**8. Conclusions**

The introduction of the modern molecular techniques and the subsequent investigations on microbial communities in the human organisms have changed our conceptions of health and disease and our approach to infectious conditions.

It is indeed well known that health and disease status are not merely determined by the presence or the absence of a pathogen but depend on a complex balance established among pathogens, resident microbiota, and host immune response.

Investigations previously described in this review have provided novel insights on the pathogenesis of middle ear diseases and led to the identification of both possible new causative agents and of potential protective bacteria, showing that imbalances in bacterial communities of the URT and ME could influence the natural history of otitis media in children.

However, scientific data on this topic are often di fficult to compare because of methodological di fferences in specimen collection and analysis, in the site of investigation, and in data reporting. Moreover, a lack of standard diagnostic criteria for OM across countries often influences the enrollment phase and contributes to increase the heterogeneity among populations under investigation.

Another element that complicates data interpretation and deserves standardization is the use of di fferent databases during OTUs assignment. This is a relevant issue that should be taken into consideration for two main reasons: di fferent databases might lead to heterogeneous results; some taxa could be misclassified with certain databases, as reported for *A. otitidis* and *T. otitidis* [89].

We believe that future investigation should be focused on the following aspects:


**Author Contributions:** P.M. and S.T. conceived the paper; F.F. conducted the literature search and wrote the manuscript; I.C. contributed to the literature search; P.M., S.T., L.D., P.C. and S.A. revised the paper; L.R. revised the technical aspects. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The authors declare no conflict of interest.
