Between 1990 and 1995, a total of 300 children, ages 2 months to 5 years, received diagnoses of acute otitis media (AOM) in a hospital emergency room in São Paulo, Brazil, and were recruited for this study. The investigation was undertaken, first, to identify microorganisms and antimicrobial susceptibilities of pathogens from AOM in Brazilian children; next, to ascertain, by comparison, whether the isolates of Streptococcus pneumoniae have the same serotypes as those included in the new conjugated heptavalent pneumococcal vaccine; and last, to determine whether additional and/or different serotypes are needed in the vaccine to ensure an immunogenic response against pneumococcal pathogens for the indigenous children in this study. Microorganisms were isolated from ear fluid of 192 patients (64%). The 5 most prevalent pathogens were S pneumoniae (48 isolates; 16%), Haemophilus influenzae (21 isolates; 7%), Moraxella catarhalis (15 isolates; 5%), Pseudomonas aeruginosa (6 isolates; 2%), and Staphylococcus aureus (3 isolates; 1%). These 5 represented 93 of the 192 total isolates. Resistance to antibiotics was found in the 3 primary pathogens. No high-level resistance of S pneumoniae to penicillin was found; instead, there was high-level resistance to trimethoprim-sulfamethoxazole. Ten serotypes of S pneumoniae were isolated: 6B, 9V, 11A, 16, 18C, 19A, 19F, 23A, 23B, and 23F. Only 5 of the 10 serotypes isolated were included in the conjugated heptavalent pneumococcal vaccine. Therefore, the other 5 serotypes (24 of 48 strains) should be considered in selecting antigens for the new vaccine.