Background Complications of otitis media is dangerous in both adults and children, Otitis media with Effusion (OME) is common in children than adults, Isolation of Streptococcal pneumonia from middle ear with effusion is one of bacterial affecting middle ear, which is important for treatment and prognosis of the disease, Aim of the work is to investigate the effect of nested polymerase chain reaction (PCR) assay for detection of S.pneumoniae as a diagnostic element in patient with OME, and detect pathogenesis of the disease and its relation to the Organism, patients and methods: this study included 34 patents diagnosed as Otitis Media with Effusion (OME) 11 adult and 33 children, aspiration from middle ear fluids were done and subjected to cultivation OIJ selective media, and nested PCR test using specific primers directed to selected pneumolsin gene_ jragment of S.pneumoniae ,Results, S Pneumoniae detected by culture in 4/34 (11.87 %) of (OME) fluid samples; 3/4 (75%) from children, (25%) adult, S.pneumoniae in 11/34 (32.4%) of the samples detected by Nested PCR, 8/11 (72.7%) were children while 3/11 (27.3%) were adults. All all culture positive samples were PCR positive, but 7 cases of the PCR positive samples were culture negative, other factors affecting our results were age, sinusitis, previous otitis media, and allergic rhinitis, the rate of organism were (72.7%, 36.4%54.5 %,.and 27.3%) respectively, Conclusion: PCR is very sensitive tool for recognition of S. pneumoniae in fluids d