Three hundred thirty-six clinically significant Streptococcus pneumoniae isolates were collected from laboratories of different hospitals in Riyadh, Saudi Arabia. Most of these isolates were from pulmonary and otitis media (68.2%), and 31.8% were extrapulmonary (blood and CSF). Of the 336 isolates, 44.6% were susceptible to penicillin, and 55.4% were penicillin non-susceptible (35.7% were intermediate and 19.7% were fully resistant). The isolates showed 9.0% resistance to co-amoxiclav, 31.8% to cefuroxime and 39.4% to cefprozil. None of the isolates were resistant to ceftriaxone. Overall macrolide resistance rates were 22.6% to erythromycin, 18.5% to roxithromycin, 17.9% to azithromycin and 17.3% to clarithromycin. Most penicillin non-susceptible pneumococci were of serogroups/types 19 (21.0%), 6 (10.8%), 18 (8.6%), 23 (8.1%) and 14 (7.0%). Serogroups 9, 15, and 1 were found in 5.4%, 4.3%, and 2.2% of the isolates, respectively. Nontypeable strains constituted 6.5%. In exploring the mechanism of resistance to macrolides, 28 of 76 (36.8%) of isolates were erythromycin-resistant due to ribosomal mechanism (all were constitutive type, none were inducible), whereas 48 (63.2%) isolates were resistant due to an efflux mechanism. Good antibiotic control with periodical antibiotic surveillance and appropriate use of pneumococcal vaccine may improve current treatment of pneumococcal infections.