Flat-Embedding for Electron Microscopy of Organisms Grown on Solid Agar Media

1961 ◽  
Vol 36 (2) ◽  
pp. 94-95 ◽  
Author(s):  
James K. Koehler
Keyword(s):  
Electron Microscopy ◽  
Solid Agar ◽  
Agar Media ◽  
Flat Embedding

TEM processing procedure for a bacillus organism grown on solid media

1990 ◽  
Vol 48 (3) ◽  
pp. 624-625
Author(s):  
Jane Payne ◽  
Philip Coudron
Keyword(s):  
Electron Microscopy ◽  
Room Temperature ◽  
Fume Hood ◽  
Vacuum Aspiration ◽  
Solid Media ◽  
Transmission Electron ◽  
Solid Agar ◽  
Processing Procedure ◽  
Agar Media ◽  
Rat Bone

This transmission electron microscopy (TEM) procedure was designed to examine a gram positive spore-forming bacillus in colony on various solid agar media with minimal artifact. Cellular morphology and organization of colonies embedded in Poly/Bed 812 resin (P/B) were studied. It is a modification of procedures used for undecalcified rat bone and Stomatococcus mucilaginosus.Cultures were fixed and processed at room temperature (RT) under a fume hood. Solutions were added with a Pasteur pipet and removed by gentle vacuum aspiration. Other equipment used is shown in Figure 3. Cultures were fixed for 17-18 h in 10-20 ml of RT 2% phosphate buffered glutaraldehyde (422 mosm/KgH2O) within 5 m after removal from the incubator. After 3 (30 m) changes in 0.15 M phosphate buffer (PB = 209-213 mosm/KgH2O, pH 7.39-7.41), colony cut-outs (CCO) were made with a scalpel.

Aeromonas hydrophila in Shellfish Growing Waters: Incidence and Media Evaluation

1989 ◽  
Vol 52 (1) ◽  
pp. 7-12 ◽  
Author(s):  
CARLOS ABEYTA ◽  
STEPHEN D. WEAGANT ◽  
CHARLES A. KAYSNER ◽  
MARLEEN M. WEKELL ◽  
ROBERT F. STOTT ◽  
...  
Keyword(s):  
Aeromonas Hydrophila ◽  
Environmental Samples ◽  
Enterotoxin Production ◽  
Beef Extract ◽  
Solid Agar ◽  
Agar Media

Levels of Aeromonas hydrophila determined for the shellfish growing area of Grays Harbor, Washington, ranged from 3 to 4600/100 g in oysters and from 3 to 2400/100 ml in water. Of isolates tested, 80% produced a hemolysin, a trait reported to correlate with enterotoxin production and pathogenicity. Two enrichment broths, Tryptic Soy Broth with ampicillin (TSBA) and Modified Rimler Shotts Broth (MRSB) were compared in combination with three solid agar media: Rimler Shotts (RS), Peptone Beef Extract Glycogen (PBG), and MacConkey's (MCA) agars. TSBA was far superior to MRSB in isolating this species from the environmental samples tested.

scholarly journals A Novel Flat-embedding Method to Prepare Ultrathin Cryosections from Cultured Cells in Their In Situ Orientation

2002 ◽  
Vol 50 (8) ◽  
pp. 1067-1080 ◽  
Author(s):  
Viola Oorschot ◽  
Heidi de Wit ◽  
Wim G. Annaert ◽  
Judith Klumperman
Keyword(s):  
Electron Microscopy ◽  
Quantitative Method ◽  
Cultured Cells ◽  
Petri Dish ◽  
Ultrastructural Level ◽  
Immunogold Labeling ◽  
Polarized Cells ◽  
Ultrathin Cryosections ◽  
Flat Embedding

Immunogold labeling of ultrathin cryosections provides a sensitive and quantitative method to localize proteins at the ultrastructural level. An obligatory step in the routine preparation of cryosections from cultured cells is the detachment of cells from their substrate and subsequent pelleting. This procedure precludes visualization of cells in their in situ orientation and hampers the study of polarized cells. Here we describe a method to sample cultured cells from a petri dish or coverslip by embedding them in a 12% gelatin slab. Subsequently, sections can be prepared in parallel or perpendicular to the plane of growth. Our method extends the cryosectioning technique to applications in studying polarized cells and correlative light–electron microscopy.

Colony formation by Bact. lactis aerogenes on solid media containing antibacterial agents

1952 ◽  
Vol 215 (1123) ◽  
pp. 565-565
Keyword(s):  
Inoculum Size ◽  
Survival Times ◽  
Fluctuation Test ◽  
Solid Media ◽  
Statistical Variation ◽  
Toxic Concentration ◽  
Solid Agar ◽  
Agar Media ◽  
Different Cultures ◽  
The Given

A study has been made of the formation of colonies of Bact. lactis aerogenes on solid agar media containing antibacterial substances (brilliant green, 1-phenyl semicarbazide, phenyl mercuric nitrate, phenol, thymol and chloramphenicol) at such concentrations that a small fraction only of the inoculated cells develop. The pattern of behaviour varies from drug to drug and sometimes from culture to culture with a given drug. As the toxic concentration increases, colonies diminish in number, in size or in both. Anomalous dependence in some cases upon inoculum size, and the appearance in others of satellites to the main colonies, indicate the operation of co-operative effects probably depending upon diffusion of metabolites or antagonists. The statistical variation in the number of developing colonies is greater for different cultures than for samples of a given culture (as in the well-known fluctuation test for mutations), but the behaviour of a culture may depend upon the aeration, and upon the precise conditions of the test. The variances show no apparent relation to the ease of production of resistance to the given drug. Nor does the scatter of the survival times in liquid media containing phenol (no resistance developable) differ much from that in chloramphenicol (resistant forms readily produced). Consideration of the factors determining the successful formation of a colony on a drug plate suggests that the fluctuation test for the demonstration of mutations must be applied with great reserve.

A technique for preserving aerial fungal structures for scanning electron microscopy

1983 ◽  
Vol 29 (6) ◽  
pp. 653-658 ◽  
Author(s):  
E. J. King ◽  
M. F. Brown
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Vapor Diffusion ◽  
Agar Media ◽  
Scanning Electron

A technique is described that employs vapor fixation and a simple vapor diffusion dehydration apparatus to minimize the disturbance of delicate fungal structures during preparation for scanning electron microscopy. The technique is applicable to fungi grown on agar media or on natural substrates.

Effect of Abamectin on Fungal Growth and its Efficacy as a Miticide in the Laboratory

2020 ◽  
Author(s):  
Dana Sebestyen ◽  
Gabriel Perez-Gonzalez ◽  
Mrinalini Ghoshal ◽  
Barry Goodell
Keyword(s):  
Fungal Growth ◽  
Recommended Dose ◽  
Effective Control ◽  
Phaeomoniella Chlamydospora ◽  
Trunk Disease ◽  
Solid Agar ◽  
The Media ◽  
Agar Media ◽  
The Common ◽  
Mold Mite

Abamectin was tested for use with solid agar media in the laboratory to eliminate or kill the common mold mite Tyrophagus spp. in fungal cultures of Phaeomoniella chlamydospora (Pch) and Phaeoacremonium minimum (Pmin), two important grape pathogens involved in grapevine trunk disease. Abamectin concentrations tested were at or below the recommended dose for abamectin in greenhouse spray applications (≦625ug/mL) to control mites and determine if: a) fungal growth would be inhibited, and b) mites would be killed or their activity suppressed. Abamectin was added either to the media before autoclaving, or filter-sterilized and added after autoclaving, to test the effects of autoclaving on abamectin efficacy. Streptomycin (100µg/mL) was also added to a set of treatments to determine if this commonly-used antibiotic would impact abamectin efficacy against mites, or have an effect on fungal growth when in combination with abamectin. Filter-sterilized abamectin in the range of 62.5 - 312ug/mL, delivered to the media after it had been autoclaved, provided the most effective control of mites while also showing limited inhibition of fungal growth on solid agar media in the absence of streptomycin. The addition of filter-sterilized streptomycin had no significant effect on fungal growth for Pch, while for Pmin a small but significant reduction in growth with streptomycin occurred at abamectin concentrations above 62.5 ug/ml.

Electron Microscopy of A New Plant-Pathogenic Spiroplasma Isolated From Opuntia

1976 ◽  
Vol 34 ◽  
pp. 56-57 ◽  
Author(s):  
F. KONDO ◽  
A. H. MCINTOSH ◽  
S. B. PADHI ◽  
K. MARAMOROSCH
Keyword(s):  
Electron Microscopy ◽  
Thin Section ◽  
Liquid Media ◽  
Serological Tests ◽  
Spiroplasma Citri ◽  
Corn Stunt ◽  
Section Electron ◽  
Agar Media ◽  
Darkfield Microscopy ◽  
Corn Stunt Spiroplasma

Thin section electron micrographs of phloem from the ornamental cactus Opuntia tuna monstrosa revealed the presence of wall-less prokaryotic cells earlier described as mycoplasma-like bodies (1). Isolation and growth of the microorganism in liquid media yielded cultures of a spiroplasma, as ascertained by darkfield microscopy. Negatively stained spiroplasmas (Fig. 1) were reminiscent of Spiroplasma citri (2) and of corn stunt spiroplasma (3). The spirals observed in negatively stained preparations seemed attached to blebs. PAGE analysis showed distinct differences between the three species of microorganisms. Serological tests confirm ed that the spiroplasma isolated from the ornamental cactus differed from S. citri and from corn stunt spiroplasma. On agar media, “fried-egg” colonies were formed (Fig. 2).

scholarly journals Effects of inositol starvation on the levels of inositol phosphates and inositol lipids in Neurospora crassa

1993 ◽  
Vol 292 (3) ◽  
pp. 805-811 ◽  
Author(s):  
P L Lakin-Thomas
Keyword(s):  
Neurospora Crassa ◽  
Phytic Acid ◽  
Inositol Phosphates ◽  
Acid Synthesis ◽  
Extracellular Medium ◽  
Absolute Requirement ◽  
Solid Agar ◽  
Inositol Lipids ◽  
Agar Media ◽  
High Concentrations

An inositol-requiring strain of Neurospora crassa was labelled during growth in liquid medium with [3H]inositol, and the levels of inositol phosphates and phosphoinositides were determined under inositol-sufficient and inositol-starved conditions. Because the mutant has an absolute requirement for inositol, the total mass of inositol-containing compounds could be determined. Inositol-containing lipids were identified by deacylation and co-migration with standards on h.p.l.c.; PtdIns3P, PtdIns4P, and PtdIns(4,5)P2 were found in approximately equal amounts, in addition to large amounts of PtdIns. Inositol starvation decreased the level of PtdIns to 10% of the sufficient level, and decreased the levels of the other phosphoinositides to about 25%. A number of inositol phosphates were found, including several InsP3s, InsP4s and InsP5s and phytic acid. Ins(1,4,5)P3 was identified by co-migration with standards on h.p.l.c. and by digestion with inositol phosphomonoesterase. High concentrations of all inositol phosphates were found in the extracellular medium in inositol-starved cultures. Inositol starvation on both liquid and solid agar media decreased the intracellular levels of some inositol phosphates, but increased the levels of phytic acid and several other inositol phosphates which may be its precursors and/or breakdown products. These results may indicate that inositol starvation induces phytic acid synthesis as a protection against the free-radical production and lipid peroxidation characteristic of inositol-less death.

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