Inhibition of Histoplasma capsulatum by Candida albicans and Other Yeasts on Sabouraud''s Agar Media

The inhibition of growth of Histoplasma capsulatum by Candida albicans and other yeasts on Sabouraud's agar was investigated. Histoplasma (yeast-phase inoculum) was grown alone and in mixtures with yeasts at 25 C for 4-week periods. As few as 10 colonies of C. albicans completely inhibited the growth of approximately 50,000 potential colonies of Histoplasma. The pH was determined in cultures of 36 colonies of Candida on media containing 1, 2, and 4% glucose by spotting the agar with pH indicators. A drop in the pH became noticeable in all three media about the 3rd day of incubation, and a pH of 3.5 was reached in about 7 days. Subsequently, the pH remained almost stationary in the 4% glucose-agar, rose slowly in the 2% glucose-agar, and rose sharply in the 1% glucose-agar. The growth of Histoplasma was inhibited completely at pH 4 and below. When the pH was controlled in mixed cultures, some growth of Histoplasma was obtained. Substitution of maltose for glucose delayed the development of acidity and allowed the appearance of numerous mycelial colonies in the presence of Candida. This growth was arrested as soon as the medium became acid. Four other species which also acidified the Sabouraud's medium effected similar inhibition. It was thus shown that severe and prolonged acidity produced by some yeasts in the sugar-rich Sabouraud's media is alone sufficient to completely inhibit Histoplasma during the standard 4-week incubation of specimens such as sputum.     

Single-channel properties of the recombinant skeletal muscle Ca2+ release channel (ryanodine receptor).

We report transient expression of a full-length cDNA encoding the Ca2+ release channel of rabbit skeletal muscle sarcoplasmic reticulum (ryanodine receptor) in HEK-293 cells. The single-channel properties of the 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate-solubilized and sucrose gradient-purified recombinant Ca2+ release channels were investigated by using single-channel recordings in planar lipid bilayers. The recombinant Ca2+ release channel exhibited a K+ conductance of 780 pS when symmetrical 250 mM KCl was used as the conducting ion and a Ca2+ conductance of 116 pS in 50 mM luminal Ca2+. Opening events of the recombinant channels were brief, with an open time constant of approximately 0.22 ms. The recombinant Ca2+ release channel was more permeable to Ca2+ than to K+, with a pCa2+/pK+ ratio of 6.8. The response of the recombinant Ca2+ release channel to various concentrations of Ca2+ was biphasic, with the channel being activated by micromolar Ca2+ and inhibited by millimolar Ca2+. The recombinant channels were activated by ATP and caffeine, inhibited by Mg2+ and ruthenium red, and modified by ryanodine. Most recombinant channels were asymmetrically blocked, conducting current unidirectionally from the luminal to the cytoplasmic side of the channel. These data demonstrate that the properties of recombinant Ca2+ release channel expressed in HEK-293 cells are very similar, if not identical, to those of the native channel.     

Escherichia coli mutants resistant to inactivation by high hydrostatic pressure.

Alternating cycles of exposure to high pressure and outgrowth of surviving populations were used to select for highly pressure-resistant mutants of Escherichia coli MG1655. Three barotolerant mutants (LMM1010, LMM1020, and LMM1030) were isolated independently by using outgrowth temperatures of 30, 37, and 42 degrees C, respectively. Survival of these mutants after pressure treatment for 15 min at ambient temperature was 40 to 85% at 220 MPa and 0.5 to 1.5% at 800 MPa, while survival of the parent strain, MG1655, decreased from 15% at 220 MPa to 2 x 10(-8)% at 700 MPa. Heat resistance of mutants LMM1020 and LMM1030 was also altered, as evident by higher D values at 58 and 60 degrees C and reduced z values compared to those for the parent strain. D and z values for mutant LMM1010 were not significantly different from those for the parent strain. Pressure sensitivity of the mutants increased from 10 to 50 degrees C, as opposed to the parent strain, which showed a minimum around 40 degrees C. The ability of the mutants to grow at moderately elevated pressure (50 MPa) was reduced at temperatures above 37 degrees C, indicating that resistance to pressure inactivation is unrelated to barotolerant growth. The development of high levels of barotolerance as demonstrated in this work should cause concern about the safety of high-pressure food processing.     

Large Scale Isolation and Purification of Eyespot Granules from Euglena gracilis var. bacillaris

Large volumes of eyespot granules were isolated from homogenates of Euglena gracilis Klebs var. bacillaris Pringsheim by flotation centrifugation in a Beckman Ti-15 zonal rotor, and were further purified by centrifugation in a swinging bucket rotor. Examination with the electron microscope showed the eyespot granules to be free from other cellular material. Freezing had no apparent effect on the structure or on the absorption properties of the eyespots. Absorption spectra of pure fractions of eyespot granules free of chloroplast contamination showed the previously reported curves in the range of 360 to 520 nanometers, as well as a peak at 660 to 675 nanometers. The procedure for the large scale isolation of eyespot granules from Euglena gracilis is compared with other methods which have employed conventional centrifugation, and the significance of the use of zonal rotors for isolating large quantities of pure eyespot granules is discussed.     

Effect of pH, Protein Concentration, and Ionic Strength on Heat Inactivation of Staphylococcal Enterotoxin B

Heat treatment of highly purified staphylococcal enterotoxin B causes a more rapid loss of immunological activity at 70 to 80 C than at 90 to 100 C. Toxicological results based on intravenous injection of dogs paralleled the results obtained by immunological means (single gel diffusion). The loss of immunological activity did not follow first-order kinetics. Results are given on the effects on heat inactivation of changing pH, ionic strength, and initial concentration of enterotoxin. Disc-gel electrophoresis of purified enterotoxin B showed a major and minor band. The minor band was a size isomer of the major band.     

Electron Microscopy Study of Reovirus Reaction Cores

A combined staining and shadowing method has been used to enhance the contrast of both reovirus cores and the RNA product that they have synthesized. After limited synthesis when all RNA strands associated with the core are nascent, many cores are observed that have more than 10 strands associated with each particle. The lengths of individual nascent mRNA molecules are not identical: many strands are extruded in the form of loops. These observations are consistent with the notion that synthesis of individual mRNA molecules occurs simultaneously at different rates at a number of different sites. The sites of extrusion are probably the 12 hollow projections located on the surface of the core.