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Journal Article

Animals; Cattle; Escherichia coli; Female; Genitalia, Female; Optical Phenomena; Photons; Xen14

The objectives of this study were to (1) characterize the photonic properties of Escherichia coli-Xen14 and (2) conduct photonic imaging of E. coli-Xen14 within bovine reproductive tract segments (RTS) ex vivo (Bos indicus). E. coli-Xen14 was grown for 24h in Luria Bertani medium (LB), with or without kanamycin (KAN). Every 24h, for an 8-d interval, inoculums were imaged and photonic emissions (PE) collected. Inoculums were subcultured and plated daily to determine the colony forming units (CFU) and ratio of photon emitters to nonemitters. In the second objective, abattoir-derived bovine reproductive tracts (n=9) were separated into posterior and anterior vagina, cervix, uterine body, and uterine horns. Two concentrations (3.2x10(8) and 3.2x10(6) CFU/200microL for relative [High] and [Low], respectively) of E. coli-Xen14 were placed in translucent tubes for detection of PE through RTS. The CFU did not differ (P=0.31) over time with or without KAN presence; they remained stable with 99.93% and 99.98% photon emitters, respectively. However, PE were lower (P<0.0001) in cultures containing KAN than in those containing no KAN (629.8+/-117.7 vs. 3012.0+/-423.5 relative lights units per second [RLU/sec], respectively). On average, the percentage of PE between RTS, for both concentrations, was higher (P<0.05) in the uterine body. In summary, E. coli-Xen14 remained stable with respect to the proportions of photon emitters with or without KAN (used to selectively culture E. coli-Xen14). However, KAN presence suppressed photonic activity. The ability to detect PE through various segments of the reproductive tract demonstrated the feasibility of monitoring the presence of E. coli-Xen14 in the bovine reproductive tract ex vivo.

Journal Article

Colony Count, Microbial; Cuspid; Dental Pulp Cavity; Disinfectants; Drug Combinations; Genetic Engineering; Humans; Hydrogen peroxide; Incisor; Luminescent Measurements; Luminescent Proteins; Maxilla; Pseudomonas aeruginosa; Root Canal Irrigants; Root Canal Preparation; Sensitivity and Specificity; Sodium Hypochlorite; Xen5

Microbial infection plays an important role in the development of pulp necrosis and formation of periapical lesions. In vitro and in vivo research in this field, traditionally microbiological culture methods using paper point sampling and quantitative culture, faces difficulties in completely removing bacteria from the root canal system and analyzing sequential procedures. This study employed genetically engineered bioluminescent bacteria and a light-sensitive imaging system to allow real-time visualization of the infection. Ten extracted teeth incubated with P. aeruginosa were treated by mechanical instrumentation with K-files (#30 K-file, #35 K-file and #40 K-file) and chemical irrigation with sodium hypochlorite and hydrogen peroxide. Irrigation alone reduced the contamination in 18%; the first chemomechanical sequence (instrumentation with a #30 K-file + irrigation) provided 41% of reduction; the second sequence (#35 K-file + irrigation) achieved 62%; and the complete therapy (#30 K-file + #35 K-file + #40 K-file + irrigation) achieved 93% of bacterial reduction. These results suggest that the endodontic treatment is dependent on the association of a chemical and mechanical approaches and that root canal enlargement improves bacterial reduction probably because the irrigation has more access to the apical third.

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

Xen5, Xen 5, Pseudomonas aeruginosa Xen 5, Anti-Bacterial Agents/administration & dosage/*pharmacology/therapeutic; use; Antimicrobial Cationic Peptides/chemistry; Biofilms/drug effects; Cholic Acid/chemistry; Cystic Fibrosis/microbiology; Hemolysis/drug effects; Humans; *Poloxamer; Pseudomonas Infections/drug therapy; Pseudomonas aeruginosa/drug effects/growth & development; Skin Diseases, Bacterial/drug therapy; Staphylococcus aureus/drug effects; Steroids/administration & dosage/*pharmacology/therapeutic use; *Surface-Active Agents

AIMS: Ceragenin CSA-13 is a synthetic mimic of cationic antibacterial peptides, with facial amphiphilic morphology reproduced using a cholic acid scaffold. Previous data have shown that this molecule displays broad-spectrum antibacterial activity, which decreases in the presence of blood plasma. However, at higher concentrations, CSA-13 can cause lysis of erythrocytes. This study was designed to assess in vitro antibacterial and haemolytic activity of CSA-13 in the presence of pluronic F-127. METHODS AND RESULTS: CSA-13 bactericidal activity against clinical strains of bacteria associated with topical infections and in an experimental setting relevant to their pathophysiological environment, such as various epithelial tissue fluids and the airway sputum of patients suffering from cystic fibrosis (CF), was evaluated using minimum inhibitory and minimum bactericidal concentration (MIC/MBC) measurements and bacterial killing assays. We found that in the presence of pluronic F-127, CSA-13 antibacterial activity was only slightly decreased, but CSA-13 haemolytic activity was significantly inhibited. CSA-13 exhibits bacterial killing activity against clinical isolates of Staphylococcus aureus, including methicillin-resistant strains, Pseudomonas aeruginosa present in CF sputa, and biofilms formed by different Gram (+) and Gram (-) bacteria. CSA-13 bactericidal action is partially compromised in the presence of plasma, but is maintained in ascites, cerebrospinal fluid, saliva, and bronchoalveolar lavage fluid. The synergistic action of CSA-13, determined by the use of a standard checkerboard assay, reveals an increase in CSA-13 antibacterial activity in the presence of host defence molecules such as the cathelicidin LL-37 peptide, lysozyme, lactoferrin and secretory phospholipase A (sPLA). CONCLUSION: These results suggest that CSA-13 may be useful to prevent and treat topical infection. SIGNIFICANCE AND IMPACT OF THE STUDY: Combined application of CSA-13 with pluronic F-127 may be beneficial by reducing CSA-13 toxicity.

Journal Article

Aminolevulinic Acid; Animals; Biofilms; Bioware; Cell Survival; Disease Models, Animal; Drug Evaluation, Preclinical; Female; Implants, Experimental; Light; Luminescent Measurements; Methylene Blue; Osteomyelitis; Photochemotherapy; Photosensitizing Agents; Rats; Rats, Sprague-Dawley; Staphylococcus aureus; Xen29

Osteomyelitis can lead to severe morbidity and even death resulting from an acute or chronic inflammation of the bone and contiguous structures due to fungal or bacterial infection. Incidence approximates 1 in 1000 neonates and 1 in 5000 children in the United States annually and increases up to 0.36% and 16% in adults with diabetes or sickle cell anaemia, respectively. Current regimens of treatment include antibiotics and/or surgery. However, the increasing number of antibiotic resistant pathogens suggests that alternate strategies are required. We are investigating photodynamic therapy (PDT) as one such alternate treatment for osteomyelitis using a bioluminescent strain of biofilm-producing staphylococcus aureus (S. aureus) grown onto kirschner wires (K-wire). S. aureus-coated K-wires were exposed to methylene blue (MB) or 5-aminolevulinic acid (ALA)-mediated PDT either in vitro or following implant into the tibial medullary cavity of Sprague-Dawley rats. The progression of S. aureus biofilm was monitored non-invasively using bioluminescence and expressed as a percentage of the signal for each sample immediately prior to treatment. S. aureus infections were subject to PDT 10 days post inoculation. Treatment comprised administration of ALA (300 mg kg(-1)) intraperitoneally followed 4 h later by light (635 +/- 10 nm; 75 J cm(-2)) delivered transcutaneously via an optical fiber placed onto the tibia and resulted in significant delay in bacterial growth. In vitro, MB and ALA displayed similar cell kill with > or =4 log(10) cell kill. In vivo, ALA-mediated PDT inhibited biofilm implants in bone. These results confirm that MB or ALA-mediated PDT have potential to treat S. aureus cultures grown in vitro or in vivo using an animal model of osteomyelitis.