Journal Article

Xen14, Xen 14, E. coli Xen14, IVIS, Animals; Bacteroides fragilis/*isolation & purification; Diagnostic Imaging/methods; Disease Progression; Escherichia coli/*isolation & purification; Luciferases, Bacterial/*diagnostic use; Luminescent Agents/*diagnostic use; Male; Peritoneal Lavage; Peritonitis/*microbiology/pathology/therapy; Rats; Rats, Wistar

BACKGROUND: Bacterial peritonitis is a life-threatening abdominal infection associated with high morbidity and mortality. The rat is a popular animal model for studying peritonitis and its treatment, but longitudinal monitoring of the progression of peritonitis in live animals has been impossible until now and thus required a large number of animals. Our objective was to develop a noninvasive in vivo imaging technique to monitor the spatiotemporal spread of bacterial peritonitis. METHODS: Peritonitis was induced in 8 immunocompetent male Wistar rats by placing fibrin clots containing 5x10(8) cells of both Bacteroides fragilis (American Type Tissue Culture [ATCC)] 25,285 and bioluminescent Escherichia coli Xen14. After 1 or 2 days, infected clots were removed and open abdomen lavage was performed. In vivo bioluminescent imaging was used to monitor the spread of peritonitis. RESULTS: Bioluminescent in vivo imaging showed an increase in the area of spread, and the number of E. coli tripled into the rat's abdominal cavity on day 1 after clot insertion; however, on day 2, encapsulation of the clot confined bacterial spread. Bioluminescent E. coli respread over the peritoneal cavity after lavage; within 10 days, however, in vivo imaging showed a decrease of 3-4 orders of magnitude in bacterial load. CONCLUSION: Bioluminescent in vivo imaging can be effectively used to monitor the spatiotemporal behavior of the peritonitis during 3 different stages of the disease process: initiation, treatment, and follow-up. Imaging allows researchers to repeatedly image the same animal, thereby reducing variability and providing greater confidence in determining treatment efficacies for therapeutic interventions using a small number of animals.

Journal Article

Animals; Escherichia coli/immunology; Escherichia coli Infections/enzymology/immunology/*prevention & control; Extracellular Space/genetics/*immunology/metabolism; Glutathione Reductase/deficiency/genetics/*physiology; Humans; Mice; Mice, Inbred C3H; Mice, Knockout; Neutrophils/*immunology/*metabolism/microbiology; Oxidative Stress/genetics/*immunology; Phagocytosis/genetics/*immunology; Staphylococcal Infections/enzymology/immunology/*prevention & control; Staphylococcus aureus/immunology

Glutathione reductase (Gsr) catalyzes the reduction of glutathione disulfide to glutathione, which plays an important role in the bactericidal function of phagocytes. Because Gsr has been implicated in the oxidative burst in human neutrophils and is abundantly expressed in the lymphoid system, we hypothesized that Gsr-deficient mice would exhibit marked defects during the immune response against bacterial challenge. We report in this study that Gsr-null mice exhibited enhanced susceptibility to Escherichia coli challenge, indicated by dramatically increased bacterial burden, cytokine storm, striking histological abnormalities, and substantially elevated mortality. Additionally, Gsr-null mice exhibited elevated sensitivity to Staphylococcus aureus. Examination of the bactericidal functions of the neutrophils from Gsr-deficient mice in vitro revealed impaired phagocytosis and defective bacterial killing activities. Although Gsr catalyzes the regeneration of glutathione, a major cellular antioxidant, Gsr-deficient neutrophils paradoxically produced far less reactive oxygen species upon activation both ex vivo and in vivo. Unlike wild-type neutrophils that exhibited a sustained oxidative burst upon stimulation with phorbol ester and fMLP, Gsr-deficient neutrophils displayed a very transient oxidative burst that abruptly ceased shortly after stimulation. Likewise, Gsr-deficient neutrophils also exhibited an attenuated oxidative burst upon encountering E. coli. Biochemical analysis revealed that the hexose monophosphate shunt was compromised in Gsr-deficient neutrophils. Moreover, Gsr-deficient neutrophils displayed a marked impairment in the formation of neutrophil extracellular traps, a bactericidal mechanism that operates after neutrophil death. Thus, Gsr-mediated redox regulation is crucial for bacterial clearance during host defense against massive bacterial challenge.

Journal Article

Xen10, Xen 10, Streptococcus pneumoniae Xen10, IVIS, Animals; Bacterial Adhesion; Bacterial Processes; Bacterial Proteins/*physiology; *Carrier State; Colony Count, Microbial; Female; Host-Pathogen Interactions; Lung/microbiology; Meningitis, Pneumococcal/microbiology; Mice; Models, Animal; Mutation; Nasopharynx/*microbiology; Pneumonia, Pneumococcal/complications; Sepsis/*microbiology; Streptococcus pneumoniae/*pathogenicity; Virulence Factors/physiology

Summary The pneumococcal cell surface protein PavA is a virulence factor associated with adherence and invasion in vitro. In this study we show in vivo that PavA is necessary for Streptococcus pneumoniae D39 colonization of the murine upper respiratory tract in a long-term carriage model, with PavA-deficient pneumococci being quickly cleared from nasopharyngeal tissue. In a pneumonia model, pavA mutants were not cleared from the lungs of infected mice and persisted to cause chronic infection, whereas wild-type pneumococci caused systemic infection. Hence, under the experimental conditions, PavA-deficient pneumococci appeared to be unable to seed from lung tissue into blood, although they survived in blood when administered intravenously. In a meningitis model of infection, levels of PavA-deficient pneumococci in blood and brain following intercisternal injection were significantly lower than wild type. Taken collectively these results suggest that PavA is involved in successful colonization of mucosal surfaces and in translocation of pneumococci across host barriers. Pneumococcal sepsis is a major cause of mortality worldwide so identification of factors such as PavA that are necessary for carriage and for translocation from tissue to blood is of clinical and therapeutic importance.

Journal Article

Xen36, Xen 36, Staphylococcus aureus Xen36, IVIS

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Journal Article

Xen36, Xen 36, Staphylococcus aureus Xen36, IVIS, Animals; Mice; Mice, Inbred C57BL; Microbial Sensitivity Tests; Nisin/*pharmacology; Peritoneal Cavity/*microbiology; Staphylococcal Infections/*prevention & control; Staphylococcus aureus/*drug effects/growth & development

AIMS: To determine the ability of nisin F to control systematic infection caused by Staphylococcus aureus, using C57BL/6 mice as a model. METHODS AND RESULTS: Twelve mice were intraperitoneally injected with 1 x 10(8) viable cells of Staph. aureus Xen 36 containing the modified Photorhabdus luminescence luxABCDE operon on plasmid pAUL-A Tn4001. After 4 h, six mice were intraperitoneally injected with 640 arbitrary units (AU) nisin F, and six were injected with sterile saline. Six mice, not infected with Staph. aureus, were treated with nisin F, and six not infected were left untreated. The viability of Staph. aureus Xen 36 was monitored over 48 h by recording photon emission levels. Nisin F suppressed Staph. aureus for 15 min in vivo. No abnormalities were recorded in blood analyses and internal organs of mice treated with nisin F. CONCLUSIONS: Nisin F suppressed the growth of Staph. aureus in the peritoneal cavity for at least 15 min. Re-emergence of Staph. aureus bioluminescence over the next 44 h suggests that nisin F was inactivated, most probably by proteolytic enzymes. SIGNIFICANCE AND IMPACT OF THE STUDY: A single dosage of nisin F administered in the peritoneal cavity controlled the growth of Staph. aureus for at least 15 min in vivo.