Showing papers by "Timothy S. Wallis published in 2002"

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

Abstract: 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 J7742 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

Efa1 influences colonization of the bovine intestine by shiga toxin-producing Escherichia coli serotypes O5 and O111.

Abstract: Shiga toxin-producing Escherichia coli (STEC) comprises a broad group of bacteria, some of which cause attaching and effacing (AE) lesions and enteritis in animals and humans. Non-O157 STEC serotypes contain a gene (efa1) that mediates attachment to cultured epithelial cells. An almost-identical gene in enteropathogenic E. coli (lifA) encodes lymphostatin, which inhibits the proliferation of mitogen-activated lymphocytes and the synthesis of proinflammatory cytokines. We have investigated the role of the efa1 gene in colonization of 4- and 11-day-old conventional calves by STEC serotypes O5 and O111. Our findings show that Efa1 is required for efficient colonization of the bovine intestinal tract by STEC, since efa1 deletion and insertion mutants were shed in the feces in significantly lower numbers. In addition, efa1 mutations dramatically reduced the number of bacteria associated with the intestinal epithelium. Expression and secretion of locus for enterocyte effacement-encoded type III secreted proteins that are required for adhesion and AE-lesion formation were impaired by mutation of efa1 in STEC but not by mutation of lifA in enteropathogenic E. coli. However, STEC efa1 mutants retain the ability to nucleate filamentous actin under sites of bacterial attachment to cultured eukaryotic cells. Efa1 is only the second STEC factor shown to influence carriage of the bacteria in the bovine intestine. Our data may have implications for strategies to reduce the prevalence of STEC in cattle.

REVIEW ARTICLE Options for the control of enterohaemorrhagic Escherichia coli in ruminants

Abstract: 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.

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.

The role of two periplasmic copper- and zinc-cofactored superoxide dismutases in the virulence of Salmonella choleraesuis.

Abstract: Periplasmic copper- and zinc-cofactored superoxide dismutases ([Cu,Zn]-SODs, SodC) of several Gram-negative pathogens can protect against superoxide-radical-mediated host defences, and thus contribute to virulence. This role has been previously defined for one [Cu,Zn]-SOD in various Salmonella serovars. Following the recent discovery of a second periplasmic [Cu,Zn]-SOD in Salmonella, the effect of knockout mutations in one or both of the original sodC-1 and the new sodC-2 on the virulence of the porcine pathogen Salmonella choleraesuis is investigated here. In comparison to wild-type, while sodC mutants – whether single or double – showed no impairment in growth, they all showed equally enhanced sensitivity to superoxide and a dramatically increased sensitivity to the combination of superoxide and nitric oxide in vitro. This observation had its correlate in experimental infection both ex vivo and in vivo. Mutation of sodC significantly impaired survival of S. choleraesuis in interferon γ-stimulated murine macrophages compared to wild-type organisms, and all S. choleraesuis sodC mutants persisted in significantly lower numbers than wild-type in BALB/c (Ity s) and C3H/HeN (Ity r) mice after experimental infection, but in no experimental system were sodC-1 sodC-2 double mutants more attenuated than either single mutant. These data suggest that both [Cu,Zn]-SODs are needed to protect bacterial periplasmic or membrane components. While SodC plays a role in S. choleraesuis virulence, the data presented here suggest that this is through overcoming a threshold effect, probably achieved by acquisition of sodC-1 on a bacteriophage. Loss of either sodC gene confers maximum vulnerability to superoxide on S. choleraesuis.

Intimin, tir, and shiga toxin 1 do not influence enteropathogenic responses to shiga toxin-producing Escherichia coli in bovine ligated intestinal loops.

Abstract: Shiga toxin-producing Escherchia coli (STEC) comprises a group of attaching and effacing (A/E) enteric pathogens of animals and humans. Natural and experimental infection of calves with STEC may result in acute enteritis or subclinical infection, depending on serotype- and host-specific factors. To quantify intestinal secretory and inflammatory responses to STEC in the bovine intestine, serotypes that are associated with human disease (O103:H2 and O157:H7) were introduced into ligated mid-ileal loops in gnotobiotic and conventional calves, and fluid accumulation and recruitment of radiolabeled neutrophils were measured after 12 h. STEC serotype O103:H2, but not serotype O157:H7, elicited strong enteropathogenic responses. To determine if the inflammatory response to STEC O103:H2 in calves requires Shiga toxin 1 or intimate bacterial attachment to the intestinal epithelium, defined mutations were made in the stx1, eae, and tir genes. Our data indicate that some STEC induce intestinal inflammatory responses in calves by a mechanism that is independent of A/E-lesion formation, intimin, or Shiga toxin 1. This may have implications for strategies to reduce STEC carriage in cattle.