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J E Cutler

Publications and source records attributed to J E Cutler.

At least 73 records · Page 4Linked to original sources
PubMed1988

Candida albicans- and Candida stellatoidea-specific DNA fragment.

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

Animals↗
PubMed1987

Methods for DNA extraction from Candida albicans.

Three different methods are described for the extraction of total genomic DNA from the dimorphic fungus Candida albicans. One method, which enables a large number of cultures to be processed simultaneously, involves pulverizing dried cells with glass beads and then allowing the disrupted cells to break apart, autolyse, by incubation in a solution which includes sorbitol and a nonionic detergent. DNA extraction by a second method with a French pressure cell can be utilized on cultures in any phase of growth, but is not practical for processing numerous samples. The third method, which involves induction of spheroplasts, is commonly used for DNA extraction from various yeasts but is not suited for processing many samples simultaneously. The DNA extracted with the three procedures is comparable in quality; in particular, it is of high molecular size (greater than 30 kbp) and reacts readily with DNA-modifying enzymes such as restriction endonucleases.

Candida albicans↗
PubMed1987

Cell surface and intracellular expression of two Candida albicans antigens during in vitro and in vivo growth.

Intracellular and cell surface expression of two antigenic determinants of Candida albicans was studied by reacting ultrathin sections of the fungus with monoclonal antibodies and locating the sites of antibody-antigen reactions by use of colloidal gold-coupled secondary antibody and electron microscopic techniques. Comparisons were made between fungal cells allowed to develop in vitro and cells (in vivo) recovered from infected mice. Differences between in vitro antigen expression and expression in vivo were particularly noted during early germination events. Antigen expression in vivo appears to be concentrated in deeper cell wall layers and beneath the plasmalemma especially on germ tubes. Cells which develop in vivo and in vitro have antigen associated with an outer flocculent layer and on the innermost cell wall layer of mother cells and hyphae. A middle electron transparent cell wall layer is usually devoid of antigen. Comparison of antigen expression during in vivo growth in early (intraperitoneal) infection versus late disseminated (kidney) disease suggests that both of the antigens are expressed in greater quantity on the surface of germ tubes than on mother cells in kidney tissue as compared to intraperitoneally-grown cells. Results also suggest that at least one of the antigens, which is a substituted mannan, can be found on a lamello-tubular intracellular structure.

Antibodies, Monoclonal↗
PubMed1987

Use of chloroflexus-specific antiserum to evaluate filamentous bacteria of a hot spring microbial mat.

Polyclonal antiserum prepared against Chloroflexus aurantiacus reacted with all Chloroflexus strains examined but not with other morphologically or physiologically similar bacteria. Only one of three filament types in a natural hot spring cyanobacterial mat reacted with this antiserum. Reacting filaments remained antigenically positive deep within the mat in material estimated to be several years old.

Journal Article↗
PubMed1986

Ultrastructural and biochemical studies of two dynamically expressed cell surface determinants on Candida albicans.

Variability in the expression of two different cell surface carbohydrate determinants was examined with two agglutinating immunoglobulin M monoclonal antibodies (H9 and C6) and immunoelectron microscopy during growth of three strains of Candida albicans. A single strain of Candida parapsilosis did not express either antigen at any time during growth. Antigens were detected on the surface of C. albicans by agglutination tests with either H9 or C6 over a 48-h growth period. The difference in specificities of the monoclonal antibodies was demonstrated by Ouchterlony double-diffusion tests with solubilized antigens and by variabilities in the reactivity of the agglutinins among yeast strains. The antigenic determinants were isolated by specific immunoprecipitation and protease digestion and characterized by methods including high-pressure liquid chromatography, gas-liquid chromatography, and mass spectroscopy with both chemical and electron ionization. These determinants both contain mannose and glucose. In the case of antigen H9, an additional carbohydrate was detected with gas chromatography and mass spectroscopy. The location of antigens on individual cells was determined by indirect labeling of the determinants, first reacting cells with H9 or C6 followed by goat anti-mouse antibody conjugated with 20-nm colloidal gold particles. Transmission electron microscopy was used to examine cells. The antigens that were reactive with the monoclonal antibodies were associated with a flocculent surface layer. Expression of this layer and expression of the antigens is a dynamic process which is growth phase and strain dependent. The antigens were not expressed on very young cells and disappeared from the cell surface of most C. albicans strains with age. The use of monoclonal antibody to cell surface determinants may allow characterization of cell surface antigens of C. albicans and be helpful in establishing receptors which mediate adherence.

Agglutination↗
PubMed1986

Variability in expression of cell surface antigens of Candida albicans during morphogenesis.

The location and expression of two different cell surface antigens on germinating and nongerminating Candida albicans cells was examined by using transmission electron microscopy after labeling with monoclonal antibodies (H9 or C6) and immunocolloidal gold. Immunodeterminant expression of the two carbohydrate antigens was followed from early germination events through 20 h of development. The determinant detected by H9 antibody, which was initially lost from the mother cell surface and preferentially expressed only on hyphae during the first 4 h of germination, reappeared on the mother cell by 20 h, whereas the antigen detected by antibody C6 was continually expressed on mother cells and germ tubes throughout germination.

Agglutinins↗
PubMed1985

Changes in surface topography of Candida albicans during morphogenesis.

Temporal studies of germ-tube forming yeast cells of Candida albicans by scanning and transmission electron microscopy indicate that extensive vacuolation and possibly also cell wall changes may cause walls of the parent yeast cell to collapse during specimen preparation. This collapse does not occur in cells which have been grown in conditions that suppress germ tube formation and have undergone the same preparative treatment.

Candida albicans↗
PubMed1984

A simple and inexpensive method for assessing in vitro candidacidal activity of leukocytes.

Candidacidal activity of mouse neutrophils and macrophages was determined directly in microtiter plates. After a suitable period of interaction between phagocytic cells and C. albicans in the wells, the mouse cells were lysed with distilled water and corn meal agar was added to each well. Following incubation at 37 degrees C, viability was assessed using an inverted microscope and counting the number of germ tubes or microcolonies which developed. This method does not use radioisotopes or vital stains and should be applicable to other genera of yeasts.

Animals↗
PubMed1984

Variability in expression of a cell surface determinant on Candida albicans as evidenced by an agglutinating monoclonal antibody.

The expression of a surface immunodeterminant of Candida albicans was investigated with an agglutinating immunoglobulin M monoclonal antibody. The 96 strains of C. albicans tested, of which 76% were recent clinical isolates, were capable of expressing the antigen. The antigen was also produced by strains of Candida tropicalis and Torulopsis glabrata, but not by other yeast species. Expression of the surface immunodeterminant in C. albicans varied as a function of growth as indicated by agglutinin reactions and indirect immunofluorescence tests. When yeast cells were tested with the agglutinin, three patterns of reactivity were observed. In general, cells in the early logarithmic phase were less reactive than cells in the mid-logarithmic phase. Antigen expression, as determined by agglutinin reactivity, was also influenced by the nutritional composition of the growth medium, and in general, cells from broth cultures were usually more reactive than cells grown on solid media. The antigen was solubilized from the cell surface of C. albicans by hot 1 M NaCl. These water-soluble extracts were capable of binding antibody, and a single precipitin band formed when soluble antigen was reacted with the monoclonal antibody in an Ouchterlony double-diffusion test. Whole cell preparations and hot NaCl extracts from yeast strains which did not agglutinate when mixed with the antibody also did not absorb the agglutinin from solution. These data indicate that the expression of surface antigens on C. albicans is a dynamic process which may be influenced by a number of environmental factors. The use of monoclonal antibodies may allow characterization of surface antigens presented to the host during candidiasis.

Absorption↗
PubMed1983

Isolation and purification of morphogenic autoregulatory substance produced by Candida albicans.

The human pathogenic, dimorphic fungus Candida albicans produces in vitro a factor designated morphogenic autoregulatory substance (MARS) which suppresses yeast to mycelial transition. MARS is best produced when a high concentration of yeast cells (2 X 10(8)/ml) in a chemically defined medium is incubated at 37 degrees C for 9-15 h. MARS was isolated to apparent homogeneity by the following successive steps: charcoal absorption; pyridine elution; ether extraction; high pressure liquid chromatography, and Sephadex LH-20 chromatography. A 469-fold increase in purity was obtained by these procedures. The use of purified MARS to study C. albicans yeast to mycelial transition may lead to a better understanding of morphogeneis of this important opportunistic pathogen.

Candida albicans↗
PubMed1983

Candida albicans-induced agglutinin and immunoglobulin E responses in mice.

Mice varied in their ability to make detectable antibody responses to cell surface determinants of Candida albicans depending upon the antigen preparation and the immunization schedule used. Immunoglobulin M (IgM) appeared to be the major class of antibody responsible for the C. albicans-agglutinating activity of the immune sera. Various inbred strains of mice injected with a ribosomal fraction from C. albicans produced a low titer (average, 4 to 8) of yeast cell agglutinins and a higher titer (64 to 512) of IgE antibodies detected by passive cutaneous anaphylaxis (PCA) in rats. The two kinds of antibodies appeared to be specific for different antigens because the agglutinin, but not IgE, could be removed by absorbing the serum with a polysaccharide from the cell wall of C. albicans, but the polysaccharide did not provoke the PCA reaction. C. albicans-specific IgE antibodies showed cross-reactivity (PCA) with ribosomal antigens from a strain of C. albicans and C. tropicalis, but PCA reactions could not be elicited with similar antigen preparations from other yeast species. IgE responses were also detected in over 20% of the mice infected intravenously or intraperitoneally with live C. albicans.

Agglutinins↗
PubMed1982

Enhanced antibody responses induced by Candida albicans in mice.

Candida albicans may immunopotentiate antibody responses in mice to antigens unrelated to the fungus. This effect occurred best with cell-associated, rather than soluble, antigens. When dead yeasts, cell walls, or a water-soluble candidal polysaccharide were used, immunopotentiation was most dramatic when the antigen and fungal materials were given concomitantly via an intraperitoneal injection. However, mice infected with viable yeasts several days before antigen administration also developed heightened responses to the antigen. The mechanism of the C. albicans-induced adjuvanticity was not defined, but the effect seemed to correlate with induction of inflammation. The presence of C. albicans or other inflammatory agents in the peritoneal cavity caused a more rapid uptake of particulate antigen by the liver. The relationship between this event and immunopotentiation is not known. These studies demonstrate that C. albicans may have profound effects on host immune responses. Because immunological aberrations are commonly found in patients with candidiasis it may be important to determine whether some of these aberrations result from, rather than precede candidiasis.

Adjuvants, Immunologic↗
PubMed1981

Analysis of an in vivo model to study the interaction of host factors with Candida albicans.

Conditions were established under which membrane diffusion chambers surgically implanted into mice could be used to study interactions between host defense factors and Candida albicans within the chambers. Depending on the size of membrane pores, soluble host substances and phagocytic cells entered the chambers during the first 24 h after chamber implantation. By 7 days in vivo, the membranes of chambers appeared impermeable to these host factors. Mouse phagocytic cells were found to be functional within the in vivo chambers whether the cells emigrated to the chambers on their own accord or were placed there before chamber implantation. Opsonic factors such as antibody and complement appeared to be biologically functional within the in vivo chambers. Conditions suitable for harvesting C. albicans from the chambers also were determined.

Animals↗
PubMed1981

Effect of mouse phagocytes on Candida albicans in in vivo chambers.

Mouse phagocytic cells in in vivo diffusion chambers had either candidacidal or candidastatic activity depending on the kind of phagocyte studied, the type of mouse whence the phagocytes came, and whether the chamber inoculum of Candida albicans consisted primarily of yeasts or hyphae. Killing of C. albicans occurred when yeasts were placed into chambers with membranes with 3.0-micrometer pores and implanted intraperitoneally into normal mice or thymus-deficient (nude) mice. Although C. albicans remains in chambers with 3.0-micrometer pores, host phagocytic cells can migrate into the chambers. Killing also occurred when yeasts were combined with normal or nude mouse neutrophils in chambers made with membranes with 0.45-micrometer pores, which restrict migration of host cells, but not diffusion of soluble factors. Populations of cells rich in macrophages were candidastatic for yeasts when the phagocytes came from normal mice but candidacidal when obtained from nude mice. Results of gradient fractionations of peritoneal exudates indicated that more than one cell type may be responsible for candidacidal activity by nude mouse macrophage-rich cells. Hyphal-phase cells of C. albicans appear to be more resistant than yeast-phase cells to killing by normal and nude mouse phagocytic cells.

Animals↗
PubMed1979

Autoregulation of germ tube formation by Candida albicans.

Germ tube formation by Candida albicans is at least partially controlled by a product(s) of the yeast phase of the organism which is released from cells upon incubation at 37 degrees C in tissue culture medium or fetal calf serum. This germination regulatory substance is stable under conditions of lyophilization and heating of 70 degrees C, but becomes inactivated at pH values of 4.0 and 9.5. A germination regulatory substance was produced by both strains of C. albicans tested and by a strain of C. tropicalis. Production does not appear to be a universal characteristic of yeasts because the factor could not be recovered from either Cryptococcus laurentii or Candida parapsilosis. Previously described C. albicans germination inhibitors such as cysteine, tryptophol, and phenylethyl alcohol appear not to be the substance described here. Because of the ability of the factor to influence C. albicans morphology, we have designated it morphogenic autoregulatory substance.

Candida albicans↗
PubMed1979

Partially purified antibodies used in a solid-phase radioimmunoassay for detecting candidal antigenemia.

The development of a solid-phase radioimmunoassay procedure for the detection of Candida albicans antigens in serum of mice is described. Antibodies against C. albicans that were used in the radioimmunoassay procedure were partially purified from immune serum by a C. albicans antigen-coupled affinity column. Elution of anti-C. albicans antibodies from the column was by glucose and mannose; 4 mg of protein was recovered per ml, which contained 50% of the candidal agglutinin activity of immune serum. Also, 81% of the protein (partially purified antibody) recovered was adsorbed by whole C. albicans cells. Anti-C. albicans antibodies were either coupled to Sepharose 4B for use as the solid phase to bind candidal antigen in serum of infected animals, or radioiodinated (125I) for use as a tracer molecule to bind to the candidal antigen solid-phase complex. Although control experiments indicated that at least 100 ng of candidal antigen should be present in a serum specimen for a positive radioimmunoassay test, candidal antigenemia was detected in 70.4% of infected mice even in cases where blood cultures for C. albicans were negative. With further refinement and adaptability to human serum, the radioimmunoassay test may become a helpful tool for use in the diagnosis of systemic candidiasis.

Animals↗