Streptococcus pneumoniae is one of the primary pathogens that are associated with acute respiratory infections (ARI) that cause high rates of morbidity and mortality among children under five years of age in developed and developing countries. This study aimed to determine the prevalence of nasopharyngeal colonization, the antimicrobial resistance profile, and the capacity for biofilm formation by S. pneumoniae isolated from children aged 0-6 years with ARI throughout the Porto Velho-RO. A total of 660 swabs were collected from children with ARI. Molecular and biochemical tests were performed to characterize the isolates. The disk-difusion method and the E-test were used for antimicrobial sensitivity testing (TSA). Biofilm formation capacity was assessed using microtiter plate assays, and serotype detection was acheived using polymerase chain reaction (PCR) analyses. The colonization rate for S. pneumoniae was 8.9% (59/660) and exhibited a high prevalence in children under 23 months of age 64.4% (38/59). The observed serotypes were 9V and 19F with frequencies of 1.7% (1/59) and 13.6% (8/59), respectively. The antimicrobial susceptibility test revealed 100% (59/59) sensitivity to vancomycin. In contrast, trimethoprim and oxacillin exhibited high resistance rates of 76.3% (45/59) and 52.5% (31/59), respectively. Of the biofilm-forming isolates, 54.8% (23/42) possessed resistance to some antimicrobials. In this study, S. pneumoniae showed high rates of antimicrobial resistance and the ability to form biofilms, as these are factors that favor bacterial persistence and can cause serious damage to the host.