Serological Comparison of the Three Morphological Phases of Coccidioides immitis by the Agar Gel Diffusion Method

Hyperimmune sera against spherules and against arthrospores of Coccidioides immitis were prepared by inoculation of rabbits. The antibody content of these sera was studied by the agar gel diffusion method. It was observed that antispherule pooled sera formed multiple precipitin bands with extracts of spherules and of arthrospores. The antiarthrospore pooled serum, however, failed to precipitate with the spherule extract, and formed a single band in the presence of an arthrospore solution. When the spherule and the arthrospore extracts were tested with a variety of different antisera, it was observed that the spherule preparation formed bands only in combination with anti-purified spherule pooled serum, whereas the arthrospore extract precipitated with anti-purified spherule, antiarthrospore, and anti-Histoplasma capsulatum pooled sera. It was also observed that a spherule culture supernatant solution formed five precipitin bands in combination with anti-spherule pooled sera, formed one band with pooled antiserum from rabbits with coccidioidomycosis, and did not precipitate in the presence of antiarthrospore pooled serum. Coccidioidin, however, formed two bands in the presence of any of these antisera. It was therefore concluded that extracts from the spherule phase of C. immitis differed from solutions obtained from the arthrospore and mycelial phases.     

Competition between soil bacteria and fusarium

Summary The phenomenon of competition has been characterized in liquid medium and sterile soil systems using a variety of soil bacteria andFusarium oxysporum f.cubense as test organisms. For most of the bacteria, suppression of the fungus was the result of a biologically induced nitrogen deficiency, this effect being reversed by the addition of excess inorganic nitrogen. High populations of competitors were found in two soils of neutral pH, but no isolates competed in the acid San Alejo loam.Agrobacterium radiobacter was able to compete when San Alejo loam was limed to about pH 6.6. Inhibition of the fungus by a number of gram-positive, spore-forming rods could not be accounted for in terms of competition for nutrients or by antibiotic production in artificial media.The competitive ability ofA. radiobacter when tested in twelve Central American soils was found to be related to pH in acid and neutral environments but was correlated with texture, organic-matter content and total nitrogen in soils of intermediate pH. In all soils where inhibition occurred, the competitive effect was overcome by additions of inorganic nitrogen. Excluding the group ofBacillus spp., the competitive ability of soil bacteria was related to the ability to develop in the absence of amino acids and growth factors but could not be correlated with growth rates of the bacteria in soil or liquid medium.It is suggested that competition for nutrients is a significant means of ecological control among members of the soil microflora and, together with competitive interactions for space and oxygen, may be the major factors governing the biological control of soil-borne fungi.The investigation was supported in part by a grant from the United Fruit Company. Agronomy Paper No. 471.     

Factors Affecting the Development of Clostridium botulinum in Whole Milk

This study was undertaken to determine the effect of the sterilization process and the age of the medium at the time of inoculation on the development of Clostridium botulinum type 62A. Whole milk was autoclaved at 121 C for 18 or 30 min and inoculated with C. botulinum so as to contain 2,000 to 5,000 spores per milliliter. No effort was made to remove dissolved oxygen or to reduce the oxidation-reduction (O/R) potential of the medium by adding sodium thioglycolate. A 3- and a 5-day-old medium were used to study the influence of aging. Eh determinations were made periodically on inoculated and uninoculated samples. Culture development was followed by use of an oval tube counting procedure. The technique used to sterilize the milk influenced the initial O/R potential as well as the autoreductive capacity of the medium. The initial Eh of whole milk sterilized in 500-ml volumes for 18 min was 234 mv. In milk sterilized for 30 min it was 192 mv. The lag phase was 7 days in the former and 5 days in the latter. The exponential growth phases were similar. Autoreduction occurred in sterilized milk. The Eh of uninoculated milk sterilized for 18 min decreased 45 mv in 6 days. In milk sterilized for 30 min the decrease was 63 mv. In milk inoculated 3 or 5 days after sterilization the lag phase was shorter than when the medium was inoculated within 2 hr after autoclaving, regardless of the sterilization procedure employed. The autoreductive property of sterilized whole milk plays a major role in the development of C. botulinum type 62A.     

A serological comparison of the three morphological phases of Coccidioides immitis; complement fixation tests with anti-Histoplasma capsulatum serum

Summary The results of this study indicated that antigens prepared from the three morphological phases ofCoccidioides immitis differed in their complement fixing activity with anti-Histoplasma capsulatum pooled serum. Spherule antigens were serologically less active in tests with the anti-H. capsulatum pooled serum than antigens prepared from arthrospores and from mycelium.Antigenic determinants which are common toC. immitis andH. capsulatum appeared to be located on the intact arthrospore cellular surface but not on the surface of spherule cells.Part of a dissertation submitted to the Graduate School of Duke University in partial fulfillment of requirements for the Ph. D. degree.This work was supported by contract with the Department of the Army, Fort Detrick, Frederick, Maryland.In conducting the research reported herein, the investigators adhered to Guide for Laboratory Animal Facilities and Care established by the Committee on the Guide for Laboratory Animal Facilities and Care of the Institute of Laboratory Animal Resources, NAS-NRC.     

Ras interaction with the GTPase-activating protein (GAP)

Biologically active forms of Ras complexed to GTP can bind to the GTPase-activating protein (GAP), which has been implicated as possible target of Ras in mammalian cells. In order to study the structural features of Ras required for this interaction, we have evaluated a series of mutant ras proteins for the ability to bind GAP and a series of Ras peptides for the ability to interfere with this interaction. Point mutations in the putative effector region of Ras (residues 32-40) that inhibit biological activity also impair Ras binding to GAP. An apparent exception is the Thr to Ser substitution at residue 35; [Ser-35]Ras binds to GAP as effectively as wild-type Ras even though this mutant is biologically weak in both mammalian and S. cerevisiae cells. In vitro, [Ser-35]Ras can also efficiently stimulate the S. cerevisiae target of Ras, adenylyl cyclase, indicating that other factors may influence Ras/protein interactions in vivo. Peptides having Ras residues 17-44 and 17-32 competed with the binding of Ras to E. coli-expressed GAP with IC50 values of 2.4 and 0.9 microM, respectively, whereas Ras peptide 17-26 was without effect up to 400 microM. A related peptide from the yeast GTP-binding protein YPT1 analogous to Ras peptide 17-32 competed with an IC50 value of 19 microM even though the YPT1 protein itself is unable to bind to GAP. These results suggest that determinants within Ras peptide 17-32 may be important for Ras binding to GAP.