- After the introduction of the seven valent-pneumococcal conjugated vaccine into our National Immunization Program, it is important to establish and track local serotype distribution in order to evaluate its impact specially because serotype replacement phenomena has been described.To describe the clinical, epidemiological and antimicrobial resistance patterns of Costa Rican children with otitis media caused by Streptococcus pneumoniae serotype 3. Middle ear fluid samples were obtained from Costa Rican children with otitis media who participated in various antimicrobial clinical trials between 1992 and 2007. Streptococcus pneumoniae was identified according to laboratory standard procedures. Strains were serotyped and antimicrobial susceptibility to penicillin, amoxicillin, cefuroxime, ceftriaxone, azithromycin and levofloxacin was determined by E-test. Throughout 1992-2007 a total of 1919 tympanocentesis were performed in children with otitis media (median age: 19 months) and yielded a total of 1208 middle ear isolates. The most common pathogens were: Streptococcus pneumoniae, 511 isolates (49%); Non-Typable Haemophilus influenzae, 386 isolates (37%); Moraxella catarrahalis, 100 isolates (9.5%); and Streptococcus pyogenes, 54 isolates (5%). Streptococcus pneumoniae serotyping was performed in 346/511 isolates (68%) recovered during years 1999-2006. The most common serotypes were 19F (101/30.0%), 14 (46/13.7%), 3 (34/10.1%), 6B (30/8.9%) and 23F (23/6.8%). Analysis performed per years showed a higher prevalence of serotype 3 Streptococcus pneumoniae during the study period 2004 and 2005. During the entire study period (1999-2006) serotype 3 was most commonly isolated in children older than 24 months (61.2% vs 40.6%;P = 0.05) and showed a lower rate of penicillin non-susceptibility (4.0% vs 18%; P = 0.003). Streptococcus pneumoniae serotype 3 is an important pathogen in Costa Rican children with otitis media, especially in children older than 24 months of age (P = 0.05). Most serotype 3 isolates were susceptible to penicillin, cephalosporins, macrolides and quinolones.