SummaryThe severity of infections caused by Salmonella enterica serovar Typhimurium varies depending on the host species. Numerous virulence genes have been identified in S . Typhimurium, largely from studies in mice, but their roles in infections of other species remain unclear. In the most comprehensive survey of its kind, through the use of signaturetagged mutagenesis of S. Typhimurium we have identified mutants that were unable to colonize calf intestines, mutants unable to colonize chick intestines and mutants unable to colonize both species. The type three secretion systems encoded on Salmonella pathogenicity islands (SPIs) 1 and 2 were required for efficient colonization of cattle. However, disruption of these secretion systems only caused a minor defect in S . Typhimurium colonization of chicks. Transposon insertions in SPI-4 compromised S . Typhimurium colonization of cattle, but not chicks. This is the first data confirming a role for SPI-4 in pathogenesis. We have also been able to ascribe a role in colonization for cell surface polysaccharides, cell envelope proteins, and many 'housekeeping' genes and genes of unknown function. We conclude that S . Typhimurium uses different strategies to colonize calves and chicks. This has major implications for vaccine design.

The expression of Escherichia coli group 2 capsules (K antigens) is temperature dependent, with capsules only being expressed at temperatures above 20°C. Thermoregulation is at the level of transcription, with no detectable transcription at 20°C. Using the E. coli K5 capsule gene cluster as a model system, we have shown that the nucleoid-associated protein H-NS plays a dual role in regulating transcription of group 2 capsule gene clusters at 37 and 20°C. At 37°C H-NS is required for maximal transcription of group 2 capsule gene clusters, whereas at 20°C H-NS functions to repress transcription. The BipA protein, previously identified as a tyrosinephosphorylated GTPase and essential for virulence in enteropathogenic E. coli, was shown to play a similar role to H-NS in regulating transcription at 37 and 20°C. The binding of integration host factor (IHF) to the region 1 promoter was necessary to potentiate transcription at 37°C and IHF binding demonstrated by bandshift assays. The IHF binding site was 3 to the site of transcription initiation, suggesting that sequences in the 5 end of the first gene (kpsF) in region 1 may play a role in regulating transcription from this promoter at 37°C. Two additional cis-acting sequences, conserved in both the region 1 and 3 promoters, were identified, suggesting a role for these sequences in the coordinate regulation of transcription from these promoters. These results indicate that a complex regulatory network involving a number of global regulators exists for the control of expression of group 2 capsules in E. coli.

Many vaccines have been developed from live attenuated forms of bacterial pathogens or from killed bacterial cells. However, an increased awareness of the potential for transient side-effects following vaccination has prompted an increased emphasis on the use of sub-unit vaccines, rather than those based on whole bacterial cells. The identification of vaccine sub-units is often a lengthy process and bioinformatics approaches have recently been used to identify candidate protein vaccine antigens. Such methods ultimately offer the promise of a more rapid advance towards preclinical studies with vaccines. We have compared the properties of known bacterial vaccine antigens against randomly selected proteins and identified differences in the make-up of these two groups. A computer algorithm that exploits these differences allows the identification of potential vaccine antigen candidates from pathogenic bacteria on the basis of their amino acid composition, a property inherently associated with sub-cellular location.