Investigation of biofilms associated with chronic otitis media with effusion and adenoids hypertrophy

Ph. D. ThesisChronic otitis media with effusion (COME) is the most common cause of acquired hearing loss in young children. Bacterial biofilm is an important contributor to the aetiopathogenesis of COME, although conventional culture generally recovers few microorganisms from the middle ear. Extracellular DNA (eDNA) is a key structural component within the matrix of many microbial biofilms including those associated with COME. The aims of this study were to characterise microbial populations associated with COME, and to explore the efficacy of the DNase, NucB, to control in vitro biofilms associated with COME. Methods were established to culture biofilms in vitro and to challenge with NucB. NucB efficiently disrupted biofilms of clinically isolated Staphylococcus aureus strains either alone or in combination with the antibiotic Co-amoxiclav. Concentrations of NucB 100-fold higher than those required for biofilm inhibition had no toxic effects on human epithelial cells. Twenty-seven bacterial species were isolated from middle ear effusion fluids (MEEFs) of 34 patients with COME. Culture-positive MEEFs were increased two-fold following optimisation of the culture methods. Microbiome analysis of MEEFs by 16S rDNA sequencing identified the majority of the cultured species and several additional species. Similar species were also detected by 16S rRNA gene sequencing of microbial DNA from adenoids. The ability of 23 bacterial isolates from MEEF to form biofilm was assessed. Twenty strains formed biofilms, and 16 of these were sensitive to NucB. Imaging analysis showed significant structural alterations in biofilms of the selected COME isolates after NucB treatment. In conclusion, this study has provided further insights into the microbiology of middle ear infections and has shown that many bacteria from this environment are capable of forming biofilms. NucB alone or in combination with antibiotics may potentially be a potent and safe agent to control biofilm-associated conditions including COME.Iraqi Ministry of Higher Education and Scientific Research, Kirkuk Universit