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Enteropathogenic strains of Yersinia enterocolitica harbor a virulence plasmid (70 kilobases) which specifies, at 37°C, a calcium requirement for growth, autoagglutinability, resistance to the bactericidal activity of human serum, and the expression of some outer membrane proteins (OMPs). To map the genes encoding these properties, the virulence plasmid of a serogroup 9 strain (W22708) was subjected to transposon mutagenesis. A set of 68 independent mutations was obtained in Escherichia coli by transposon Tn813 (a tnpR mutant of Tn2l)-mediated cointegration with the self-transmissible R388 plasmid. The resulting cointegrates were introduced and studied in Y. enterocolitica W22708. One mutant lost the calcium dependence property. Two other mutants presented a peculiar phenotype: they grew poorly at 37°C, especially in the presence of calcium. Lastly, two mutants were affected in the properties of autoagglutination and resistance to human serum. Analysis of the OMP pattern of these two mutants revealed the absence of the largest OMP, called P1 (I. Bolin, and H. Wolf-Watz, Infect. Immun. 43:72-78, 1984). Complementation of one of these mutations with the cloned structural gene of OMP P1 restored the wild-type phenotype. However, OMP P1 was not sufficient by itself to specify the serum resistance property and a rapid autoagglutination of the host.

SUMMARY. The virulence of Yersinia enterocolitica depends on the presence of a 70-kilobase plasmid, called the Vwa plasmid. This situation is particularly favourable for studies of the mechanism of pathogenicity, but these are hindered by the lack of a suitable animal test to monitor the virulence of the human-pathogenic strains isolated outside the USA which belong to serogroups 0:3, 0:9 and 0:5,27. We observed that, after oral administration to the mouse, the Vwa-positive strains of these serogroups produce a discrete systemic infection while the Vwa-negative strains do not. We present here a simple mouse-virulence test based on this observation.