Bacterial Profile, Molecular Serotyping, and Key Genetic Determinants for Adhesion, Immune Evasion, and Tissue Spread Among Bulgarian Children with Acute Otitis Media

<b>Background</b>: Acute otitis media (AOM) is one of the most common pediatric infections. We aimed to investigate the bacterial profile of AOM in children, the serotype distribution, and the main genetic virulence factors involved in adhesion, immune evasion, and tissue spread. <b>Methods</b>: In total, 121 AOM cases involving children aged 0 to 14 years were studied. Middle ear fluids (MEF) (n = 42) and nasopharyngeal samples (n = 79) were collected. All strains were identified using routine microbiological tests, conventional PCRs and real-time PCR methods. Molecular serotyping was performed for <i>S. pneumoniae</i> and <i>H. influenzae</i> isolates. An immunofluorescence serotyping technique was employed for <i>M. catarrhalis</i>. Target genetic factors were determined for all involved bacterial agents using singleplex or multiplex PCRs. <b>Results</b>: We analyzed 148 nasopharyngeal and MEF. Among 121 AOM cases, a total of 127 bacterial agents were identified, including <i>S. aureus</i> (n = 41), <i>S. pneumoniae</i> (n = 28), <i>H. influenzae</i> (n = 23), <i>M. catarrhalis</i> (n = 19), and <i>S. pyogenes</i> (n = 16). The leading three serotypes among <i>S. pneumoniae</i> were: 19A (18.0%), 6A (14.3%), and 15B (14.3%). 91.3% of <i>H. influenzae</i> isolates were non-typeable (lacking a capsule-NTHi). The <i>M. catarrhalis</i> isolates were distributed in serotypes A (57.9%), B (26.3%), and C (15.8%). Presence of pili type 1 was detected in 21.4% pneumococci, and the fimbrial gene <i>hifA</i> was found in 34.8% of the <i>H. influenzae</i> strains. In 73.6% of the <i>M. catarrhalis</i> strains, <i>ompCD</i> was identified, while 84.2% contained <i>ompE</i>. 62.5% of the <i>S. pyogenes</i> isolates harbored the <i>sdc</i> gene, and 56.2% possessed the <i>sdaD</i> gene, predominantly in the MEF isolates. The <i>cna</i> adhesin was found in 28.0% of the <i>S. aureus</i> strains. <b>Conclusions</b>: The monitoring of bacterial pathogens responsible for otitis media, along with their serotype distribution and the prevalence of genetic factors involved in disease pathogenesis, is essential for public health and can help predict disease severity and treatment options.