Fourteen pneumococcal strains isolated in three nationwide studies were characterized for amino acid changes in the enzymes GyrA, GyrB, ParC and ParE, and for the in vitro activity of eight fluoroquinolones and the new non-fluorinated quinolone BMS 284756. Gemifloxacin and BMS 284756 exhibited the best in vitro activity against all 14 isolates tested. In nine of the 14 isolates mainly classical alterations in ParC (D83N/Y, S79Y/F), as well as rarer alterations such as S80P and D78N, contributed to the decreased susceptibility to fluoroquinolones. In two of the 14 isolates the classical alteration in GyrA (S81F) was found. In only one isolate did alterations in ParC and GyrA exist in parallel.