Low-Dose Nitric Oxide as Targeted Anti-biofilm Adjunctive Therapy to Treat Chronic Pseudomonas aeruginosa Infection in Cystic Fibrosis.

Despite aggressive antibiotic therapy, bronchopulmonary colonization by Pseudomonas aeruginosa causes persistent morbidity and mortality in cystic fibrosis (CF). Chronic P. aeruginosa infection in the CF lung is associated with structured, antibiotic-tolerant bacterial aggregates known as biofilms. We have demonstrated the effects of non-bactericidal, low-dose nitric oxide (NO), a signaling molecule that induces biofilm dispersal, as a novel adjunctive therapy for P. aeruginosa biofilm infection in CF in an ex vivo model and a proof-of-concept double-blind clinical trial. Submicromolar NO concentrations alone caused disruption of biofilms within ex vivo CF sputum and a statistically significant decrease in ex vivo biofilm tolerance to tobramycin and tobramycin combined with ceftazidime. In the 12-patient randomized clinical trial, 10 ppm NO inhalation caused significant reduction in P. aeruginosa biofilm aggregates compared with placebo across 7 days of treatment. Our results suggest a benefit of using low-dose NO as adjunctive therapy to enhance the efficacy of antibiotics used to treat acute P. aeruginosa exacerbations in CF. Strategies to induce the disruption of biofilms have the potential to overcome biofilm-associated antibiotic tolerance in CF and other biofilm-related diseases.

A quantitative analysis of Ureaplasma urealyticum and Ureaplasma parvum compared with host immune response in preterm neonates at risk of developing bronchopulmonary dysplasia.

Multiplex, real-time PCR for the identification of Ureaplasma urealyticum and Ureaplasma parvum was performed on nucleic acids extracted from sequential endotracheal aspirates obtained from preterm neonates born at <29 weeks of gestation and ventilated for more than 48 h admitted to two level 3 neonatal intensive care units. Specimens were obtained shortly after birth and sequentially up until extubation. One hundred fifty-two specimens (93.8%) contained material suitable for analysis. Ureaplasma spp. were identified in 5 of 13 neonates studied. In most cases, the DNA load of the detected Ureaplasma species was low and decreased over time. In addition, changes in detectable Ureaplasma species DNA did not relate to changes in the inflammatory marker C-reactive protein (CRP) or respiratory status. All but two blood samples obtained at times of suspected sepsis were culture positive for other microorganisms; the species cultured were typically coagulase-negative staphylococci and were associated with increased levels of CRP (>10 mg/liter). This study was limited by the small number of patients examined and does not have the power to support or contradict the hypothesis that postnatal lung infection with Ureaplasma parvum is causally related to bronchopulmonary dysplasia (BPD) or adverse respiratory outcomes after preterm birth. However, in this study, increases in CRP levels were not associated with patients in whom Ureaplasma parvum was detected, in contrast to the detection of other bacterial species.

Characterization of bacterial community diversity in chronic rhinosinusitis infections using novel culture-independent techniques.

BACKGROUND: Chronic rhinosinusitis (CRS) with or without polyps is a common chronic upper airway condition of multifactorial origin. Fundamental to effective treatment of any infection is the ability to accurately characterize the underlying cause. Many studies have shown that only a small fraction of the total range of bacterial species present in CRS is detected through conventional culture-dependent techniques. Consequently, culture data are often unrepresentative of the true diversity of the microbial community within the sample. These drawbacks, along with the length of time required to complete the analysis, strongly support the development of alternative means of assessing which bacterial species are present. As such, molecular microbiological approaches that assess the content of clinical samples in a culture-independent manner could significantly enhance the range and quality of data obtained routinely from such samples. We aimed to characterize the bacterial diversity present in tissue and mucus samples taken from the CRS setting using molecular nonculture-dependent techniques. METHODS: Through 16S ribosomal RNA (rRNA) gene clone sequencing and terminal restriction fragment length polymorphism (T-RFLP) analysis, the bacteria present in 70 clinical samples from 43 CRS patients undergoing endoscopic sinus surgery were characterized. RESULTS: Bacterial T-RFLP profiles were generated for 70 of 73 samples and a total of 48 separate bands were detected. Species belonging to 34 genera were identified as present by clone sequence analysis. Of the species detected, those within the genera Pseudomonas, Citrobacter, Haemophilus, Propionibacterium, Staphylococcus, and Streptococcus were found numerically dominant, with Pseudomonas aeruginosa the most frequently detected species. CONCLUSION: This study has validated the use of the culture-independent technique T-RFLP in sinonasal samples. Preliminary characterization of the microbial diversity in CRS suggests a complex range of common and novel bacterial species within the upper airway in CRS, providing further evidence for the polymicrobial etiology of CRS.