Genetic analysis of the plasmid region controlling virulence in Yersinia enterocolitica 0:9 by Mini-Mu insertions and lac gene fusions

Enteropathogenic strains of Yersinia enterocolitica harbor a virulence plasmid (pYVe plasmid) of 70 kilobases (kb) which specifies, at 37 degrees C, a calcium requirement for growth, autoagglutinability, resistance to the bactericidal activity of human serum, and the expression of outer membrane proteins (OMP). Some mutations suppress the calcium requirement for growth while others make the bacteria unable to grow at 37 degrees C, even in the presence of calcium. To analyse the genes involved in these latter phenotypes, the plasmid of a serogroup 0:9 strain was subjected to transposon mutagenesis with a Mini-Mu (Kan, lac) element. The mapping of 15 insertions and the analysis of transcription showed that at least four transcription units, spanning 22 kb, are involved in the phenomenon of calcium dependence. Mutations in two divergent units (virA and virB) suppressed the requirement for calcium at 37 degrees C. When insertions occurred in the other units (virC and virD), the Y. enterocolitica host became thermosensitive for growth. VirA, B and C mutants did no longer express the pYVe dependent OMPs. VirD mutants expressed and released these proteins save two of mol. wt 37,400 and 40,800 daltons. Transcription of the lac genes in the four groups of mutants was dependent on temperature.