No need to do animal experiments for that. To check your culture for the
presence of the virulence plasmid, use the congo-red magnesium-oxalate
agar described in G. Riley & S. Toma, J. Clin. Microbiol. (1989)
27:213-4. The minute red colonies are the ones carrying the plasmid. If
there are no minute red colonies left (also check the plate in the region
where you streaked the inoculum), the plasmid is gone. Pick and restreak
one of the minute colonies, avoiding those splitting off white-cream
coloured growth (needs some practice!), on nutrient agar with sufficient
amounts (mM range) of calcium and incubate at room temperature. Such a
culture will contain mainly plasmid-bearing cells (check again on
congo-red medium, if necessary).
High temperature and low calcium during growth of yersiniae selects for
loss of the plasmid. We have been able to maintain the virulence plasmid
in all our strains of yersinia over many years simply by avoiding
incubation temperatures above 30°C at all times.
--
Andre P. Burnens MD
Institute for Veterinary Bacteriology, University of Berne
Laenggass-Strasse 122, CH-3012 Berne, Switzerland
Tel. +41 31 631 24 85 Fax. +41 31 631 26 34
