Background Otitis media with effusion (OME) is a commonest cause of hearing loss in childhood especially in developing countries. Objectives and methodology the study included 50 patients with OME. In all cases, myringotomy operation was done. It aimed to investigate the prevalence of aerobic and anaerobic bacteria in children with OME, to determine the presence of H. pylori in middle ear effusions (MEE) using polymerase chain reaction (PCR) and to examine the antibiotic susceptibility profile of isolated bacterial strains. Results Staphylococcus aureus (28%) was the most prevalent, followed by Peptostreptococcus species (19.2%), Coagulase negative staphylococci (17.5%), Klebsiella species (8.7%), E. coli (5.2%), and Pseudomonas aeruginosa (3.2%). H. pylori could be detected in (5.2%) using PCR. Regarding antibiotic sensitivity, most isolated strains were sensitive for ciprofloxacin and chloramphenicol while they were resistant to cephalothin except Peptostreptococcus species. Conclusion the obtained results indicate that H. pylori may play a role in the pathogenesis of OME.