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Life sciences · Journal article
Microbiology Spectrum · September 30, 2026
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ABSTRACT Acute otitis media (AOM) is one of the most prevalent infectious diseases in childhood, with Streptococcus pneumoniae and Haemophilus influenzae being the most commonly involved pathogens. Following the coronavirus disease 2019 (COVID-19) pandemic and the incorporation of 13-valent pneumococcal conjugate vaccine (PCV13) into the national immunization program, changes in pneumococcal invasive disease have been described, but data on AOM remain limited. This study aimed to evaluate the current bacterial etiology of pediatric AOM. Spontaneous otorrhea fluid (SOF) samples and optional nasopharyngeal samples (NP) were collected from children with AOM attending 13 primary care centers in southern Barcelona. Conventional culture and PCR were used for pathogen detection. Most frequent pathogens ( Streptococcus pyogenes, S. pneumoniae, and H. influenzae ) underwent antimicrobial susceptibility testing and whole-genome sequencing. A total of 145 SOF and 83 NP samples were obtained. Bacterial growth in SOF samples was detected by culture in 80.7% of the samples and by molecular methods in 91.7%. The predominant pathogens were H. influenzae (40.7%), S. pyogenes (35.2%), and S. pneumoniae (33.8%). Serotype 3 (mainly GPSC12-CC180) was the most frequent pneumococcal serotype (15.6%), and 23.8% of identified serotypes were included in PCV13 despite high vaccination coverage. Most of the S. pyogenes analyzed (43.1%) belonged to the emm1 -ST28 genotype, and 94.1% of H. influenzae isolates analyzed were non-typeable. These findings demonstrate a marked shift in AOM etiology in the post-PCV13 era, with H. influenzae emerging as the leading pathogen and an increase in S. pyogenes AOM. Continued surveillance is essential to inform future vaccination and prevention strategies. IMPORTANCE This multicenter study provides new insights into the current epidemiology of acute otitis media (AOM) in children vaccinated with the 13-valent pneumococcal conjugate vaccine. First, it underscores the role of molecular methods in pathogen detection. Second, it adds new evidence of the lack of concordance between nasopharyngeal and spontaneous otorrhea fluid samples, particularly for S. pneumoniae. Third, whole-genome analysis confirmed the predominance of the GSPC12-clade Ia as the major contributor to the vaccine failures due to serotype 3 pneumococci. Fourth, we characterized the S. pyogenes lineages associated with its recent upsurge in AOM, which differs from those causing invasive disease in our area. Finally, our findings confirm the current dominance of non-typeable H. influenzae as the leading cause of AOM in vaccinated children.