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ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
2. To obtain a biphasic culture, add 0.4 ml of the suspension to a #260 slant. Add remaining 0.1 ml of the suspension to a #260 plate and streak for isolation.
3. Incubate at 37oC under microaerophilic conditions using an anaerobe jar with an active catalyst and a microaerophilic gas generator pack, or other acceptable method, to obtain microaerophilic conditions. Incubate slant with cap loose.
4. Within 3 days of incubation, good growth should be obtained in the broth pool at the bottom of the slant. Additional incubation may be required for colonies to appear on the plate. Further subcultures can be made using broth pool as the inoculum source.
The organism is a short, curved, motile bacillus.
It does not readily form colonies on an agar surface. Growth on agar takes longer than with the biphasic culture. Colonies are small, circular, entire, convex, smooth, and gray.
The cells do not Gram stain well using traditional procedures. To obtain the best results, use a basic fuchsin counterstain in place of the safranin.
Once good growth in obtained, transfer or freeze the culture. Adding an equal amount of 20% sterile glycerol to pooled broth from several biphasic slants, followed by freezing in liquid nitrogen or ultra-low temperature freezer is recommended.
See also Helicobacter species, General Procedures in ATCC Bacteria & Bacteriophages, 19th edition, 1996, p.471.
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Dewhirst FE, et al. Phylogeny of Helicobacter isolates from bird and swine feces and description of Helicobacter pametensis sp. nov.. Int. J. Syst. Bacteriol. 44: 553-560, 1994. PubMed: 7520743
Seymour C, et al. Isolation of Helicobacter strains from wild bird and swine feces. Appl. Environ. Microbiol. 60: 1025-1028, 1994. PubMed: 8161169