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PubMed2006

Modification of adherence to plastic and to human buccal cells of Candida albicans and Candida dubliniensis by a subinhibitory concentration of itraconazole.

Exposure to subinhibitory concentrations of antifungal agents can influence the adherence of Candida spp. to the host cell. In this study the adherence of Candida albicans ATCC 10231 and Candida dubliniensis CECT 11455 to plastic and to human buccal epithelial cells was evaluated following pre-exposure to 0.5 x minimum inhibitory capacity (MIC) of itraconazole and compared with the corresponding cellular surface hydrophobicity. The yeasts were grown in Sabouraud broth or RPMI-1640 with itraconazole (0.5 x MIC) for 24-26 h at 37 degrees C and the drug was then removed. The adhesion capacity to plastic was studied by turbidimetry in a polystyrene microtiter plate. The adhesion of the yeast to buccal epithelial cells was determined using microscopy techniques. The cellular surface hydrophobicity levels were determined by the microbial adhesion hydrocarbons test. Pre-exposure to itraconazole decreased plastic adherence and cellular surface hydrophobicity in both species when grown in RPMI. When C. albicans was grown in Sabouraud broth, it was nonhydrophobic and did not adhere and therefore no change was detected with the antibiotic. Itraconazole increased adherence to buccal epithelial cells in both species and media studied, as compared to controls without antifungal agents. To study the effects of these antifungal agents on pathogenicity mechanisms, it will be necessary to standardize the methodology for evaluation to determine their in vivo therapeutic efficacy.

Adhesiveness↗
PubMed1997

[Fluconazole-resistant Candida species from HIV infected patients with recurrent Candida stomatitis: cross resistance to itraconazole and ketoconazole].

In vitro susceptibility to fluconazole of Candida species isolated from 83 HIV-infected patients treated with fluconazole because of recurrent Candida stomatitis was monitored over a period of two years. A microdilution assay with high-resolution antifungal assay (HR) medium and RPMI 1640-medium were compared. In vitro less susceptible and fluconazole resistant C. species were observed in 23 patient at the end of the study. The Candida isolates recovered from oral rinsing fluids at the beginning and the end of study were tested for crossresistance to itraconazole and ketoconazole. Susceptibility to ketoconazole and to itraconazole was reduced using RPMI 1640-medium. Susceptibility of the isolates to fluconazole was not influenced by the assay medium. In 21 patients in vitro fluconazole resistant or less susceptible C. albicans were observed. 9 of 21 isolates showed crossresistance to itroconazole and ketoconazole, in 10 isolates only crossresistance to itraconazole was observed. During fluconazole treatment double infections due to combination of C. albicans and C. glabrata or C. krusei increased from 20% to 78% C. krusei was resistant to the three azoles. C. glabrata was less susceptible (4-8 mg/l) or resistant (> 8 mg/l) to fluconazole and resistant to itraconazole and ketoconazole High dosed intravenous fluconazole treatment with 400 to 600 mg daily failed in 11 patients with fluconazole resistant C. albicans and in 3 (3/10) patients with les susceptible C. albicans isolates.

AIDS-Related Opportunistic Infections↗
PubMed1998

Typing of Candida albicans and Candida parapsilosis: species-related limitations of electrophoretic karyotyping and restriction endonuclease analysis of genomic DNA.

Species-related discrimination limits of electrophoretic karyotyping (EK) and restriction endonuclease analysis of genomic DNA (REAG), using pulse-field gel electrophoresis, in typing Candida albicans (CA) and Candida parapsilosis (CP) were compared. Eleven CA and 12 CP isolates from individual neonates and three CA and three CP control isolates were used. For CA, EK and REAG with sfiI displayed seven and six banding-patterns, respectively. One karyotype and two SfiI banding-patterns were observed among the control-isolates. Combining EK/REAG (SfiI) demonstrated nine composites and three distinct control-composites. For CP, EK displayed nine karyotypes, REAG (SfiI) demonstrated four banding-patterns, and REAG (BssHII) yielded six banding-patterns. EK and REAG/SfiI failed to distinguish any CP-controls whereas REAG/BssHII distinguished 2/3 CP-controls. Combining EK/REAG (SfiI) showed 10 composites indistinguishable from CP-controls whereas EK/REAG (BssHII) demonstrated 11 composites and three distinct control-composites. These results illustrate that singly, EK and REAG have significant limitation in typing Candida species though EK is more precise. Combining both methods yields better results but the appropriate restriction endonuclease may vary by strains or species. These findings underscore the importance of combining multiple typing methods, testing several control isolates, and correlating the results with careful epidemiological assessment.

Adult↗
PubMed1998

Candida-associated denture stomatitis. Aetiology and management: a review. Part 1. Factors influencing distribution of Candida species in the oral cavity.

Candida species are yeasts and within the oral cavity, Candida albicans is the most frequently isolated. There is clear evidence that C. albicans adheres to oral surfaces including acrylic dentures and mucosa. The mechanisms of attachment differ, with candidal adhesion to inert surfaces under the control of hydrophobic and electrostatic forces and adhesion to mucosa dependent on a number of complex ligand-recognition systems. Other factors within the oral environment such as saliva, pH, bacteria and hyphal formation have been shown to influence adhesion of candida species to surfaces in the mouth.

Acrylic Resins↗
PubMed2006

Effects of caspofungin against Candida guilliermondii and Candida parapsilosis.

The in vitro activity of caspofungin (CAS) was investigated against 28 yeast isolates belonging to Candida albicans (n = 5), Candida guilliermondii (n = 10), and Candida parapsilosis (n = 13). CAS MICs obtained by broth dilution and Etest methods clearly showed a rank order of susceptibility to the echinocandin compound with C. albicans > C. parapsilosis > C. guilliermondii. Similarly, time-kill assays performed on selected isolates showed that CAS was fungistatic against C. albicans and C. parapsilosis, while it did not exert any activity against C. guilliermondii. In a murine model of systemic candidiasis, CAS given at doses as low as 1 mg/kg of body weight/day was effective at reducing the kidney burden of mice infected with either C. albicans or C. guilliermondii isolates. Depending on the isolate tested, mice infected with C. parapsilosis responded to CAS given at 1 and/or 5 mg/kg/day. However, the overall CFU reduction for C. guilliermondii and C. parapsilosis was approximately 100-fold less than that for C. albicans. Our study shows that CAS was active in experimental systemic candidiasis due to C. guilliermondii and C. parapsilosis, but this activity required relatively high drug dosages.

Animals↗
PubMed2006

Candida albicans and Candida glabrata clinical isolates exhibiting reduced echinocandin susceptibility.

A recognized hotspot for mutations conferring reduced echinocandin susceptibility (RES) is residue S645 of Candida albicans Gsc1(Fks1). We report that the mutation F641Y is associated with RES in a C. albicans isolate. The analogous Fks2 residue is mutated F to V in a Candida glabrata RES isolate; the introduction of this mutation into susceptible C. glabrata confirmed its role in RES. Y641-equivalent Fks residues were identified in intrinsically RES Fusarium species and Candida guilliermondii.

Amino Acid Sequence↗
PubMed1998

Decreased accumulation or increased isoleucyl-tRNA synthetase activity confers resistance to the cyclic beta-amino acid BAY 10-8888 in Candida albicans and Candida tropicalis.

BAY 10-8888, a cyclic beta-amino acid, exerts its antifungal activity by inhibition of isoleucyl-tRNA synthetase activity after accumulation to a millimolar concentration inside the cell. We have selected and characterized BAY 10-8888-resistant Candida albicans mutants. Reduced BAY 10-8888 accumulation as well as increased isoleucyl-tRNA synthetase activity was observed in these mutants. Some of the mutants were cross-resistant to cispentacin, a structurally related beta-amino acid, while sensitivities to 5-fluorocytosine and fluconazole remained unchanged in all mutants. All except two in vitro-resistant mutants were pathogenic in a murine candidiasis model, and BAY 10-8888 failed to cure the infection. Furthermore, we have characterized BAY 10-8888 transport and isoleucyl-tRNA synthetase activity in several Candida tropicalis strains which showed MICs higher than those of other Candida strains. An analysis of the C. tropicalis strains revealed that intracellular concentrations of BAY 10-8888 were in the millimolar range, comparable to those for C. albicans. However, these isolates expressed isoleucyl-tRNA synthetase activities about fourfold higher than those for C. albicans. To test the possibility of resistance modeling, we determined the correlations between the intracellular concentration of BAY 10-8888, the specific activity of isoleucyl-tRNA synthetase, the number of free, i.e., noninhibited, isoleucyl-tRNA synthetase molecules/cell, and growth, assuming a linear relation. We found significant correlations between growth and the intracellular concentration of BAY 10-8888 and between growth and the number of free isoleucyl-tRNA synthetase molecules/cell, but not between growth and the specific activity of isoleucyl-tRNA synthetase.

Candida↗
PubMed2005

Activity of Hoechst 33258 against Pneumocystis carinii f. sp. muris, Candida albicans, and Candida dubliniensis.

Hoechst 33258 is a compound that binds nucleic acids. We report that Hoechst 33258 exhibits antimicrobial activity against Pneumocystis carinii f. sp. muris in a mouse model for P. carinii pneumonia and against Candida albicans and Candida dubliniensis in vitro. Relative to saline treatment, a 14-day, daily treatment of mice with 37.5 mg of Hoechst 33258/kg of body weight after inoculation with P. carinii reduced by about 100-fold the number of P. carinii organisms detected by either PCR or by microscopy after silver staining. For comparison, treatment based on a dose of 15 to 20 mg of the trimethoprim component in trimethoprim-sulfamethoxazole/kg reduced the number of P. carinii by about fourfold. In vitro inhibition of P. carinii group I intron splicing was observed with a 50% inhibitory concentration (IC50) of 30 microM in 2 or 4 mM Mg2+, suggesting RNA as a possible target. However, Hoechst 33258 inhibits growth of Candida strains with and without group I introns. IC50s ranged from 1 to 9 microM for strains with group I introns and were 12 and 32 microM for two strains without group I introns. These studies demonstrate that compounds that bind fungal nucleic acids have the potential to be developed as new therapeutics for Pneumocystis and possibly other fungi, especially if they could be directed to structures that are not present in mammalian cells, such as self-splicing introns.

Animals↗
PubMed1992

Gastrointestinal colonization and systemic dissemination by Candida albicans and Candida tropicalis in intact and immunocompromised mice.

Gastrointestinal colonization and systemic dissemination by Candida albicans and Candida tropicalis were compared in intact and immunocompromised mice. Five-day-old CFW mice were inoculated by the oral-intragastric route with 1.0 x 10(7) CFU of two C. albicans and two C. tropicalis strains isolated from the blood of patients with acute leukemia and with C. albicans 4918 and its cerulenin-resistant mutant 4918-10. C. albicans and C. tropicalis spread to the lungs, liver, and kidneys within 30 min postinoculation, and organ CFU of the two species were comparable over the following 10 days. Close association of blastoconidia with the villous surface of the small intestine resulted in lysis of microvilli and then progressive invasion of villi. Blastoconidia within villi were surrounded by a conspicuous zone of clearing. Persistent colonization of the small and large intestines by C. albicans blood isolates and strains 4918 and 4918-10 was similar for 31 days after inoculation, but consistently exceeded that of C. tropicalis. In mice colonized with C. albicans, immunosuppression with cortisone acetate and cyclophosphamide on days 30 and 33 after inoculation increased stomach CFU 40- to 370-fold and intestinal CFU 30- to 80-fold. In contrast, persistent colonization by C. tropicalis was undetectable before immunosuppression and only became apparent after treatment. C. albicans disseminated more frequently and with higher organ CFU than C. tropicalis. Despite this fact, 20% of mice infected with C. tropicalis died, compared with 4% infected with C. albicans blood isolates. Indirect immunofluorescence revealed penetrative growth by Candida hyphae exclusively in the mucosa and submucosa of the stomach from immunosuppressed, persistently colonized mice. Taken together, the data indicate that C. tropicalis appears to be more virulent than C. albicans and that factors responsible for gastrointestinal colonization, systemic dissemination, and mortality in immunocompromised mice may not be identical.

Animals↗
PubMed1961

Antigenic studies of Candida. I. Observation of two antigenic groups in Candida albicans.

Hasenclever, H. F. (U. S. Public Health Service, Bethesda, Md.), and William O. Mitchell. Antigenic studies of Candida. I. Observations of two antigenic groups in Candida albicans. J. Bacteriol. 82:570-573. 1961.-Two distinct antigenic groups, detected by tube agglutination, have been observed among strains of Candida albicans. Adsorption of an antiserum with a heterologous strain did not remove the agglutinating properties for the homologous strain. Homologous adsorption of the antiserum did remove its agglutinating properties. Seventy-one isolates of C. albicans have been screened with this adsorbed antiserum and 38 were agglutinated at a serum dilution of 1:240 to 1:480 (group A), whereas 33 were not agglutinated at a serum dilution of 1:30 (group B). Thirty-five of the strains were studied with rabbit antisera prepared against each of six strains (three from each group). Agglutination reactions of these strains with samples of each antiserum adsorbed individually with each immunizing strain verified the results of the screening agglutination reactions. All the agglutinating properties of antisera prepared against the three group B strains were removed by adsorption with suspensions of either group A or group B strains. The identity of all isolates used in this study was confirmed by chlamydospore formation, fermentation, and carbohydrate assimilation reactions.

Agglutination Tests↗
PubMed1988

Candida albicans- and Candida stellatoidea-specific DNA fragment.

DNA was isolated from whole cells of Candida albicans and digested with MspI restriction enzyme. In addition to the expected large number of low-molecular-weight DNA pieces resulting from the digestion, multiple high-molecular-weight (greater than 3.0 kilobase pairs) fragments were generated by this enzyme, which cleaves DNA at CCGG sequences. Some of these fragments appeared highly repeated. An MspI fragment which was similar in size to one of the repeat elements (2.9 kilobase pairs) was cloned into the ClaI site of the plasmid vector pBR322 and replicated in a suitable Escherichia coli host strain. The candidal fragment was radiolabeled and used to probe Southern blots of DNA from several Candida species, various other fungi, and mouse and human cells. Only DNA from C. albicans and a strain of Candida stellatoidea was found to contain sequences of significant homology for hybridization. The cloned fragment may possibly be of use as a DNA probe for detection of the presence of C. albicans.

Animals↗
PubMed2000

Growth competition between Candida dubliniensis and Candida albicans under broth and biofilm growing conditions.

Seven isolates each of Candida albicans and Candida dubliniensis were paired (11 pairs) and examined for competitive interaction. Equal numbers of CFU of each competitor were inoculated into Sabouraud dextrose broth and incubated at 37 degrees C with vigorous shaking under conditions favorable to either broth or biofilm growth. Surviving proportions of each competitor were calculated from the broth culture at 24 and 96 h and the biofilm culture at 96 h, with species differentiation done on CHROMagar Candida medium. C. albicans had a competitive advantage over C. dubliniensis in broth culture and under biofilm growing conditions; however, with the presence of a supporting structure for biofilm formation, C. dubliniensis was able to better withstand the competitive pressures from C. albicans.

AIDS-Related Opportunistic Infections↗
PubMed2001

Discrimination between Candida albicans and other pathogenic species of the genus Candida by their differential sensitivities to toxins of a panel of killer yeasts.

The differential sensitivities to toxins produced by a short panel of four killer yeasts allowed discrimination between 91 strains of the yeast Candida albicans and 223 non-C. albicans Candida strains. One hundred percent of C. albicans isolates exhibited negative results to the toxin panel, while 100% of non-C. albicans cultures gave well-defined and reproducible positive results to at least one of the four killer toxins. Among C. albicans strains only 96 and 87% gave germ tube (GT)- and chlamydospore-positive results, respectively. In addition a few GT-false-positive strains were detected among non-C. albicans isolates. Susceptibility to the toxin panel is apparently expressed more consistently than either GT or chlamydospore production and may constitute a promising basis for a new simple and easy-to-use procedure for routine discrimination between the species C. albicans and other species of the genus Candida.

Candida↗
PubMed2003

Contribution of the Platelia Candida-specific antibody and antigen tests to early diagnosis of systemic Candida tropicalis infection in neutropenic adults.

The Platelia Candida-specific antigen and antibody assays (Bio-Rad Laboratories) were used to test serial serum samples from seven neutropenic adult patients with hematological malignancies who had developed systemic Candida tropicalis infections. The diagnosis of candidiasis was based on a positive blood culture (all seven patients) and the isolation of C. tropicalis from a normally sterile site (six patients). All patients received early antifungal therapy with amphotericin B and/or an azole derivative and had successful outcomes. When the combined assays were applied to sera collected at different time points before and after the first positive blood culture, all patients tested positive. In six patients, at least one positive test was obtained with sera collected, on average, 5 days (range, 2 to 10 days) prior to the first positive blood culture, while blood cultures were constantly negative. High and persistent mannanemias were detected in all patients during the neutropenic period. In five patients, an increased antibody response was detected when the patients recovered from aplasia. Controls consisted of 48 serum samples from 12 febrile neutropenic patients with aspergillosis (n = 4), bacteremia (n = 4), or no evidence of infection (n = 4). A low level of mannanemia was detected in only one serum sample, and none showed significant Candida antibody titers. Our data thus confirm the value of the combined detection of mannanemia and antimannan antibodies in individuals at risk of candidemia and suggest that in neutropenic patients, an approach based on the regular monitoring of both markers could contribute to the earlier diagnosis of C. tropicalis systemic infection.

Adult↗
PubMed2004

Evaluation of a rapid immunochromatographic assay for identification of Candida albicans and Candida dubliniensis.

Candida dubliniensis was first established as a novel yeast species in 1995. It is particularly associated with recurrent episodes of oral candidosis in human immunodeficiency virus (HIV)-infected patients, but it has also been detected at other anatomical sites and at a low incidence level in non-HIV-infected patients. It shares so many phenotypic characteristics with C. albicans that it is easily misidentified as such. No rapid, simple, and commercial test that allows differentiation between C. dubliniensis and C. albicans has been developed, until now. Accurate species identification requires the use of genotype-based techniques that are not routinely available in most clinical microbiology diagnostic laboratories. The present study was designed to evaluate the efficiency of a new test (the immunochromatographic membrane [ICM] albi-dubli test; SR2B, Avrille, France) to differentiate between C. albicans and C. dubliniensis. The organisms evaluated were strains whose identities had previously been confirmed by PCR tests and freshly isolated clinical strains and included 58 C. albicans isolates, 60 C. dubliniensis isolates, and 82 isolates belonging to other species of yeast. The ICM albi-dubli test is based on the principle of immunochromatographic analysis and involves the use of two distinct monoclonal antibodies that recognize two unrelated epitopes expressed by both species or specific to only one species. The assay requires no complex instrumentation for analysis and can be recommended for routine use in clinical microbiology laboratories. Results are obtained within 2 h and 30 min and are easy to interpret. This evaluation demonstrated the good performance of this immunochromatographic test for C. albicans and C. dubliniensis isolated on Sabouraud dextrose agar, CHOROMagar Candida, and CandidaSelect, with sensitivities and specificities ranging from 93.1 to 100%. These parameters decreased, however, to 91.4% when the test was performed with yeast isolated with Candida ID.

Antibodies, Fungal↗
PubMed1986

Integrative transformation of Candida albicans, using a cloned Candida ADE2 gene.

Candida albicans is a diploid dimorphic yeast with no known sexual cycle. The development of a DNA transformation system would greatly improve the prospects for genetic analyses of this yeast. Plasmids were isolated from a Candida Sau3A partial library which complements the ade2-1 and ade2-5 mutations in Saccharomyces cerevisiae. These plasmids contain a common region, part of which, when subcloned, produces ade2 complementation. Among the small number of auxotrophs previously isolated in C. albicans, red adenine-requiring mutants had been identified by several groups. In two of these strains, the cloned Candida DNA transformed the mutants to ADE+ at frequencies of 0.5 to 5 transformants per micrograms of DNA. In about 50% of the transformants, plasmid DNA sequences became stably integrated into the host genome and, in the several cases analyzed by Southern hybridization, the DNA was integrated at the site of the ADE2 gene in one of the chromosomal homologs.

Candida↗
PubMed2000

When to suspect fungal infection in neonates: A clinical comparison of Candida albicans and Candida parapsilosis fungemia with coagulase-negative staphylococcal bacteremia.

OBJECTIVES: To determine the epidemiology of candidemia in our neonatal intensive care unit; to compare risk factors, clinical presentation, and outcomes for neonates infected with Candida albicans, Candida parapsilosis, and coagulase-negative staphylococcus (CoNS); and to suggest a rational approach to empiric antifungal therapy of neonates at risk for nosocomial infection. DESIGN: Retrospective chart review of all neonatal intensive care unit patients with systemic candidiasis or CoNS infection between January 1, 1995 and July 31, 1998 at Duke University Medical Center. RESULTS: Fifty-one patients were reviewed. Nine of 19 patients infected with C parapsilosis and 5 of 15 patients infected with C albicans died of fungemia. Seventeen neonates had >2 positive cultures for CoNS obtained within 96 hours and 1 died. There was no statistically significant difference in birth weight, gestational age, or age at diagnosis between patient groups; however, candidemic patients had a sevenfold higher mortality rate. Before diagnosis, candidemic patients had greater exposure to systemic steroids, antibiotics, and catecholamine infusions. Of the 51 patients, 32 received third-generation cephalosporins in the 2 weeks before diagnosis and 19 did not. Twenty-nine of the 32 who were treated with third-generation cephalosporins subsequently developed candidemia, while candidemia occurred in only 5 of 19 patients who were not treated with cephalosporins. At the time of diagnosis, candidemic patients were more likely to have required mechanical ventilation and were less likely to be tolerating enteral feeding. Multivariate clustered logistic regression analysis revealed that candidemic patients had more exposure to third-generation cephalosporins. Once the clinician was notified of a positive blood culture for Candida, patients infected with C parapsilosis retained their central catheters longer than patients infected with C albicans. CONCLUSIONS: In this retrospective review, we were able to identify aspects of the clinical presentation and medication history that may be helpful in differentiating between candidemia and CoNS bacteremia. Those key features may be used by clinicians to initiate empiric amphotericin B therapy in premature neonates at risk for nosocomial infections. Prolonged use of third-generation cephalosporins was strongly associated with candidemia. There was no statistically significant difference in the morbidity and mortality between patients infected with C parapsilosis and those infected with C albicans. Observed delays in removal of the central venous catheter may have contributed to finding a mortality rate from C parapsilosis that was higher than was previously reported.

Amphotericin B↗
PubMed2000

New PCR primer pairs specific for Candida dubliniensis and detection of the fungi from the Candida albicans clinical isolates in Japan.

The genetic patterns of Candida dubliniensis, including type strain, were analyzed by random amplified polymorphic DNA (RAPD) method in comparison with those of reference Candida albicans genotypes A, B and C strains. This RAPD pattern analysis showed that the patterns of two strains of C. dubliniensis including the type strain are very similar to the three oligo PCR primers used, but different from other C. albicans genotypes A, B and C strains with most of the PCR primers tested. The RAPD band considered to be specific for C. dubliniensis in one oligo primer was extracted and sequenced. Based on the sequence information (417-bp), two new PCR primer pairs specific for C. dubliniensis were prepared. The survey of 58 strains of Candida clinical isolates using these newly prepared PCR primer pairs identified as C. albicans in Japan suggested the presence of one strain of C. dubliniensis. The phenotypic characteristics and the sequence analysis of the variable D1/D2 domain of the large subunit (26S) ribosomal DNA of the strain confirmed that it belongs to C. dubliniensis. The usefulness and specificity of these PCR primer pairs for the identification of C. dubliniensis are discussed.

Base Sequence↗