Identification of essential biofilm proteins in middle ear fluids of otitis media with effusion patients

Abstract

Objectives: Otitis media with effusion (OME) is a common disease of childhood that is largely asymptomatic. However, middle ear fluid can persist for months and negatively impact a child's quality of life. Many cases of OME remain chronic and require surgical intervention. Because biofilms are known to contribute to the persistence of many diseases, this study examined effusions collected from children with chronic OME for the presence of essential biofilm structural components, members of the DNABII family of bacterial DNA-binding proteins.

Methods: Middle ear effusions were recovered from 38 children with chronic OME at the time of tympanostomy tube insertion. A portion of each specimen was submitted for microbiology culture. The remaining material was assessed by immunoblot to quantitate individual DNABII proteins, integration host factor (IHF), and histone-like protein (HU).

Results: Sixty-five percent of effusions (24 of 37) were culture-positive for bacterial species or yeast, whereas 35% (13 of 37) were culture-negative. IHF was detected in 95% (36 of 38) at concentrations from 2 to 481 ng/μL effusion. HU was detected in 95% (36 of 38) and quantitated from 13 to 5,264 ng/μL effusion (P ≤ 0.05 compared to IHF).

Conclusion: Because DNABII proteins are essential structural components of bacterial biofilms, these data lend further support to our understanding that biofilms are present in the vast majority of chronic middle ear effusions, despite negative culture results. The presence and ubiquity of DNABII proteins in OME specimens indicated that these proteins can serve as an important clinical target for our novel DNABII-directed strategy to treat biofilm diseases such as chronic OME.

Level of evidence: NA Laryngoscope, 130:806-811, 2020.

Keywords: DNABII proteins; HU; IHF; chronic otitis media with effusion; pediatric otolaryngology.

Figure 1.. Quantitation of IHF and HU in effusions recovered from children with chronic OME.

Caption: Panel A, representative immunoblot to demonstrate varied amount of IHF detected in a panel of 16 OME specimens. Panel B, quantitation of IHF (blue circles) and HU (red circles). Among the 38 OME specimens collected, 95% were positive for both IHF and HU proteins by quantitative immunoblot analysis, and overall, a significantly greater amount of HU was detected, compared to IHF (P≤ 0.05). These data indicated that the immunoblot was highly sensitive in ability to both reveal the presence of DNABII proteins within the chronic OME samples, as well as quantitate them. *, P≤ 0.05

Figure 2.. Lack of correlation between culture status of each chronic OME specimen and quantity of IHF (Panel A) or HU (Panel B) as determined by immunoblot assay.

Caption: Of 13 samples that yielded negative culture results, all 13 were positive for the DNABII proteins IHF and HU by quantitative immunoblot assay, as depicted by the blue circles. Red squares indicate the concentration of IHF or HU detected by quantitative immunoblot in the 24 specimens that yielded positive culture results. White squares depict specimens wherein the quantitative immunoblot assay yielded zero IHF or HU, but positive culture results for bacteria or yeast were obtained. No significant difference in quantity of either IHF or HU was calculated between specimens that were culture-positive versus those that were culture-negative. In sum, DNABII proteins were shown to be present by objective immunoblot assay even in the 13 specimens that had been deemed culture-negative.