The cultivation of Treponema microdentium as surface colonies

Socransky, S. S.; Macdonald, James B.; Sawyer, Sylvia J.

doi:10.1016/0003-9969(59)90009-3

Cited by 41 publications

(29 citation statements)

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“…Investigators who attempted to grow anaerobic spirochetes on solid media used media containing 1% or less agar (4,11,12) or preincubated the plates in anaer obicallyconditioned chambers (13). We also incubated the solid media contain ing0.9 to 3.1% agar for 2 days in an anerobic chamber and succeeded in colonizing the human oral treponeme, strain G7201, as well as Kazan and Reiter treponemes.…”

Section: Resultsmentioning

confidence: 99%

Colonial Morphology of Treponemes Observed by Electron Microscopy

Umemoto¹,

Namikawa²,

Yamamoto³

1984

Microbiology and Immunology

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The colonial morphology of three strains of cultivable, nonpathogenic treponemes including a human oral treponeme was examined by light and electron microscopy. Treponema phagedenis strains Kazan and Reiter produced large white colonies on the surface of solid media composed of sterility test broth, 0.9 to 3.1% agar, rifampin, and 12.5% rabbit or horse serum. A human oral treponeme, strain G7201, grew as diffused white zones on 0.9 to 3.1% agar plates. Under the cultural conditions employed agar concentrations slightly affected the time of appearance of colonies of the three strains of treponemes. When the colonies of these three strains were viewed by scanning electron microscopy, differences in their colonial morphology were observed. The 11‐day‐old colonies of human oral strain G7201 were very small, 5 to 15 μm in diameter, and had a slight irregular border. Kazan treponemes developed circular, entire and low convex colonies. Scanning and transmission electron microscopy revealed that the colonies of Reiter treponemes contained spherical forms almost up to 5 μm in diameter, each consisting of an outer membrane and a treponemal main body. They were very similar to the spherical bodies produced by strain G7201 in sucrose‐containing broth.

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Section: Resultsmentioning

confidence: 99%

Colonial Morphology of Treponemes Observed by Electron Microscopy

Umemoto¹,

Namikawa²,

Yamamoto³

1984

Microbiology and Immunology

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“…This observation has been made repeatedly in the past, and dilution to extinction has already led to the cultivation of spectacular environmental species (for examples, see references 9 and 39). In addition to the above results, and when designing our cultivation strategy, we took into account earlier findings that strict anaerobic incubation often increases microbial recovery (42,48,49,52). Finally, we hypothesized that conventional cultivation with nonselective media rich in sugars, such as Trypticaseglucose-yeast extract (TGY), brain heart infusion (BHI), Lactobacillus MRS, Wilkins-Chalgren, and many others may select for fast-growing species, thus masking growth of other, rarely cultivated or uncultivated strains (15,56).…”

Section: Discussionmentioning

confidence: 99%

New Approaches for Isolation of Previously Uncultivated Oral Bacteria

Sizova

Hohmann

Hazen

et al. 2012

Appl Environ Microbiol

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A significant number of microorganisms from the human oral cavity remain uncultivated. This is a major impediment to the study of human health since some of the uncultivated species may be involved in a variety of systemic diseases. We used a range of innovations previously developed to cultivate microorganisms from the human oral cavity, focusing on anaerobic species. These innovations include (i) in vivo cultivation to specifically enrich for species actively growing in the oral cavity (the "minitrap" method), (ii) single-cell long-term cultivation to minimize the effect of fast-growing microorganisms, and (iii) modifications of conventional enrichment techniques, using media that did not contain sugar, including glucose. To enable cultivation of obligate anaerobes, we maintained strict anaerobic conditions in most of our cultivation experiments. We report that, on a per cell basis, the most successful recovery was achieved using minitrap enrichment (11%), followed by single-cell cultivation (3%) and conventional plating (1%). Taxonomically, the richest collection was obtained using the single-cell cultivation method, followed by minitrap and conventional enrichment, comprising representatives of 13, 9, and 4 genera, respectively. Interestingly, no single species was isolated by all three methods, indicating method complementarity. An important result is the isolation and maintenance in pure culture of 10 strains previously only known by their molecular signatures, as well as representatives of what are likely to be three new microbial genera. We conclude that the ensemble of new methods we introduced will likely help close the gap between cultivated and uncultivated species from the human oral cavity.

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“…Therefore, a controlled, anaerobic environment is needed for its successful manipulation. The use of an anaerobic chamber (Figure 1A) provides the most stable environment and the ideal conditions for effective cultivation of C. difficile and other anaerobic bacteria 14 . Here, an atmosphere containing a gas mixture (5% CO 2 , 10% H 2 , 85% N 2 ) can be stably maintained.…”

Section: Protocolmentioning

confidence: 99%

Culturing and Maintaining <em>Clostridium difficile</em> in an Anaerobic Environment

Edwards

Suárez

McBride

2013

JoVE

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Clostridium difficile is a Gram-positive, anaerobic, sporogenic bacterium that is primarily responsible for antibiotic associated diarrhea (AAD) and is a significant nosocomial pathogen. C. difficile is notoriously difficult to isolate and cultivate and is extremely sensitive to even low levels of oxygen in the environment. Here, methods for isolating C. difficile from fecal samples and subsequently culturing C. difficile for preparation of glycerol stocks for long-term storage are presented. Techniques for preparing and enumerating spore stocks in the laboratory for a variety of downstream applications including microscopy and animal studies are also described. These techniques necessitate an anaerobic chamber, which maintains a consistent anaerobic environment to ensure proper conditions for optimal C. difficile growth. We provide protocols for transferring materials in and out of the chamber without causing significant oxygen contamination along with suggestions for regular maintenance required to sustain the appropriate anaerobic environment for efficient and consistent C. difficile cultivation.

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The cultivation of Treponema microdentium as surface colonies

Cited by 41 publications

References 0 publications

Colonial Morphology of Treponemes Observed by Electron Microscopy

Colonial Morphology of Treponemes Observed by Electron Microscopy

New Approaches for Isolation of Previously Uncultivated Oral Bacteria

Culturing and Maintaining <em>Clostridium difficile</em> in an Anaerobic Environment

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