An aroA mutant of Yersinia pestis is attenuated in guinea-pigs, but virulent in mice.
This study describes a PCR-based approach for the production of a rationally attenuated mutant of Yersinia pestis. Degenerate primers were used to amplify a fragment encoding 91.45% of the aroA gene of Y. pestis MP6 which was cloned into pUC18. The remainder of the gene was isolated by inverse PCR. The gene was sequenced and a restriction map was generated. The Y. pestis aroA gene had 75.9% identity with the aroA gene of Yersinia enterocolitica. The cloned gene was inactivated in vitro and reintroduced into Y. pestis strain GB using the suicide vector pGP704. A stable aro-defective mutant. Y. pestis GB aroA, was isolated and its virulence was examined in vivo. The mutant was attenuated in guinea-pigs and capable of inducing a protective immune response against challenge with the virulent Y. pestis strain GB. Unusually for an aro-defective mutant, the Y. pestis aroA mutant was virulent in mice, with a median dose which induced morbidity of death similar to that of the wild-type, although time to death was significantly prolonged.
Oyston
P C
PC
Chemical and Biological Defence Establishment, Salisbury, Wiltshire, UK. 100432.3200@compuserve.com
Russell
P
P
Williamson
E D
ED
Titball
R W
RW
D051229:3-Phosphoshikimate 1-Carboxyvinyltransferase; D019883:Alkyl and Aryl Transferases; D000595:Amino Acid Sequence; D000818:Animals; D001428:Bacterial Vaccines / Q000302:isolation & purification; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004195:Disease Models, Animal; D005260:Female; D006168:Guinea Pigs; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D010930:Plague / Q000209:etiology / Q000276:immunology / Q000517:prevention & control; D017386:Sequence Homology, Amino Acid; D014166:Transferases / Q000235:genetics* / Q000276:immunology; D014613:Vaccines, Attenuated / Q000302:isolation & purification; D014774:Virulence / Q000235:genetics; D015010:Yersinia pestis / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity*
D001428:Bacterial Vaccines; D017931:DNA Primers; D004269:DNA, Bacterial; D014613:Vaccines, Attenuated; D014166:Transferases; D019883:Alkyl and Aryl Transferases; D051229:3-Phosphoshikimate 1-Carboxyvinyltransferase