activation or inactivation of the carriers.

Binding Sites↗