Biofilms of pathogenic bacteria are present on the middle ear mucosa of children with chronic otitis media (COM) and may contribute to the persistence of pathogens and the recalcitrance of COM to antibiotic treatment. Controlled studies indicate that adenoidectomy is effective in the treatment of COM, suggesting that the adenoids may act as a reservoir for COM pathogens. To investigate the bacterial community in the adenoid, samples were obtained from 35 children undergoing adenoidectomy for chronic OM or obstructive sleep apnea. We used a novel, culture-independent molecular diagnostic methodology, followed by confocal microscopy, to investigate the in situ distribution and organization of pathogens in the adenoids to determine whether pathogenic bacteria exhibited criteria characteristic of biofilms. The Ibis T5000 Universal Biosensor System was used to interrogate the extent of the microbial diversity within adenoid biopsy specimens. Using a suite of 16 broad-range bacterial primers, we demonstrated that adenoids from both diagnostic groups were colonized with polymicrobial biofilms. Haemophilus influenzae was present in more adenoids from the COM group (P ‫؍‬ 0.005), but there was no significant difference between the two patient groups for Streptococcus pneumoniae or Staphylococcus aureus. Fluorescence in situ hybridization, lectin binding, and the use of antibodies specific for host epithelial cells demonstrated that pathogens were aggregated, surrounded by a carbohydrate matrix, and localized on and within the epithelial cell surface, which is consistent with criteria for bacterial biofilms.

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Objectives/Hypothesis. Mucin genes MUC2, MUC5AC, and MUC5B have been identified as major gel-forming mucins in the middle ear (ME). This study compared polymorphisms in MUC2, MUC5AC, and MUC5B genes in otitis media (OM) patients and controls.Study Design. Cross-sectional case-control study. Patients age 6 months to 14 years undergoing routine tympanostomy tube insertion for recurrent otitis media (RecOM) or chronic otitis media with effusion (OME) were compared to control patients with no history of OM undergoing an unrelated procedure.Methods. Genomic DNA extracted from peripheral blood samples was analyzed by Southern blots using gene-specific probes to determine sizes of MUC2 and MUC5AC genes. Polymerase chain reaction was used to determine the size of MUC5B genes.Results. Twenty RecOM patients, 20 OME patients, and 40 control patients were analyzed. There were no statistically significant differences in polymorphism expression identified between groups for MUC2 and MUC5B genes. OME patients were more likely than controls or RecOM patients to carry the longer MUC5AC-b allele.Conclusions. Differences in mucin polymorphisms have been demonstrated in other diseases. In otitis media, OME patients are more likely than controls or RecOM to carry the longer MUC5AC-b allele. MUC5AC is a primary innate defense mechanism for ME epithelium, and alterations in protein structure have the potential to affect that defense and predispose patients to disease. This study demonstrates that the properties of post-transcriptionally modified MUC5AC deserve further study, and specific pathogen-host interactions studies should explore the impact of this polymorphism.

Objective Mucin gene 5AC (MUC5AC) has been identified as a major secretory mucin in the middle ear (ME). MUC5AC is fundamentally important in the development of ME mucoid effusions, hearing loss and also provides ME mucosal protection and bacterial clearance. The objective of this study is to compare levels of ME MUC5AC expression in patients with otitis media (OM) to patients without OM. Subjects Patients from 9 months to 7 years undergoing routine tympanostomy tube (TT) insertion for recurrent otitis media (RecOM) or chronic otitis media with effusion (COME) were compared to control patients without a history of otitis media undergoing cochlear implantation. Methods During routine TT placement or cochlear implantation, a 1-mm biopsy of the ME epithelium was obtained. RNA was extracted, and real time RT-PCR was used to quantify levels of MUC5AC expression. Results Twenty-three OM patients (12 RecOM and 11 COME) were analyzed using 5 controls. Mean age was not different between groups. In RecOM samples, mean expression of MUC5AC was 25.92 times greater than controls. In COME samples, mean expression was 155.40 times greater than controls. Conclusions Levels of MUC5AC expression in the human ME are significantly increased in both RecOM and COME patients compared to controls. This study demonstrates MUC5AC gene changes in patients with OM and highlights the need for greater understanding of the molecular responses in OM; particularly that of mucin. A thorough exploration of these factors will provide opportunities to develop novel interventions for the extremely common problem of OM.

Objective A novel mouse model with a specific genetic mutation in a G protein coupled receptor (GPCR) encoded by the oxgr1 gene results in a predisposition to spontaneous otitis media with effusion. As a primary component of interest in OME, mucin expression was examined in this model to assess expression as compared to wild type animals and suitability as a murine model of OME. Method Mutant (oxgr1−/−) and wild-type (oxgr1+/+) mice between ages of 2–5 months were examined by otoscopy and auditory brainstem response (ABR). Histology changes in the middle ear were evaluated. Expression of mucin genes in the middle ear epithelium was determined using RT-PCR and quantitative PCR. Result Otoscopic exam showed signs of inflammation in 82% of mutant mice. Significant elevated ABR thresholds were detected in mutant mice indicating hearing loss. Histology analysis of the middle ears demonstrated the presence of inflammatory cells, changes in the mucosal epithelium, and middle ear fluid. RT PCR using universal primers for bacterial 18s rRNA suggested the absence of bacteria in the middle ear. The knockout mice demonstrated expression of muc1, muc2, muc3, muc4, muc5AC, muc5B, muc9, muc10, muc13, muc15, muc16, muc18, muc19 and muc20. There was a trend of increase in muc5B and muc19 expression in the middle ear of the knockout mice compared to that of wild-type. There was no significant change in the level of muc2, and muc5AC was expressed at a level below the detection limit of quantification. Conclusion Development of a murine model with genetic defect has several attractive features. The rate of OME in these animals is high at 82%. It is clear that this OME is related to histopathologic changes in the middle ear epithelium of these knock-out mice. Induction of mucus effusion is evident though the viation in dysregulation of GFM does exist in this non-challenge study condition. The underlying cause of these differences between individual animal requires further investigation. Given this, the oxgr1−/− model is likely to be an ideal model to examine mucin regulation in MEE and potentially develop novel GPCR-specific targeted interventions to regulate these processes.