Development and validation of a rabbit model of Pseudomonas aeruginosa non-ventilated pneumonia for preclinical drug development.

Background: New drugs targeting antimicrobial resistant pathogens, including Pseudomonas aeruginosa, have been challenging to evaluate in clinical trials, particularly for the non-ventilated hospital-acquired pneumonia and ventilator-associated pneumonia indications. Development of new antibacterial drugs is facilitated by preclinical animal models that could predict clinical efficacy in patients with these infections. Methods: We report here an FDA-funded study to develop a rabbit model of non-ventilated pneumonia with Pseudomonas aeruginosa by determining the extent to which the natural history of animal disease reproduced human pathophysiology and conducting validation studies to evaluate whether humanized dosing regimens of two antibiotics, meropenem and tobramycin, can halt or reverse disease progression. Results: In a rabbit model of non-ventilated pneumonia, endobronchial challenge with live P. aeruginosa strain 6206, but not with UV-killed Pa6206, caused acute respiratory distress syndrome, as evidenced by acute lung inflammation, pulmonary edema, hemorrhage, severe hypoxemia, hyperlactatemia, neutropenia, thrombocytopenia, and hypoglycemia, which preceded respiratory failure and death. Pa6206 increased >100-fold in the lungs and then disseminated from there to infect distal organs, including spleen and kidneys. At 5 h post-infection, 67% of Pa6206-challenged rabbits had PaO2 <60 mmHg, corresponding to a clinical cut-off when oxygen therapy would be required. When administered at 5 h post-infection, humanized dosing regimens of tobramycin and meropenem reduced mortality to 17-33%, compared to 100% for saline-treated rabbits (P<0.001 by log-rank tests). For meropenem which exhibits time-dependent bactericidal activity, rabbits treated with a humanized meropenem dosing regimen of 80 mg/kg q2h for 24 h achieved 100% T>MIC, resulting in 75% microbiological clearance rate of Pa6206 from the lungs. For tobramycin which exhibits concentration-dependent killing, rabbits treated with a humanized tobramycin dosing regimen of 8 mg/kg q8h for 24 h achieved Cmax/MIC of 9.8 ± 1.4 at 60 min post-dose, resulting in 50% lung microbiological clearance rate. In contrast, rabbits treated with a single tobramycin dose of 2.5 mg/kg had Cmax/MIC of 7.8 ± 0.8 and 8% (1/12) microbiological clearance rate, indicating that this rabbit model can detect dose-response effects. Conclusion: The rabbit model may be used to help predict clinical efficacy of new antibacterial drugs for the treatment of non-ventilated P. aeruginosa pneumonia.

Cytocompatibility and antimicrobial activity of a novel endodontic irrigant combining citric acid and chlorhexidine.

OBJECTIVES: This study aimed to evaluate the antibacterial ability and cytocompatibility of a new irrigant solution for endodontic treatment composed of 10% citric acid (CA) and 1% chlorhexidine (CHX). METHODS: Thirty-five extracted single-canal human teeth were selected and de-crowned. Canal systems (n = 7/group) were infected with Enterococcus faecalis for 4 weeks and subject to irrigation with 1% CHX; 10% CA; irrigating solution 10% CA associated with 1% CHX (CACHX); 2.5% NaOCl or sterile water (control). Microbiological samples were collected immediately and 18 h after irrigation (enriched samples). The canals were filled with culture medium post irrigation to verify the bacterial presence/absence qualitatively and quantitatively through colony counting (log10 CFU/mL). A multiparametric assay was performed after exposure of human periodontal ligament fibroblasts (HPdLF) to the test solutions. The Kruskal-Wallis test with Dunn´s post-test and Fisher's exact test were employed at the 95% confidence level to compare differences among groups. RESULTS: All tested solutions were cytocompatible with human periodontal ligament fibroblasts. No difference was observed on antibacterial activity between 1% CHX, 10% CA, CACHX and 2.5% NaOCl (p > 0.05). Eighteen hours after irrigation, CACHX samples were the only that did not present E. faecalis in the root canal system. CONCLUSIONS: The demonstrated good in vitro biocompatibility and elimination of E. faecalis suggest a potential use of 10% CA associated with 1% CHX as a solution for microbiological control during endodontic treatment. CLINICAL RELEVANCE: Irrigants play an essential role during endodontic therapy. This irrigating solution, based on the association of 10% citric acid with 1% chlorhexidine, seems viable for clinical procedures.

Antimicrobial Photodynamic Therapy Associated with Conventional Endodontic Treatment: A Clinical and Molecular Microbiological Study.

This study evaluated antimicrobial photodynamic therapy (aPDT) as an adjunct to endodontic treatment. Ten uniradicular teeth (control group (CG) = 4 (2 and test group (TG) = 6) with primary endodontic infections, from both genders, between 17 and 65 years old, were analyzed. Microbiological samples were collected before and after chemical-mechanical instrumentation (CMI), after aPDT (for the TG), and after the removal of the temporary restorations (second session). In TG, the aPDT was performed with 100 µg mL-1 methylene blue and irradiated with low power laser (InGaAIP, 660 nm; 100 mW; 40 s) with a fiber-coupled optical laser. Another irradiation (3 J; 30 s; spot size of 3 mm2 ) was performed in the gingiva close to the apical foramen. The PCR was performed, after previous whole-genome amplification, for Enterococcus faecalis, Candida genus and Bacteria domain. For TG, a positive tooth for Candida spp. before of the CMI presented negative results in subsequent samples. Additionally, E. faecalis species was present in four samples before CMI, two after CMI, in one after the aPDT and was not detected at the second session. aPDT may be an effective adjunct therapy, resulting in a reduction (P = 0.0286) of the incidence of E. faecalis before root canal obturation.