J Infect Dis, 1996 Dec, 174(6), 1369 - 72 Inhibition of Candida albicans growth by calprotectin in the absence of direct contact with the organisms; Sohnle PG et al.; Calprotectin is a calcium- and zinc-binding protein that is present in neutrophil cytoplasm and abscess fluid supernatants . This protein appears to inhibit microbial growth through competition for zinc; however, experiments to show that calprotectin can inhibit growth of microorganisms across filter membranes have yielded conflicting results to date . To prevent recontamination of the filtrate by zinc in this type of experiment, Candida albicans was cultured on filter membranes placed on top of an agarose gel containing calprotectin . In these studies, calprotectin in the gels underneath did suppress growth on top of the filters, an effect reversible by 30 microM ZnSO4 . In other experiments, the protein did not adhere to the organisms and later suppress their growth . These results indicate that calprotectin inhibits C . albicans growth in the absence of direct contact with the organisms; the findings support a zinc-deprivation mechanism of antimicrobial activity for this protein. Gene, 1996 Nov 21, 180(1-2), 189 - 96 A novel group I intron in Candida dubliniensis is homologous to a Candida albicans intron; Boucher H et al.; In the present study, we determined the sequence of group I self-splicing introns found in the large ribosomal RNA subunit of Candida albicans, Candida stellatoidea and the recently-described species Candida dubliniensis . It was found that both the intron and ribosomal RNA nucleotide sequences are almost perfectly identical between different C . albicans strains as well as between C . albicans and C . stellatoidea strains . Comparisons of ribosomal RNA sequences suggest that local isolates of atypical C . albicans from individuals infected with human immunodeficiency virus can be assigned to the C . dubliniensis species . C . dubliniensis strains also harbor a group I intron in their ribosomal RNA, as observed in about 40% of C . albicans strains and all C . stellatoidea strains . This novel C . dubliniensis group I intron is identical to the C . albicans and C . stellatoidea intron, except for two widely divergent stem-loop regions . Despite these differences, the C . dubliniensis intron possesses self-splicing ability in an in vitro assay . Taken together, these data support the idea that C . albicans and C . stellatoidea should be joined together as variants of the same species while C . dubliniensis is a distinct but closely related microorganism . To our knowledge, the C . albicans and C . dubliniensis introns are the first example of a pair of homologous group I introns differing only by the presence of apparently facultative sequences in some stem-loops suspected to be involved in stabilization of tertiary structure. Eur J Biochem, 1996 Nov 15, 242(1), 29 - 35 Exchange of regions of the carboxypeptidase Y propeptide . Sequence specificity and function in folding in vivo; Ramos C et al.; The propeptide of carboxypeptidase Y from Saccharomyces cerevisiae is important for folding of the enzyme . Previous work {Ramos, C., Winther, J.R . & Kielland-Brandt, M . C . (1994) J . Biol . Chem . 269, 7006-7012} suggested that the sequences essential for in vivo folding were situated in the COOH-proximal third of the propeptide . Concentrating on this region we have investigated the functionality of propeptide variants . Using a random mutagenesis approach we found that two segments can be defined: one in which there is a fairly high tolerance for substitution with unrelated sequences and another that has a more strict requirement for sequence conservation . Nevertheless, an overall lack of requirement for propeptide sequence conservation was found by substitution of the carboxypeptidase Y propeptide with that of a highly divergent propeptide sequence from an otherwise similar carboxypeptidase from Candida albicans . This propeptide was partially functional when combined with carboxypeptidase Y . Analysis of the biosynthesis of the mutant forms of the zymogen showed that a fraction of the molecules proceeded from the endoplasmic reticulum with fairly rapid kinetics, while the rest was degraded. Anal Biochem, 1996 Nov 15, 242(2), 165 - 71 Discrimination between acid and alkali-labile phosphorylated residues on Immobilon: phosphorylation studies of nucleoside diphosphate kinase; Biondi RM et al.; We have critically analyzed current methodologies for distinguishing histidine and serine phosphorylated residues in proteins and report a simple technique that assures a reliable discrimination . Electro-transfer of a phosphorylated enzyme to Immobilon membranes and its treatment at pH 1 and 14 in buffers containing 5% methanol allows unambiguous distinction between serine/threonine and histidine phosphorylation (O-phosphomonoesters and phosphoramide, respectively) since under these conditions only one type of residue is dephosphorylated . The addition of 5% methanol to all buffers was indispensable to deplete phosphate from membranes incubated successively under acid and basic conditions . The technique was applied to the study of nucleoside diphosphate kinase (NDP kinase) phosphorylation . In this enzyme, autophosphorylation of active site histidine is an accepted intermediate step in the catalytic phosphate transfer activity of nucleoside diphosphate kinase (NDP kinase) . Nonetheless, a significant degree of autophosphorylation on other residues has been reported by several laboratories, and the hypothesis has been advanced that this nonhistidine phosphorylation may play an important role in NDP kinase cellular function, signaling the suppression of metastasis in the case of human NDP kinase A . Using this improved method, we show that human, Escherichia coli and Candida albicans NDP kinases are only autophosphorylated on histidine residues . In addition, we present evidence that the presence of phosphoserine after strong acid hydrolysis of the histidine autophosphorylated enzyme is in fact a nonenzymatic transphosphorylation from phosphohistidine due to the harsh acid treatment . This methodology was also applied to in vivo phosphorylation studies of C . albicans NDP kinase . We believe that the technique will be generally useful in histidine phosphorylation screenings. Proc Natl Acad Sci U S A, 1996 Nov 12, 93(23), 13223 - 8 Candida albicans strains heterozygous and homozygous for mutations in mitogen-activated protein kinase signaling components have defects in hyphal development; Kohler JR et al.; The Candida albicans genes, CST20 and HST7, were cloned by their ability to suppress the mating defects of Saccharomyces cerevisiae mutants in the ste20 and ste7 genes, which code for elements of the mating mitogenactivated protein (MAP) kinase pathway . These Candida genes are both structural and functional homologs of the cognate Saccharomyces genes . The pattern of suppression in Saccharomyces is related to their presumptive position in the MAP kinase cascade . Null alleles of these genes were constructed in Candida . The Candida homozygous null mutants are defective in hyphal formation on some media, but are still induced to form hyphae by serum, showing that serum induction of hyphae is independent of the MAP kinase cascade . The Candida heterozygotes CST20/cst20 and HST7/hst7 are also defective in hyphal formation . This lack of dominance of the wild-type allele suggests that gene dosage is important in Candida. Proc Natl Acad Sci U S A, 1996 Nov 12, 93(23), 13217 - 22 Signal transduction through homologs of the Ste20p and Ste7p protein kinases can trigger hyphal formation in the pathogenic fungus Candida albicans; Leberer E et al.; The CST20 gene of Candida albicans was cloned by functional complementation of a deletion of the STE20 gene in Saccharomyces cerevisiae . CST20 encodes a homolog of the Ste20p/p65PAK family of protein kinases . Colonies of C . albicans cells deleted for CST20 revealed defects in the lateral formation of mycelia on synthetic solid "Spider" media . However, hyphal development was not impaired in some other media . A similar phenotype was caused by deletion of HST7, encoding a functional homolog of the S . cerevisiae Ste7p protein kinase . Overexpression of HST7 partially complemented the deletion of CST20 . Cells deleted for CST20 were less virulent in a mouse model for systemic candidiasis . Our results suggest that more than one signaling pathway can trigger hyphal development in C . albicans, one of which has a protein kinase cascade that is analogous to the mating response pathway in S . cerevisiae and might have become adapted to the control of mycelial formation in asexual C . albicans. Neuroimmunomodulation, 1996 Nov-Dec, 3(6), 333 - 5 Interaction of cyclophosphamide and ketamine in vivo; Rojavin MA et al.; Cyclophosphamide 100 mg/kg i.p . increased the duration of ketamine-induced anesthesia in BALB/c mice by 39% . However, combined action of these two substances did not change the number of splenocytes, proliferation of T cells, or phagocytic activity of murine peritoneal macrophages against Candida albicans. J Investig Allergol Clin Immunol, 1996 Nov-Dec, 6(6), 392 - 4 Candidin in immediate hypersensitivity . Comparison of two antigens; Netto CF et al.; One hundred outpatients in a Clinic of Allergy were submitted to intradermic tests with two types of candidins . The main focus of the research was the comparison of two antigens obtained from the same strains of Candida albicans: one a suspension of yeast cells and the other, a polysaccharide . The readings, taken 20 minutes after the intradermic injections, with positive results were considered as hypersensitivity of the immediate type . Positive results were obtained in 74% of the patients with the yeast cell antigen and in 73% with the polysaccharide antigen . This research mainly deals with the advantages that can be obtained by using the polysaccharide antigen in intradermic tests for evaluating hypersensitivity of the immediate type. Appl Microbiol Biotechnol, 1996 Nov, 46(4), 400 - 4 Evaluation of 2-{N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino}-2-deoxy-D-glucose, a new fluorescent derivative of glucose, for viability assessment of yeast Candida albicans; Yoshioka K et al.; A new fluorescent derivative of D-glucose, 2-{N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino}-2-deoxy-D-glucose (2-NBDG), which had been previously developed for the analysis of glucose uptake activity by living cells, was investigated to evaluate its applicability for assaying the viability of yeast Candida albicans . Lineweaver-Burk plots showed to uptake of 2-NBDG to be competitively inhibited by D-glucose and not by L-glucose, which suggested the involvement of the glucose transporting system of C . albicans in the uptake of 2-NBDG . A good correlation was obtained between the yeast viability, determined by the plate-count method, and the 2-NBDG uptake activity of yeast cells (correlation constant: r = 0.97) . This is expected to lead to the development of a new fluorescent probe for the determination of yeast cell viability. J Med Vet Mycol, 1996 Nov-Dec, 34(6), 401 - 6 Development of a method to detect secretory mucinolytic activity from Candida albicans; Colina AR et al.; Ultrastructural examinations of sites where Candida albicans invaded the bowel wall after oral intragastric inoculation of infant mice suggested that blastoconidia are capable of progressive extracellular digestion of the intestinal mucus barrier . Microplate assay methods, based on biotin or digoxigenin-labelling systems, were therefore devised for quantitation of protease and glycosidase activities against the glycoprotein mucin . Labelled mucin was adsorbed on microplate wells, incubated with sample to be assayed for enzyme activity, and the remaining labelled mucin was quantitated by spectrophotometry . Proteolytic activity against mucin was demonstrated using concentrated culture filtrate of C . albicans strain LAM-1, grown in yeast nitrogen base medium containing mucin as sole nitrogen source . The activity was inhibited by boiling for 10 min or by incubation with the aspartyl proteinase inhibitor pepstatin A. J Med Vet Mycol, 1996 Nov-Dec, 34(6), 393 - 400 Molecular cloning of a gene encoding translation initiation factor (TIF) from Candida albicans; Mirbod F et al.; The differential display technique was applied to compare mRNAs from two clinical isolates of Candida albicans with different virulence; high (potent strain, 16240) and low (weak strain, 18084) extracellular phospholipase activities . Complementary DNA fragments corresponding to several apparently differentially expressed mRNAs were recovered and sequenced . A complementary DNA fragment seen distinctly in the potent phospholipase producing strain was highly homologous to the yeast translation initiation factor (TIF) . The selected DNA fragment was then used as a probe to isolate its corresponding complementary DNA clone from a library of C . albicans genomic DNA . The sequence of isolated gene revealed an open reading frame of 1194 nucleotides with the potential to encode a protein of 397 amino acids with a predicted molecular weight of 43 kDa . Over its entire length, the amino acid sequence showed strong homology (78-89%) to Saccharomyces cerevisiae TIF and (63-80%) to mouse eIF-4A proteins . Therefore, our C . albicans gene was identified to be TIF (Ca TIF) . Northern blot analysis in the two strains of C . albicans revealed that Ca TIF expression is 1.5-fold higher in the potent phospholipase producing strain . The restriction endonuclease digestion of genomic DNA from this potent strain revealed at least two hybridized bands in Southern blot analysis, suggesting two or more closely related sequences in the C . albicans genome. Microbiology, 1996 Nov, 142 ( Pt 11), 3171 - 80 Changes in cell morphology and carnitine acetyltransferase activity in Candida albicans following growth on lipids and serum and after in vivo incubation in mice; Sheridan R et al.; Candida albicans C316, maintained in the yeast form, showed a proliferation of peroxisomes when grown on triolein or serum as sole carbon source but these structures were absent from glucose-grown cells . Peroxisomes were also apparent in C . albicans obtained after injection into mice and recovery from intraperitoneal washings and kidneys; they may therefore be useful markers to assess a potential in vivo response in cells that are growing in vitro . Transcell-wall structures also occurred in C . albicans grown on triolein or serum, and in cells cultured in vivo, but were not seen in cells grown on glucose . These structures consisted of electron-dense fibrillar material penetrating through the cell wall from the plasmalemma side and protruded out to the exterior of the cell . Endoplasmic reticulum, located at the periphery of the cell, was found to be in close proximity with these cell wall structures . Carnitine acetyltransferase (CAT; EC 2.3.1.7), the key enzyme for the translocation of acetyl units between intracellular compartments, was present in low activities in glucose-grown cells; its activity was increased some 100-fold in triolein-grown cells but only 4-fold in serum-grown cells . It was not possible to assess this activity in the in vivo-cultured cells . Two separate CAT proteins, partially purifed from isolated microchondria and peroxisomes, respectively, were identified, with different specificities and kinetic properties. J Infect, 1996 Nov, 33(3), 221 - 6 Identification of an amphotericin B resistant strain of Candida albicans using a rapid 3H-glucose incorporation microassay; Sweeney JF et al.; Using a 3H-glucose incorporation assay, antifungal sensitivity testing undertaken on an isolate of Candida albicans cultured from the blood of a bone marrow transplant patient documented resistance to amphotericin B but sensitivity to fluconazole and itraconazole . Information obtained from in vitro antifungal sensitivity testing can be used to direct in vivo antifungal therapy . Widespread application of standardized in vitro antifungal sensitivity testing is needed. Immunopharmacol Immunotoxicol, 1996 Nov, 18(4), 549 - 69 Impairment of non-specific immunity in patients in persistent vegetative state; Munno I et al.; In fourteen patients in persistent vegetative state (PVS) immune responsiveness was investigated . In particular, we studied the relationship between brain lesions following traumatic injury and immune system . In this respect, phagocytosis and killing of Candida albicans by polymorphonuclear cells (PMN) and monocytes were tested . In addition serum levels of Interferon-gamma (IFN-gamma) were evaluated . The patients come out from PVS by 3-4 month were used as control group . Data shown a profound impairement of phagocytosis and killing of monocytes and low serum levels of IFN gamma when compared with normal values . Taken together, these findings suggest that brain lesions, may affect non-specific immune response. Curr Genet, 1996 Nov, 30(5), 423 - 31 Molecular cloning of a cDNA encoding enolase from the filamentous fungus, Aspergillus oryzae; Machida M et al.; A 1.6-kbp full-length cDNA for the Aspergillus oryzae enolase gene (enoA) was cloned . The sequenced insert contained a continuous open reading frame of 1314 bp encoding a protein of molecular weight 47 405 . Among all enolases sequenced to-date, the deduced amino-acid sequence showed the highest homology (74.9%) with Candida albicans enolase (ENO1) . Strong codon biases and multiple transcription start sites downstream from CT-blocks in the 5'-flanking region suggested strong expression . enoA mRNA was found to occupy approximately 3% (w/w) of total mRNA of A . oryzae by quantitative RT-PCR . This strong transcription was dependent on the carbon source in the medium and correlated with the growth rate of the mycelium. Arch Microbiol, 1996 Nov, 166(5), 327 - 35 Study of supramolecular structures released from the cell wall of Candida albicans by ethylenediamine treatment; Mormeneo S et al.; Candida albicans cell wall components were analyzed by ethylenediamine (EDA) treatment . Based on their different solubility properties, the cell wall components produced three fractions (A, B, and C) . Fractions B (EDA-soluble, water-insoluble) and C (EDA-insoluble) contained glucan, chitin, and protein in different proportions . After zymolyase (mainly a beta-glucanase complex) or chitinase treatment of fractions B and C, more polysaccharides and proteins were solubilized by a second EDA treatment, suggesting that the solubility of the polymers in EDA depends on the degree of polymer interactions . Western blot analysis using two monoclonal antibodies (1B12 and 4C12) revealed electrophoretic patterns that were similar in mycelial and yeast morphologies, except that in material obtained from mycelial walls, an additional band was detected with MAb 1B12 . Fluorescence microscopy of cell wall fractions treated with FITC-labeled Con-A, Calcofluor white, and FITC-labeled agglutinin showed that glucan and mannoproteins are uniformly distributed in fractions B and C, while chitin is restricted to distinct patches . Transmission electron microscopy demonstrated that fraction C maintained the original shape of the cells, with an irregular thickness generally wider than the walls . When fraction C was treated with chitinase, the morphology was still present and was maintained by an external glucan layer, with an internal expanded fibrillar material covering the entire cellular lumen . Degradation of the glucan skeleton of fraction C with zymolyase resulted in the loss of the morphology. J Pediatr, 1996 Nov, 129(5), 688 - 94 Candida albicans arthritis one year after successful treatment of fungemia in a healthy infant; Swanson H et al.; Fungal arthritis in pediatric patients is rare and is most often associated with hematogenous spread to the affected joint . It is generally seen concomitant with, or shortly after, fungemia . We report a case of an immunocompetent patient in whom candidal arthritis developed 1 year after initial fungemia . The initial candidiasis was considered to be adequately treated with amphotericin B . The Candida isolates from the neonatal fungemia and subsequent arthritis were the some as identified by electrophoretic karyotype, restriction fragment length polymorphism analysis, and antifungal susceptibility testing . Pediatric candidal fungemia, arthritis, and their treatments are discussed. Clin Diagn Lab Immunol, 1996 Nov, 3(6), 740 - 5 Production of T-helper cell subsets and cytokines by lymphocytes from patients with chronic mucocutaneous candidiasis; Kobrynski LJ et al.; Chronic mucocutaneous candidiasis (CMC) is a heterogeneous group of disorders characterized by recurrent and persistent superficial candidal infections . Cytokine-induced dysregulation of T-helper cell function has been described in other immune-deficient states but has not been studied in CMC patients . We studied T-helper cell subsets by flow cytometry and cytokine production by stimulated lymphocytes in six CMC patients, two healthy pediatric controls, and five healthy adult controls . Peripheral blood lymphocytes were stimulated in vitro with phytohemagglutinin or Candida albicans extract, and the production of interleukin-2R (IL-2R), IL-4, IL-10, and gamma interferon in the supernatants was measured by enzyme-linked immunosorbent assay . CMC patients had a decrease in the CD29+/CD29+ cell population compared with the numbers in controls (P < 0.02) . The percentage of CD4+/CD45RA+ cells was greater in patients than in controls, but the difference was not significant . There was no difference in the production of IL-10 or gamma interferon by the patient lymphocytes . CMC patients produced more IL-4 than the controls (P < 0.001), whereas the controls tended to produce more IL-2R than the patients (P = 0.19) . These findings support the concept that a decrease in CD4+/CD29+ T-helper inducer cells along with T-helper cell dysregulation may lead to defective memory responses to antigens in CMC patients and a decrease in cell-mediated immunity due to inhibition of TH1 cells by increased levels of IL-4. Clin Diagn Lab Immunol, 1996 Nov, 3(6), 645 - 50 Evidence for the presence of immunoglobulin E antibodies specific to the cell wall phosphomannoproteins of Candida albicans in patients with allergies; Kanbe T et al.; To determine the major antigenic component of Candida albicans against immunoglobulin E (IgE) antibodies in the sera of patients with allergies who were positive for IgE antibodies to C . albicans crude antigen in a CAP system, phosphomannoproteins (CAMP/A or CAMP/B for serotype A or B strain, respectively) and their acid-stable portions (CAMP-S/A or CAMP-S/B) were isolated from beta-mercaptoethanol (2-ME) extracts of C . albicans cells of serotypes A and B, and IgE antibodies against these components were compared with those against protein complex and enolase (CAE) fractions isolated from C . albicans cells . The dot blot test, which was used to detect IgE antibodies to the C . albicans antigens, showed that IgE antibodies to the 2-ME extract and phosphomannoprotein fractions were present in the sera of 98.0% (2-ME extract), 96.8% (CAMP/A), 93.2% (CAMP-S/A), 97.2% (CAMP/B), and 81.5% (CAMP-S/B) of the patients, whereas IgE antibodies to the protein complex and CAE fractions were found in the sera of 73.6 and 48.8% of the patients, respectively . The extent of IgE binding to the 2-ME extract and phosphomannoproteins was well correlated with the fluorescence intensities estimated with the CAP system . Furthermore, the results obtained from the inhibition experiment with the CAP system indicated that the binding of IgE antibodies to Candida antigens is strongly inhibited by the phosphomannoprotein fraction and is an indication that the serum of the patients contained IgE antibodies specific to the cell wall phosphomannoproteins of C . albicans . Finally, an initial chemical analysis indicated that the epitopes for IgE antibodies on the phosphomannoproteins is a carbohydrate portion, since the ability of CAMP/A to inhibit the binding of IgE antibodies to the homologous CAMP/A was destroyed after oxidation by sodium periodate but not after digestion with proteinase K. Genetics, 1996 Nov, 144(3), 991 - 1003 Asm-1+, a Neurospora crassa gene related to transcriptional regulators of fungal development; Aramayo R et al.; This report describes the identification, cloning, and molecular analysis of Asm-1+ (Ascospore maturation 1), the Neurospora crassa homologue of the Aspergillus nidulans stuA (stunted A) gene . The Asm-1+ gene is constitutively transcribed and encodes an abundant, nucleus-localized 68.5-kD protein . The protein product of Asm-1+ (ASM-1), contains a potential DNA-binding motif present in related proteins from A . nidulans (StuA), Candida albicans (EFGTF-1), and Saccharomyces cerevisiae (Phd1 and Sok2) . This motif is related to the DNA binding motif of the Swi4/Mbp1/Res family of transcription factors that control the cell cycle . Deletion of Asm-1+ destroys the ability to make protoperithecia (female organs), but does not affect male-specific functions . We propose that the APSES domain (ASM-1, Phd1, StuA, EFGTF-1, and Sok2) defines a group of proteins that constitute a family of related transcription factors involved in the control of fungal development. Antimicrob Agents Chemother, 1996 Nov, 40(11), 2622 - 5 Effect of fluconazole on viability of Candida albicans over extended periods of time; Sohnle PG et al.; The treatment of chronic mycoses may expose the infecting organisms to antimicrobial agents for extended periods of time . It is possible that an azole antifungal drug such as fluconazole, with primarily fungistatic activity in standard in vitro susceptibility tests, might be able to damage the fungal cells and reduce their viability over prolonged incubations under nonproliferating conditions . To test this possibility, Candida albicans yeast cells were exposed to various concentrations of fluconazole in RPMI 1640 tissue culture medium for 4 h at 37 degrees C, washed free of the drug, and then incubated at 37 degrees C for a 28-day period; enumeration of the remaining CFU at various times during this period revealed no increased loss of viability for the fluconazole-exposed organisms . However, when fluconazole was added to the organisms maintained in distilled water (with or without pretreatment with the drug), a marked reduction of viability was found . At 14 days of incubation with two strains of C . albicans, negative cultures were found for 7 of 10 and 10 of 11 samples, respectively, containing 1.0 microgram of fluconazole per ml versus 0 of 10 and 1 of 11 control samples (P of < 0.01 and 0.001, respectively) . The effect of fluconazole on fungal viability under these conditions became noticeable at approximately 7 days and was greater when the samples were incubated at 37 degrees C rather than 25 degrees C . These findings suggest that fluconazole may have fungicidal effects on fungal cells during prolonged exposures under conditions in which the organisms are prevented from proliferating by lack of nutrients. Antimicrob Agents Chemother, 1996 Nov, 40(11), 2511 - 6 In vitro interaction between amphotericin B and azoles in Candida albicans; Vazquez JA et al.; The use of azole prophylaxis as a measure to prevent invasive fungal infections in high-risk patients is increasing and is now the standard of care in many institutions . Previous studies disagree on whether preexposure of Candida albicans to azoles affects their subsequent susceptibility to amphotericin B (AmB) . The present in vitro study indicates that azole exposure generates a subpopulation of cells that are not affected by subsequent exposure to AmB . These cells that are phenotypically resistant to AmB tolerated by most cells not exposed to azole . The percentage of cells that convert to phenotypic resistance to AmB varies with the concentration and the azole . Itraconazole is more effective than fluconazole in generating cells that are phenotypically resistant to AmB and that tolerate an otherwise lethal transient exposure to AmB . Until cells that are not exposed to fluconazole are simultaneously challenged with AmB, they are not protected to a significant extent from killing by AmB . Cells that are challenged with continuous exposure to AmB also acquire phenotypic resistance to AmB at increased frequencies by azole preexposure, but this requires that the azole be continuously present during incubation with AmB . In addition, Candida cells taken from mature colonies that are not actively growing are not susceptible to the short-term killing effects of AmB without azole preexposure . The adaptive responses of C . albicans to AmB and potentially other antifungal agents that may result from prior exposure to azoles in vitro or potentially in microenvironments in vivo that induce physiological changes may have major clinical implications. Artif Organs, 1996 Nov, 20(11), 1191 - 5 Antimicrobial activities of iodinated polystyrene derivatives; Lin KJ et al.; The antimicrobial activities of insoluble halogenated acetamidomethy1-styrene polymers (prepared by covalent bonding of iodine to polystyrene) were assessed as were the factors determining antimicrobial efficacy . The most active materials were selected from chlorinated or iodinated polymers . Antimicrobial activities were assessed for Escherichia coli (ATCC 25922; American Type Culture Collection, Rockville, MD, U.S.A.), Saccharomyces cerevisiae, and Candida albicans by determining time-course changes in microbial counts in vitro . A 2-iodoacetamidomethylstyrene polymer (No.6-I:-CH2I) was found to have the greatest antimicrobial activity against both bacteria and fungi . No.6-I is the first antimicrobial material that did not make an inhibition hollow in the conventional diffusion test or for which conjugated iodine showed antibacterial activity . This material can be introduced into styrene units on the surface of devices by chemical modification . This material was most active at 37 degrees C . For coated dishes, antimicrobial activity depended on the depth or swollen character of the reactive layer . NO.6-I requires not only a minimum width of polymer layer, but also frequent contact with microbes to have an antimicrobial effect . No.6-I is valuable as a new material because it has strong antimicrobial activity by itself but does not release active iodine . This material is expected to have various applications in implantable clinical devices. South Med J, 1996 Nov, 89(11), 1104 - 7 Infection of a pancreatic pseudocyst due to Candida albicans; Foust RT; A 40-year-old man with diabetes mellitus, congestive heart failure, alcoholic cirrhosis, and chronic pancreatitis had an exacerbation of pancreatitis due to alcohol abuse . His condition deteriorated rapidly with development of apparent sepsis; cultures were negative . He slowly improved with multiple antibiotic therapy and total parenteral nutrition . Serial imaging of the pancreas revealed edematous pancreatitis that evolved initially into a phlegmon and later into multiple pseudocysts . Intermittent fever prompted computed-tomography-directed percutaneous aspiration of the largest pancreatic fluid collection, yielding purulent material that grew only Candida albicans . Subsequently, disseminated candidiasis developed . Despite therapy with amphotericin B and aggressive supportive care, the patient died from multiple organ system failure. J Am Vet Med Assoc, 1996 Nov 1, 209(9), 1604 - 7 Primary photosensitization related to ingestion of alfalfa silage by cattle; House JK et al.; A herd of 650 Holstein cows was examined for skin disease . Approximately 400 of the lactating adults were affected, but heifers, calves, and nonlactating cows were clinically normal . The condition was characteristic of primary photosensitization . Milk production of the affected cows was normal . Affected cows did not appear to be ill, and none of the cows was icteric . Three of 7 cows had high serum gamma-glutamyltransferase activities, but in the other 4 cows, activity was within the reference range . Serum activities of other hepatic enzymes were within reference ranges in the 7 cows that were examined . Hepatic biopsy specimens from 3 cows were normal . Specimens from 4 other cows had changes that ranged from minimal to mild, chronic, lymphoplasmacytic periportal hepatitis to acute, random, necrotizing hepatitis . Development of photosensitivity was related to ingestion of alfalfa silage . Acetone extracts of the alfalfa silage, but not of other feedstuffs, were found to inhibit growth of Candida albicans under ultraviolet light . Cows experimentally fed a diet composed exclusively of the alfalfa silage developed skin lesions after 6 days, but did not have detectable serum concentrations of phylloerythrin. J Bacteriol, 1996 Nov, 178(21), 6223 - 6 Isolation and composition of inositolphosphorylceramide-type sphingolipids of hyphal forms of Candida albicans; Wells GB et al.; Hyphal forms of the human pathogen Candida albicans have been found to contain substantial quantities of phosphosphingolipids . These lipids were fractionated into three classes by normal-phase high-performance liquid chromatography . The first class contained equimolar amounts of phosphorus, inositol, phytosphingosines, and fatty acids; their composition and chromatographic behavior suggest that these compounds are inositolphosphorylceramides . The second class contained equimolar amounts of phosphorus, mannosylinositol, phytosphingosines, and fatty acids; their composition and chromatographic behavior indicate that these compounds are mannosylinositolphosphorylceramides . The third class of compounds contained phosphorus, mannosylinositol, inositol, phytosphingosines, and fatty acids in a molar ratio of 2:1:1:1:1; their composition and chromatographic behavior indicate that these compounds are mannosyldiinositolphosphorylceramides . Molecular species in each class differ in the composition of long chain bases and fatty acids; the most abundant long chain bases were C18 and C20 phytosphingosines, and the most abundant fatty acids were hydroxy and nonhydroxy C24-26 . The array of sphingolipids in C . albicans is similar to that of Saccharomyces cerevisiae . Sphingolipids have been shown to be essential in S . cerevisiae, thus these lipids, which are not present in animals, offer a potentially unique target for antifungal chemotherapy against C . albicans. Infect Immun, 1996 Nov, 64(11), 4714 - 8 Gamma interferon protects endothelial cells from damage by Candida albicans by inhibiting endothelial cell phagocytosis; Fratti RA et al.; Once Candida albicans comes in contact with endothelial cells, it induces cellular injury . This endothelial cell injury may be a mechanism by which blood-borne organisms escape from the intravascular compartment and invade the tissue parenchyma during hematogenous infection . We have been investigating the ability of cytokines to modulate endothelial cell injury caused by C . albicans . Previously we reported that pretreatment of endothelial cells with gamma interferon (IFN-gamma) protects these cells from candidal injury in vitro . In the current study, we examined potential mechanisms of the cytoprotective effects of IFN-gamma . Time course experiments demonstrated that maximal reduction in candidal injury of endothelial cells occurred after the endothelial cells had been exposed to IFN-gamma for at least 72 h . In other studies, we determined that IFN-gamma reduced endothelial cell phagocytosis of C . albicans by 41.3% compared with that of untreated endothelial cells (P < 0.01) . Since endothelial cell phagocytosis of C . albicans is required for damage to occur, inhibition of phagocytosis is likely a mechanism by which IFN-gamma protects endothelial cells from candidal injury . We also found that the cytoprotective effect of IFN-gamma is not mediated by reducing access of the organisms to intracellular endothelial cell iron or by upregulating the synthesis of reactive oxygen intermediates (which could potentially reduce the ability of C . albicans to injure endothelial cells) . Thus, inhibiting endothelial cell phagocytosis of C . albicans may be a mechanism by which IFN-gamma augments the host defense against hematogenously disseminated candidal infections. Infect Immun, 1996 Nov, 64(11), 4561 - 6 Intravenous injection of Candida-derived mannan results in elevated tumor necrosis factor alpha levels in serum; Garner RE et al.; Intravenous injection of Candida albicans into mice produced elevated serum tumor necrosis factor alpha (TNF-alpha) levels . We hypothesized that immunostimulants released in vivo from C . albicans during fungal sepsis might contribute to the elevated levels of TNF-alpha in serum . We tested this hypothesis in mice with C . albicans mannan (CAM) . Increased serum TNF-alpha levels were observed following intravenous and intraperitoneal injections of CAM . Injection of CAM into mice resulted in increased serum TNF-alpha concentrations that reached 1,200 pg/ml of blood, compared with 2,400 microg/ml of blood following injection of 10 microg of endotoxin . The response to CAM was concentration dependent, requiring a minimum dose of 20 microg of CAM per g of body weight . Sera from mice were tested 30, 60, 90, and 120 min after intravenous injections with CAM . TNF-alpha concentrations were minimal 30 and 120 min after intravenous injection and maximal 60 and 90 min after CAM injection . The relative distribution of CAM in vivo in decreasing order was determined to be as follows: blood > liver > lung > spleen, 90 min following injection of a single 5-mg dose of CAM . CAM was confirmed as the stimulating substance by utilizing anti-CAM antibodies in vivo to block the response . Rabbit anti-mannan antibodies administered by intraperitoneal injection 24 h before CAM injection significantly suppressed (P < 0.05) the accumulation of TNF-alpha in the sera . Dexamethasone administered to mice before intravenous injection of mannan significantly reduced (40 to 90% reduction; P < 0.05) the concentrations of TNF-alpha in the sera of treated mice . Thus, when in vivo CAM clearance mechanisms are exceeded, sufficient CAM may become available to stimulate TNF-alpha production, making CAM an important part of pathogenesis in Candida sepsis. Infect Immun, 1996 Nov, 64(11), 4514 - 9 Evidence for degradation of gastrointestinal mucin by Candida albicans secretory aspartyl proteinase; Colina AR et al.; A zone of extracellular digestion of the mucin layer around Candida albicans blastoconidia was observed by transmission electron microscopy in the jejunum of mice inoculated intragastrically (G . T . Cole, K . R . Seshan, L . M . Pope, and R . J . Yancey, J . Med . Vet . Mycol . 26:173-185, 1988) . This observation prompted the hypothesis that a putative mucinolytic enzyme(s) may contribute to the virulence of C . albicans by facilitating penetration of the mucus barrier and subsequent adherence to and invasion of epithelial cells . Mucinolytic activity was observed as zones of clearing around colonies of C . albicans LAM-1 grown on agarose containing yeast nitrogen base, glucose, and hog gastric mucin . In addition, concentrated culture filtrate obtained after growth for 24 h in yeast nitrogen base, supplemented with glucose and mucin as the sole nitrogen source, contained proteolytic activity against biotin-labelled mucin which was inhibited by pepstatin A . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the culture filtrate revealed two components of 42 and 45 kDa, with pIs of 4.1 and 5.3, respectively . A zymogram showed that mucin was degraded only by the 42-kDa component, which was also recognized by immunoblotting with an anti-secretory aspartyl proteinase (anti-Sap) 2p monoclonal antibody . The N-terminal sequence of the first 20 amino acids matched that reported for Sap2p . These results demonstrate that Sap2p is responsible for proteolysis of mucin by C . albicans in vitro and may be involved as a virulence factor in the breakdown of mucus and penetration of the mucin barrier by C . albicans. Proc Natl Acad Sci U S A, 1996 Oct 29, 93(22), 12473 - 7 Molecular markers reveal that population structure of the human pathogen Candida albicans exhibits both clonality and recombination; Graser Y et al.; The life history of Candida albicans presents an enigma: this species is thought to be exclusively asexual, yet strains show extensive phenotypic variation . To address the population genetics of C . albicans, we developed a genetic typing method for codominant single-locus markers by screening randomly amplified DNA for single-strand conformation polymorphisms . DNA fragments amplified by arbitrary primers were initially screened for single-strand conformation polymorphisms and later sequenced using locus-specific primers . A total of 12 single base mutations and insertions were detected from six out of eight PCR fragments . Patterns of sequence-level polymorphism observed for individual strains detected considerable heterozygosity at the DNA sequence level, supporting the view that most C . albicans strains are diploid . Population genetic analyses of 52 natural isolates from Duke University Medical Center provide evidence for both clonality and recombination in C . albicans . Evidence for clonality is supported by the presence of several overrepresented genotypes, as well as by deviation of genotypic frequencies from random (Hardy-Weinberg) expectations . However, tests for nonrandom association of alleles across loci reveal less evidence for linkage disequilibrium than expected for strictly clonal populations . Although C . albicans populations are primarily clonal, evidence for recombination suggests that sexual reproduction or some other form of genetic exchange occurs in this species. Transplantation, 1996 Oct 27, 62(8), 1182 - 4 Candida albicans osteomyelitis in a liver transplant recipient: a case report and review of the literature; Jonnalagadda S et al.; A 51-year-old man developed fever and back pain 2 months after orthotopic liver transplantation for end-stage liver disease secondary to chronic hepatitis C infection . CT scan demonstrated destructive lesions in T12 suggestive of osteomyelitis . Aspiration biopsy of the vertebra revealed granulomatous inflammation and yeast forms; culture yielded Candida albicans . The patient improved with intravenous amphotericin B and 5-fluorocytosine and did not require surgical intervention . Candida osteomyelitis is a rare condition and to our knowledge it has not been reported before in liver transplant recipients . Awareness of this potential complication may shorten the delay in making the definitive diagnosis, which in turn may increase the likelihood of a response without sequela. J Med Chem, 1996 Oct 25, 39(22), 4471 - 7 Structure-activity relationships in a series of substituted indolocarbazoles: topoisomerase I and protein kinase C inhibition and antitumoral and antimicrobial properties; Pereira ER et al.; A series of compounds structurally related to staurosporine, rebeccamycin, and corresponding aglycones was synthesized, and their activities toward protein kinase C and topoisomerases I and II were tested together with their in vitro antitumor efficiency against murine B16 melanoma and P388 leukemia cells . Their antimicrobial activities were also examined against a Gram-negative bacterium (Escherichia coli), a yeast (Candida albicans), and three Gram-positive bacteria (Bacillus cereus, Streptomyces chartreusis, and Streptomyces griseus) . To avoid side effects expected with protein kinase C inhibitors, we introduced substitution on the maleimide nitrogen and/or a sugar moiety linked to one of the indole nitrogens to obtain specific inhibitors of topoisomerase I with minimal activities on protein kinase C . As expected, these structures were inefficient on topoisomerase II, and some of them exhibited a strong activity against topoisomerase I . Generally, dechlorinated compounds were found to be more active than chlorinated analogues against both purified topoisomerase I and protein kinase C . On the other hand, opposite results were obtained in the cell antiproliferative assays . These results suggest lack of cell membrane permeability in the absence of the chlorine residue or cleavage of carbon-chlorine bonds inside the cell. Gene, 1996 Oct 24, 177(1-2), 29 - 34 Candidacidal activity of human salivary histatin recombinant variants produced by site-directed mutagenesis; Driscoll J et al.; Histatin 5 (Hst5) is a 24-amino acid (aa) member of the Hst family that is found in human salivary secretions and exhibits candidacidal activity . Hst5 contains a 13-aa region that alone is capable of killing fungal pathogens and is referred to as the functional domain . To investigate the role of specific aa located within the functional domain, the pRSET bacterial expression system was used to produce recombinant Hst5 (re-Hst5) and several re-variants that were generated by site-directed mutagenesis . The vector pRSETC expresses genes of interest as fusion proteins attached to the carboxy end of an N-terminal His6 tag that binds to nickel (Ni2+) . The re-variants were generated using the polymerase chain reaction (PCR) and had Gly substituted for either the His, Glu or Lys/Arg within the functional domain . PCR products that encoded either the wild-type or variant forms of re-Hst5 were inserted into pRSETC and produced as fusion proteins which were affinity purified from cell lysates by Ni(2+)-Sepharose chromatography . Fusion proteins were digested with CNBr and re-Hsts were purified by reversed-phase high performance liquid chromatography (RP-HPLC) . Re-Hsts were tested in bioassays to measure the ability to kill both Candida albicans (C . albicans) blastoconidia and spheroplasts which were generated by removal of the cell wall . In both assays, re-Hst5 displayed dose-dependent candidacidal activity that was nearly identical to that of native Hst5 purified from human salivary secretions . Re-Hst5 variants with either Glu or Lys/Arg substitutions demonstrated significantly lower candidacidal activity in both assays, while the variant with His mutated showed essentially no activity at physiological concentrations . These results indicate that acidic and basic aa within the functional domain contribute to candidacidal activity and that the His are essential for candidacidal activity . Additionally, since C . albicans spheroplasts were also susceptible to Hsts, the cell wall is not an essential component in the Hst mechanism of candidacidal action. Biochim Biophys Acta, 1996 Oct 23, 1284(2), 181 - 90 Functional complementation between transmembrane loops of Saccharomyces cerevisiae and Candida albicans plasma membrane H(+)-ATPases; Mason AB et al.; Saccharomyces cerevisiae PMA1 sequences encoding a putative antifungal target site comprising transmembrane loops 1 + 2 and/or 3 + 4 were replaced with the homologous sequences from Candida albicans PMA1 by using PCR-mediated domain transfer . The chimeric pma1 mutants and an isogenic wild type S . cerevisiae strain had similar growth rates, growth yields, glucose-dependent proton pumping rates, acid-activated omeprazole sensitivities, salt tolerances and antifungal sensitivities . The yields and kinetic properties of H(+)-ATPases in plasma membranes of mutant and wild type strains were comparable . Single heterologous transmembrane loops caused deleterious phenotypes at low pH and elevated temperature . Inclusion of both heterologous transmembrane loops fully suppressed the temperature sensitivity caused by heterologous transmembrane loop 1 + 2, partially suppressed the pH sensitivity and gave Candida-like in vitro sensitivity to vanadate, suggesting that the loops operate as a domain . The fully functional chimeric H(+)-ATPase containing C . albicans transmembrane loops 1 + 2 and 3 + 4 demonstrates this domain's complementarity to the equivalent region of the S . cerevisiae enzyme and validates the wild type S . cerevisiae H(+)-ATPase as an antifungal screening target. Indian J Biochem Biophys, 1996 Oct, 33(5), 420 - 4 Changes in the cellular composition of Candida albicans resistant to miconazole; Sharma S et al.; Biochemical changes that accompany acquisition of miconazole resistance in a single step mutant of C . albicans 3153 were analyzed . Experiments show that resistance to this drug was associated with decrease in total lipids, phospholipids and sterol content . Fluorescence polarization studies with 1,6-diphenyl-1,3,5-hexatriene (DPH) showed decrease in polarization value (p) in resistant cells, thus indicating changes in membrane fluidity . Uptake of {3H} proline by intact cells revealed decrease in K(m) and Vmax of high affinity system (S1) of proline transport in cells resistant to miconazole . Results of this study suggest that membrane sensitivity of miconazole is determined by overall membrane organisation rather than by affinity of antifungal drug(s) for a single membrane component. J Can Dent Assoc, 1996 Oct, 62(10), 808 - 12 {Denture stomatitis: etiology and clinical considerations}; Girard B Jr et al.; Wearing removable dental prosthesis causes an alteration in the oral microflora . For certain individuals, this new environment is responsible for the development of a particular condition: prosthetic stomatitis . This article reviews the pertinent literature regarding the main predisposing factors causing the disease . It targets the different risk groups and identifies the proposed mechanism for the proliferation of Candida albicans on the palatal side of the prosthesis . Various treatments depending on the severity of the disease are also mentioned. J Chemother, 1996 Oct, 8(5), 351 - 7 In vitro inhibitory activity of citreoviridin against HIV-1 and an HIV-associated opportunist: Candida albicans; Vieta I et al.; Citreoviridin, a mycotoxin produced by some molds of the genera Penicillium and Aspergillus, inhibits the growth of bacteria of the Bacillus genus . Since significant information was not available on the effects of citreoviridin on eukaryotic cells and viruses, this molecule was tested on CD4+ T-lymphoid cell lines, on HIV-1 and on Candida albicans, which sometimes complicates HIV-infection . Antiviral activity was detected in H9 HTLV IIIB cells, a clone chronically infected by HIV-1 . Citreoviridin reduced p24 in the supernatant of H9 HTLV IIIB in a dose-dependent manner with a pharmacological selectivity index of 2.6 . In C . albicans, the effects of the mold-derivative were evaluated on some parameters associated with pathogenicity and virulence: cellular proliferation, germ tube production, expression of heat shock mannoproteins, release of proteases and phospholipases . At a 12.5 microM dose, citreoviridin showed a marked inhibitory effect on all parameters analyzed . As regards the mechanism of action, it is possible to hypothesize that the effects of citreoviridin may be due to a reduction of protein synthesis, since it inhibited the replication of HIV-1 at post-integrational stages and reduced the intracellular RNA and protein content in C . albicans. J Clin Pathol, 1996 Oct, 49(10), 861 - 3 Sensitive and universal method for microbial DNA extraction from blood products; Golbang N et al.; A new method of extracting bacterial and yeast DNA from blood products dependent on guanidinium thiocyanate acid extraction and proteinase K treatment is described . In spiked samples the sensitivity per 0.1 ml of serum and blood, respectively, was 26 and 150 colony forming units (cfu) for Escherichia coli, 80 and 120 cfu for Staphylococcus aureus and 20 and 26 cfu for Candida albicans . This compared well with existing methodologies, worked on limited clinical samples and was not pathogen specific. Immunology, 1996 Oct, 89(2), 295 - 300 The involvement of nitric oxide in the anti-Candida albicans activity of rat neutrophils; Fierro IM et al.; Rat peritoneal neutrophils (PMN) spontaneously release nitric oxide (NO) when incubated in vitro . Addition of the NO synthase inhibitor L-monomethylarginine (L-NMMA) to the PMN reduces NO production and impairs the killing of the yeast Candida albicans, both effects being reversed by L-arginine . These data strongly suggest that oxidative metabolism of L-arginine by PMN is involved in the candidacidal activity of these cells . Rat blood PMN, which do not produce significant amounts of NO, exhibit a reduced killing capacity compared with peritoneal cells, except when they are obtained from lipopolysaccharide (LPS)-treated rats . In this case they produce measurable amounts of nitrite and express high fungicidal activity in vitro . Confirming the candidacidal activity of NO, the exposure of the C . albicans cultures to different concentrations of NO donors leads to a reduction in their survival . The candidacidal activity related to the NO pathway in rat PMN is phagocytosis dependent, since the activity can be inhibited by cytochalasin B . However, the oxidative products of oxygen released by rat PMN do not seem to be involved in their candidacidal activity, as incubation of the cells with phorbol myristate acetate (PMA) increases release of superoxide anion but does not affect the pattern of killing . Our results suggest that NO could be an important candidacidal pathway in rat neutrophils. J Antimicrob Chemother, 1996 Oct, 38(4), 671 - 7 ZD0870 treatment of murine candidiasis caused by fluconazole resistant isolates of Candida albicans; Najvar LK et al.; Mice were infected intravenously with three fluconazole susceptible and ten fluconazole resistant isolates of Candida albicans then treated with escalating doses of 0.25, 0.5, 1, 2.5, 10 and 40 mg/kg/day of the new antifungal triazole, ZD0870, for 10 days . A minimum protective dose of < 0.25 mg/kg was determined for infections introduced by the three fluconazole susceptible C . albicans and one of the fluconazole resistant isolates whereas doses ranging from 2.5 to 10 mg/kg/day were required for infections induced by seven of the resistant isolates and > or = 40 mg/kg/day for the remainder . Thus, infections caused by fluconazole resistant C . albicans may be successfully treated with ZD0870, though higher doses than those used to treat infections due to susceptible yeast may be required. J Antimicrob Chemother, 1996 Oct, 38(4), 579 - 87 The effects of antifungal agents on the morphogenetic transformation by Candida albicans in vitro; Hawser S et al.; The effects of antifungal agents, with different mechanisms of action, on the morphogenetic transformation by synchronised yeast-phase Candida albicans cells in vitro and their respective anti-Candida activities are described . MIC data demonstrated that the azoles, amphotericin B and echinocandin were the most active agents against four C . albicans strains . Morphogenetic transformation experiments demonstrated that amphotericin B was significantly better at preventing the transformation, under a variety of test conditions, than the azoles and flucytosine: amphotericin B abolished the transformation at low concentrations while the azoles only prevented the morphogenetic transformation at much higher concentrations (> 100 x MIC). Infect Immun, 1996 Oct, 64(10), 4406 - 8 Ubiquitin-like epitopes associated with Candida albicans cell surface receptors; Sepulveda P et al.; We have recently reported the cloning of a Candida albicans polyubiquitin gene and the presence of ubiquitin in the cell wall of this fungus . The polyubiquitin cDNA clone was isolated because of its reactivity with antibodies generated against the candidal 37-kDa laminin-binding protein . In the present study, we have further investigated the relationship between ubiquitin and cell wall components displaying receptor-like activities, including the 37-kDa laminin receptor, the 58-kDa fibrinogen-binding mannoprotein, and the candidal C3d receptor . Two-dimensional electrophoretic analysis and immunoblot experiments with antibodies against ubiquitin and the individually purified receptor-like molecules confirmed that these cell surface components are ubiquitinated . In an enzyme-linked immunosorbent assay, polyclonal antisera to each receptor reacted with ubiquitin, thus demonstrating that the purified receptor preparations used as immunogens contained ubiquitin-like epitopes . It is proposed that ubiquitin may play a role in modulating the activity of these receptors and in the interaction of C . albicans cells with host structures. New Microbiol, 1996 Oct, 19(4), 335 - 43 Morphotypes of Candida albicans and their associations with underlying diseases and source of samples; Riviera L et al.; The morphotyping method was applied to differentiate a series of 276 Candida albicans strains recovered from hospitalized patients, by using a three-digit code based on the characteristics of the fringe outgrowth . By this scheme 32 different morphotypes were identified . An 86% reproducibility was achieved . The aim of this study was to investigate whether significant correlations exist between the morphotypes and: i) some underlying diseases of the patients, ii) the anatomical sources of the samples . The most striking associations were observed between fringeless strains and non-neoplastic diseases of the respiratory tract, and again between continuous filamentous fringe with parallel outgrowth and AIDS . With a significantly high frequency, samples from the genitourinary tract had a very coarse fringe texture. New Microbiol, 1996 Oct, 19(4), 327 - 34 Observation on the nucleic acids in the chlamydospores of Candida albicans; Vidotto V et al.; The presence of red (RNA) and green (DNA) fluorescent material identifying nucleic acids in the chlamydospores of Candida albicans has been well documented . Red fluorescence in chlamydospores is relatively diffused and ranges from small spots, observed in hyphal cells, to the entire protoplasmic content . Green fluorescence is rarely visible in these structures and, when present it can be observed next to the plasmalemma . The initial percentage values of the two curves related to the cell counts of red fluorescence of the suspensor cells and chlamydospores showed remarkable differences between the two structures . About 54% of the chlamydospores showed red fluorescence compared to about 28% of the suspensor cells . It seems from the results obtained in this study that much RNA was produced and/or accumulated in the chlamydospores and suspensor cells, rather than in mycelium where red fluorescence was not observed . The results obtained sustain the hypothesis that a chlamydospore should he considered a fully functional cell that is morphologically and physiologically active and independent from mycelium. AIDS, 1996 Oct, 10(12), 1369 - 76 Itraconazole cyclodextrin solution for fluconazole-refractory oropharyngeal candidiasis in AIDS: correlation of clinical response with in vitro susceptibility; Phillips P et al.; OBJECTIVE: To evaluate the efficacy of itraconazole cyclodextrin solution in fluconazole-refractory oropharyngeal candidiasis (OPC), and to correlate clinical outcome with in vitro susceptibility and serum azole levels . DESIGN: A prospective, open-label, intervention study . SETTING: A university hospital, which serves as the provincial HIV referral center . PATIENTS AND INTERVENTIONS: Thirty six HIV-infected individuals referred for fluconazole-refractory OPC were evaluated prospectively between May 1993 and March 1995, including clinical assessment, serum azole levels, and susceptibility testing of Candida spp, isolates . Itraconazole solution was administered orally at a daily dose of 200 mg for 14 days, followed by suppressive therapy . Thirty-four patients were evaluable . MAIN OUTCOME MEASURE: Resolution of oral pseudomembranous lesions . RESULTS: Initial isolates were Candida albicans (n = 33), C . glabrata (n = 1), C . krusei (n = 1), and mixed infection with C . albicans and C . krusei (n = 1) . Fluconazole serum levels obtained at the time of failed therapy ranged from 4.7 to 40 mg/l (median, 12.9 mg/l) . Itraconazole was generally well tolerated . Clinical responses were observed in 65% (22 out of 34) of evaluable cases . Among the responders, relapse had occurred within 2 months for four (36%) out of 11 cases who continued with follow-up . The median fluconazole minimal inhibitory concentration (MIC) was 64 mg/l for isolates from fluconazole-refractory cases, compared with a median of 0.5 mg/l for control isolates (P = 0.002) . The median itraconazole MIC for isolates from fluconazole-refractory cases was 1.25 mg/l, compared with a median of 0.078 mg/l for controls (P = 0.011) . CONCLUSION: A correlation between clinical response and in vitro susceptibility was clearly demonstrated for fluconazole, but not for itraconazole . Itraconazole cyclodextrin solution may be effective for fluconazole-refractory OPC and should be considered prior to salvage therapy with intravenous amphotericin B. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 1996 Oct, 82(4), 402 - 7 Clinical characteristics and management responses in 85 HIV-infected patients with oral candidiasis; Silverman S Jr et al.; Eighty-five consecutively seen HIV-positive persons with oral candidiasis were evaluated for clinical characteristics, staging of HIV disease, quantitation of candidal colony formation, and response to systemic antifungal treatment with Nizoral (ketoconazole) . Fifty-five had CD4 counts less than 200 . There was an inconsistent association between clinical signs, patient symptoms, CD4 counts, and candidal colony-forming units . However, there was a trend toward higher colony-forming unit counts (> 500) in patients with lower CD4 cells (< 200) . Sixty-five patients had a complete clinical response to the ketoconazole treatment (200 mg daily for 7 days), even though 81% of posttreatment cultures remained positive . Nonsmokers were more likely to respond to antifungal treatment when compared with smokers, and there was a slight tendency for complete responses when colony-forming unit counts were low . The most common lesion presentation was a combination of the white (pseudomembranous) and red (erythematous) forms . Forty-nine percent had complaints of pain . The variable responses indicated the importance of flexible dose-time and drug considerations in antifungal management . Candida albicans was the predominant species. Antimicrob Agents Chemother, 1996 Oct, 40(10), 2300 - 5 Susceptibilities of Candida albicans multidrug transporter mutants to various antifungal agents and other metabolic inhibitors; Sanglard D et al.; Some Candida albicans isolates from AIDS patients with oropharyngeal candidiasis are becoming resistant to the azole antifungal agent fluconazole after prolonged treatment with this compound . Most of the C . albicans isolates resistant to fluconazole fail to accumulate this antifungal agent, and this has been considered a cause of resistance . This phenomenon was shown to be linked to an increase in the amounts of mRNA of a C . albicans ABC (ATP-binding cassette) transporter gene called CDR1 and of a gene conferring benomyl resistance (BENr), the product of which belongs to the class of major facilitator multidrug efflux transporters (D . Sanglard, K . Kuchler, F . Ischer, J . L . Pagani, M . Monod, and J . Bille, Antimicrob . Agents Chemother . 39:2378-2386, 1995) . To analyze the roles of these multidrug transporters in the efflux of azole antifungal agents, we constructed C . albicans mutants with single and double deletion mutations of the corresponding genes . The mutants were tested for their susceptibilities to these antifungal agents . Our results indicated that the delta cdr1 C . albicans mutant was hypersusceptible to the azole derivatives fluconazole, itraconazole, and ketoconazole, thus showing that the ABC transporter Cdr1 can use these compounds as substrates . The delta cdr1 mutant was also hypersusceptible to other antifungal agents (terbinafine and amorolfine) and to different metabolic inhibitors (cycloheximide, brefeldin A, and fluphenazine) . The same mutant was slightly more susceptible than the wild type to nocodazole, cerulenin, and crystal violet but not to amphotericin B, nikkomycin Z, flucytosine, or pradimicin . In contrast, the delta ben mutant was rendered more susceptible only to the mutagen 4-nitroquinoline-N-oxide . However, this mutation increased the susceptibilities of the cells to cycloheximide and cerulenin when the mutation was constructed in a delta cdr1 background . The assay used in the present study could be implemented with new antifungal agents and is a powerful tool for assigning these substances as putative substrates of multidrug transporters. Microbiology, 1996 Oct, 142 ( Pt 10), 2741 - 6 Candida albicans adherence to a human oesophageal cell line; Enache E et al.; The oesophageal epithelium appears to be one of the primary cell targets of Candida albicans in AIDS patients . To study this interaction, we have established an in vitro adherence assay using a human epithelial oesophageal cell line (HET1-A) . When yeast cells were grown in 500 mM D-galactose, adherence increased significantly over cultures prepared in 500 mM D-glucose . In addition to HET1-A cells, adherence of the organism grown in D-galactose to human buccal epithelial cells and a murine alveolar macrophage cell line was also higher . Adherence of yeast cells to HET1-A cells was partially inhibited in the presence of D-glucosamine or N-acetyl-D-glucosamine, but not with D-mannose, D-glucose, L-fucose or D-galactose . Attachment to HET1-A cells was studied using scanning and transmission electron microscopy . Partial phagocytosis of adhering yeast cells was observed occasionally within the first 90 min following infection, as evidenced by the formation of HET1-A pseudopodia in instances of close contact with yeast cells . The influence of D-galactose on cell surface proteins was studied by analysing beta-mercaptoethanol-extracted proteins from yeast cells grown in either 500 mM D-galactose or D-glucose . From D-galactose-grown cells only, a glycoprotein of approximately 190 kDa was observed in Aurodye-stained SDS-PAGE gels and in Western blots using an immunoglobulin fraction (IgG) prepared from sera of rabbits infected with the organism . These studies demonstrate that C . albicans adheres to human oesophageal cells and may utilize cell surface proteins whose synthesis is nutritionally regulated. J Clin Microbiol, 1996 Oct, 34(10), 2408 - 10 Evaluation of the Microbial Identification System for identification of clinically isolated yeasts; Crist AE Jr et al.; The Microbial Identification System (MIS; Microbial ID, Inc., Newark, Del.) was evaluated for the identification of 550 clinically isolated yeasts . The organisms evaluated were fresh clinical isolates identified by methods routinely used in our laboratory (API 20C and conventional methods) and included Candida albicans (n = 294), C . glabrata (n = 145), C . tropicalis (n = 58), C . parapsilosis (n = 33), and other yeasts (n = 20) . In preparation for fatty acid analysis, yeasts were inoculated onto Sabouraud dextrose agar and incubated at 28 degrees C for 24 h . Yeasts were harvested, saponified, derivatized, and extracted, and fatty acid analysis was performed according to the manufacturer's instructions . Fatty acid profiles were analyzed, and computer identifications were made with the Yeast Clinical Library (database version 3.8) . Of the 550 isolates tested, 374 (68.0%) were correctly identified to the species level, with 87 (15.8%) being incorrectly identified and 89 (16.2%) giving no identification . Repeat testing of isolates giving no identification resulted in an additional 18 isolates being correctly identified . This gave the MIS an overall identification rate of 71.3% . The most frequently misidentified yeast was C . glabrata, which was identified as Saccharomyces cerevisiae 32.4% of the time . On the basis of these results, the MIS, with its current database, does not appear suitable for the routine identification of clinically important yeasts. J Am Acad Dermatol, 1996 Oct, 35(4), 539 - 42 A U.S . epidemiologic survey of superficial fungal diseases; Kemna ME et al.; BACKGROUND: Large-scale studies performed outside the United States have demonstrated that most cases of onychomycosis and tinea pedis are caused by dermatophytes, primarily Trichophyton rubrum . However, other studies have suggested that yeasts and nondermatophytic molds may play a role, particularly in onychomycosis . OBJECTIVE: This study was undertaken to determine the epidemiology of superficial fungal infections in a U.S . population . METHODS: Fungal cultures were performed on patients with clinically suspected tinea cruris, tinea corporis, tinea capitis, tinea pedis, and onychomycosis . RESULTS: Dermatophytes were the most commonly isolated fungi in each type of superficial fungal disease studied . T . rubrum was the most commonly isolated dermatophyte species, although Trichophyton tonsurans was more common in tinea capitis and equally common in tinea corporis/tinea cruris . In tinea pedis and onychomycosis, dermatophytes appeared in approximately 95% and 82% of isolates, respectively . Candida albicans and nondermatophyte molds played only a minor role in onychomycosis; C . albicans was isolated in 7% of nail cultures and nondermatophytic molds were isolated in 11% . CONCLUSION: These results are in general agreement with other major epidemiologic studies performed outside the United States . Dermatophyte fungi cause most superficial fungal infections. J Infect Dis, 1996 Oct, 174(4), 888 - 90 Role of extended culture time on synergy of fluconazole and human monocyte-derived macrophages in clearing Candida albicans; Gujral S et al.; Possible enhanced candidacidal synergy of fluconazole and human monocyte-derived macrophages by extended culture time was studied . Synergistic candidacidal activity increased from 1 day to 3 days and was maximal at 6 days . Enhanced synergistic candidacidal activity at 144 h was observed over a 10-fold challenge dose range . Enhanced synergy with culture time was seen with fluconazole-susceptible and -resistant isolates of Candida albicans . These findings emphasize the importance of dosage and an adequate period of therapy. J Infect Dis, 1996 Oct, 174(4), 821 - 7 Detection and significance of fluconazole resistance in oropharyngeal candidiasis in human immunodeficiency virus-infected patients; Revankar SG et al.; The epidemiology and clinical significance of fluconazole resistance were assessed in a cohort of advanced human immunodeficiency virus (HIV)-infected patients with recurrent oropharyngeal candidiasis . Fifty patients were prospectively evaluated using a novel method of detecting fluconazole resistance with chromogenic media containing fluconazole; results were confirmed with macrobroth testing . Resistant yeasts, defined as MICs > or = 8 micrograms/mL, were detected in 16 (32%) of 50 patients: 7 (14%) had resistant Candida albicans, 7 (14%) had resistant non-C . albicans yeast, and 2 (4%) had mixed resistant yeasts . MICs were > or = 32 in 11 of 16 isolates . Previous fluconazole use and severe immunosuppression were risk factors for resistance . However, 5 of 26 patients had resistant isolates with no prior fluconazole use, and all were severely immunosuppressed . Despite the high prevalence of resistance, 48 patients clinically responded to fluconazole . Fluconazole-resistant C . albicans and non-C . albicans yeast infections are common in patients with advanced immunodeficiency, but clinical efficacy of fluconazole remains high. Obstet Gynecol, 1996 Oct, 88(4 Pt 2), 704 - 6 Recurrent vulvovaginal candidiasis associated with long-term tamoxifen treatment in postmenopausal women; Sobel JD et al.; BACKGROUND: Symptomatic vulvovaginal candidiasis is rare in postmenopausal subjects because of the estrogen-dependence of this infection . Tamoxifen, a breast-cancer cell estrogen-antagonist, has not previously been reported to predispose to vulvovaginal candidiasis . CASES: Three postmenopausal women, age range 60-81 years (mean 71), were identified with recurrent vulvovaginal candidiasis . In all three cases, new onset of recurrent vulvovaginal candidiasis followed daily tamoxifen therapy . The duration of prior tamoxifen therapy was 1-7 years (mean 3.5) . One patient had diabetes mellitus, an additional risk factor for vulvovaginal candidiasis . In all three patients, Candida glabrata was identified as the causal pathogen, although in two patients symptomatic episodes caused by Candida albicans also occurred . In all cases, diagnosis was easily established using conventional investigations, and eradication of vulvovaginal candidiasis was possible without cessation of tamoxifen . CONCLUSION: Long-term tamoxifen treatment may be complicated by recurrent vulvovaginal candidiasis in postmenopausal women. J Bacteriol, 1996 Oct, 178(19), 5850 - 2 The mitogen-activated protein kinase homolog HOG1 gene controls glycerol accumulation in the pathogenic fungus Candida albicans; San Jose C et al.; The Candida albicans HOG1 gene (HOG1CA) was cloned by functional complementation of the osmosensitive phenotype associated with Saccharomyces cerevisiae hog1 delta mutants . HOG1CA codes for a 377-amino-acid protein, 78% identical to S . cerevisiae Hog1p . A C . albicans hog1 null mutant was found to be sensitive to osmotic stress and failed to accumulate glycerol on high-osmolarity media. Arch Microbiol, 1996 Oct, 166(4), 260 - 8 Differential profiles of soluble proteins during the initiation of morphogenesis in Candida albicans; Niimi M et al.; Candida albicans is a dimorphic fungus that can grow either as yeast or as mycelia . The mycelial form may be required for tissue penetration and therefore may have a role in pathogenesis . The protein profiles of the cell-free S100 fraction from budding yeast cells and germ tube-forming cells (an early stage of the transition between yeast and mycelia) were evaluated using two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) . Yeast growth or germ tube formation was induced in carbon-starved cells at 37 degrees C by either glucose, galactose or N-acetylglucosamine at pH 4.5 or pH 6.7 . More than 400 constitutively synthesised polypeptides were identified on 2-D PAGE by silver staining . A few polypeptides which seem to reflect the release from carbon starvation were detected, but no polypeptides unique to either morphology were observed . Fractionation of S100 preparations by polyethylenimine or heparin-agarose affinity chromatography, which have been used to detect DNA-binding proteins, revealed several proteins that were synthesised on the resumption of cell growth or in response to pH difference . Heparin-agarose also bound novel polypeptides in the size range 130-200 kDa that were preferentially synthesised in germ tube-forming cells . These results suggest that any protein factors that might exert a regulatory role early in germ tube formation are of low abundance, and that a minor group of soluble proteins involved in C . albicans morphogenesis may be differentially synthesised. Br Dent J, 1996 Sep 21, 181(6), 209 - 11 Nystatin pastilles and suspension in the treatment of oral candidosis; Millns B et al.; Clinical audit revealed that the treatment of oral candidosis was more successful with nystatin pastilles than with nystatin suspension . The purpose of this investigation was to determine the reasons for this observation . The concentration of nystatin needed to kill 49 consecutive clinical isolates of Candida albicans was measured . The isolates varied in cidal concentrations from 1.875 to 30 U/ml . The time taken to kill these isolates at their cidal concentrations was found to vary from 120 to 300 min . Volunteer studies showed that antifungal activity in the oral cavity was eliminated rapidly after the use of nystatin suspension . In contrast, the polyene could be detected for at least 5 hours after use of the nystatin pastille . The nystatin pastille can be expected to be more effective at killing Candida albicans than the suspension due to its persistent effects. Fortschr Med, 1996 Sep 20, 114(26), 319 - 21 {Pathogenicity of fungi in the intestines--current status of the discussion}; Scheurlen M; The hypothesis that colonization of the intestinal tract by yeasts (e.g . Candida albicans) can lead to disease in immunocompromised individuals is currently being discussed controversially . Proponents assume that toxins produced by the fungi can trigger such complaints as irritable bowel syndrome of the chronic fatigue syndrome, and that such chronic or recurrent infections may be caused by an intestinal reservoir of yeasts . Opponents of the hypothesis, however, point out that no hard data on the pathogenetic significance of an intestinal reservoir of yeasts are available, controlled studies have failed to demonstrate the effectiveness of antifungal treatment . Discussions are however, hampered by a lack of objective data . The postulated pathomechanisms therefore need to be clarified, diagnostic criteria developed, and the efficacy of the proposed therapeutic measures shown by controlled studies . Until this has been done, assumption about the pathogenicity of yeasts in the bowel, cannot be taken as a basis for binding therapeutic recommendations. EMBO J, 1996 Sep 16, 15(18), 5060 - 8 Transfer RNA structural change is a key element in the reassignment of the CUG codon in Candida albicans; Santos MA et al.; The human pathogenic yeast Candida albicans and a number of other Candida species translate the standard leucine CUG codon as serine . This is the latest addition to an increasing number of alterations to the standard genetic code which invalidate the theory that the code is frozen and universal . The unexpected finding that some organisms evolved alternative genetic codes raises two important questions: how have these alternative codes evolved and what evolutionary advantages could they create to allow for their selection? To address these questions in the context of serine CUG translation in C.albicans, we have searched for unique structural features in seryl-tRNA(CAG), which translates the leucine CUG codon as serine, and attempted to reconstruct the early stages of this genetic code switch in the closely related yeast species Saccharomyces cerevisiae . We show that a purine at position 33 (G33) in the C.albicans Ser-tRNA(CAG) anticodon loop, which replaces a conserved pyrimidine found in all other tRNAs, is a key structural element in the reassignment of the CUG codon from leucine to serine in that it decreases the decoding efficiency of the tRNA, thereby allowing cells to survive low level serine CUG translation . Expression of this tRNA in S.cerevisiae induces the stress response which allows cells to acquire thermotolerance . We argue that acquisition of thermotolerance may represent a positive selection for this genetic code change by allowing yeasts to adapt to sudden changes in environmental conditions and therefore colonize new ecological niches. Biochim Biophys Acta, 1996 Sep 13, 1297(1), 1 - 8 D-arabinose dehydrogenase and biosynthesis of erythroascorbic acid in Candida albicans; Kim ST et al.; D-Arabinose dehydrogenase was purified 2750-fold from the cytosolic fraction of Candida albicans to apparent homogeneity, with an overall yield of 3%, by a purification procedure consisting of ammonium sulfate precipitation and DEAE-Sepharose A-50, Sephacryl S-200, Cibacron blue and phenyl-Sepharose CL-4B chromatographies . Gel-filtration chromatography gave an apparent molecular mass of 41 kDa and SDS-PAGE showed only one protein band corresponding to a molecular mass of 42 kDa, indicating that the enzyme is a single polypeptide . The enzyme was optimally active at pH 8.0 and the pI value of the enzyme was 5.0 . The enzyme was relatively stable from pH 4.5 to 7.5 . The optimal temperature for the enzyme activity was 30 degrees C . The activity of the enzyme was inhibited by Hg2+, Fe2+, Zn2+, Cu2+, Mg2+, Mn2+, N-ethylmaleimide and p-chloromercuribenzoic acid . The enzyme catalysed the oxidation of D-arabinose, L-fucose, L-xylose and L-galactose, which have the same configurations of hydroxyl groups at C2- and C3-positions, with apparent K(m) values of 29.2, 28.9, 37.1 and 91.3 mM at pH 8.0, respectively, with 50 microM NADP+ . The enzyme used NADP+ as a coenzyme . Apparent K(m) value at 60 mM D-arabinose for NADP+ was 44.6 microM . NADPH inhibited the enzyme activity competitively with respect to NADP+ (Ki = 78.6 microM) . The amino-terminal sequence of the enzyme was Met-Lys-Leu-Ala-Thr-Glu-Ile-Asp-Phe-X-Leu-Asn-Asn-Gly- . The reaction product was D-arabinono-1,4-lactone, judged from gas-liquid chromatography/mass spectrometry . In C . albicans, D-erythroascorbic acid was formed from D-arabinose by D-arabinose dehydrogenase and D-arabinono-1,4-lactone oxidase. Mycoses, 1996 Sep-Oct, 39(9-10), 353 - 6 Increased prevalence of oral Candida albicans serotype B in homosexual men: a comparative and longitudinal study in HIV-infected and HIV-negative patients; Torssander J et al.; Several investigators have shown a comparatively high prevalence of Candida albicans serotype B among HIV-infected individuals . We serotyped oral C . albicans strains from 50 HIV-infected homosexual men, 39 HIV-seronegative homosexual men and 40 clinical oral isolates of a control group . The prevalence of serotype B was significantly higher in homosexual men, regardless of HIV serostatus, than in the control subjects . We suggest that the reported high prevalence of serotype B among AIDS patients in Europe and the USA simply reflects the high proportion of homosexual men among HIV-infected patients . In 22 subjects, oral C . albicans isolates were obtained at two or more time points, up to 8 years apart . No change in serotype was observed over time . The serotype prevalences in HIV-infected patients with oral thrush or AIDS-defining illness were similar to the group of homosexual men as a whole, indicating that there is no serotype-related variation in pathogenicity. Mycoses, 1996 Sep-Oct, 39(9-10), 347 - 51 Fluorometric determination of acid proteinase activity in vulvovaginal candidosis; Kilic N et al.; Vulvovaginal candidosis is one of the most frequent disorders in obstetrics and gynaecology . Candida albicans is commonly considered to be the true vaginopathic agent . The secreted acid proteinase might be especially relevant in the pathogenesis of vulvovaginal candidosis . A fluorometric determination of acid proteinase activity of clinical C . albicans isolates was carried out during the present work using fluorescamine . L-Leucyl-L-alanine was included as an internal standard and the results were expressed as nmoles of leucylalanine equivalents h-1 per 2 x 10(4) cells . The 13 isolates were taken from non-diabetic, non-pregnant women aged 22-35 years with vulvovaginal candidosis . Candida albicans ATCC 44858 was used as a control . The proteinase activity in culture supernatants was detectable starting from the mid- to late- exponential phase of growth, peaked between 30 and 46 h, and then declined . The control had an activity of 2.72 nmol h-1 per 2 x 10(4) cells, whereas eight of the samples had a lower activity (1.05 nmol h-1 per 2 x 10(4) cells on average) and five of the samples had a higher activity (6.53 nmol h-1 per 2 x 10(4) cells on average) . The fluorometric determination of acid proteinase activity was found to be more reproducible and sensitive than the previously used spectrophotometric determinations. Mycoses, 1996 Sep-Oct, 39(9-10), 341 - 6 Detection of Candida albicans DNA with a yeast-specific primer system by polymerase chain reaction; Wildfeuer A et al.; The in vitro and in vivo selectivity and sensitivity of a yeast-specific primer system was investigated . A two-step polymerase chain reaction (PCR) was used: the first amplified a 245-bp fragment of the gene for cytochrome P450L1A1 and the second a product of 193 bp . This nested PCR produced an approximately 1000-fold increase in the sensitivity of the test for Candida albicans DNA compared with the first primer pair . The lower level of sensitivity of the test in physiological saline and tissue homogenate was about 10 C . albicans cells ml-1 . On the other hand, the sensitivity of the nested PCR method was reduced by a factor of more than 1000 when C . albicans was fixed with 4% formalin . After i.v . injection of different doses of C . albicans into mice, the yeast could be demonstrated in blood and in six different organs . The nested PCR was to some extent more sensitive than culturing for the detection of the yeast in the specimens of organs such as lung, cardiac muscle, liver, kidneys and brain . In contrast, in blood and spleen the culture was superior to the PCR technique used . Nested PCR is thus a useful additional method for the demonstration of yeasts. Mycoses, 1996 Sep-Oct, 39(9-10), 329 - 39 Clinical use of oral nystatin in the prevention of systemic candidosis in patients at particular risk; Schafer-Korting M et al.; Systemic candidosis is currently a major concern among certain groups of patients at particular risk because of recent treatment modalities . To prevent spread of Candida albicans, in particular, from the orogastrointestinal tract antimycotic treatment would appear beneficial . So far, however, suitable drugs are rare . Polyenes, and in particular oral nystatin, are the main ones considered so far . More recently, the oral azoles have provided therapeutic alternatives . In this review the current role of nystatin and, in particular nystatin tablets, which are better accepted than suspensions at higher dose levels, is described, focusing on efficacy and safety as determined in controlled trials . Recent evidence suggests that oral application of nystatin tablets can be considered both efficacious and safe in the appropriate context . The relative potency of oral nystatin and systemic azoles, particularly ketoconazole and fluconazole, awaits final determination. Minerva Med, 1996 Sep, 87(9), 433 - 5 {Symptomatic efficacy of clotrimazole combined with surgery in the treatment of recurring anal and genital condylomatosis in seropositive and AIDS patients}; Erenbourg L et al.; During their daily surgical activity in Hiv-positive and AIDS patients the authors have seen in surgical treatment of anogenital recurring acuminate condylomata (treated with diatermocoagulation) in 1995 that prescribing elotrimazole ointment in Candida albicans infection of the same regions there was and amelioration or the healing of the mycotic infection but also a net lowering of the condylomata and persistent latency (disease-free time) till to the next local recurrence . So they have hypothesized and antiviral and antiblastic effect of elotrimazole . Under this heading they have done a literature research discovering that in 1995, only some months ago, two fundamental works have been published on this intriguing topic . They demonstrate at an experimental level in mice a strong antiviral and antiblastic activity of clotrimazole . This new fascinating field is completely open to future research and the consequences at a therapeutic level will be of utmost importance. APMIS, 1996 Sep, 104(9), 623 - 8 Biotypes of oral Candida albicans isolates in a Tanzanian child population; Matee MI et al.; Although biotypes of Candida albicans from adult populations, especially in the West, have been described, there are no data either from a child population, or from the African continent . Hence a total of 200 oral C . albicans isolates from Tanzanian children aged 6-24 months were biotyped using two commercially available API micromethod kit systems and a boric acid resistance test . The predominant biotypes, which comprised two thirds of the organisms isolated, were J1S (19.5%), A1S (16.0%), J1R (14.5%), A1R (9.5%) and P1R (7.5%) . In total, 16 new biotypes comprising 44 (22%) isolates which have not hitherto been described were found in this Tanzanian population and, of these, the P1R biotype predominated with 15 (7.5%) isolates . There was no significant association between predominant biotypes (with clusters > or = 15 isolates) and age, gender, breast feeding and malnutrition . These data indicate that the biotype profile of C . albicans isolates may differ in paediatric and adult populations, and/or global distribution of various subtypes of this common opportunistic pathogen. Intern Med, 1996 Sep, 35(9), 707 - 11 Changes in clinical features of fungemia in a Japanese University Hospital over a 12-year period; Mizushima Y et al.; Forty-five patients with fungemia during 1982-1993 (periods I = 1982, II = 1986-1989, III = 1990-1993) in a Japanese university hospital were reviewed to follow changes in the clinical features of fungemia . The percentage of fungi among microorganisms isolated from blood cultures was almost constant (6-10%) throughout the study period . Fungemia was highly associated with use of intravascular catheters, and some changes in clinical features were observed: 1) Candida albicans, C . parapsilosis and C . glabrata were the main isolates, and the number of fungal species showed a tendency to increase . 2) The percentage of patients over 65 years old increased from 36 to 50% . 3) The percentage of patients who were treated with anti-fungal agents and/or removal of catheter increased from 50 to 89 and to 86% . 4) The percentage of patients who died within 28 days after isolation of fungus decreased from 64 to 27% . The improved prognosis was thought to be due to the development of new anti-fungal agents and faster removal of intravascular catheter when infection was suspected. J Med Vet Mycol, 1996 Sep-Oct, 34(5), 337 - 9 Role of antifungal drug resistance in the pathogenesis of recurrent vulvovaginal candidiasis; Lynch ME et al.; The aetiology of recurrent vulvovaginal candidiasis (RVVC) caused by Candida albicans remains unclear . To adequately address the role of antifungal resistance as a potential mechanism for RVVC, a longitudinal susceptibility analysis of 177 C . albicans isolates collected from 50 C . albicans RVVC patients over a period of 3 months to 7 years was performed . Antifungals tested included clotrimazole, ketoconazole, miconazole, itraconazole and fluconazole . Results showed that all vaginal isolates were uniformly susceptible to all drugs tested and that successive isolates from individual patients did not show increased resistance to any drug despite long-term exposure to azoles . These results suggest that episodes of RVVC caused by C . albicans are rarely of ever attributable to azole antifungal resistance. J Med Vet Mycol, 1996 Sep-Oct, 34(5), 315 - 22 Cloning and characterization of a cDNA coding for Candida albicans polyubiquitin; Sepulveda P et al.; Immunoscreening of a Candida albicans cDNA library in the expression vector lambda gt11 with rabbit polyclonal antibodies against the 37 kDa cell surface laminin receptor of C albicans resulted in the isolation of a cDNA clone of 0.9 kb . Sequencing of this clone demonstrated a full length open reading frame encoding the polyubiquitin, which contains three tandem copies, head-to-tail spacerless repeats, of the 228 nucleotides coding for the 76 amino acids of the ubiquitin protein, which is identical to that of Saccharomyces cerevisiae . The third copy possesses an extra C-terminal amino acid which is distinct to that found in S . cerevisiae . Northern blot analysis revealed a single mRNA population of about 1 kb present in similar amounts in both yeast and mycelial cells . This indicates that the C . albicans polyubiquitin gene (UBI1) encodes a polyubiquitin precursor protein containing three ubiquitin repeats . Immunofluorescence and Western immunoblotting experiments with polyclonal antibodies against mammalian ubiquitin suggest the presence of ubiquitinated protein moieties in the wall of C . albicans cells. Microbiologia, 1996 Sep, 12(3), 443 - 8 A Candida albicans gene expressed in Saccharomyces cerevisiae results in a distinct pattern of mRNA processing; Iborra A et al.; Two plasmids (derived from YCplac22 and YEplac112) carrying a Candida albicans gene (including the 5' non-coding promoter sequences) coding for a 30 kDa membrane-bound protein, were used to transform Saccharomyces cerevisiae cells . A 30 kDa protein was immunodetected by Western blot in the membrane fraction of transformants . Northern analysis showed the presence of three mRNA species (of about 1.1, 0.7 and 0.5 kb) hybridizing with the C . albicans gene as a probe . The same result was obtained using the 5' and 3' regions of the gene as probes, whereas only a 1.1 kb mRNA was found in C . albicans and none was detected in S . cerevisiae control transformants . Thus, heterologous expression of this gene in S . cerevisiae results in a distinct pattern of mRNA processing, either due to the location on plasmid vectors and/or to differences in the mRNA processing systems in the two microorganisms. Comp Immunol Microbiol Infect Dis, 1996 Sep, 19(4), 267 - 74 Effect of beta-endorphin on adherence, chemotaxis and phagocytosis of Candida Albicans by peritoneal macrophages; Ortega E et al.; There is growing evidence about the role of neuroendocrine hormones in the regulation of the immune system . In the present study we have examined effects on different stages of phagocytic function of peritoneal macrophages from BALB/c mice induced by beta-endorphin . Peritoneal macrophages were incubated (30 min at 37 degrees C) in vitro with 0.22, 0.5 or 2200 ng/ml of this hormone . Adherence capacity was evaluated by means of a substrate adherence technique, chemotaxis in Boyden chambers, and ingestion of Candida albicans on migration inhibitory factor (MIF) dishes . No changes in adherence capacity were found . Chemotaxis, however, increased, and concentration of beta-endorphin correlated directly with stimulation . Incubation of macrophages with 0.5 ng/ml of beta-endorphin also stimulated phagocytosis of Candida albicans . These results indicate that beta-endorphin acts on peritoneal marine macrophages, stimulating some stages of their phagocytic function. J Antimicrob Chemother, 1996 Sep, 38(3), 485 - 97 In-vitro and in-vivo evaluation of a new amphotericin B emulsion-based delivery system; Tabosa Do Egito ES et al.; The in-vitro and in-vivo toxicity and activity of a new emulsion-based delivery system for amphotericin B (AmB-E) and of deoxycholate-amphotericin B (Fungizone) were studied . In vitro, Candida albicans and human red blood cells (RBCs) were treated with either product and dose-response curves for various cellular effects (changes in potassium cell content, haemoglobin leakage from RBCs and colony-forming ability of fungal cells) were obtained . AmB-E was less toxic than Fungizone against human RBCs and equally active against C . albicans cells . In-vivo studies showed that the LD50 of AmB-E and Fungizone in noninfected OF1 mice were 7.24 and 3.46 mg/kg, respectively . The therapeutic efficacy of AmB-E was assessed in murine candidiasis . Firstly, the efficacy of equal doses (0.8 mg/kg) of AmB-E and Fungizone was evaluated in infected mice . Both formulations increased the survival time compared to the control and were equally effective in reducing the cfu counts in the kidney . In the same model of infection, the maximum tolerated doses (MTD) of Fungizone and AmB-E were determined in order to study the efficacies of Fungizone and AmB-E at their respective MTD . AmB-E significantly increased the number of long-term survivors compared with Fungizone (MTD:2 and 1 mg/kg, respectively) . Thus, AmB-E was more effective than Fungizone for treatment of systemic mycoses at the MTD. J Ethnopharmacol, 1996 Sep, 53(3), 143 - 7 Antimicrobial evaluation of some plants used in Mexican traditional medicine for the treatment of infectious diseases; Navarro V et al.; Twelve methanolic plant extracts from botanical species used in traditional medicine in Morelos, Mexico to cure infectious diseases have been subjected to a screening study to detect potential antimicrobial activity against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Candida albicans . The antimicrobial activity of the products was evaluated using colonies growing in solid medium, establishing the minimal concentration required to inhibit their in vitro growth (MIC) . The results showed that extracts from Eucalyptus globolus Labill, Punica granatum L., Artemisia mexicana Wild., and Bocconia arborea Watt . possess strong in vitro antimicrobial activity against the tested microorganisms. Eur J Obstet Gynecol Reprod Biol, 1996 Sep, 68(1-2), 209 - 12 Fetal Candida infection associated with an intrauterine contraceptive device; Marelli G et al.; Fetal Candida infection is rarely described but is often associated with a retained intrauterine contraceptive device (IUCD) . A case of abortion due to Candida infection in a patient wearing an IUCD is reportedPIP: Although fetal intra-amniotic infection with Candida albicans is a rare event, a retained IUD during pregnancy is a major risk factor . Reported in this paper is the case of a 25-year-old woman admitted at 15 weeks' gestation with uterine contractions and vaginal discharge . She reported a history of genital condylomata . Examination revealed premature rupture of the membranes and evidence of an abruptio placentae . An IUD was visible in the uterine cavity . Histologic examination of the placenta revealed growth of Candida pseudohyphae on the surface of the amniotic membranes and at the fetal side of the chorion . A heavy polymorphonuclear infiltrate with areas of necrosis and hemorrhage was noted . Vaginal cultures revealed Candida albicans infection . Most other cases of IUD-associated intra-amniotic candidiasis reported in the literature failed to note whether Candida was present in the vagina or cervix as well . Diagnostic amniocentesis is recommended in all pregnant women with preterm labor and a retained IUD; also indicated is prompt topical antifungal treatment to prevent the spread of Candida to the amniotic cavity . FEMS Immunol Med Microbiol, 1996 Sep, 15(2-3), 73 - 9 Use of polymorphic short and clustered coding-region microsatellites to distinguish strains of Candida albicans; Field D et al.; We describe the identification of polymorphic microsatellite loci in the pathogenic yeast, Candida albicans . A search for all coding-region microsatellites with more than four repeats that can be found in Candida sequences in GenBank was conducted . Nine such microsatellite sequences consisting of trinucleotide motifs were found . Three of these were perfect microsatellites while the remaining six sequences were found in one imperfect microsatellite and two compound microsatellites . Because of the close proximity of some of these repeats, all could be assayed with six PCR primer pairs . All of these microsatellite sequences were found in five nuclear genes, ZNF1, CCN1, CPH1, EFG1, and MNT2 . Except for a single (CTT)5 serine tract, all coded for polyglutamine tracts . Another locus with seven alleles, a region of the ERK1 protein kinase gene, was also examined, and may be a representative of a new class of highly polymorphic "clustered' microsatellites . Such loci, in which several non-contiguous but closely linked microsatellites are clustered together, may be a useful source of DNA polymorphisms in microorganisms in which long microsatellite sequences are unavailable . All seven regions amplified were polymorphic, having between two and seven variable length alleles in the 11 strains of Candida albicans examined . The results of this and similar searches will facilitate epidemiological and evolutionary studies of Candida and other microorganisms. Bone Marrow Transplant, 1996 Sep, 18(3), 533 - 40 Bone marrow transplantation for the treatment of haematological disorders in Down's syndrome: toxicity and outcome; Rubin CM et al.; We report 18 patients with Down's syndrome who underwent bone marrow transplantation, and review nine previously published patients . The indications for transplant in the combined group of 27 patients were acute lymphoblastic leukaemia in 14 cases (52%), acute myeloid leukaemia in 11 cases (41%) and aplastic anemia in two cases (7%) . Transplants were autologous in five cases (19%) and allogeneic in 22 cases (81%); of the 22 allogeneic transplants, 16 donors were HLA-matched siblings . In all patients the conditioning regimen included total body irradiation of 7.5 Gy or more, and/or contained cyclophosphamide of 120 mg/kg or more . Seven patients (26%) had fatal pulmonary disease including pneumonitis and pulmonary hemorrhage . Five patients (19%) had significant airway problems including three with severe mucositis who required intubation for airway protection, one with severe mucositis with partial airway obstruction that required observation in the intensive care unit but did not require intubation, and one with Candida albicans laryngitis with development of a glottic web . Nineteen patients (70%) survived beyond 100 days post-transplant . There was no clear association between 100-day survival and the use of any particular agent or regimen used for conditioning or graft-versus-host disease prophylaxis, and the majority of patients tolerated high-dose cyclophosphamide, high-dose cytosine arabinoside, high-dose busulfan, total body irradiation, cyclosporin A, and methotrexate . There appeared to be more early deaths in patients who received the combination of cyclophosphamide and total body irradiation, compared with those receiving the combination of busulfan and cyclophosphamide or those receiving the combination of cytosine arabinoside and total body irradiation . Also, the use of methotrexate was associated with a greater number of early deaths, compared with cyclosporin A . At 3 years, life table estimates of freedom from relapse, relapse-free survival and survival were 75%, 44% and 48%, respectively . The estimated cumulative risk of death due to a non-leukaemic cause at 3 years was 39% . The data show that Down syndrome patients can tolerate the commonly used transplant conditioning regimens with acceptable toxicity; however, there is a strong suggestion in the data that the rates of life-threatening and fatal toxicity are higher than would be expected to occur in patients without Down's syndrome . Patients with Down's syndrome may have a predisposition to fatal pulmonary complications and reversible airway problems during the immediate post-transplant period. Antimicrob Agents Chemother, 1996 Sep, 40(9), 2221 - 3 Effects of cyclophosphamide and ceftriaxone on gastrointestinal colonization of mice by Candida albicans; Samonis G et al.; Adult male Crl:CD1 (ICR) mice were fed chow containing Candida albicans to induce sustained gastrointestinal colonization by the yeast . Groups of mice were rendered neutropenic with cyclophosphamide and subsequently received ceftriaxone, while other groups received normal saline and served as controls . Stool cultures were obtained immediately before and at the end of treatment . The administration of cyclophosphamide substantially increased the C . albicans counts in the stools of mice . The addition of ceftriaxone to the cyclophosphamide regimen did not significantly increase the level of gastrointestinal colonization by C . albicans . There was no evidence of Candida dissemination to internal organs. Antimicrob Agents Chemother, 1996 Sep, 40(9), 1998 - 2003 Comparison of a spectrophotometric microdilution method with RPMI-2% glucose with the National Committee for Clinical Laboratory Standards reference macrodilution method M27-P for in vitro susceptibility testing of amphotericin B, flucytosine, and fluconazole against Candida albicans; Rodriguez-Tudela JL et al.; The National Committee for Clinical Laboratory Standards has proposed a reference broth macrodilution method for in vitro antifungal susceptibility testing of yeasts (the M27-P method) . This method is cumbersome and time-consuming and includes MIC endpoint determination by visual and subjective inspection of growth inhibition after 48 h of incubation . An alternative microdilution procedure was compared with the M27-P method for determination of the amphotericin B, flucytosine, and fluconazole susceptibilities of 8 American Type Culture Collection strains (6 of them were quality control or reference strains) and 50 clinical isolates of candida albicans . This microdilution method uses as culture medium RPMI 1640 supplemented with 18 g of glucose per liter (RPMI-2% glucose) . Preparation of drugs, basal medium, and inocula was done by following the recommendations of the National Committee for Clinical Laboratory Standards . The MIC endpoint was calculated objectively from the turbidimetric data read at 24 h . Increased growth of C . albicans in RPMI-2% glucose and its spectrophotometric reading allowed for the rapid (24 h) and objective calculation of MIC endpoints compared with previous microdilution methods with standard RPMI 1640 . Nevertheless, good agreement was shown between the M27-P method and this microdilution test . The MICs obtained for the quality control or reference strains by the microdilution method were in the ranges published for those strains . For clinical isolates, the percentages of agreement were 100% for amphotericin B and fluconazole and 98.1% for flucytosine . These data suggest that this microdilution method may serve as a less subjective and more rapid alternative to the M27-P method for antifungal susceptibility testing of yeasts. Arzneimittelforschung, 1996 Sep, 46(9), 934 - 6 In vitro synergistic activity of ketoconazole with valproic acid against Candida species; Krajewska-Kulak E et al.; The susceptibility of 148 strains of Candida albicans and 20 strains of Candida species from patients was tested against ketoconazole (CAS 65277-42-1, KTZ), valproic acid (CAS 99-66-1, VPA) and the combination of KTZ and VPA, using Sabouraud's and YNB (yeast nitrogen base) media . Minimal inhibitory concentrations (MIC) with regard of C . albicans determined on diagnostic plates which contained Sabouraud's medium gave values of 49.84 +/- 5.83 mg/l (KTZ) and 202.97 +/- 17.70 mg/l (VPA), and on plates which contained YNB agar, values of 8.06 +/- 0.99 mg/l (KTZ) and 122.57 +/- 12.08 mg/l (VPA) . The combination of KTZ and VPA in various ratios (1:1, 1:2, 2:1) was found to exert synergistic effects against C . albicans on Sabouraud's medium and the mean values of the combinations were: 4.29 +/- 0.37, 5.29 +/- 0.37, 5.02 +/- 0.38 mg/l . These results were significantly different (p < 0.001) compared with KTZ . The findings indicate that VPA, an antiepileptic drug, increases the antifungal activity of KTZ against C . albicans and Candida species in vitro. Eur J Pediatr, 1996 Sep, 155(9), 756 - 8 Tribological and mycological consequences of the use of a miconazole nitrate-containing paste for the prevention of diaper dermatitis: an open pilot study; Pierard-Franchimont C et al.; The diaper environment increases the coefficient of skin friction and compromises the function of the stratum corneum . In this study, the tribological and mycological benefit of the use of a miconazole nitrate-containing paste on diapered skin was evaluated . A total of 135 instrumental measurements of both erythema and stratum corneum alterations were made on alternate days for 3 weeks in 15 infants . Biometrological parameters were the chromacity a* of the skin and the squamometry index . Cultures testing for Candida albicans were also performed . In the days following the use of the paste, the chromacity a*, the squamometry index and the number of positive cultures of C . albicans were significantly reduced compared to the same evaluations made off treatment . Conclusion: Miconazole nitrate-containing paste reduces the tribological interference between cloth diapers and skin as well as providing the diapered skin with an improved microbial environment. J Clin Microbiol, 1996 Sep, 34(9), 2246 - 54 Frequency, intensity, species, and strains of oral Candida vary as a function of host age; Kleinegger CL et al.; While the age of the host has been suggested as a determining factor in yeast carriage, no studies in which the genetic relatedness of isolates has been assessed in combination with the frequency and intensity of carriage as a function of host age have been performed in a single geographical locale and over a short time window . Therefore, by using a simple plating protocol to determine the frequency and intensity of carriage, sugar assimilation patterns to determine species, and Southern blot hybridization with the DNA fingerprinting probe Ca3 combined with computer-assisted analysis to determine the genetic relatedness of strains of Candida albicans, yeast carriage was analyzed as a function of age . All test individuals lived in the Iowa City, Iowa, locale and, except for some of the 0.5- to 1.5-year-olds, were dentate . The results demonstrate that for this test population, the frequency, average intensity, predominant species, and genetic relatedness of C . albicans strains varied as a function of host age . In addition, comparison with oral commensal organisms from the Ann Arbor, Mich., locale confirms the geographical specificity of C . albicans strains and the existence of an Iowa City-enriched strain which is most prevalent in elderly individuals. J Clin Microbiol, 1996 Sep, 34(9), 2106 - 12 Candida albicans serotype analysis by flow cytometry; Mercure S et al.; Candida albicans strains can be assigned to either of two major serogroups, A or B . Antigenic surface determinants present only in serotype A strains allow such a distinction, which has epidemiologic relevance . Reports have established that the relative distributions of the two serotypes can vary depending on the geographic origin of the isolates . A prevalence of susceptibility to an antifungal agent, flucytosine, was also observed with isolates of serotype A . More recently, it was suggested that the occurrence of serotype B isolates in various clinical forms of candidiasis is increasing . However, this latest finding remains controversial since serotyping results vary widely from one laboratory to another because of the lack of standardized methodologies . Difficulty in interpretation of results, which may lead to erroneous serotype identification, is the major setback associated with current methods . For this study, we thus devised a procedure that relies on flow cytometry and that may eliminate ambiguities in serotype determination . The validation of results was achieved with two types of serotype A-specific antisera, Iatron Factor 6 antiserum and an anti-C . albicans antiserum adsorbed on serotype B yeast cells . Agreement between results obtained with these two reagents was 100% with a wide array of Candida strains . These results confirmed the potential of the flow cytometric procedure as a reliable and reproducible method to establish the serotypes of C . albicans strains . Furthermore, some applications of this procedure to the epidemiological study of this human pathogen are presented. Phytochemistry, 1996 Sep, 43(2), 513 - 20 Xanthones from Hypericum roeperanum; Rath G et al.; Four new xanthones have been isolated from the roots of Hypericum roeperanum . Their structures have been established by a combination of spectroscopic and chemical methods as 1,6-dihydroxy-5-methoxy-4',5'-dihydro-4',4',5'-trimethylfurano- (2',3':3,4)-xanthone (5-O-methyl-2-deprenylrheediaxanthone B),