Effect of calcium on endospore ultrastructure

Clostridium sporogenes has been designated as a test species for the closely related but pathogenic Cl. perfringens. Cl. sporogenes possess a unique morphology. High resolution EM has not been applied and little is known about its calcium distribution, thus this study presents a morphological evaluation as well as data on calcium localization, which is considered important for endospore resistance.Cl. Sporogenes was grown on cooked meat/media. Sporulation was monitered using the malachite green method. After maximum sporulation the spores were harvested in deionized distilled water, heated for 20 minutes at 80°C to kill any remaining vegetative cells, washed 3 times with sterile deionized water and the stock suspension was stored at 2-8°C until needed. Aliquots of the spores were placed momentarily in warm agar prior to fixation then fixed in 3% gluteraldehyde (GCHO) in 0.1 M cacodylate buffer pH 7.4 to which Ca was deleted or with 1% CaCl.

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Clostridium Spore Attachment to Human Cells

Microscopy and Microanalysis ◽

10.1017/s1431927600013143 ◽

1997 ◽

Vol 3 (S2) ◽

pp. 1251-1252

Author(s):

B. Panessa-Warren ◽

G. Tortora ◽

J. Warren

Keyword(s):

High Resolution ◽

Cell Surface ◽

Human Cells ◽

Carcinoma Cells ◽

Embryonic Fibroblasts ◽

Clostridium Sporogenes ◽

Colon Carcinoma Cells ◽

Spore Attachment ◽

Cooked Meat ◽

Commercial Field

This paper uses high resolution scanning electron microscopy (SEM) with a LaB6 gun and the newest commercial field emission guns, to obtain high magnification images of intact clostridial spores throughout the activation/germination/outgrowth process. by high resolution SEM, the clostridial exosporial membrane can be seen to produce numerous delicate projections (following activation), that extend from the exosporial surface to a nutritive substrate (agar), or cell surface when anaerobically incubated in the presence of human cells (embryonic fibroblasts and colon carcinoma cells). Magnifications of 20,000 to 200,000Xs at accelerating voltages low enough to minimize or eliminate specimen damage (1-5 kV) have permitted the entire surface of C.sporogenes and C.difficile endospores to be examined during all stages of germination. The relationships between the spore and the agar or human cell surface were also clearly visible.Clostridium sporogenes ATCC 3584 and C.difficile ATCC 43594 and 9689 were grown in cooked meat media, filtered and cleaned.

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High Resolution Stereography of Complementary Freeze-Fracture Replicas Reveals the Presence of Depressions Opposite Membrane Associated Particles of Split Membranes

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100066267 ◽

1971 ◽

Vol 29 ◽

pp. 442-443

Author(s):

Russell L. Steere

Keyword(s):

High Resolution ◽

Cardiac Muscle ◽

Electron Micrographs ◽

Chromic Acid ◽

Freeze Fracture ◽

Distilled Water ◽

Fine Scale ◽

Acid Cleaning ◽

Cleaning Solution

Complementary replicas have revealed the fact that the two common faces observed in electron micrographs of freeze-fracture and freeze-etch specimens are complementary to each other and are thus the new faces of a split membrane rather than the original inner and outer surfaces (1, 2 and personal observations). The big question raised by published electron micrographs is why do we not see depressions in the complementary face opposite membrane-associated particles? Reports have appeared indicating that some depressions do appear but complementarity on such a fine scale has yet to be shown.Dog cardiac muscle was perfused with glutaraldehyde, washed in distilled water, then transferred to 30% glycerol (material furnished by Dr. Joaquim Sommer, Duke Univ., and VA Hospital, Durham, N.C.). Small strips were freeze-fractured in a Denton Vacuum DFE-2 Freeze-Etch Unit with complementary replica tooling. Replicas were cleaned in chromic acid cleaning solution, then washed in 4 changes of distilled water and mounted on opposite sides of the center wire of a Formvar-coated grid.

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Conversion of Strontium Hydroxyapatite Nanopowders to Porous Scaffolds for Bone Implant Application

Applied Mechanics and Materials ◽

10.4028/www.scientific.net/amm.607.3 ◽

2014 ◽

Vol 607 ◽

pp. 3-6

Author(s):

Che Murad Mardziah ◽

Iis Sopyan ◽

Koay Mei Hyie ◽

N.R. Nik Roselina

Keyword(s):

Compressive Strength ◽

Cancellous Bone ◽

Compression Test ◽

Ambient Air ◽

Porous Scaffolds ◽

Distilled Water ◽

Bone Implant ◽

Morphological Evaluation ◽

Dispersing Agent ◽

Xrd Patterns

The fabrication of Sr hydroxyapatite (HA) porous scaffolds was done by using polymeric sponge method. To prepare the porous samples, the synthesized SrHA nanopowders were mixed with distilled water and appropriate amount of dispersing agent followed by drying in the ambient air and sintering at 1300°C. The compressive strength of the materials was strongly influenced by the porosity, while there was almost no dependence on the crystallinity of the powders since XRD patterns showed high crystallinity of HA phase for all porous samples. Morphological evaluation by FESEM revealed that the SrHA scaffolds were characterized by macro-micro interconnected porosity, which replicates the morphology of the cancellous bone. Compression test on the porous scaffolds demonstrated that doping 10 mol% of strontium in HA has increased the compressive strength by a factor of two compared to the undoped HA with 1.81±0.26 MPa at 41% porosity.

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Improvement of the methodology of the tetrazolium test using different pretreatments in seeds of the genus Epidendrum (Orchidaceae)

Journal of Seed Science ◽

10.1590/2317-1545v42231028 ◽

2020 ◽

Vol 42 ◽

Cited By ~ 2

Author(s):

Seir Antonio Salazar Mercado ◽

Jesús David Quintero Caleño ◽

Laura Yolima Moreno Rozo

Keyword(s):

Deionized Water ◽

Control Group ◽

Distilled Water ◽

In Vitro Germination ◽

Test Results ◽

Germination Test ◽

Viability Test ◽

Cloth Filter ◽

Tetrazolium Test

Abstract: The aim of the present study was to determine the most suitable pretreatment to enhance the tetrazolium test in seeds of the Epidendrum genus. Initially, mature capsules were harvested at El Escorial village, in the municipality of Pamplona, Colombia. Subsequently, the seeds were subjected to five pretreatments: deionized water, 0.5% NaClO, 1.0% NaClO, 10.0% sucrose and a control group. Using the syringe method with cloth filter, the seeds were rinsed with distilled water and subjected to two concentrations of tetrazolium solution (0.25%, 1.0%) and three exposure times (6 h, 24 h and 48 h). Finally, the tetrazolium viability test results were correlated with the in vitro germination test. It was found that the use of deionized water improves the efficiency of the tetrazolium test in seeds of Epidendrum fimbriatum and E. microtum; as in seeds of E. elongatum when using 1.0% tetrazolium for 24 h. Similarly, a high correlation was found between viability and germination, using deionized water and 10.0% sucrose, with homogeneous results with each other.

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High-resolution SEM of clostridial attachment structures

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100140841 ◽

1995 ◽

Vol 53 ◽

pp. 894-895

Author(s):

B. Panessa-Warren ◽

G.T. Tortora ◽

J.B. Warren

Keyword(s):

High Resolution ◽

Light Microscopy ◽

Lanthanum Hexaboride ◽

Specimen Preparation ◽

Initial Attachment ◽

Clostridium Sporogenes ◽

The Past ◽

Attachment Structures ◽

Plate Counts ◽

Colonization Process

Clostridium sporogenes forms small (1 μm), dehydrated endospores that are resistant to the effects of solvents, acids, alkali, pressure, UV radiation and most sporocides. In the past it was felt that the exosporial membrane served no function in the infection and colonization process, however our recent work on intact spores has demonstrated that the exosporial membrane seems to play a significant role in the initial attachment of the spore to a nutritive substrate. With the improved resolution of the SEM, it has been possible to examine intact spores attached to various substrates and characterize each stage of attachment. This paper examines the exosporial membranes of activated C.sporogenes spores using conventional specimen preparation with imaging using a lanthanum hexaboride gun, and specimens imaged with a field emission SEM. Spores were grown on Trypticase Soy agar with 0.5% glucose (TSA) added and blood CDC (BCDC) agar and attachments and viability monitored by plate counts and light microscopy of unfixed spores (both phase and malachite green/safranin staining).

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The Influence of Dentine on the pH of Calcium Hydroxide, Chlorhexidine Gel, and Experimental Bioactive Glass-Based Root Canal Medicament

The Scientific World JOURNAL ◽

10.1155/2015/686259 ◽

2015 ◽

Vol 2015 ◽

pp. 1-5 ◽

Cited By ~ 2

Author(s):

Ceci Nunes Carvalho ◽

Laila Gonzales Freire ◽

Alexandre Pinheiro Lima de Carvalho ◽

Evandro Luiz Siqueira ◽

José Bauer ◽

...

Keyword(s):

Calcium Hydroxide ◽

Statistical Difference ◽

External Solution ◽

Deionized Water ◽

Distilled Water ◽

Time Points ◽

Tukey Test ◽

Ph Values ◽

The Mean ◽

Chlorhexidine Gel

Objectives. To evaluate the influence of dentine on the pH of different medications in standardized simulated canals.Materials and Methods. Forty resin blocks were divided into groups with and without dentine powder, as follows: 2% chlorhexidine gel; 2% chlorhexidine gel associated with calcium hydroxide PA; calcium hydroxide PA delivered in propylene glycol 600; and NPG delivered in distilled water. The dentine powder was obtained from the root dentine of bovine teeth and added to the medications. The simulated canals were placed in containers with 1.5 mL of deionized water and pH was monitored in multiple intervals, up to 30 days. The mean pH values were calculated and submitted to statistical analysis using paired Student’st-test and ANOVA complemented by the Tukey test (p<0.05).Results. There was no statistical difference between the groups with and without dentine powder (p>0.05). The pH values of calcium hydroxide were significantly higher than those of NPG in the first 24 hours (p<0.05). After 7 days, both behaved in a similar manner.Conclusion. The addition of dentine powder to the medications evaluated did not alter the pH of the external solution in any of the time points tested.

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Method to Biomonitor the Cooked Meat Carcinogen 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine in Dyed Hair by Ultra-Performance Liquid Chromatography–Orbitrap High Resolution Multistage Mass Spectrometry

Analytical Chemistry ◽

10.1021/acs.analchem.5b01129 ◽

2015 ◽

Vol 87 (12) ◽

pp. 5872-5877 ◽

Cited By ~ 8

Author(s):

Jingshu Guo ◽

Kim Yonemori ◽

Loïc Le Marchand ◽

Robert J. Turesky

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

High Resolution ◽

Ultra Performance Liquid Chromatography ◽

Cooked Meat

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Evaluation of Four Pea (Pisum sativum) Cultivars in PE-2 Medium for the MPN Enumeration of Anaerobic Spore-Forming Organisms

Journal of Food Protection ◽

10.4315/0362-028x-60.12.1574 ◽

1997 ◽

Vol 60 (12) ◽

pp. 1574-1576 ◽

Cited By ~ 3

Author(s):

O. A. OGUNRINOLA ◽

C. G. EDWARDS ◽

P. M. DAVIDSON

Keyword(s):

Pisum Sativum ◽

Anaerobic Bacteria ◽

False Positives ◽

Clostridium Butyricum ◽

The Other ◽

Most Probable Number ◽

Clostridium Sporogenes ◽

Probable Number ◽

Cooked Meat

Untreated ‘Alaska’ seed peas have traditionally been used to prepare PE-2 medium, a medium used to recover anaerobic bacteria. Three cultivars of seed peas, ‘Columbian’, ‘Yellow’, and ‘Scotch’ were compared to the cultivar ‘Alaska’ peas in PE-2 for recovery of Clostridium butyricum ATCC 860, Clostridium sporogenes ATCC 7955/NCA 3679, and Thermoanerobacterium thermosaccharolyticum ATCC 25773 determined by using the most probable number (MPN) technique. Organisms were grown in cooked meat medium (CMM) and enumerated by the three-tube MPN method in PE-2 media incubated at 37°C for 48 h. Recovery of C. butyricum and T. thermosaccharolyticum grown in homogenates of commercial cream-style corn and vegetable beef soup were also evaluated. Similar recovery of organisms from stock cultures or from foodstuffs were observed in PE-2 prepared with three of the pea cultivars (‘Alaska’, ‘Columbian’, and ‘Yellow’). While the ‘Scotch’ cultivar PE-2 medium yielded recoveries comparable to PE-2 prepared with the other pea cultivars, turbid tubes were occasionally observed from which viable microorganisms could not be recovered. This cultivar is therefore not recommended due to the probability of yielding false positives in the MPN technique.

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