Changes in the proportion of penicillin-non-susceptible Streptococcus pneumoniae (PNSP) isolates during an intervention programme were evaluated by phenotypic analysis of all initial isolates with penicillin MIC > or =0.5 microg/ml (n=1248) collected 1995-2004. During the study period, the proportion of such isolates was fairly constant (12-19%), and there was no statistically significant variation in the proportion of total PNSP cases (MIC > or =0.12 microg/ml) or PNSP with MIC > or =0.5 microg/ml, with the exception of an increase in 2004. Analysis restricted to clinical cases revealed no statistically significant changes. 23 different serogroups were found, and serogroup 9 isolates accounted for almost half of the PNSP cases. Only minor changes in phenotypic characteristics occurred in the other serogroups, which indicates that the increase in PNSP in 2004 was not due to import of a new resistant clone. Antibiotic consumption is considered to be an important risk factor for penicillin resistance in S. pneumoniae. After initiation of the intervention programme in Malmö, overall prescribing of antibiotics decreased 28%, and the reduction was even greater among children (52%). In conclusion, the proportion of PNSP isolates in Malmö has remained stable, despite the intervention programme and decreased consumption of antibiotics.