Philip W. Jones

An Inv/Mxi-Spa-like type III protein secretion system in Burkholderia pseudomallei modulates intracellular behaviour of the pathogen

Burkholderia pseudomallei is the causative agent of melioidosis, a serious infectious disease of humans and animals that is endemic in subtropical areas. B. pseudomallei is a facultative intracellular pathogen that may invade and survive within eukaryotic cells for prolonged periods. After internalization, the bacteria escape from endocytic vacuoles into the cytoplasm of infected cells and form membrane protrusions by inducing actin polymerization at one pole. It is believed that survival within phagocytic cells and cell‐to‐cell spread via actin protrusions is required for full virulence. We have studied the role of a putative type III protein secretion apparatus (Bsa) in the interaction between B. pseudomallei and host cells. The Bsa system is very similar to the Inv/Mxi‐Spa type III secretion systems of Salmonella and Shigella. Moreover, B. pseudomallei encodes proteins that are very similar to Salmonella and Shigella Inv/Mxi‐Spa secreted proteins required for invasion, escape from endocytic vacuoles, intercellular spread and pathogenesis. Antibodies to putative Bsa‐secreted proteins were detected in convalescent serum from a melioidosis patient, suggesting that the system is functionally expressed in vivo. B. pseudomallei mutant strains lacking components of the Bsa secretion and translocation apparatus were constructed. The mutant strains exhibited reduced replication in J774.2 murine macrophage‐like cells, an inability to escape from endocytic vacuoles and a complete absence of formation of membrane protrusions and actin tails. These findings indicate that the Bsa type III secretion system plays an essential role in modulating the intracellular behaviour of B. pseudomallei.

Options for the control of enterohaemorrhagic Escherichia coli in ruminants

Enterohaemorrhagic Escherichia coli (EHEC) comprise an important group of zoonotic enteric pathogens. In humans, some EHEC infections result in bloody or nonbloody diarrhoea, which may be complicated by haemorrhagic colitis and severe renal and neurological sequelae, including haemolytic uraemic syndrome (HUS). Ruminants are an important reservoir of EHEC and human infections are frequently associated with direct or indirect contact with ruminant faeces. Strategies to reduce the prevalence of EHEC in ruminants should lower the incidence of human infection. However, little is currently known about the mechanisms of intestinal colonization of ruminants by EHEC and no eective vaccines have yet been developed. Here we review current knowledge on the host responses to EHEC infection and the factors mediating EHECintestinal interactions and survival in cattle and sheep. Perspectives for the use of vaccines, probiotics and other measures to control EHEC in farm animals are discussed.

Salmonella enterica Serovars Typhimurium and Dublin Can Lyse Macrophages by a Mechanism Distinct from Apoptosis

ABSTRACT Salmonella enterica serovars Typhimurium and Dublin lysed primary bovine alveolar macrophages and immortalized J774.2 macrophage-like cells in the absence of either the morphological changes or DNA fragmentation characteristic of apoptosis. Macrophage lysis was dependent on a subset of caspases and an intactsipB gene.

Interaction of Salmonella serotypes with porcine macrophages in vitro does not correlate with virulence.

The interaction between Salmonella serotypes and macrophages is potentially instrumental in determining the outcome of infection. The nature of this interaction was characterized with respect to virulence and serotype-host specificity using pigs as the infection model. Experimental infection with Salmonella typhimurium, Salmonella choleraesuis or Salmonella dublin resulted in enteric, systemic or asymptomatic infection, respectively, which correlates well with the association of S. choleraesuis with systemic disease in pigs in epidemiological studies. Persistence within porcine alveolar macrophages in vitro did not directly correlate with virulence since S. typhimurium persisted in the highest numbers, and S. choleraesuis in the lowest. Comparison to other studies revealed that the relatively high persistence of S. typhimurium in macrophages correlates with its virulence in a broad range of animals: this could be a virulence mechanism for broad-host-range serotypes. There were little or no significant differences in the induction of pro-inflammatory cytokines by macrophages infected with the three serotypes. S. typhimurium and S. dublin, but not S. choleraesuis, damaged porcine macrophages, and the mechanism of damage did not resemble apoptosis. In conclusion, the virulence of Salmonella serotypes in pigs did not directly correlate with their interaction with porcine macrophages in vitro. The interaction of Salmonella and macrophages in vitro may not accurately model their interaction in vivo, and this will form the basis of further study.

Analysis of Salmonella enterica Serotype-Host Specificity in Calves: Avirulence of S. enterica Serotype Gallinarum Correlates with Bacterial Dissemination from Mesenteric Lymph Nodes and Persistence In Vivo

ABSTRACT Host and bacterial factors that determine whether Salmonella serotypes remain restricted to the gastrointestinal tract or penetrate beyond the mucosa and cause systemic disease remain largely undefined. Here, factors influencing Salmonella host specificity in calves were assessed by characterizing the pathogenesis of different serotypes. Salmonella enterica serotype Dublin was highly virulent intravenously, whereas S. enterica serotype Choleraesuis was moderately virulent. Both serotypes were virulent in calves infected orally. In contrast, S. enterica serotypes Gallinarum and Abortusovis were avirulent by either route. Serotypes Dublin, Gallinarum, and Abortusovis colonized the intestinal tract 24 h after oral inoculation, yet only serotype Dublin was consistently recovered from systemic tissues. Serotypes Dublin and Gallinarum invaded bovine intestines in greater numbers and induced greater enteropathogenic responses than serotypes Choleraesuis and Abortusovis. However, only serotype Dublin was able to persist within the intestinal mucosa, and use of a novel cannulation model demonstrated that serotype Dublin was able to pass through the mesenteric lymph nodes in greater numbers than serotype Gallinarum. Together, these results suggest that initial interactions with the intestinal mucosa do not correlate with host specificity, although persistence within tissues and translocation via efferent lymphatics appear to be crucial for the induction of bovine salmonellosis.

Mutation of waaN Reduces Salmonella enterica Serovar Typhimurium-Induced Enteritis and Net Secretion of Type III Secretion System 1-Dependent Proteins

ABSTRACT Mutation of waaN, a gene involved in lipid A biosynthesis, reduced enteropathogenic responses induced bySalmonella enterica serovar Typhimurium in bovine ligated ileal loops. However, the secretion of key virulence determinants was also reduced, and therefore the reduction in enteropathogenicity cannot be solely attributed to a reduction in biological activity of lipid A.

Plasmid Genes Involved in Virulence in Salmonella

Salmonella virulence plasmids were first recognised in 1970 (Dowman and Meynell, 1970), but it was 10 years before a role in virulence was identified (Jones et al., 1982). A series of papers followed which indicated that several serotypes contained such plasmids and that curing reduced virulence in mice or chickens, which could be restored by reintroduction of the plasmid (Table 1: Chikami et al., 1985; Gulig and Curtiss, 1987; Barrow et al., 1987; Barrow and Lovell; 1988, Kawahara et al. 1988; Hovi et al., 1988). Originally referred to as “cryptic” or serotype-specific, they are now generally and more accurately referred to as virulence plasmids. Although virulence plasmids are generally serotype-specific, there are exceptions. The closely related S. rostock and S. dublin carry an identical plasmid (Platt, D.J., personal communication), and different virulence plasmids are found within both S. enteritidis and S. pullorum (Williamson et al., 1988a). The virulence plasmids range in size from about 54kb to 98kb, and therefore represent up to 2% of the genetic information of the Salmonella genome. So far only 12 serotypes have been found to contain virulence plasmids (Table 1), and within a serotype there are phage-type differences in plasmid carriage (Brown et al., 1986; Woodward et al., 1989). The relative importance of these plasmids for virulence in mice varies in different serotypes (Williamson et al. 1988a). Discussion of the role of the plasmid genes in virulence must also take into account that serotypes without virulence plasmids are not avirulent.