Experimental Shigella Infections: Characteristics of a Fatal Infection Produced in Guinea Pigs

Formal, Samuel B.; Dammin, Gustave J.; LaBrec, E. H.; Schneider, Herman

doi:10.1128/jb.75.5.604-610.1958

Cited by 170 publications

(59 citation statements)

References 3 publications

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“…Wild-type (WT) S. flexneri corresponds to the serotype 2a 2457T strain originally isolated in 1954. 48 The S. flexneri spa47 null strain was engineered by Abdelmounaa€ ım Allaoui as described in Jouihri et al 21 E. coli strains and 2X ligation mix were from Novagen (Madison, WI). Restriction enzymes, the pTYB21 protein expression plasmid, PCR buffer, Phusion High-Fidelity polymerase, and chitin resin were purchased from New England Biolabs (Ipswich, MA).…”

Section: Methodsmentioning

confidence: 99%

Spa47 is an oligomerization‐activated type three secretion system (T3SS) ATPase from Shigella flexneri

et al. 2016

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Gram-negative pathogens often use conserved type three secretion systems (T3SS) for virulence. The Shigella type three secretion apparatus (T3SA) penetrates the host cell membrane and provides a unidirectional conduit for injection of effectors into host cells. The protein Spa47 localizes to the base of the apparatus and is speculated to be an ATPase that provides the energy for T3SA formation and secretion. Here, we developed an expression and purification protocol, producing active Spa47 and providing the first direct evidence that Spa47 is a bona fide ATPase. Additionally, size exclusion chromatography and analytical ultracentrifugation identified multiple oligomeric species of Spa47 with the largest greater than 8 fold more active for ATP hydrolysis than the monomer. An ATPase inactive Spa47 point mutant was then engineered by targeting a conserved Lysine within the predicted Walker A motif of Spa47. Interestingly, the mutant maintained a similar oligomerization pattern as active Spa47, but was unable to restore invasion phenotype when used to complement a spa47 null S. flexneri strain. Together, these results identify Spa47 as a Shigella T3SS ATPase and suggest that its activity is linked to oligomerization, perhaps as a regulatory mechanism as seen in some related pathogens. Additionally, Spa47 catalyzed ATP hydrolysis appears to be essential for host cell invasion, providing a strong platform for additional studies dissecting its role in virulence and providing an attractive target for anti-infective agents.

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Section: Methodsmentioning

confidence: 99%

Spa47 is an oligomerization‐activated type three secretion system (T3SS) ATPase from Shigella flexneri

et al. 2016

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“…Indeed, Shigella invades the corneal epithelium and spreads to contiguous cells, with the more virulent strains causing ulcerative keratoconjunctivitis. 30,31 Despite its usefulness for immunogenicity studies and protective efficacy of Shigella vaccine candidates, a detailed analysis of host immune responses remains difficult in this model.…”

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confidence: 99%

Shigella’s ways of manipulating the host intestinal innate and adaptive immune system: a tool box for survival?

2007

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Shigella, a Gram-negative invasive enteropathogenic bacterium, causes the rupture, invasion and inflammatory destruction of the human colonic epithelium. This complex and aggressive process accounts for the symptoms of bacillary dysentery. The so-called invasive phenotype of Shigella is linked to expression of a type III secretory system (TTSS) injecting effector proteins into the epithelial cell membrane and cytoplasm, thereby inducing local but massive changes in the cell cytoskeleton that lead to bacterial internalization into non-phagocytic intestinal epithelial cells. The invasive phenotype also accounts for the potent proinflammatory capacity of the microorganism. Recent evidence indicates that a large part of the mucosal inflammation is initiated by intracellular sensing of bacterial peptidoglycan by cytosolic leucine-rich receptors of the NOD family, particularly NOD1, in epithelial cells. This causes activation of the nuclear factor kappa B and c-JunNH 2 -terminal-kinase pathways, with interleukin-8 appearing as a major chemokine mediating the inflammatory burst that is dominated by massive infiltration of the mucosa by polymorphonuclear leukocytes. Not unexpectedly, this inflammatory response, which is likely to be very harmful for the invading microbe, is regulated by the bacterium itself. A group of proteins encoded by Shigella, which are injected into target cells by the TTSS, has been recently recognized as a family of potent regulators of the innate immune response. These enzymes target key cellular functions that are essential in triggering the inflammatory response, and more generally defense responses of the intestinal mucosa. This review focuses on the mechanisms employed by Shigella to manipulate the host innate response in order to escape early bacterial killing, thus ensuring establishment of its infectious process. The escape strategies, the possible direct effect of Shigella on B and T lymphocytes, their impact on the development of adaptive immunity, and how they may help explain the limited protection induced by natural infection are discussed.

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“…The keratoconjunctivitis assay is regarded as the gold standard for protective immunity in guinea-pigs (Sereny, 1955), but its limitation lies in the difficulty in the quantification of an inflammatory response and the irrelevance of the target organ. Guinea-pigs that were administered wildtype S. flexneri 2a and treated with opium post 4 days starvation developed fatal enteric infections (Formal et al, 1958). Because of the fatal effects at a relatively early stage of infection, this model was not ideal for the purpose of screening vaccine candidates.…”

Section: Discussionmentioning

confidence: 99%

Development of a new guinea-pig model of shigellosis

Barman

Saha

Ramamurthy

et al. 2011

FEMS Immunol Med Microbiol

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Shigellosis is a major form of bacillary dysentery caused by Shigella spp. To date, there is no suitable animal model to evaluate the protective efficacy of vaccine candidates against this pathogen. Here, we describe a successful experimental shigellosis in the guinea-pig model, which has shown the characteristic features of human shigellosis. This model yielded reproducible results without any preparatory treatment besides cecal ligation. In this study, guinea-pigs were discretely infected with virulent Shigella dysenteriae type 1 and Shigella flexneri type 2a into the cecocolic junction after ligation of the distal cecum. All the experimental animals lost ∼10% of their body weight and developed typical dysentery within 24-h postinfection. In the histological analysis, distal colon showed edema, hemorrhage, exudation and inflammatory infiltrations in the lamina propria. Orally immunized animals with heat-killed S. dysenteriae type 1 and S. flexneri type 2a strains showed high levels of serum immunoglobulin G (IgG) and mucosal IgA antibodies and conferred significant homologous protective immunity against subsequent challenges with the live strains. The direct administration of shigellae into the cecocolic junction induces acute inflammation, making this animal model useful for assessing shigellosis and evaluating the protective immunity of Shigella vaccine candidates.

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Experimental Shigella Infections: Characteristics of a Fatal Infection Produced in Guinea Pigs

Cited by 170 publications

References 3 publications

Spa47 is an oligomerization‐activated type three secretion system (T3SS) ATPase from Shigella flexneri

Spa47 is an oligomerization‐activated type three secretion system (T3SS) ATPase from Shigella flexneri

Shigella’s ways of manipulating the host intestinal innate and adaptive immune system: a tool box for survival?

Development of a new guinea-pig model of shigellosis

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