Helicobacter pullorum Stanley et al.
contents of vial with 0.5 ml of Trypticase Soy Broth.
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 37°C 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 to 7 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.
Growth on agar takes longer than with the biphasic culture. Colonies show as a haze to pinpoint in size. More mature colonies will appear slightly larger with some spreading Once good growth is present, these organisms tend to lose viability, especially if exposed to air for lengthy periods. Viability also decreases with repeated subculturing. 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 is 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.
Additional information on this culture is available on the ATCC web site at www.atcc.org.
Stanley J, et al. Helicobacter pullorum sp. nov.-genotype and phenotype of a new species isolated from poultry and from human patients with gastroenteritis. Microbiology 140: 3441-3449, 1994. PubMed: 7533595